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Figure 10.8. Alignment of the amino acid sequences of seven S-proteins of Nicotiana alata. The five conserved (C1 to C5) and five hypervariable (HV1 to HV5) regions are enclosed in solid lines. Small boxes within the conserved regions are residues that are different from the conserved residues at those sites. Open circles enclose residues that are different between the two homology classes but are conserved within one or two homology classes. The nine cysteine residues are indicated by asterisks. The circled asparagine residues are potential N-glycosylation sites. (From Kheyr-Pour et al 1990. Sex. Plant Reprod. 3:88-97. © Springer-Verlag, Berlin.)
1989; Kheyr-Pour et al 1990)). The occurrence of conserved cysteine residues and glycosylation sites in the alleles suggests specific structural roles for these molecules in the stability, conformation, and biological activity of the glycoproteins. Is the SLG gene of N. alata a member of a multigene family? When Southern blots of endonuclease-digested genomic DNA from S-2, S-2, S-3, S-6, and S-7 allelic plants were probed with cDNAs corresponding to the alleles, the fragments hybridizing from different alleles were found to be dissimilar in size, and, in most cases, single diagnostic restriction fragments hybridized intensely with the probe. Each cDNA probe bound strongly to a DNA fragment of that allele and weakly or not at all to DNA fragments of other S-alleles. These
286
observations are compatible with the view that the isolated cDNA clones are allelic products of the same S-locus and that this is a single-copy genetic system. Additional data confirming that the cDNA sequences are allelic products of the same S-locus have come from breeding experiments combined with DNA blot analysis, two-dimensional electrophoresis, and cation exchange liquid chromatography of style glycoproteins. It was found that the S-genotypes of individual plants determined from conventional breeding tests correlate precisely with their DNA blot analysis using S-allele-specific DNA probes and with the presence of S-allele-specific glycoprotein spots and protein elution profiles (Bernatzky, Anderson and Clarke 1988; M. A. Anderson et al 1989; Jahnen et al 1989). Of further interest is a report that a small fragment located at the 5'-end of the coding sequence of S-2 DNA shows homology with mitochondrial DNA in Southern blots (Bernatzky, Mau and Clarke 1989). How the transfer of sequences between nuclear and mitochondrial DNA might have occurred, and what the role of this sequence is in the function and expression of self-incompatibility, remain to be explored. (i) Ribonucleases as S-glycoproteins A feature of the amino acid sequences of S-glycoproteins isolated from N. alata was their homology to extracellular fungal RNase. The homology itself was not particularly striking, as only 30 of the 122 amino acids conserved among S-2, S-3, and S-6 allelic glycoproteins were aligned with identical molecules in the fungal enzyme. Another 22 amino acids in the glycoproteins aligned with closely related amino acids in the RNase, and 5 out of the 9 or 10 cysteine residues and 2 catalytic histidine residues were conserved between the two proteins. Nonetheless, this finding led to intensive research on the RNase activity of glycoproteins and eventually to the identification of glycoproteins as RNases. That RNase activity is an inherent component of the S-glycoproteins became clear from the fact that the enzyme is found to co-elute perfectly with the individual S-glycoproteins by ion-exchange chromatography and that it has a specific activity close to that of purified RNase T2. In physiological terms, the specific activity of the enzyme from stylar extracts of the self-incompatible N. alata is found to be 100-1,000-fold higher than that of the self-compatible N. tabacum. This correlation implies a possible role for the stylar product of the self-incompatibility gene, designated as S-RNase, in self-pollen rejection reactions (McClure et al 1989). S-RNase also
Pollination and Fertilization
appears to have a bearing in pollen rejection associated with unilateral incompatibility in certain species combinations in Nicotiana. This was shown by up-regulating RNase expression in transgenic plants that acquire the capacity to reject normally compatible pollen grains (Murfett et al 1996). A general mechanism by which RNase might function in the self-incompatibility response is by the degrading of pollen RNA, which results in pollen tube death in an incompatible pollination. Support for this view has come from the work of McClure et al (1990), who followed the integrity of RNA extracted from the style of N. alata of a defined genotype after compatible and incompatible pollinations with 32P-labeled pollen grains. The results were straightforward in showing extensive RNA breakdown in incompatible styles but not in compatible ones (Figure 10.9). Furthermore, the results suggest that pollen tube rRNA could have suffered degradation in the incompatible style due to RNase cytotoxicity, because RNA appeared as a Style Fraction RNAs
Stigma Fraction RNAs
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Figure 10.9. Agarose gel electrophoresis of 32P-labeled RNA from compatible and incompatible crosses in Nicotiana alata. RNAs from style and stigma parts of the flower were treated separately. Compatible crosses represented are 5,53 X S2S2 and S2S2 X 5,S3; incompatible crosses represented are S2S2 X S2S2 and S^S^ X S.Sy Mostly intact RNAs are seen in the styles of compatible crosses and in the stigmas of all crosses, whereas RNAs of the styles in the incompatible crosses are degraded, which results in the accumulation of low-molecular-mass species. (From McClure et al 1990. Reprinted with permission from Nature 347:757-760. © Macmillan Magazines Ltd. Photograph supplied by Dr. A. E. Clarke.)
Molecular Biology of Self-Incompatibility
smear of low-molecular-mass species in an agarose gel run under denaturing conditions. Results from other experiments have been marshalled to show that the inhibition of pollen tube growth in the style results from the entry of S-RNase and the consequent degradation of pollen tube RNA. For example, following autoradiography of 3H-labeled S2-RNAse incorporation, the label is present in the pollen grain cytoplasm and, to a lesser extent, in the pollen tube. Immunogold electron microscopy using an antibody raised to a synthetic peptide corresponding to an allele-specific region of S2-RNase showed labeling in the walls of the germinated pollen grain and in the pollen tube cytoplasm. Treatment of pollen grains with S2-RNase inhibited pollen tube growth and, concomitantly, protein synthesis. S2-RNase also inhibited cell-free translation of proteins by self-pollen RNA to about 70% of the control rate (Gray et al 1991). In the absence of data on the comparative effects of S2-RNase on compatible pollen grains, the significance of these results leaves much to be desired. Despite this criticism, the results lead to the inference that RNA degradation is instituted soon after the incompatible pollen tubes reach the style, as a result of which their growth is stymied. Analyses of S-RNases recovered separately from the stigma and style have shown that the two enzymes are identical, although their respective transcripts differ in the length of the polyadenylate tail. This has raised the obvious question of why pollen tube growth is arrested in the style rather than in the stigma after self-pollination. Different mechanisms of enzyme uptake by the pollen tube from the stigma and stylar regions might underlie their differential sensitivity, but this remains to be further investigated (McClure et al 1993). In one working model proposed to explain the operation of the system, it is believed that in an incompatible pollination, pollen tubes carry S-locus-specific receptors that bind SRNases. How these molecules find their way into the pollen tube is a mystery, but once inside, they decimate the endogenous RNA of the pollen tubes and inhibit their growth. Compatible pollen tubes survive the RNase onslaught since they have some way of preventing the uptake of the enzyme. Alternatively, it has been postulated that RNases are taken up by pollen tubes regardless of their specificity but, inside the pollen tube, specificity of action prevails due to an intracellular inactivation of the enzyme or a compartmentalization of the enzyme that makes the RNA pool inaccessible (Franklin-Tong and Franklin 1993; Dodds, Clarke and Newbigin 1996).
287
Attractive though these models are, the mechanism by which allelic specificity is controlled during the incompatibility episode remains unclear. Since the gametophytic self-incompatibility system is based on the expression of the S-gene in the pollen as well as in the style, the problem is compounded by the fact that S-gene products of the pollen have not been identified. The dilemma has been heightened with the discovery that the SRNase gene is also expressed at a low level in the pollen grains of N. alata; it is difficult to visualize an incompatible reaction as a result of the interaction of two identical molecules (Dodds et al 1993). Based on the fine structural analysis of S-glycoproteins, N-linked carbohydrate chains are thought to be involved through posttranslational modification in the function of the S-alleles (Woodward et al 1992). This implies that the choice between the fates of S-alleles in the pollen grain and style is predetermined rather than stochastic. From observations showing that S-RNases are effective substrates for protein kinases of pollen tubes, a role for phosphorylation of S-RNase involving Ca2+-dependent protein kinases in the signal transduction pathway of self-incompatibility appears as a distinct possibility (Kunz et al 1996). (ii) Expression of 5-Locus Glycoprotein Genes
An important aspect of the cloning of S-locus glycoprotein genes from N. alata concerns their temporal and spatial appearance. At what period are the transcripts of the gene expressed relative to the stage of floral development at which overt selfincompatibility syndromes appear? Does the accumulation of transcripts coincide with the site of pollen tube inhibition? Fortunately, these questions have generated complementary sets of data that indicate that the pattern of gene expression reflects the age of the flower when it becomes self-incompatible and the age of the tissues where the reactions occur. From Northern blot analysis of stylar RNA from flower buds of different ages probed with S-2 cDNA, it would appear that gene expression is most intense in mature styles, weak in the ovary, and absent in immature styles. At the cellular level, the S-2 gene is expressed in the stigmatic papillae and stylar transmitting tissue of mature flowers as a pathfinder for the pollen tube before it succumbs, but the gene is not prevalent in the immature stigma-style complex before it becomes self-incompatible. The weak expression of the gene detected in the ovary in Northern blot is seen by in
288 situ hybridization as labeling in the epidermis of the placenta. The epidermal cells are probably engaged in arresting the growth of self-pollen tubes that escape inhibition in the style (Anderson et al 1986; Cornish et al 1987). Immunocytochemical studies have shown that S-glycoproteins accumulate in the same tissues in which the gene transcripts are found (M. A. Anderson et al 1989). In an approach using transgenic plants to localize the sites of S-gene expression, self-compatible N. tabacum plants were transformed with constructs encoding the S2-glycoproteins/ and gene expression was followed by protein gel blot and Northern blot analyses and by immunolocalization. The aspects of gene expression in transgenic tobacco plants that can be correlated with self-incompatibility in N. alata are the localization of S2-glycoproteins in the stylar transmitting tissues, an increase in the level of the glycoprotein with maturity of the style, and RNase activity of S2-glycoproteins. However, no change in the self-incompatibility phenotype of transformants was observed as they spontaneously self-pollinated (Murfett et al 1992). In another work, prompt rejection of self-pollen was seen when a high level of RNase expression was engineered in transgenic self-compatible hybrids between N. alata and N. langsdorfii. Conversely, hybrids of N.
Pollination and Fertilization
and P. inflata. In studies of P. hybrida, cDNA libraries made to mRNA of mature styles of different S-alleles were screened with an oligonucleotide homologous to the conserved N-terminal region of N. alata glycoprotein to generate style-specific clones. These clones reacted specifically with RNA from the style, where the levels of mRNA homologous to the cloned cDNAs increased dramatically with the transition of the flower from self-compatibility to self-incompatibility. The clones were of low copy number and, consistent with their allelic nature, hybridized to polymorphic DNA restriction fragments (Clark et al 1990). A cDNA clone encoding S-locus glycoprotein in a S-2 homozygote of P. inflata was selected from a cDNA library using N. alata cDNA as a probe; subsequently, cDNA clones corresponding to the glycoproteins of other S-locus alleles of P. inflata were obtained by probing with S2 cDNA clones (Ai et al 1990). Antisense technology has been successfully used to demonstrate a correlation between inhibition of S-protein synthesis in the style and enforced breakdown of self-incompatibility in transgenic P. inflata (Lee, Huang and Kao 1994).
Combining the predicted amino acid sequences of SLGs of P. hybrida and P. inflata allows the following conclusions to be drawn about their plumbaginifolia X N. alata transformed with anti- sequence conservation and variability. Like N. alata sense construct targeted to the S2-RNase gene of N. glycoproteins, Petunia proteins are about 220 amino alata had low RNase activity and were compatible acids in length and include a 20-amino acid signal with N. alata pollen. These ingenious crosses were peptide characteristic of secreted proteins. The necessary to overcome the problems of the low sequences reveal a general pattern of several highly expression of S-RNase genes in transgenic N. alata conserved domains, interspersed by regions of and the poor regeneration of this species in tissue sequence variability. Among the individual culture (Murfett et al 1994, 1995; Murfett, Bourque residues that are highly conserved are eight cysand McClure 1995). Such problems will no doubt teine clusters. Although SLGs of N. alata have sevbe examined critically as more S-RNase expressions eral N-glycosylation sites, S-glycoproteins of are monitored in transgenic plants with the insight Petunia have only one or, at best, two conserved gained from the knowledge of the structure of residues that constitute potential N-glycosylation RNase genes and their promoter sequences sites. The importance of RNase to the self-incom(Matton et al 1995). For the present, these experi- patibility mechanism in Petunia is underscored by ments come close to providing a direct demonstra- the presence of conserved domains in the S-glycotion that the S-RNase gene is sufficient for proteins with strong homology to active site self-pollen rejection. regions of fungal RNases (Broothaerts et al 1989, 1991; Ai et al 1990; Clark et al 1990; Ioerger et al 1991; Lee, Singh and Kao 1992). Additionally, SLGs 3. PETUNIA AND OTHER SOLANACEAE from self-incompatible genotypes of P. hybrida and Petunia enjoys the distinction as one of the first P. inflata have been purified and shown to be genera to be used in self-incompatibility research, RNases, as are the SLGs of N. alata (Broothaerts et yet our knowledge of the molecular biology of self- al 1991; Singh, Ai and Kao 1991). A further point of incompatibility in this genus lags behind that of interest is that the carbohydrate moiety of the SBrassica and Nicotiana. cDNA clones encoding S- RNase is not required for the self-incompatibility allele-associated proteins were isolated and char- reaction in P. inflata, suggesting that the S-allele acterized almost simultaneously from P. hybrida specificity of the protein is determined by its amino
Molecular Biology of Self-Incompatibility
acid sequence and not by the glycan chain (Karunanandaa, Huang and Kao 1994). Direct proof implicating RNase in self-incompatibility in P. inflata has come from an experiment in which a mutant s-gene that lacked RNase activity due to a site-directed mutagenesis of one of the catalytic histidines was introduced into transgenic P. inflata and failed to reject self-pollen bearing the same Sallele as the transgene (Huang et al 1994). However, the extent to which stylar RNase activity in the self-incompatible lines of Petunia inhibits self-pollen tube growth is at present uncertain, and this gap in our knowledge has thwarted attempts to formulate a unitary concept of S-gene action in the gametophytic systems. The uncertainty stems from an examination of the levels of Sproteins and RNases in pseudo-self-compatible and self-compatible lines of P. hybrida. When RNase activity of these lines is compared, equal levels of activity are found in both (Clark et al 1990). Similarly, the S-proteins of self-compatible lines of P. hybrida display the same spatial and temporal pattern of expression as the SLGs of self-incompatible P. inflata lines. Proteins of both lines also have comparable levels of RNase activity (Ai, Kron and Kao 1991). This evidence begs the question of how S-RNase distinguishes between self- and nonselfpollen and how the recognition reaction leads to growth inhibition of the pollen tube. Conflicting data exist about the expression of SRNase transcripts in pollen grains of Petunia. Microprojectile bombardment showed that chimeric S-RNase-GUS gene accumulates to low levels in developing anthers of P. hybrida prior to anthesis (Clark and Sims 1994). However, the selfincompatibility trait of pollen grains of transgenic P. inflata is unaffected in transgenic plants whose phenotype is transformed from self-incompatibility to self-fertility (Lee, Huang and Kao 1994). Some insight into the structure of the S-gene of P. inflata is provided by an analysis of genomic clones of two S-locus alleles. The structure associates both alleles with a single, short intron that is embedded in the region identified with high interallelic variability in SLGs and that shows small variations in size and position between alleles. The clones of both alleles share the same TATA-box structure and have short, untranslated leader sequences. In both, there is a lack of interallelic homology in the flanks of the coding regions, which themselves contain repeated sequences. Despite these and other general similarities showing that the clones are alleles of the same S-gene, the clones appear to be basically heterogeneous,
289
except for conserved sites within the coding regions (Coleman and Kao 1992). The sequence divergence in the flanking regions might reflect an inevitable effect of an independent evolution of the two alleles. Attempts to clone self-incompatibility genes from other solanaceous genera have used oligonucleotide probes or heterologous probes to screen cDNA libraries. Since conserved regions of SLGs thus far characterized display some homology, it might be reasonable to expect heterologous probes to identify sequences similar to the primary structural features of S-locus proteins from cDNA libraries of related genera. In one round of experiments, cDNA or genomic clones encoding S-locus proteins were isolated from Solanum chacoense (Xu et al 1990b; Despres et al 1994; Saba-El-Leil et al 1994), S. tuberosum (Kaufmann, Salamini and Thompson 1991), and Lycopersicon peruvianum (Tsai et al 1992; Rivers et al 1993; Chung et al 1994). A rare, variant self-compatible line of L. peruvianum was used to illustrate the essentiality of RNases in preserving self-incompatibility in this species. Crosses between the variant line and a self-incompatible line showed that the mutation to self-compatibility is located at the S-locus and is associated with the absence of stylar RNase activity. The derived amino acid sequence for the glycoprotein from the compatible line was found to differ from that of the incompatible line by the loss of a histidine residue; as histidines are essential for the catalytic activity of the enzyme, this leads to the production of a protein that lacks RNase activity (Kowyama et al 1994; Royo et al 1994). Comparison of the deduced amino acid sequences of S-locus proteins of S. chacoense, S. tuberosum, and L. peruvianum with SLGs of Nicotiana alata, Petunia
hybrida, and P. inflata has shown a set of five conserved regions, including domains similar to those in the active site of fungal RNases. Equally significant is the presence of eight conserved cysteine residues in all S-locus proteins. Extreme sequence diversity between alleles of the same, as well as of different species, is also a fundamental property of the proteins characterized from the four genera of Solanaceae (Tsai et al 1992). Comparison of the amino acid sequences of S-locus proteins of 11 alleles from N. alata, P. inflata, and S. chacoense has
revealed the interesting fact that sequence similarity is relatively low within species, whereas some interspecific homologies are higher than intraspecific homologies (Ioerger, Clark and Kao 1990). That extreme sequence diversity can be pervasive between populations of even a single species is
290 apparent from a study in which the S-allele from L. peruvianum was used to probe DNA from several natural populations of the species (Rivers et al 1993). These and other observations have been put together in the form of a hypothesis about the phylogenetic relationships of the S-alleles that states that S-allele polymorphism has a common ancestry and that it arose prior to speciation in the four genera of Solanaceae (Ioerger, Clark and Kao 1990; Rivers et al 1993). In common with the structure of the S-gene of P. inflata, genomic clones corresponding to two S-alleles of S. tuberosum have a single intron and a short, untranslated leader sequence (Kaufmann, Salamini and Thompson 1991). From the data available, no major differences are discernible between the gene structures of the alleles. Expression of an S-RNase gene from S. tuberosum under the control of the tomato pollen-specific LAT-52 gene is developmentally regulated in the pollen grains of transgenic tobacco, but that alone does not appear sufficient to influence the compatibility phenotype of the transformants (Kirch et al 1995).
Pollination and Fertilization
In seeking a common denominator with other gametophytic self-incompatibility systems, it was found that the focus on RNase as a candidate for stigmatic S-locus protein in P. rhoeas was misplaced. This summation is based on several lines of evidence, such as the presence of negligible amounts of RNase in the stigma of P. rhoeas, the lack of correlation of enzyme activity with the development of incompatibility, the lack of RNase activity in purified S-locus protein, and the insensitivity of pollen grains to RNase in in vitro bioassays (Franklin-Tong et al 1991). On the other hand, the rapidity of the self-incompatibility reaction in P. rhoeas has led to the view that this is mediated by cytosolic Ca2+ acting as a second messenger. In support of this view, using laser scanning confocal microscopy and fluorescence ratio imaging of pollen tubes stained with the fluorescent dye Calcium Green-1 to localize Ca2+ fluxes, researchers have visualized transient increases in cytosolic Ca2+ levels in growing pollen tubes after the addition of partially purified, incompatible stigmatic protein extract or the recombinant S-protein. Evidently, changes in Ca2+ levels are farreaching, for the growth of pollen tubes that generate a rise in Ca2+ when challenged with the stigmatic extract or S-protein is inhibited (Figure 10.10). There 4. PAPAVER RHOEAS A N D OTHER PLANTS is no rise in the Ca2+ levels of pollen tubes after Molecular biology of self-incompatibility in addition of either compatible or heat-denatured, Papaver rhoeas may provide a testing ground for incompatible stigmatic extract. Even when the level concepts applied to gametophytic and sporophytic of endogenous Ca2+ is raised artificially by UV actisystems of self-incompatibility. Although the selfvation of caged Ca2+, an inhibition of pollen tube incompatibility system in this species is of the growth is noted in the absence of incompatible stiggametophytic type, inhibition of self-pollen occurs matic extract (Franklin-Tong et al 1993; Franklinin the stigma, as in the sporophytic type. What Tong, Ride and Franklin 1995). Conceivably, the makes this system interesting is that it is possible to transcriptional activation of genes necessary for the reproduce the self-incompatibility reaction in vitro initiation of self-incompatibility reactions in P. and that identification of the putative S-gene prod- rhoeas pollen might be the consequence of a rise in ucts has been made through their biological activ- Ca2+ level that is induced by incompatible S-locus ity of the inhibition of pollen tube growth proteins and is accompanied by an increased phos(Franklin-Tong, Lawrence and Franklin 1988; phorylation of proteins, but we have a long way to Franklin-Tong et al 1989). Transcript expression of go before the signal transduction pathway and the the S-gene cloned from P. rhoeas correlates well spa- mode of action of gene products are defined tially and temporally with the appearance of (Franklin-Tong, Lawrence and Franklin 1990; Rudd S-protein in the stigma and the onset of self-incom- et al 1996). patibility. The functional significance of this gene is underscored by the fact that the recombinant proBrassicaceae, Solanaceae and Papaveraceae thus tein isolated from Escherichia coli expressing the represent the three families in which S-genes and gene inhibits pollen tube growth in culture, as does their products have been intensively studied at the the stigmatic extract (Foote et al 1994). Finally, molecular level; very little information is available some preliminary evidence has been obtained for on other plants. A beginning has been made toward the presence in mature P. rhoeas pollen grains of a the molecular analysis of self-incompatibility in plasma membrane-associated glycoprotein that members of Rosaceae by characterizing cDNA binds to the stigma S-proteins in a nonspecific clones representing S-locus alleles of apple and manner (Hearn, Franklin and Ride 1996). As is evi- Pyrus serotina. As is the case in the S-locus genes of dent by now, these observations hold the key to the other plants, specificity of expression of transcripts basic reactions in self-incompatibility in all plants. of apple cDNA clones in the stylar tissues is inher-
Molecular Biology of Self-Incompatibility
291
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Figure 10.10. Changes in Ca2+ and growth of pollen tubes of Papaver rhoeas microinjected with Calcium Green-1. The filled triangles represent fluorescence (pixel values) from the first 100 urn of the pollen tube; the open squares indicate the relative length of the pollen tube at the corresponding time point, (a) Addition of compatible stigmatic S-glycoproteins (first arrow) followed by addition of incompatible stigmatic S-glycoproteins (second arrow). No
increase in Ca2+ and no significant inhibition of pollen tube growth are observed after treatment with the compatible protein; after addition of the incompatible protein, there is a transient rise in Ca2+, indicated by the two peaks, and a cessation of pollen tube growth around 2,000 seconds, (b) Photoactivation of caged Ca2+ (arrow) and cessation of pollen tube growth almost immediately. (From Franklin-Tong et al 1993.)
ent to their function, and, similar to S-proteins of members of Solanaceae, purified S-proteins from apple and P. serotina correspond to RNases (Broothaerts et al 1995; Sassa et al 1996). It appears that RNases are likely to mediate the self-incompatibility syndrome in certain lines of Antirrhinum majus also (Xue et al 1996). So, if a single regulatory molecule is required to activate the stylar self-pollen rejection mechanism, it would probably be the same molecule in members of Solanaceae, Rosaceae, and Scrophulariaceae. There is preliminary evidence showing that SLG and SRK-like genes modulate the sporophytic type of self-incompatibility in Ipomoea trifida (Kowyama et al 1995). What appears to be the pollen component of the self-incompatibility allele has been cloned from the
grass Phalaris coerulescens (X. Li et al 1994). Unlike the other gametophytic systems studied, selfincompatibility in P. coerulescens is under the control of two unlinked loci, S and Z, and self-pollen rejection is the rule when both S and Z alleles are present in the pollen as well as in the style. A cDNA clone identified by differential screening was found to be expressed only in mature pollen grains and was absent from all other tissues tested, including the stigma. The deduced amino acid sequence showed that the protein has a variable N-terminus that has no homology to known proteins and a conserved Cterminus with a thioredoxin domain. The absence of a signal sequence and the presence of thioredoxin sequences probably indicate that, unlike other Slocus proteins, this is a cytoplasmic protein. How
292 this protein controls both recognition specificity • and inhibition of pollen tube growth in the selfincompatibility reaction remains to be elucidated. Considering the self-incompatibility systems for which good molecular data are available, it is clear the pollen self-rejection system evolved independently several times in the evolution of angiosperms. Millions of years have probably separated the different incompatibility systems. The molecular data from Brassicaceae, Solanaceae, and Papaveraceae show that there are some obvious, basic similarities, as well as differences, in the operation of signaling systems in the three families. A viewpoint that has gained momentum in recent times is that cellular responses unleashed during sporophytic self-incompatibility may share some components of host-pathogen responses in plants and that self-incompatibility is evolutionarily related to the system that conditions resistance to invading pathogens. Besides superficial similarities between the genetics, physiology, and biochemistry of the self-incompatibility and host-pathogen responses, the possible involvement of a receptor kinase in both strengthens the view that there is a connection between the two responses (Hodgkin, Lyon and Dickinson 1988; Lamb 1994). 5. GENERAL COMMENTS
Molecular analysis shows that specific characteristics of the self-incompatibility genes are varied in the four systems in which they have been studied
Pollination and Fertilization
intensively, and this makes it difficult to interpret the mechanism of action of the encoded proteins in the rejection of self-pollen. In a general sense, we know that in Brassica the structure of the proteins suggests a role in the signal transduction mechanism, whereas in Nicotiana and Petunia the protein that shows similarity to RNases functions by disabling incompatible pollen tubes. Particularly intriguing is that in Papaver the protein elicits a cellular response that results in a transient increase in Ca2+ concentration. Dramatic progress is currently being made in understanding the way the different proteins work in the model incompatibility systems. Identification of the recognition sequences expressed by the pollen grains assumes an obvious significance in the expected flow of information. At this stage, we do not know whether the recognition sequences expressed by pollen grains are different from those expressed by the female part of the flower. In analyzing the conditions that lead to the incompatibility reactions, delineation of the parts of the incompatibility molecules important for recognition begs resolution. Equally significant is the role of other parts of the incompatibility protein, perhaps in the initiation of events that subsequently result in pollen tube collapse. It is encouraging to note that more than one model system of self-incompatibility has been investigated to a level insightful enough that realistic progress in our understanding of the molecular basis of the recognition reaction and its consequences can be expected in the foreseeable future.
Chapter 11
Fertilization: The Beginning of Spomphytic Growth Cellular Nature of the Sperm (i) The Sperm Cytoskeleton (ii) The Male Germ Unit (iii) Isolation and Characterization of the Sperm Odyssey of the Sperm and Double Fertilization (i) Growth of the Pollen Tube through the Style (ii) Passage of Sperm into the Embryo Sac (iii) Fusion of Nuclei (iv) Cytoplasmic Inheritance In Vitro Approaches to the Study of Fertilization (i) Intraovarian Pollination (ii) Test-Tube Fertilization (iii) In Vitro Fertilization (iv) Gynogenesis 4. General Comments
In the life cycle of plants, fertilization is invariably associated with sexual reproduction and therefore represents the genetic switch that initiates the diploid, or the sporophytic, phase of development. The history of research into fertilization in angiosperms may be said to have begun with the discovery of syngamy, or the actual fusion of the male and female gametes, by Strasburger (1884) and of double fertilization by Nawaschin (1898) and Guignard (1899). The latter two investigators called attention to the streamlining of the male gametophyte's economy by providing light microscopic evidence for the fusion of one sperm with the egg cell to form the diploid zygote and of the other sperm with the polar fusion nucleus of the central cell to generate the triploid endosperm. In spite of an abundance of studies on the development of the male and female gametophytes and on
postfertilization events in angiosperms in the years following these discoveries, relatively few investigators have ventured into the area of fertilization research. This has led to the unenviable situation that, as a biological process, fertilization in angiosperms has remained as an unknown black box. The paucity of information on fertilization is undoubtedly related to the fact that events leading to the egg-sperm encounter take place in the privileged interior of the multicellular ovule, where, hidden from the eye, they are not prone to experimental assault and to the technical difficulties that have so far prevented the development of an ex ovulo in vitro fertilization system. However, during the past three decades, there has been considerable progress in our understanding of many facets of fertilization, particularly the ultrastructure of the male and female gametes and the sequence of events leading to gametic fusion. Extensive use of the electron microscope, experiments involving immunological and molecular approaches, and nondestructive isolation techniques have all contributed to this progress. Chapter 6 described in some detail the structure of the egg at the time of fertilization, and so this material will not be considered here again except as it relates to the male gametophyte preparatory to gametic fusion. The primary focus of the present chapter will be on the structure of the sperm cells, their fate in the pollen tube, and their eventual fusion with the cells of the embryo sac. The final section of this chapter is devoted to a review of the work done on the manipulation of the fertilization process under controlled conditions. Until recently, the subject of fertilization has been reviewed almost always within the larger context of male and female gametogenesis (Linskens 1969; Jensen 293
294 1973; Kapil and Bhatnagar 1975; van Went and Willemse 1984), but more focused reviews (Knox and Singh 1987; Knox, Southworth and Singh 1988; Russell, Cresti and Dumas 1990; Russell 1992) are beginning to appear. These should serve as excellent sources of information on various aspects of the biology of fertilization.
Pollination and Fertilization
appears as densely staining granular chromatin (Knox and Singh 1987; Knox, Southworth and Singh 1988). The nucleus is enclosed in a nuclear envelope in which the usual pores appear to be lacking or are few in number (Mogensen and Wagner 1987; Southworth, Platt-Aloia and Thomson 1988; Taylor et al 1989; Southworth 1990). Not much should be read into this last observation except that the nucleus of the mature sperm is tran1. CELLULAR NATURE OF THE SPERM scriptionally inactive and negotiates a limited trafOne point of special concern in the early studies fic with the surrounding cytoplasm. on the structure of the sperm was whether the sperm It is within the framework of the subcellular is a true cell with all the trappings or just an structure that factors responsible for the differentiorphaned nucleus. Although some light microscopic ation of cells reside; however, in all cases examined, investigations on this question remained inconclu- the cytoplasm of the male gamete appears truly sive, the cellular nature of the sperm was clearly undistinguished and does not give any hints about established by the early 1940s. Subsequently, elec- the specialized character of the cell (Figure 11.1). In tron microscopy, particularly the pioneering investi- classical descriptions, limited amounts of ER, ribogations of Jensen and Fisher (1968b) on bicellular somes, polysomes, lipid and protein bodies, mitopollen grains of Gossypium hirsutum, and of Hoefert chondria, dictyosomes, microtubules, and vesicles (1969b) and Cass (1973) on tricellular pollen grains of appear to make up the bulk of the sperm cytoBeta vulgaris and Hordeum vulgare, respectively, pro- plasm, with the mitochondrion varying with vided the first direct evidence that the male gamete respect to its morphology in certain species in angiosperms is not a naked nucleus but is a rela- (Dumas, Knox and Gaude 1985; McConchie, tively simple cell, with its own cytoplasmic sheath Hough and Knox 1987; Charzynska, Ciampolini containing a reduced complement of organelles. and Cresti 1988; Hu and Yu 1988; Southworth and Exhibiting great simplicity, the sperm in G. hirsutum is Knox 1989; Taylor et al 1989; Yu, Hu and Zhu 1989; reduced to little more than a bag for a prominent Cresti, Murgia and Theunis 1990; Theunis 1990; P. nucleus bathed in a thin layer of cytoplasm contain- E. Taylor et al 1991; Theunis, Cresti and Milanesi ing dictyosomes, ER, polysomes, vesicles, and 1991; van der Maas et al 1994). Mitochondria are organelles resembling mitochondria (Jensen and the most prominent organelles of the sperm of Zea Fisher 1968b). Besides these cytoplasmic organelles, mays and, as seen in three-dimensional reconstrucsperm cells of B. vulgaris and H. vulgare also harbor tion, they form a filamentous complex, each indifiles of microtubules along their length, suggesting vidual filament partially wrapping around itself that the cytoskeletal elements might play a role in (McConchie, Hough and Knox 1987; Mogensen, maintaining the shape of the sperm or in affecting Wagner and Dumas 1990). At the other extreme are sperm motility (Hoefert 1969b; Cass 1973). the conventional, small, ellipsoidal mitochondria found in the sperm cells of Brassica campestris and Further information about the more intimate B. oleracea (McConchie et al 1987). An important details of the structure of sperm cells has come question raised in these studies relates to the inherfrom electron microscopy combined with threeitance of plastids - whether plastids are paternally dimensional reconstruction, freeze-fracture analyinherited or not - and hence to their presence in the sis, confocal laser scanning fluorescent microscopy, cytoplasm of the sperm cells. The answer to this and immunochemistry. From these investigations question appears equivocal, because sperm cells of emerges the following summary picture. Sperm some plants, like Plumbago zeylanica (Russell and cells generally extend in length to about 35 fim and 1981; Russell 1983), Rhododendron laetum, and Cass 3 are approximately 1-3 fjm in width and 7-133 ^m R. macgregoriae (Taylor et al 1989), come equipped in volume. The characteristic shape of sperm cells, with plastids in their cytoplasm, whereas those of as seen in mature pollen grains or in pollen tubes, Spinacia oleracea (Wilms, Leferink-ten Klooster and is elongate or spindle shaped. The most conspicuvan Aelst 1986), Z. mays (McConchie, Hough and ous organelle of the sperm is the nucleus, which Knox 1987), B. campestris, B. oleracea (McConchie occupies an enlarged region at one end of the cell; et al 1987), B. napus (Murgia et al 1991b), and one or more tapering cytoplasmic extensions are Nicotiana tabacum (Yu, Hu and Zhu 1989) lack found at the other end. The nucleus of the mature plastids. sperm contains paternal genetic material, which
Fertilization: The Beginning of Sporophytic Growth
295 (i) The Sperm Cytoskeleton
Analysis of the cytoskeletal machinery of the sperm has followed a two-tier course, analogous to that employed in the study of the cytoskeleton of the vegetative and generative cells (Chapter 3). Early studies were conducted at the ultrastructural level, whereas later studies enlisted immunofluorescence methods. These investigations have provided a surprisingly uniform picture of the organization of sperm microtubules, which attain conformations compatible with the function of the sperm cells. Commonly, microtubules, singly or in bundles, are aligned longitudinally or helically in relation to the axis of the sperm (Hoefert 1969b; Cass 1973; Karas and Cass 1976; Zhu et al 1980; Dumas, Knox and Gaude 1985; Pierson, Derksen and Traas 1986; Charzynska, Ciampolini and Cresti 1988; Taylor et al 1989; Cresti, Murgia and Theunis 1990; Del Casino et al 1992; Palevitz and Liu 1992). A striking feature of the sperm of Tradescantia virginiana is the presence of numerous interconnected helical to longitudinal branches of microtubules that appear to arise from the main microtubule bundle and invest the entire cell (Palevitz and Cresti 1988). Whether the microtubule elements of the sperm are inherited from the interphase cytoskeletal network of the generative cell at the time of division or are assembled anew from stable precursors at the end of division is not clear. According to Palevitz and Cresti (1989), the entire array of microtubules of the sperm of T. virginiana is inherited from those seen in the late anaphase of the generative cell mitosis; these microtubule bundles are derived from the kinetochore fibers, which remain enmeshed with each other and with the surrounding microtubules during anaphase. Consistent with this view is the observation that the precise, stepwise aggregation of the kinetochore fibers at the poles of the generative cell is also reflected in the cytoskeleton of the young sperm (Palevitz 1990). However, circumstantial evidence such as the presence of fewer and lessprominent microtubule bundles in sperm cells compared to their progenitor cell, as seen in Rhododendron laetum and R. macgregoriae (Taylor et al
Figure 11.1. Structure of the sperm cell in the pollen tube of Populus deltoides. Two elongate sperm cells seen in the longitudinal section of the pollen tube are connected by narrow cytoplasmic projections. PT, pollen tube; SC, sperm cell. Scale bar = 1 urn. (From Rougier et al 1991; photograph supplied by Dr. C. Dumas.)
1989), argues against the inheritance of microtubules from the previous generation. On the whole, the situation presents an invitation for more experimentation about the origin of microtubules in the sperm cells of angiosperms. In ultrastructural investigations of sperm cells, not much attention has been paid to the presence of actin-containing microfilaments, and, if microfila-
296 ments are present, they have clearly remained elusive. Experiments using rhodamine-phalloidin as a fluorescent detection probe have also failed to reveal the presence of microfilaments in sperm cells
Pollination and Fertilization
Plumbago zeylanica (Russell and Cass 1981), and barley (Mogensen and Wagner 1987). The contacts and physical associations of the two sperm cells born out of the division of a generative of Lilium longiflorum, Nicotiana tabacum, Petunia cell were occasionally commented upon in some of hybrida, and Tradescantia virginiana (Pierson, the early light and electron microscopic studies. Are Derksen and Traas 1986; Palevitz and Cresti 1988, the two sperm cells closely linked together as a pair, 1989). As indicated in Chapter 3, ultrastructural or are they aligned with the vegetative cell nucleus, and immunofluorescence studies have failed to or do they float aimlessly in the pollen grain or in detect microfilaments in the generative cell; in this the pollen tube cytoplasm? Although the occurrespect, sperm cells appear to share a unique prop- rence of paired sperm cells and their closeness to erty with their progenitor cell. The only reports of the vegetative cell nucleus in the pollen tube of cotthe presence of microfilaments are in sperm cells of ton was noted by Jensen and Fisher (1968b, 1970), it wheat (Zhu et al 1980) and Plumbago zeylanica was the work of Russell and Cass (1981) that intro(Russell and Cass 1981), but these observations duced a new level of understanding of the charachave not been corroborated by other researchers. teristics of the sperm cell association and the One study using antiactin immunofluorescence as sperm-vegetative cell package. This work showed a specific probe has reported an impressive local- that in the tricellular pollen grain of P. zeylanica, the ization of actin in the generative cells and sperm two sperm are directly linked to each other by a cells of Rhododendron laetum and R. macgregoriae common lateral wall transgressed by plasmodes(Taylor et al 1989), but a subsequent work using the mata. It was also found that while both sperm are same and other materials and the same antiactin enclosed in the inner plasma membrane of the vegprobe failed to substantiate this observation etative cell, one of them is interlocked with the (Palevitz and Liu 1992). On the weight of the evi- nucleus of the vegetative cell in a complex way. The dence, the latter investigators have concluded that conformational relationship of the sperm cell to the vegetative cell nucleus is traced to a cytoplasmic microfilaments do not form part of the cytoskeleton extension that wraps around the periphery of the of the sperm cells of angiosperms. vegetative cell nucleus and is partially enclosed by convolutions of this nucleus (Figure 11.2). The sperm cell remains associated with the nucleus of (ii) The Male Germ Unit the vegetative cell during the period of active The electron microscopic image of the outer growth of the pollen tube, but the connection is sevboundary of the male gametes in the pollen grain is ered following the discharge of the pollen tube conthat of protoplasm covered by a plasma membrane, tents. Although it has been suggested that the with two sperm cells enclosed together in the inner sperm-vegetative cell nucleus assembly might facilplasma membrane of the vegetative cell. The mem- itate gamete delivery, its importance is not fully branes may be tightly appressed or may form a understood because the association of the naked loose association with intramembrane pockets that nucleus and the adhering cells cannot be easily enclose vesicles and osmiophilic droplets (Zhu et al manipulated or examined in situ. 1980; Dumas, Knox and Gaude 1985; McConchie, Hough and Knox 1987; McConchie et al 1987; An exciting concept demonstrated in a later Southworth and Knox 1989). The paired nature of work (Russell 1984) is that the two sperm cells in the outer membrane system of the sperm has been the generative cell, rather than being isomorphic, confirmed by visualizing cells by freeze-fracture, in display varying degrees of dimorphism in size and which the frozen membranes are cleaved along organelle content. This conclusion was made possitheir hydrophobic interior and the resulting surface ble by an examination of serial, ultrathin sections of is observed in the electron microscope; this sperm cells, by computer-aided three-dimensional approach did not reveal any connections or bridges reconstruction of their structure, and by the actual between the membranes (Southworth, Platt-Aloia counting of organelles in sections. The dimorphism and Thomson 1988; Southworth 1990; van Aelst, in shape is due to a membranous extension from Theunis and van Went 1990). This observation the one sperm of the pair that superficially undoubtedly relegates the male gamete to the sta- embraces the nucleus of the vegetative cell. This tus of a natural protoplast; however, a rudimentary sperm appears to be the larger of the two; the wall has been reported to be present in the sperm smaller one, which lacks a cellular projection, is of wheat (Zhu et al 1980; Hu, Zhu and Xu 1981), unassociated with the vegetative cell nucleus.
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Figure 11.2. Reconstruction of the sperm cells and the male germ unit of Plumbago zeylanica. (a) The two sperm cells; the sperm cell associated with the vegetative cell nucleus is Svn; the other sperm cell, not associated with the vegetative cell nucleus, is Sua. (b) The physical association between Svn and the nucleus of the vegetative cell (VN). The cellular
protrusion of the sperm cell (arrowheads) is found within the clasping region of the lobed vegetative cell nucleus and extends through a shallow hole in the vegetative cell nucleus at one location (arrow). (From Russell 1984. Planta 162:385-391. © Springer-Verlag, Berlin.)
Perhaps the most dramatic difference between the two sperm lies in their relative content of heritable organelles. The large sperm is rich in mitochondria and is typically impoverished of plastids, whereas the small one is poor in mitochondria but has numerous plastids. Since P. zeylanica is unusual in lacking synergids in the embryo sac, sperm-vegetative cell nucleus association and sperm dimorphism seen in this species raised objections that they may not be representative of the wide range of angiosperms characterized by the presence of two synergids flanking the egg cell. This objection was soon dispelled when similar features were found in sperm cells of
lacks a cytoplasmic outgrowth, is linked to the larger one by a common cell junction. The small sperm cells in both species of Brassica also appear unique in their paucity of mitochondria, with 4 in B. campestris and 6 in B. oleracea, compared to a maximum of 15 and 34 in the large sperm cells of B. oleracea and B. campestris, respectively. Neither
Brassica campestris and B. oleracea (Dumas, Knox
and Gaude 1985; McConchie, Jobson and Knox 1985; McConchie et al 1987), both of which possess synergids. In both, dimorphism in shape is due to the presence of an evagination on the sperm that is attached to the vegetative cell nucleus, which thus makes it the larger of the two sperm cells. The distinction between the two species lies in the fact that in B. oleracea the evagination on the sperm surrounds the surface of the vegetative cell nucleus superficially, whereas in B. campestris it penetrates the enclaves of the vegetative cell nucleus and emerges at the other end as a protuberance (Figure 11.3). In both species, the smaller sperm, which
MT MT
Fig. 11.3. A schematic diagram of the male germ unit of Brassica campestris. M, mitochondria; MT, microtubules; N, nucleus; VN, nucleus of the vegetative cell; SC1, SC2, sperm cells 1 and 2. (From McConchie, Jobson and Knox 1985.)
298
Pollination and Fertilization
sperm cell contains any plastids. The published This attention to detail seems justified because other accounts of the structure of the sperm of Spinacia electron microscopic studies, with or without threeoleracea are much less detailed than those of Brassica dimensional reconstruction of serial sections, have sp., although differences in the sperms' size and demonstrated a physical relationship between the association with the nucleus of the vegetative cell sperm and the vegetative cell nucleus; included in are obvious in the electron micrographs (Wilms, these studies are male germ units identified in pollen Leferink-ten Klooster and van Aelst 1986; Theunis, tubes of bicellular pollen grains of Hippeastrum vittaCresti and Milanesi 1991). tum (Mogensen 1986), Petunia hybrida (Wagner and Thanks to the skills of the electron microscopist Mogensen 1988), Rhododendron laetum, R. macgregriae and the availability of techniques for computer (Taylor et al 1989), Cyphomandra betacea (Solanaceae) simulation of the data, these observations add a (Hu and Yu 1988), Nicotiana tabacum (Yu, Hu and new dimension to our knowledge of sperm struc- Zhu 1989; Yu, Hu and Russell 1992; Yu and Russell ture and organelle inheritance in angiosperms. 1994b), and Populus deltoides (Rougier et al 1991), Despite the minimal structural attributes displayed and in tricellular pollen grains of Spinacia oleracea by the sperm, this cell appears to be highly differ- (Wilms, Leferink-ten Klooster and van Aelst 1986), entiated in relation to its developmental tasks, Catananche caerulea (Asteraceae) (Barnes and especially to respond with preestablished speci- Blackmore 1987), and B. napus (Murgia et al 1991b). ficity and to discriminate between the egg nucleus In C. betacea, N. tabacum, and P. hybrida, the two and the polar fusion nucleus during fertilization. sperm are linked together by a common transverse The presence of significantly different numbers of wall as in Plumbago zeylanica. In N. tabacum, the wall heritable organelles in the two sperm cells of a pair appears to be further specialized by the presence of suggests the possibility of sophisticated mecha- fibrils, small tubules, and callose (Yu, Hu and nisms of organelle transmission at fertilization. The Russell 1992). differences in the sizes of the two sperm and in The importance of three-dimensional reconstructheir organelle contents pose some important ques- tion of the sperm-vegetative cell nucleus assembly tions on differential gene activity during sperm cell lies in the demonstration that complex association differentiation. between cells can be understood in simple graphic The novel features of the structure of the sperm pictures. The impressive precision of these studies provided the impetus for further work, which caused many to believe that it was only a matter of brings the observations in line with the current time before the concept of the male germ unit could thinking on the nature of the male gamete. Dumas be considered as a secure generalization applicable et al (1984) designated the functional complex of the universally to angiosperms. This did not prove to two sperm cells of common origin plus the nucleus be the case. From light and electron microscopic of the vegetative cell in the pollen grain, or in the observations of pollen grains and pollen tubes of pollen tube, as the male germ unit; the rationale for wheat (Chandra and Bhatnagar 1974), Helleborus this concept is the view that all the nuclear and foetidus (J. Heslop-Harrison, Heslop-Harrison and cytoplasmic DNA of male heredity is linked Heslop-Harrison 1986), Euphorbia dulcis (Murgia together, forming a single functional unit for the and Wilms 1988), and Alopecurus pratensis (J. recognition of target female cells, the transmission Heslop-Harrison and Heslop-Harrison 1984), it of hereditary materials, and fusion during double appears that there is a spatial or temporal separafertilization. Some investigators (Kaul et al 1987; Yu tion of the sperm pair from the nucleus of the vegeand Russell 1992) have referred to a complex asso- tative cell. In live pollen tubes of Secale cereale, not ciation formed between the vegetative cell nucleus only is there no clear-cut evidence of linkage and the generative cell as the male germ unit; in between the sperm cells, but the cells have been other cases, the transient union noted between the observed to shoot past each other and move in two sperm after they are formed from the genera- opposite directions (J. Heslop-Harrison and tive cell has been identified as the male germ unit Heslop-Harrison 1987). According to Weber (1988), (Levieil 1986; Charzynska et al 1989; Charzynska who followed the division of the generative cell in and Lewandowska 1990). It is a moot point whether tricellular pollen grains of Galium mollugo, such associations, however complex they are, Trichodiadema setuliferum, and Avena sativa, there is should be included within the concept of the male no linkage between sperm cells that separate from germ unit, which was coined to convey a precise one another soon after they are formed. These functional union involving the two sperm and the observations, though drawing attention to the indevegetative cell nucleus prior to double fertilization. pendent existence of sperm cells and the nucleus of
Fertilization: The Beginning of Sporophytic Growth
299
the vegetative cell, do not take into account the fact try. Further work has shown that the high degree of that an association between them may be formed at cytoplasmic difference displayed by the sperm of a later stage, and so further work is called for before Plumbago zeylanica, characterized by the presence of the issue can be considered settled. There is strong significantly different numbers of plastids and support for the view that in some plants the physi- mitochondria, is somewhat unique and that far cal connection between sperm cells of a pair and more widespread are cases in which organelle difbetween one sperm and the vegetative cell nucleus ferences are restricted to mitochondrial number or is ephemeral and would have been completely nuclear size (McConchie, Hough and Knox 1987). missed in routine electron microscopic observa- Moreover, pollen grains of some plants with pertions. For example, by three-dimensional analysis of fectly standard male germ units have revealed no ultrathin sections, it was found that in the pollen appreciable differences in organelle contents by grains of barley (Mogensen and Rusche 1985) and serial thin-sectioning, computer reconstruction, maize (McConchie, Hough and Knox 1987), the and quantification. Such isomorphic sperm cells sperm cells are discrete and do not exist in physical have been found in barley (Mogensen and Rusche association with the vegetative cell nucleus as the 1985), Petunia hybrida (Wagner and Mogensen male germ unit. The major effort in a subsequent 1988), Rhododendron sp. (Taylor et al 1989), Medicago study on barley was to look for evidence of a phys- sativa (Zhu, Mogensen and Smith 1992), and ical association between sperm cells and the vegeta- Nicotiana tabacum (Yu, Hu and Russell 1992). In the tive cell nucleus during hydration and germination most detailed quantitative analysis of the sperm in of the pollen grain (Mogensen and Wagner 1987). N. tabacum, no statistically significant differences This work showed that following controlled polli- were found between the two sperm cells in several nation, before sperm cells move into the emerging cellular parameters (Yu, Hu and Russell 1992; Yu pollen tube, there is a close association between and Russell 1994b). sperm cells and between them and the nucleus of In summary, it appears that in many angiothe vegetative cell. Although the vegetative cell sperms, part of the preprogramming of sperm cells nucleus is not seen to migrate into the pollen tube, involves the formation of the male germ unit with sperm cells remain attached in the pollen tube; as or without a concurrent asymmetry in the distribusperm cells approach the embryo sac, there is a furtion of heritable organelles. How this specialized ther intimacy in their association and they appear to packaging of the sperm influences the fertilization be connected at both ends. In maize pollen also, the process is not known at present. Dimorphism vegetative cell nucleus is found to be separated raises questions about the identity of the sperm cell from sperm cells, but it remains closely associated that fuses with the egg and of the one that fuses with sperm cells as a male germ unit in the course with the polar fusion nucleus, with perhaps the of postpollination journey (Rusche and Mogensen hardest question to answer being, what is the 1988; Mogensen 1992). In Medicago sativa, the assonature of the factors that endow the specificity? We ciation of sperm cells with the nucleus of the vegealso need to know the mechanisms that cause the tative cell is tenuous in the pollen grain, but is well unequal partitioning of organelles and the trigger defined in the pollen tube throughout its growth that induces it. (Zhu, Mogensen and Smith 1992). It has been argued from these observations that the male germ unit is not the developmental consequence of divi(iii) Isolation and Characterization of the sion of the generative cell; rather, it is induced as a Sperm result of pollen germination (Mogensen 1992). Since To gain insight into the molecular biology of male germ unit formation is a morphogenetic process, it is also possible that, in some plants, the double fertilization, it is necessary to know somestructure formed in the pollen grains is not stabi- thing about the dynamics of the participating lized before germination; germination is the trigger gametes. Although it is relatively easy to monitor that stabilizes the male germ unit in the pollen sperm cells microscopically, it is hard to investigate their metabolic activities experimentally. This objectubes. tive is best achieved by isolating sperm from pollen It was mentioned earlier that in certain plants grains or from pollen tubes. As the following survey that possess a male germ unit, the sperm cells show shows, from a modest beginning using sperm cells dimorphism in size and in the content of heritable extruded from pollen grains, the stage is set now for organelles. But the presence of a male germ unit is the large-scale isolation of viable sperm cells from not always associated with cytoplasmic asymme- pollen grains and pollen tubes of various plants. A
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brief review of the present state of the art is provided by Theunis, Pierson and Cresti (1991). The first attempts to isolate sperm used tricellular pollen grains of barley (Cass 1973) and Plumbago zeylanica (Russell and Cass 1981). In recognition of the fact that the milieu of the pollen grain is in a state of high osmotic pressure, pollen contents were released into a medium of high osmolarity secured by the addition of 30% sucrose. Although the number of sperm cells isolated by this method was not large, some basic observations, such as the change in the shape of extruded sperm from spindle shaped to round or spherical, suggested a role for the cytoskeletal framework in the process. Similar methods, with variations in the concentrations of sucrose, were used subsequently for the small-scale isolation of sperm from tricellu-
Pollination and Fertilization
is necessary when pollen cannot be burst osmotically; however, germinating the pollen grains for a short period of time to generate tubes was sufficient to allow sperm release by osmotic shock (P. E. Taylor et al 1991). All these methods for the largescale isolation of sperm have used tricellular pollen grains; an osmotic shock combined with treatment with pectinase and cellulase has been devised to obtain high sperm yields from pollen tubes of several bicellular species (Mo and Yang 1992). Viability of isolated sperm cells is routinely assessed by using Evans-blue as an exclusion dye (Russell 1986; Southworth and Knox 1989) or by fluorochrome reaction (Dupuis et al 1987; Yang and Zhou 1989; Mo and Yang 1991). Zhang et al (1992a, b) have advocated the use of flow cytometry as a highly sensitive method to evaluate the viability of lar pollen grains of Brassica oleracea, Triticum aes- isolated sperm cells. Not unexpectedly, viability was tivum (Matthys-Rochon et al 1987), and Zea mays found to decrease with time of storage of isolated (Cass and Fabi 1988). The first successful isolation sperm cells (Russell 1986; Dupuis et al 1987; Theunis of sperm from bicellular types of pollen grains was and van Went 1989; Yang and Zhou 1989; Mo and accomplished by enzymatic hydrolysis of the Yang 1991). Among the measures recommended to pollen tube wall of Rhododendron macgregoriae andenhance the longevity and viability of isolated Gladiolus gandavensis (Shivanna et al 1988). For this sperm of maize are the use of a buffered galactose purpose, pollen tubes allowed to grow through the solution or the use of a dilute mineral-salt solution style into a block of agar were treated with a mix- especially low in calcium and magnesium (Zhang et ture of macerozyme and cellulase, and sperm cells al 1992b; Roeckel and Dumas 1993; G. Zhang et al were separated by microfiltration. 1995). A procedure for increasing the yield and viaGiven the differences in density between the bility of sperm cells isolated from Lolium perenne has sperm and the rest of the pollen grain or pollen used vitamin-C and fetal calf serum during isolation tube contents, one might expect to enrich a pollen and storage (van der Maas et al 1993b). Maintaining fraction for sperm by differential centrifugation. isolated sperm in a prime condition is an issue to This was achieved by subjecting osmotically which attention needs be focused for sperm to fulfill shocked pollen grains of P. zeylanica to centrifuga- their potential use in in vitro fertilization and as tion in a sucrose gradient (Russell 1986). gene vectors in genetic manipulation. In other Moderately high yields of sperm essentially free of respects, such as in the absence of the common contamination by vegetative cell nuclei and starch plasma membrane of the vegetative cell, isolated and suitable for physiological manipulations were sperm cells resemble true protoplasts (Russell 1986; obtained by this method. Dupuis et al (1987) and Dupuis et al 1987; Cass and Fabi 1988; Theunis Theunis and van Went (1989) used Percoll gradi- 1990). Morphometric analysis and three-dimenents to obtain high sperm yields from pollen grains sional reconstruction of serial sections of isolated of maize and spinach, respectively. Sperm cells at a sperm of Z. mays have revealed that there is considconcentration of 20,000/ml were obtained from erable heterogeneity in the volume and number of pollen grains of Gerbera jamesonii by gentle homog- the complement of organelles in a given population enization followed by microfiltration (Southworth of sperm (Wagner, Dumas and Mogensen 1989; and Knox 1989). Both grinding and a two-step Mogensen, Wagner and Dumas 1990). From a osmotic shock procedure have been claimed to give heuristic standpoint, this shows that strengthening high yields of sperm from pollen grains of B. the differences between two groups of sperm in a campestris, B. napus, Z. mays, and Secale cereale (Yang population is not an easy task. and Zhou 1989; Mo and Yang 1991). In the latter Except in two instances (see Section 3), it has not procedure, pollen grains hydrated in a high con- been determined whether isolated sperm cells are centration of sucrose were shocked to burst and functionally competent to fuse with an egg cell or release the sperm by dilution of the medium. This with a central cell in an in vitro environment.
Fertilization: The Beginning of Sporophytic Growth
However, a few reported cases of fusion between isolated sperm cells of the same species and between sperm of different species and between isolated sperm and isolated microspore protoplasts have provided some hopeful hints of a wider application of the technique. This curious behavior has been described among sperm cells of Gladiolus gan-
301
both transcription and translation (Zhang, Gifford and Cass 1993; Matthys-Rochon et al 1994). But the evidence is not really conclusive as to whether this is a reflection of the synthetic activities of sperm cells or is a reaction to isolation stress. It is clear from this account that in recent years a major effort in the study of male gametes of davensis, Zephyranthes Candida, Hemerocallis minor, angiosperms has been directed toward isolation proH. fulva, and Hippeastrum vittatum; between sperm cedures. Despite the fact that well-established procecells of G. gandavensis and H. vittatum; between dures are already at hand for the biochemical sperm and microspore protoplasts of H. minor; and characterization of sperm, such studies are limited. between sperm of H. vittatum and microspore pro- The special appeal of male gametes derives from the toplasts of H. fulva (Mo and Yang 1992). fact that each sperm of a pair in the pollen grain The metabolic and biochemical activities of iso- is programmed for different fusion events. lated sperm cells have hardly been investigated. Characterization of the recognition factors in the Preliminary attempts have been made to character- sperm cells is an essential prerequisite to underize the surface antigens of sperm cells by employ- standing the physiological and molecular basis of ing monoclonal antibodies. Hough et al (1986) double fertilization. raised hybridoma cell lines against isolated male gametes of Brasska campestris and, by screening them for antibody specificity by a simplified 2. ODYSSEY OF THE SPERM AND DOUBLE FERTILIZATION immunofluorescence technique, identified a few sperm surface-specific antigens. In another investiThe fate of male gametes in the pollen tube gation, Pennell et al (1987) screened hybridoma growing in the style is certainly the most critical antibodies elicited by Plumbago zeylanica sperm issue to be considered now, as it lies at the heart of against both sperm and pollen fractions. Whereas a much of our understanding of double fertilization. number of antibodies were found to bind to The pollen tube makes its way into the embryo sac, contaminating fractions from the vegetative cell where it discharges its cargo for fertilization events. cytoplasm and soluble macromolecules, a high per- The tasks involved here are far too prodigious to be centage of the reactants were also sperm specific. accomplished by sperm cells without some outside Although much remains to be done to characterize help. For convenience, the events leading up to the antibodies in both species, this is a task that pollen tube discharge are designated as the might eventually lead to insights into the basic progamic phase; this is followed by the syngamic problems of sperm recognition. phase, in which the predominant event is a set of In isolated sperm cells of P. zeylanica, a beginning nuclear fusions. has also been made to study the protein profile of the sperm by SDS-PAGE with a view toward identi(i) Growth of the Pollen Tube through fying recognition molecules during egg-sperm the Style interaction (Geltz and Russell 1988). Obtaining evidence for the existence of sperm-specific proteins In species with tricellular pollen grains, the was difficult in this work because of contamination sperm cells and the vegetative cell nucleus migrate of the preparation with vegetative cell nuclei, into the emerging pollen tube, whereas in plants organelles, and water-soluble proteins. Nonetheless, with bicellular pollen grains the generative cell about 14% of the most conspicuous polypeptides divides in the pollen tube to give rise to sperm were specific to sperm cells; whether these include cells. The stigmatic tissue on which pollen grains cell surface determinants of male gametes remains land is composed of glandular cells that facilitate to be investigated. In similar studies with isolated germination of pollen grains and growth of pollen sperm cells of maize, the protein synthetic profile tubes (see Chapter 7). These cells lead to the transwas found to change with time after isolation. The mitting tissue of the style, which is also made up of increased incorporation of radioactive precursors of cells with glandular properties. What is the mechaRNA and protein synthesis into isolated sperm cells nism by which the pollen tube is guided toward the that was observed in these investigations evidently ovary? It is now established that pollen tubes grow shows that changes in protein synthesis involve through the intercellular matrix of the style, and
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indications are that a variety of control mechanisms operate within the style to maintain the downward directionality of the pollen rube. It has been argued quite convincingly that in some plants the guidance of the pollen tube is mechanical or electrical and that such cues interact with the intercellular growth machinery of the stylar matrix. Mechanical guidance begins to take hold when the tip of the pollen tube is properly oriented in the stylar matrix. To examine this proposition closely, Iwanami (1953) dusted the stigma of Lilium auratum with pollen and followed the course of the pollen tube. The finding in this work that the normal site of pollen tube extension was along the longitudinally oriented cells surrounding the stylar canal led to the view that the pollen tube is directed mechanically by these cells. That the pollen tube is not guided in this situation by some polarized gradient of a chemical substance was subsequently established in an extensive range of delicate surgical experiments on the styles of L. auratum and L. longiflorum. In one experiment, when cut ends of the style were grafted to its morphologically inverted pieces, the artificial style was found to serve admirably as a medium for the growth of pollen rubes (Iwanami 1959). This could not have happened if chemical gradients were at operation in directing pollen tube growth. Some investigations on pollen-stigma interaction in maize and other Poaceae are also consistent with the view that pollen rubes are oriented in the direction of the ovary by mechanical constraints (Y. HeslopHarrison, Reger and Heslop-Harrison 1984b). Following germination of the pollen grain on the receptive stigmatic trichome in the species of Poaceae studied, the orientation of the pollen tube tip in the stylar tract seems to determine its unhindered growth toward the ovary. This occurs even in the absence of any visible structural adaptations directing the pollen tube toward the transmitting tracts. A long-distance influence, probably of a mechanical nature, on the direction of growth of the pollen tube through the style has also been indicated in Nicotiana alata (Mulcahy and Mulcahy 1987). Reports of the responses of pollen tubes to applied electric fields have nurtured the view that endogenous currents may direct pollen tube growth. Since directional growth of pollen tubes in vitro toward both the anode (Wulff 1935; Wang, Rathore and Robinson 1989; Nozue and Wada 1993) and the cathode (Marsh and Beams 1945; Zeijlemaker 1956; Nakamura et al 1991) has been observed, it is difficult to evaluate the significance
Pollination and Fertilization of the data. In Narcissus pseudonarcissus, the ovary is
electrically negative by about 100 mV with respect to the stigma; the potential voltage difference may be the guiding force of the pollen tube to the ovary (Zeijlemaker 1956). Given the delicate nature of the pollen tube and the need for micromanipulative procedures, the mechanism directing electrical field-induced pollen tube growth is likely to remain mysterious for some time. The attraction of pollen rubes to a source of Ca2+, to be described later, coupled with the evidence that calcium ions constitute one component of the endogenous current in the pollen tube (Weisenseel, Nuccitelli and Jaffe 1975; Weisenseel and Jaffe 1976) and that Ca2+ accumulates at the tip of the pollen tube (Jaffe, Weisenseel and Jaffe 1975; Reiss, Herth and Nobiling 1985), has engendered much speculation about the possible role of redistribution of these ions in influencing the direction of pollen rube growth through applied electrical fields. The most commonly mentioned mechanism guiding the direction of growth of pollen tube through the style is the attraction toward unspecified chemical substances in the style itself. The glandular cells of the style have drawn attention as a possible source of chemotropic substances. Following upon the work of several earlier investigators going back to the end of the 19th century, Tsao (1949) demonstrated chemotropism of pollen tubes to pistil parts of 9 out of 36 species tested on artificial media; it was further shown that the active factor from the pistil of L. superbum was diffusible into agar blocks, which then attracted pollen tubes. A later study by Miki (1964) confirmed and extended these results using L. longiflorum and L. japonicum and showed that the chemotropic factor is a heat-labile, diffusible, small molecule. Linck and Blaydes (1960) demonstrated chemotropism in four additional species, of which Clivia nobilis was particularly noteworthy for the attraction of pollen tubes not only by slices of the carpel but also by those of sepal, petal, and leaf. Attempts to elucidate the mechanism regulating pollen tube pathway in other species of Lilium have also been made. For example, pollen tubes of several hybrids of L. longiflorum are found to respond positively to the upper part of the style, although the activity is weak or absent in the mid and lower segments; the ovary displays no activity at all (Rosen 1961). In L. leucanthum and L. regale, the glandular cells lining the stylar canal have been identified as the source of the putative chemotropic factor, which is apparently distributed in a basipetal gradient (Welk, Millington and Rosen 1965). An influence of the ovary on
Fertilization: The Beginning of Sporophytic Growth
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lowing the early studies and, until recently, other investigations on this topic have not been directed at chemotropic agents in the style. Pollen tubes of Pennisetum glaucum have been shown to respond positively to diffusates of the pistil tissue as well as to glucose; a totally different understanding of the in vivo metabolism of the style has emerged from the discovery that the pistil diffusate response is due to glucose produced by tissue-bound invertase action rather than to free glucose (Reger, Chaubal and Pressey 1992; Reger, Pressey and Chaubal 1992). Instead of positing the growth of pollen tubes to be under a chemotropic influence, some investigations have led to a model of pollen tube growth in the style as a directional cell migration along a gradient of a factor in the secretory matrix of the style; this model is analogous to a biochemical recognition-adhesion system. The initial experimental support for this model came from the observation Progress in our understanding of the interaction that latex beads approximating the size of pollen between the stylar matrix and the growing pollen tube tips, when introduced into the stylar transmittube has been bolstered by attempts to identify the ting tracts of certain plants, traveled to the site of chemotropic substance. A key piece of information the ovary at rates similar to those of pollen tubes in this context came from the work of Mascarenhas regardless of the stylar gravitational orientation and Machlis (1962b), who showed that in (Sanders and Lord 1989). Since substrate adhesion Antirrhinum majus the positive chemotropic factor is molecules probably confer this ability on the beads a small, heat-stable, water-soluble molecule associ- to travel in the stylar matrix, attempts have been ated with larger molecules. This finding led to the made to identify these substrate molecules and search for inorganic ions and to the discovery that their receptors in the style. A candidate molecule Ca2+ is chemotropically active for pollen tubes of A. identified in the substrate adhesion role is vitmajus, Narcissus pseudonarcissus, and Clivia miniata ronectin. Immunolocalization of vitronectin-like (Mascarenhas and Machlis 1962a, 1964). It was also proteins in cross sections of the pistil of Vicia faba found that the levels of Ca2+ present in chemotropi- showed strong reaction in the stylar transmitting cally active parts of the pistil of A. majus were well tract, suggesting a possible role for these proteins within the physiological range of the ion active in in guiding pollen tube growth in a manner similar bioassays of chemotropism. In the pistil of Petunia to cell migration in animals (Sanders et al 1991). hybrida, membrane-bound Ca2+ is found in a Jauh and Lord (1995) have further shown that by decreasing gradient from the stigma to the base of excising different lengths of the profusely pollithe style before pollination, and in the reverse direc- nated style of lily, it is possible to cause the tip of tion after pollination (Bednarska 1995). However, the pollen tube, in the absence of the remains of the there is some skepticism as to whether Ca2+ is a uni- pollen grain and the spent pollen tube, to convey versal chemoattractant for pollen tubes. In this con- the sperm cells to the ovules and effect fertilization. text, in vitro bioassays showing that pollen tubes of The tip of the pollen tube can be considered analoOenothera longiflora and other species grow in a ran- gous to a population of migrating cells that adhere dom manner in the presence of calcium compounds to one another and to the cells of the transmitting have had a particularly powerful impact on the tissue of the style during its journey to the ovule. skeptics (Glenk, Schimmer and Wagner 1970). It is The role of adhesive substrates in pollen tube also relevant to point out that there is a lack of a gra- growth has unfolded further with the demonstradient of ionic calcium in the style of A. majus, which tion that glycoproteins isolated from the stylar seems to argue for the presence of an additional fac- transmitting tissue of tobacco adhere to the surface tor as a chemotropic agent (Mascarenhas 1966a). of pollen tubes and are incorporated into the wall There has not been much research output on pollen of the pollen tubes growing in the style. It appears tube chemotropism in the style in the period fol- that within the style, pollen tube-bound enzymes pollen tube growth in Petunia hybrida has been demonstrated using a "semi-vivo" technique in which the ovary is removed at different times after pollination. Whereas the growth of pollen tubes in the style detached from the ovary was inhibited irrespective of the time after pollination, their growth in the intact pistil occurred normally toward the ovary (Mulcahy and Mulcahy 1985). Despite the fact that most of these observations are based on in vitro experiments that may have little or no parallel to events in vivo, they nonetheless suggest a complex chemical interaction between the growing pollen tube, the stylar matrix, and an ovule-derived guidance mechanism controlling pollen tube growth (Hiilskamp, Schneitz and Pruitt 1995). It is worth noting that use of the same methods described here has not led to the demonstration of pollen tube chemotropism in certain other plants (Cook and Walden 1967; Hepher and Boulter 1987; Kandasamy and Kristen 1987b).
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deglycosylate the proteins and thus release sugar (Tilton 1980). The filiform apparatus is believed to molecules that are presumably utilized by the produce most of the protein- and carbohydrate-rich growing pollen tubes (Wu, Wang and Cheung exudate in G. verrucosa, with minor contributions 1995). These observations suggest that stylar glyco- from the micropylar cells of the nucellus (Franssenproteins of tobacco serve an important role in Verheijen and Willemse 1993). The characteristics matrix adhesion and in cell metabolism. of the chemotropic systems guiding the pollen tube The general path of pollen tube entry into the to the ovule that do not involve Ca2+ provide interovule has been described convincingly in numer- esting comparisons with the growth of the pollen ous classical studies. These works showed that tube in the style, considered earlier, and its final after reaching the ovary, the pollen tube almost push into the embryo sac to be considered next. always grows downward over the outer integument, then between the inner ovary wall and the (ii) Passage of Sperm into the Embryo Sac inner integument of the ovule, toward the placenta. When the placental region is reached, the pollen This part begins with a brief account of the tube executes a sharp turn and continues its jour- ovary. As is well known, the ovary not only proney between the placenta and the outer integument vides the physical protection for the enclosed to the micropyle. At the micropyle, the tube grows ovules, but also harbors the tissues through which intercellularly through the nucellus to the embryo the pollen tube grows during the final phase of its sac. Observations of this path reported from con- existence. As described in Chapter 6, the embryo ventional histological studies were later confirmed sac is formed within the ovule, which itself is born in research using fluorescent microscopy to trace on the placenta. Before the pollen tube reaches the the pollen tube in the ovary (Y. Heslop-Harrison, embryo sac, it encounters the placenta, funiculus, Heslop-Harrison and Reger 1985; Reger, Chaubal and integuments of the ovule. Light microscopic and Pressey 1992). How does the pollen tube find studies have revealed three general pathways its way to the micropyle from the base of the style? through which the pollen tube enters the ovule and From indirect lines of evidence, the basic plan races to the embryo sac. In the most common route, appears to be one guided by a chemotropic attrac- known as porogamy, the pollen tube enters the tant produced at a site close to the micropyle. The ovule through the micropyle. In some plants, presence of such substances has been demon- mucilaginous canals and ancillary structures such strated in the micropylar region of ovules of as obturator, embellum, ponticulus, and papillae Paspalum orbiculare (Poaceae) (Chao 1971), that aid in the entry of the pollen tube into the Ornithogalurn caudatum (Tilton 1980), maize (Y. micropyle have evolved (Kapil and Vasil 1963; Heslop-Harrison, Heslop-Harrison and Reger Kapil and Bhatnagar 1975; Busri, Chapman and 1985), beet root (Bruun and Olesen 1989), Gasteria Greenham 1993; Martinez-Palle and Herrero 1995). verrucosa (Franssen-Verheijen and Willemse 1993), Two other preferred sites of entry of the pollen tube and Asclepias exaltata (Asclepiadaceae) (Sage and are at the chalazal end (chalazogamy) and on one Williams 1995), although the origin of the sub- side of the ovule (mesogamy), but they are used stances has not been clearly determined in all cases. only sporadically (Maheshwari 1950). These obserThe substance found in P. orbiculare is PAS positive vations suggest that pollen tubes may be directed and proteinaceous; it consists of secretion products to specific parts of the ovule by a gradient of chemof the nucellus and integuments, as well as the ical substances. Irrespective of the pathway taken product of dissolution of cells of the outer and by the pollen tube to the ovule, the tip of the pollen inner integuments close to the micropyle (Chao tube invariably makes contact with the embryo sac 1971, 1977). The substance in maize stains for pro- at its micropylar end. teins and pectic polysaccharides, although in the This leads to the consideration of the entry of light microscope its origin is not easily resolved (Y. the pollen tube into the embryo sac and the disHeslop-Harrison, Heslop-Harrison and Reger charge of sperm. It is an interesting commentary on 1985). In beet root, the middle lamella of the nucellar cells in the micropylar region comes apart dur- the state of uncertainty in this field that up to the ing the maturation process and these cells probably early 1960s, based on light microscopy, as many as secrete PAS-positive substances into the micropyle six different modes of entry of the pollen tube into (Bruun and Olesen 1989). Proteins and RNA are the the embryo sac involving one or both synergids main components of the exudate secreted by the were described. As summarized by Kapil and nucellar cap and inner integument of O. caudatum Bhatnagar (1975), the pollen tube might (a) enter between the two intact synergids, (b) enter between
Fertilization: The Beginning of Sporophytic Growth
the egg and the synergid, (c) enter between the embryo sac wall and one or both synergids, (d) penetrate one of the synergids, destroying it in the process, (e) destroy the synergid on contact, or (f) enter the cytoplasm of the synergid, which has degenerated before the arrival of the pollen tube. Route (d), by which the pollen tube enters the embryo sac and destroys one of the synergids in the process, has been described most frequently, whereas the other modes of entry have been documented in only a few cases. It should be pointed out that in the light microscope it is often difficult to distinguish between a healthy and a degenerating synergid in sections of the embryo sac clouded by pollen tube discharge. The era of electron microscopy led to a new level of understanding of the passage of the pollen tube into the embryo sac. What appeared as a basic pattern of pollen tube entry at the electron microscope level was described by Jensen and Fisher (1968a) in Gossypium hirsutum, which to date remains as the one species in which events associated with fertilization are best known. In G. hirsutum, as in many other species, the two synergids are identical before pollination. However, one of the synergids begins to disintegrate prior to the entry of the pollen tube into the embryo sac. In fact, pollination triggers the degeneration process, which is most frequently initiated when the pollen tube is in the style. The first signs of synergid collapse are irregular thickening and darkening of the plasma membrane and of the outer membranes of adjacent plastids and mitochondria, accompanied by a general increase in the electron density of the cytoplasm. When the pollen tube reaches the embryo sac, the tip of the tube grows through the filiform apparatus into the degenerating synergid. The degeneration of the synergid initiated before pollen tube entry into the embryo sac is accelerated after entry of the pollen tube. Signs of accelerated degeneration of the synergid are transformation of the plastids and mitochondria into ghosts of their original structure, collapse of vacuoles, and breakdown and disappearance of the plasma membrane. As to the fate of the pollen tube, after passage through the cytoplasm of the degenerating synergid, pollen tube growth comes to a standstill and then the pollen tube discharges its contents of sperm and cytoplasm. The observations on G. hirsutum imply that rather than being the result of mechanical intrusion of the pollen tube, synergid degeneration is caused by pollination and is actually required for normal pollen tube entry into the embryo sac. Fascinating
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though this thesis is, its possible significance in the fertilization process was not widely appreciated until there was a long list of plants in which it was established that synergid degeneration begins before the arrival of the pollen tube and that the pollen tube consistently favors the degenerating synergid. In subsequent years, other species, such as barley (Cass and Jensen 1970), Zea mays (Diboll 1968; van Lammeren 1986), Epidendrum scutella (Cocucci and Jensen 1969c), Linum usitatissimum (Vazart 1969; Russell and Mao 1990), Quercus gambelii (Mogensen 1972), Proboscidea louisianica (Mogensen 1978b), Nicotiana tabacum (Mogensen and Suthar 1979; Huang and Russell 1992), Spinacia oleracea (Wilms 1981b), Crepis capillaris (Kuroiwa
1989), Glydne max (Dute, Peterson and Rushing 1989), Populus deltoides (Russell, Rougier and Dumas 1990; Rougier et al 1991), and Arabidopsis (Murgia et al 1993), have gained general acceptance as examples in which synergid degeneration occurs in apparent response to pollination. The pattern of cytoplasmic reorganization observed in one of the synergids following pollination varies widely, from a series of disordered changes in the organelles, as in G. hirsutum (Jensen and Fisher 1968a), to only a difference in the density of the cytoplasm due to an increase in the number of ribosomes, as in N. tabacum (Mogensen and Suthar 1979). That the loss of synergid integrity is not an artifact of the chemical fixatives used in these investigations has been confirmed by observing similar changes in synergids of cryofixed ovules of N. tabacum, although the full extent of cellular degeneration is delayed until pollen tube penetration (Huang, Strout and Russell 1993). Considerable variation in the condition of the synergid in relation to the timing of pollination and pollen tube growth has been observed in other plants. For example, in wheat, degeneration of both synergids is initiated in embryo sacs collected just prior to pollination; the degeneration cycle is completed in one synergid after entry of the pollen tube, whereas the other appears unaffected (You and Jensen 1985). In Brassica campestris, both synergids show degenerative changes in pollinated as well as in unpollinated flowers; this implies that degeneration of the synergid does not depend upon pollination (van Went and Cresti 1988a; Sumner 1992). In Beta vulgaris (Bruun 1987) and Oryza sativa (Dong and Yang 1989), one of the synergids readies to receive the pollen tube and begins to display signs of degeneration before anthesis of the flower. There are also reports of synergids remaining healthy even after pollination and
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pollen tube growth; apparently, when the pollen been employed to localize gross Ca2+ and loosely tube makes contact with the synergid, the degener- sequestered exchangeable Ca2+, respectively, in the ative program of the cell is set in motion. Examples embryo sac cells of cereal grains. The synergids in which this situation is found are Capsella bursa- were found by both methods to display high conpastoris (Schulz and Jensen 1968b), Petunia hybrida centrations of Ca2+, suggesting an active role for the (van Went 1970c), Ornithogalum caudatum (van ion as an erotropic substance for the attraction of Rensburg and Robbertse 1988), and Lilium longiflo- pollen tubes (Chaubal and Reger 1990,1992a). Since rum (Janson and Willemse 1995). Although the the pollen tube moves directionally to the synergid, 2+ paired synergids of Helianthus annuus do not show the existence of a continuous Ca gradient from the placenta to the synergid has been postulated any structural differences before pollination, degen(Chaubal and Reger 1990). In other cases, PAS-stainerative changes begin to surface in one of them gradually after pollination, whereas the other one ing substances secreted by the synergid probably remains perfectly healthy until after fertilization function as part of the pollen tube guidance system (Newcomb 1973a; Yan, Yang and Jensen 1991a). (Chao 1971; Bruun 1987; Willemse, Plyushch and Despite the fact that ultrastructural analysis has not Reinders 1995). However, as is true of most cells been carried out to the same depth in all of the growing in isolation within a tissue of different oriaforementioned cases, sufficient details are at hand gin, the movement of the pollen tube may be condito establish the essential validity of the hypothesis tioned by the physical state of the neighboring cells. that irrespective of the prior response of the syn- For example, compared to the intact cells of the ergid to pollination stimulus, the pollen tube grows female gametophyte, the degenerating synergid into one of the two synergids and then, as the pollen might offer a lower resistance to the growing pollen tube discharge occurs, degenerative changes set in. tube (Russell 1992). Moreover, when penetrating It is not, however, easy to fit into this model genera the synergid, the pollen tube conforms in a regithat lack synergids, such as Plumbago and mented way to the filiform apparatus. Since microtubules are associated with the filiform apparatus, Plumbagella. According to Russell (1982), arrival of the possibility that they may somehow be involved the pollen tube at the micropylar end of the embryo in giving directionality to the pollen tube also sac in Plumbago zeylanica is not heralded by the disdeserves attention (Cass and Karas 1974; B.-Q. array of a receptor cell; rather, the pollen tube peneHuang et al 1990). trates the wall ingrowths of the filiform apparatus at the base of the egg and grows in the intercellular The degeneration of the synergid before or after space between the egg and the central cell until it entry of the pollen tube is obviously an important reaches the vicinity of the nucleus near the chalazal step in the fertilization process. We do not yet end of the egg. A similar sequence of events occurs understand how this happens, even in plants in in Plumbagella micrantha (Russell and Cass 1988). It which pollination triggers the degeneration prois clear from this that, whatever else is going on, cel- gram. One common view of the nature of the stimlular degeneration is not an indispensable part of ulus is that it is a hormonal substance. In support of this, it has been shown that in cotton ovules grown successful pollen tube penetration. By and large, the signal for pollen tube penetra- in culture, synergid degeneration occurs when a tion into the embryo sac is provided by a synergid. mixture of IAA and GA is added to the medium. As The question arises, what is the nature of this sig- in ovules of pollinated flowers, only one synergid nal? The most persistent hypothesis to account for in cultured, unpollinated ovules responds to the the directional growth of the pollen tube is that it is hormonal signal Qensen, Schulz and Ashton 1977). attracted to a chemical substance. In the absence of It has also been shown that supplementation of the direct evidence for the presence of a chemotactic medium with GA alone triggers the degeneration substance in the synergids, a speculative view is of the synergid in ovules of unpollinated flowers that the release of Ca2+, especially from the degener- (Jensen, Ashton and Beasley 1983). These experiating synergid, is associated with pollen tube pene- ments by themselves do not prove that the hortration. Jensen (1965b) visualized inorganic ash in mone directly mediates the synergid response, but the vacuoles and cytoplasm of microincinerated they alert us to such a possibility in any hypothesis sections of cotton ovules. The ash, which is largely to explain synergid2+degeneration. Another suggesabsent from the egg, is assumed to contain Ca2+. tion involving Ca and related elements is that their loss after pollen tube discharge reduces the More recently, high-energy X-ray dispersion analyosmotic pressure of the synergid, leaving the way sis and antimonate precipitation methods have
Fertilization: The Beginning of Sporophytic Growth
open for an efflux of water from the cell and for subsequent shrinkage of the cell (Chaubal and Reger 1992b). Since there are two synergids in most species investigated, the next question is whether degeneration is a random event or whether one of the synergids is more predisposed to degenerate than the other. Based on examination of a limited number of species, indeed it appears that synergid degeneration is a preprogrammed, nonrandom event. In Helianthus annuus, the synergids are isomorphic; however, one is found closer to the placenta than the other. Using this feature for orientation purposes, it was found that in 81% of the ovules examined, the degenerating synergid was the one aligned with the placenta (Yan, Yang and Jensen 1991a). In barley, Mogensen (1984) found that in one sampling there was a greater tendency for the synergid closer to the placental attachment of the ovule (proximal) to degenerate than for the distal one. In these instances, therefore, the force for degeneration is derived from the sporophytic tissues due to physiological differences between the two synergids, such as the transport of nutrients or erotropic substances through the placental vasculature to one of the synergids. On the other hand, in Linum usitatissimum, some kind of gametophytic control seems to be operating in determining which one of a pair of synergids degenerates (Russell and Mao 1990). Based on the orientation of synergids relative to the egg, there is a greater preference for the left than the right synergid to degenerate. Since the volume and the surface area of the filiform apparatus of the left synergid are significantly lower than those of the right, the model predicts a degree of control by intrinsic factors that affect synergid function. A particularly fascinating and important aspect of the race of the pollen tube into the synergid is the fact that sperm cells keep pace with the distal end of the growing pollen tube. The mechanism by which sperm cells move down the pollen tube continues to attract the attention of investigators even to this day, but no satisfactory hypothesis has emerged. Because sperm cells share a number of properties with the generative cell, including shape and the cytoskeletal framework, comments made in Chapter 8 about the movement of the generative cell in the pollen tube are germane here. Although sperm cells have their own endowment of microtubules, it is doubtful whether they function as a motility system. Cass (1973) found that isolated sperm cells of barley show little or no directional movement, which should not be the case if their
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microtubules are called into service. In view of the absence of microfilaments in the sperm, it comes as no surprise that no one has implicated them in gamete movement. Because of the negative evidence, it has been speculated that, like the generative cell, the sperm cells are propelled along the longitudinally disposed actin cytoskeletal elements of the pollen tube cytoplasm (J. Heslop-Harrison and Heslop-Harrison 1987). It is hard to justify the presence of microtubules in the short-lived sperm cells if they do not have a role in locomotion; it might be that sperm movement in the pollen tube is due to the combined efforts of the sperm microtubules and the actin system of the pollen tube. Once the pollen tube has made contact with the synergid, it grows some distance into this cell and readies itself to release sperm. Based on an examination of the cytoskeletal organization of the embryo sac of Plumbago zeylanica at various stages of pollen tube penetration, it has been proposed that positions of both microtubules and actin bundles change significantly to guide the pollen tube to the site of sperm delivery (B.-Q. Huang et al 1993). In other plants, the electron microscope has revealed several features of the final hours of the pollen tube and the life of the sperm as free cells. It is no accident that the first detailed description of the fate of the pollen tube contents in the synergid was provided in cotton, for this plant has proved to be remarkable for the highly regulated way in which fertilizationrelated events occur. In cotton, the arrest of pollen tube growth in the degenerating synergid is followed by the formation of a thick cap at the tip. The most dramatic event at this juncture is the appearance of a pore at the back of the tip through which the male gametes and the vegetative cell nucleus are released along with a generous amount of the pollen tube cytoplasm. Adding to the chaotic environment of the synergid is the discharge of polysaccharide spheres presumably constituting unused cell wall material (Jensen and Fisher 1968a). The force of the discharge, combined with changes in the physical properties of the discharged cytoplasm, apparently propels the vegetative cell nucleus and sperm cells to the chalazal end of the synergid, close to the egg nucleus. The end of the discharge is signaled by the formation of a callose plug over the pore, effectively preventing any cytoplasmicflowbetween the pollen tube and the synergid. In other plants, such as Capsella bursa-pastoris (Schulz and Jensen 1968b), Epidendrum scutella (Cocucci and Jensen 1969c), Petunia hybrida (van Went 1970c), Epilobium palustre (Bednara 1977),
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Quercus gambelii (Mogensen 1972), Plumbago zeylan-accomplished between the plasma membranes of the ica (Russell 1982), Plumbagella micrantha (Russell sperm and of the egg or the central cell, leading to the and Cass 1988), and Brassica campestris (Sumner formation of a bridge through which the sperm 1992), the discharge of the pollen tube occurs slides. The chalazal part of the egg, where the cell through a terminal aperture. In P. zeylanica, growth wall is incomplete, serves as the window where the of the pollen tube to the summit of the egg is often sperm fuses (Jensen and Fisher 1968a). In Petunia the prelude to pollen tube rupture. Sperm cells, hybrida, contact between the sperm and the female vegetative cell nucleus, and a small amount of cyto- reproductive cells is followed by cell fusion someplasm, which are released from the pollen tube, what analogous to the fusion of protoplasts (van find a place between the egg and the central cell Went 1970c). Regarding the fate of the male nucleus, (Russell 1982). Various mechanisms have been pro- some observations suggest that the chromatin of the posed to account for the rupture of the pollen tube, male nucleus becomes dispersed and indistinguishbut they seem incomplete in their explanations. able from the chromatin of the egg nucleus (Hu and One view is that as the pollen tube stops growing, Zhu 1979). A specific role for the degenerating synosmotic pressure built within it causes rupture at ergid in transporting sperm to the site of the egg and the weakest point. In P. zeylanica the pollen tube central cell has been described in Triticum aestivum. wall is relatively thin at the tip near the point of Transport is accomplished by a tubelike extension of rupture. Moreover, fibrillar elements near the aper- the degenerating synergid containing the pollen tube ture are less organized than in the proximal region, discharge and the sperm nuclei, which penetrates indicating both an apparent constraint in the syn- into the space between the egg, central cell, and the thesis of wall material and the onset of stress persistent synergid (Gao et al 1992a). (Russell 1982). Another view is that changes in oxySeveral events are played out in a compressed gen tension stop the growth of the pollen tube and time frame during gametic fusion in the synergidcause its rupture (Stanley and Linskens 1967). This less angiosperm Plumbago zeylanica. Immediately is based on the observation that low levels of oxy- after their release from the pollen tube, sperm cells gen promote bursting of pollen tubes in vitro and shed their investment of the inner plasma memin the style; whether pollen tubes experience a sim- brane of the vegetative cell and the incipient cell ilar stress in the embryo sac is not known. It has wall materials. At the same time, cell wall compoalso been suggested that the cessation of pollen nents of the egg are dispersed, thus exposing the tube growth and the rupture of its tip may be egg cell membrane. The stage is now set for caused by supraoptimal concentrations of Ca2+ in gametic fusion in which plasma membranes of the sperm cells become directly appressed to the the synergid (Chaubal and Reger 1990). plasma membranes of the egg and central cell. Fusion initiated at one location between the partic(iii) Fusion of Nuclei ipating cells spreads in a wavelike fashion through The observations terminating in sperm discharge the membrane and results in the transmission of lead to the consideration of the mechanism by which both nucleus and cytoplasm. Aggregates of actin gametic fusion is accomplished. The range of the found at the sperm delivery site between the egg search for this mechanism is wide and involves such and the central cell, designated as a corona, have aspects of cell behavior as movement, recognition, been assumed to cause the two sperm nuclei to and adhesion. The first issue concerns the actual align with their respective female nuclei (Russell transfer of sperm to the egg or the central cell. The 1983; B.-Q. Huang et al 1993). In the final act of two sperm cells are presumed to move separately, fusion, the sperm cell is incorporated into the cytoone to the egg and the other to the central cell. plasm of the egg or the central cell, and the plasma Electron microscopy has provided some pointers on membrane remaining from the fusion events dissithis, although it is not possible to determine the pates as fragments in the cytoplasm of the female mechanism that underlies the movement by inspec- cells involved. The events described here occur tion of electron micrographs alone. In Torenia fournieri within a time span of about 30 minutes. The actin (van der Pluijm 1964), the receptive synergid opens corona is also present between the synergids, egg, up a passage at a specific site at its chalazal end, pro- and central cell, and near the chalazal end of the viding direct access of the sperm to the egg and the degenerating synergid, during fertilization in IV. central cell. In terms of the end result, the transfer of tabacum (Huang and Russell 1994). The corona has the sperm cells to the egg and the central cell in cot- no counterpart in the embryo sac before one of the ton falls into the same type as T. fournieri. Fusion is synergids begins to degenerate.
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tribute to the nuclear envelope. During nuclear fusion, pockets of cytoplasm containing fragments of organelles are trapped in the nuclear material, but they are probably forced out prior to the complete fusion of the nuclei (Jensen 1964; Jensen and Fisher 1967). Membrane fusion events similar to those observed during sexual karyogamy have been described during the fusion of the male nucleus with the polar fusion nucleus in Capsella bursa-pastoris (Schulz and Jensen 1973), Spinacia oleracea (Wilms 1981b), and Triticale (Hause and Schroder 1987). As shown in maize, karyogamy in the central cell occurs between the sperm nucleus and the diploid polar fusion nucleus or with one of the haploid polar nuclei (M61, Matthys-Rochon and Dumas 1994). Tangible evidence for the completion of double fertilization in conventional embryo sacs is the presence of dark-staining oval structures, referred to as X-bodies, in the degenerate synergid. Although these structures were variously interpreted by early embryologists, present opinion favors the view that they are nuclei or fragments of nuclei, presumably of the vegetative cell and of the degenerate synergid (Fisher and Jensen 1969; Cass and Jensen 1970; van Went 1970c; Huang, Strout and Russell 1993). A vexing question in angiosperm embryology is whether cell fusions during double fertilization are random or preferential events. In other words, are there traceable differences between sperm cells that fuse with the egg, on the one hand, and with the polar fusion nucleus, on the other? Based on light microscopic studies, the idea that both sperm cells Impatiens glandulifera, and Tradescantia paludosa, the are structurally identical and, hence, there is no sperm nucleus is already in the interphase stage at preferential fusion, held center stage for several the time of fusion, and the first zygotic division fol- decades. The first dent in this model was adminislows without delay (Kapil and Bhatnagar 1975). In tered by Roman (1948), who provided genetic eviGnetum gnemon (Gnetales), the gametic nuclei pass dence for the existence of nuclear dimorphism in through the S phase and double their DNA content the sperm cells of maize. It was found that when Bbefore they begin to court each other and fuse chromosomes do not separate during sperm cell (Carmichael and Friedman 1995). formation, the sperm containing an extra set of BElectron microscopy has added a wealth of new chromosomes fuses in the majority of cases with details about karyogamy in a selected list of plants. the egg. Although not striking at first glance as a In cotton, the sperm nucleus may enter the egg and revolutionary insight, the significance of this obsertravel the circumference of the egg before align- vation in the context of double fertilization is that ment with its nucleus preparatory to karyogamy. in maize, the presence of supernumerary B-chroKaryogamy is initiated when the two nuclei come mosomes seems to confer a competitive edge on together and the bordering ER fuse at several certain sperm cells as true male gametes. A fluorespoints and join their outer nuclear membranes. cence in situ hybridization method developed by This results in the formation of bridges between the Shi et al (1996) using a B-chromosome-specific nuclei. Next, the inner layers of the nuclear DNA sequence to identify and track maize sperm envelopes also fuse and the nucleoplasm becomes cells containing B-chromosomes now makes it poscontinuous at several points. When a fusion sible to follow up this question. nucleus is formed, membranes of both nuclei conOne of the most provocative pieces of evidence Nuclear fusions, or karyogamy, between the sperm and egg nuclei and between the sperm and central cell nuclei are at the heart of double fertilization and constitute a major reason for sustained interest in the process itself. GerassimovaNavashina (1960) used the cyclic state of the nuclei as an elegantly simple criterion to identify two types of karyogamy involving the egg and sperm. In the most common type, known as the premitotic type, the sperm nucleus fuses immediately with the egg nucleus and, after a short hiatus, the zygote nucleus passes through S phase and continues through G2 and mitosis to complete the first zygotic division. Illustrative examples of this type of karyogamy are found in members of Asteraceae and Poaceae. Premitotic karyogamy has been nicely demonstrated recently (Figure 11.4) in isolated egg and zygote protoplasts of barley (Mogensen and Holm 1995). In the postmitotic type, frequently observed in members of Liliaceae, the male and female nuclei approach one another and remain in close proximity; then follows an increase in the size of the nucleus and nucleolus of the sperm. Although the nuclei enter mitosis independently, a common metaphase plate is formed, leading to the formation of a zygote nucleus. The underlying basis of postmitotic karyogamy in the gymnosperm Ephedra trifurca (Gnetales) entails only the completion of S phase of the cell cycle by the gametic nuclei during the period of courtship before fusion occurs (Friedman 1991). Intermediate situations also prevail, depending upon the duration of karyogamy. For example, in Mirabilis jalapa,
Pollination and Fertilization
Figure 11.4. Premitotic karyogamy in isolated egg and zygote protoplasts of barley, (a) Nomarski optics photograph of an unfertilized egg. (b) Unfertilized egg, showing the fluorescence of DAPI-stained nucleus, (c) Egg showing the sperm nucleus beginning to fuse, (d) Fertilized egg showing the brightly fluorescing male chromatin. (e) Zygote showing the dispersion of the male and female chromatin.
in support of preferential fusion was provided in Plumbago zeylanica (Russell 1985). It was pointed out earlier that the paired sperm cells of P. zeylanica display heteromorphism in size, shape, and organelle contents. Because plastids are virtually absent from the cytoplasm of the sperm that is in
(f) Zygote before beginning of mitosis, (g) Zygote at the early prophase stage, (h) Zygote at the metaphase stage. (i) Late telophase stage of zygote mitosis, en, egg nucleus; sn, sperm nucleus. Scale bar = 5 |jm. (From Mogensen and Holm 1995. Plant Cell 7:487-494. © American Society of Plant Physiologists. Photographs supplied by Dr. H. L. Mogensen.)
physical association with the nucleus of the vegetative cell, it is possible to use paternal plastids as markers to follow the fate of the two male gametes in the embryo sac. By painstaking identification and counting in electron microscopic profiles of the number of plastids of paternal origin in the egg after
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fertilization, it was established that the plastid-rich, (iv) Cytoplasmic Inheritance mitochondrion-poor sperm fuses with the egg cell. Because fusion of the plastid-impoverished sperm The transfer of male cytoplasm along with the with the central cell is as frequent as fusion of the nucleus during fusion of the sperm with the egg plastid-rich sperm with the egg, this brings us tan- raises an important question: Is the inheritance of talizingly close to considering the occurrence of a plastids and mitochondria uniparental or putative recognition event at the gametic level. In biparental? Chapter 3 described several instances of analogy to animal systems, this might involve the the first haploid pollen mitosis, during which the presence of different cell surface molecules on the generative cell is cut off with few or no plastids or two sperm cells that will recognize the egg and the mitochondria. This led to the view that in the majorpolar fusion nucleus, respectively. Although this ity of plants, cytoplasmic male DNA is not inherappears to be an attractive hypothesis, we should ited, which results in uniparental maternal not lose sight of the fact that it is based on the study organelle inheritance. In a few instances in which of one unusual species and that a great many prob- sperm cells are endowed with plastids and mitolems have to be overcome before gamete surface chondria, mechanisms also operate to exclude them recognition substances are identified. P. zeylanica from the egg prior to fertilization. Rarely, plastids has made an interesting contribution to our under- and mitochondria are transferred from the sperm to standing of the structural basis of double fertiliza- the egg during fertilization; however, as the fertiltion, one that is far richer than has been apparent ized egg begins to divide, it displays characteristic from studies of other angiosperms. uniparental inheritance. How does this happen? Mogensen (1996) has reviewed the elaborate Little is known about the existence of any cryptic pattern of recognition system on the female side. ways by which cytoplasmic organelle transmission Pennell et al (1991) have demonstrated the presence is restricted or enhanced during and following of a plasma membrane-bound arabinogalactan pro- genetic recombination in angiosperms. The classitein epitope in the paired sperm cells and in the egg cal method of excluding sperm cytoplasm from the cell, but not in the central cell, of Brassica napus; the egg at fertilization is by allowing passage of only differential accumulation of the marker protein in the nucleus into the egg cell, leaving the cytoplasm the egg and central cell may suggest a role for the outside. This is probably the case in cotton, where arabinogalactan protein in female gametic cell no trace of the sperm cytoplasm in any form or shape is detected in the egg (Jensen and Fisher recognition. To round out this account, brief mention should 1967,1968a). Ultrastructural evidence for the exclube made of reports of fusion of a second sperm sion of sperm cytoplasm and organelles during nucleus with egglike counterparts in the female karyogamy has also been reported in barley gametophytes of Ephedra and Gnetum. These fusion (Mogensen 1982, 1988). In the barley embryo sac, events have been billed as double fertilization; in E. an enucleate cytoplasmic body, approximating the trifurca, in which it has been examined in detail, the size of a prefusion sperm cell and containing a genproduct of second fertilization yields an embryo erous number of mitochondria and a limited numafter a brief free-nuclear interlude (Friedman 1990, ber of plastids, is frequently observed within the 1992; Carmichael and Friedman 1995). The direct degenerate synergid adjacent to a recently fertilanalogy of the situation in the gymnosperms to ized egg cell. This body has been interpreted as the that in angiosperms may be too facile a compari- entire male cytoplasm left out after fusion of the son, and the view of a likely origin of the egg and sperm. Since only one cytoplasmic body is endosperm in angiosperms from an embryolike observed in each postfertilization embryo sac structure in gymnosperms may be far-fetched. It examined, it is believed that uniparental inheridoes not take into account the fact that the product tance is restricted to the zygote and that the of the second fertilization event does not produce endosperm receives the paternal cytoplasm. What the nutritive tissue that is the hallmark of the causes the separation of the intact cytoplasm from angiosperms. In the absence of evidence to refute the nucleus is difficult to comprehend, although that the second fusion event is another manifesta- cytoplasmic degeneration may certainly contribute tion of polyembryony, for which gymnosperms are to this effect. In Populus deltoides, the sperm cells notorious, the thesis that double fertilization, in develop large vacuoles during their descent into the sense we understand the process in the pollen tube, and it is believed that this is the sigangiosperms, originated in gymnosperms cannot nal that leads to the separation of the cytoplasm from the nucleus at the time of fertilization be supported.
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(Russell, Rougier and Dumas 1990). An unusual way of eliminating the male cytoplasm from the egg probably occurs in spinach in which plasma membranes of the two sperm cells initially fuse together to form a binucleate cell. It has been claimed that fusion of the male nuclei with the egg nucleus and with the polar fusion nucleus of the central cell is followed by sealing of the plasma membranes, thus excluding any remaining cytoplasm from the female cells (Wilms 1981b). Just as the complete elimination of the sperm cytoplasm at the time of fertilization offers an explanation for maternal organelle inheritance, so also might a process of progressive cytoplasmic diminution during sperm cell maturation explain maternal inheritance. Mogensen and Rusche (1985) first reported that during maturation of barley sperm cells, lateral projections containing mitochondria are frequently pinched off, resulting in a conspicuous reduction in the surface area, volume, and mitochondrial number of the sperm. In Nicotiana tabacum, cytoplasmic diminution, which occurs in the sperm cells as they descend into pollen tubes, is accomplished by the formation of enucleate cytoplasmic bodies, the spontaneous vesiculation of the tip of cytoplasmic extensions, and the accumulation of vesiculate bodies in the periplasm of the sperm cells (Yu, Hu and Russell 1992). It is difficult to reconcile these observations with those of a subsequent work, which showed that during fertilization in N. tabacum, male cytoplasm including heritable organelles is transmitted into the female target cells (Yu, Huang and Russell 1994). Since sperm cells do not produce nuclei denuded completely of their enveloping cytoplasm, it is likely that processes such as those described in barley and tobacco, combined with cytoplasmic elimination at the time of fertilization, might offer an explanation for maternal organelle inheritance in the majority of plants in both species, with the caveat that occasional biparental inheritance also occurs. Transmission of a part or all of the cytoplasm of the sperm cell in the act of fertilization certainly is the basis of biparental or paternal inheritance of organelles. In Chapter 3 it was reported that in some plants the generative cell is cut off with an appreciable number of plastids, which suggests that the plastids are transmitted to the sperm and subsequently into the egg cell during gametic fusion. This seems to be true where the fate of the sperm cell has been followed electron microscopically. Very clearcut, indeed, is the evidence for the transmission of sperm cytoplasm into the egg and central cell in
Pollination and Fertilization
Plumbago zeylanica, in which, based on size and fluorescent-staining differences, it was possible to discriminate between plastids and mitochondria of sperm cell origin from those present in the egg at the time of fertilization (Russell 1980, 1987; Sodmergen et al 1995). Electron microscopic evidence has augmented genetic data to help explain certain cases of uniparental, paternal inheritance of organelles. One such case is Medicago sativa, in which RFLP for plastid DN A has indicated a predominant paternal inheritance of this organelle in reciprocal crosses between two subspecies (Schumann and Hancock 1989). Paternal inheritance is also correlated with a high plastid content in the generative cell, although there are no significant differences in plastid abundance between genotypes known to differ in their male plastid transmission pattern (Zhu, Mogensen and Smith 1990, 1991). Likewise, there is no consistent correlation among genotypes between the mean number of plastid DNA aggregates (nucleoids) in the generative cell and the strength of male plastid transmission (Shi et al 1991). A quantitative, threedimensional analysis of the plastid distribution patterns in egg cells of genotypes designated as weak plastid transmitters or strong plastid transmitters has provided an explanation of this paradox. Apparently, eggs of weak plastid transmitters are characterized by a largely micropylar plastid distribution, whereas plastids of the strong females are positioned in the chalazal part of the egg. This would ensure that following the first division of the zygote, the plastid population of the weak females is segregated to the basal micropylar cell, which becomes the transient suspensor, and the apical organogenetic part of the embryo will be blessed with the plastids delivered by the sperm. On the other hand, in strong plastid transmitters, the apical cell, which inherits the plastids at the time of the first division of the zygote, will perpetuate maternal or biparental plastid transmission (Zhu, Mogensen and Smith 1993; Rusche et al 1995). This viewpoint assumes that fertilization is a passive event and that organelles stay put in whichever locations they find themselves at the time of nuclear fusion. As described in Chapter 12, electron microscopic studies have revealed wholesale rearrangement in the distribution of organelles in the egg cells of several plants following fertilization. A strong bias in favor of paternal inheritance of plastids also prevails in the interspecific cross Daucus muricatus X D. carota. The basis for this is
the presence of an exact replica of restriction fragments of D. carota plastid DNA in self-pollinated Fl
Fertilization: The Beginning of Sporophytic Growth
plants (Boblenz, Nothnagel and Metzlaff 1990). This is attributed to the presence of relatively few intact plastids in the egg of D. muricatus compared to the other species and to the presence of degenerate plastids (Hause 1991). However few in number the plastids may be, how these plastids are excluded from the zygote is an unsolved problem. Among the ideas suggested are the domination of paternal plastids and eventual elimination of the maternal plastid genome, differential plastid replication with the paternal plastids' holding an advantage, and the wholesale degeneration of maternal plastids. Investigations that take these ideas into account seem to be necessary to provide complete structural evidence for the genetic data for paternal plastid inheritance. In recognition of the fact that some male plastids invariably find their way into the egg cytoplasm at the time of fertilization, efforts have been made to explain plastid behavior in the zygote that results in uniparental inheritance. According to Zhu, Mogensen and Smith (1991), who have collated this information, elimination of male plastids from the zygote occurs by (a) differential degradation of male plastid DNA within the zygote; (b) recombination between different plastomes in the zygote, with subsequent DNA repair using the strong plastome of the female parent as the template; (c) differential multiplication of plastids of the two parents within the zygote and proembryo, with the maternal plastids' holding an edge; and (d) segregation of maternal plastids to the apical cell and paternal plastids to the basal cell in embryo types in which the apical cell of the two-celled embryo becomes the organogenetic part and the basal cell becomes a short-lived suspensor. Perhaps symptomatic of the primitive state of our understanding of the fate of the paternal plastids following fertilization is the fact that there are no markers to follow them in the chaotic environment of the zygote. 3. IN VITRO APPROACHES TO THE STUDY OF FERTILIZATION In the reproductive phase of angiosperms, fertilization and seed set are modulated by several external environmental and internal genetic factors. Among the former are female receptivity, the age and quality of pollen grains, and properties of the ambient atmosphere. In traditional plant breeding programs involving incompatible pollinations, prefertilization blocks such as the failure of pollen grains to germinate on the stigma, physiological disturbances in the directional growth of pollen
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tubes through the style toward the embryo sac, and failure of fertilization itself often thwart seed set. When transfer of beneficial alien genes across interspecific and intergeneric barriers is attempted, such as in wide hybridizations, it is seen that in spite of normal pollination and fertilization, deleterious genes carried forward into the zygote cause embryo lethality and seed collapse. Since success in agricultural practices would depend upon devising techniques to control reproductive processes in plants to our advantage, methods to overcome these natural bottlenecks to sexual recombination have attracted a great deal of attention. As described in Chapter 10, advances made in our understanding of the molecular biology of incompatibility offer potential for the development of methods for overcoming this barrier to crossability in plants. Methods to overcome embryo abortion in wide hybrids are briefly considered in Chapter 12. The purpose of this section is to provide an overview of the in vitro methods that have been developed to study fertilization in angiosperms under controlled conditions. Detailed protocols for most of the in vitro approaches currently in use are given by Yeung, Thorpe and Jensen (1981). (i) Intraovarian Pollination The primary objective of in vitro approaches to study fertilization is to secure the encounter of the egg and sperm in a relatively uncomplicated way that bypasses pollination and pollen tube growth in the style. An extensive discussion would be required to recount all of the early attempts in the experimental manipulation of the fertilization process. Only the highlights can be described here; refer to the brief review by Tilton and Russell (1984) for a historical perspective of this work. In one of the first published full investigations of this kind, Bosio (1940) was able to induce fertilization in ovules of three species of Paeonia by combining excision of the style with actual deposition of pollen grains on the upper part of the ovary. Because it effectively bypasses stigmatic pollination and stylar growth of pollen tubes in a distinctive way, the technique is christened as "intraovarian pollination." Unfortunately, neither the technique nor the observations sparked much interest until Kanta (1960) used a modified method to induce fertilization and seed set in Papaver rhoeas. The experimental design involved emasculating receptor flowers before anthesis and injecting into the ovary small quantities of a suspension of pollen grains prepared in boric acid. This
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resulted in the germination of pollen grains, in the the filiform apparatus of the synergid as it does growth of pollen tubes and their entry into the during in vivo fertilization. embryo sac, and eventually, in fertilization. Results Test-tube fertilization also occurs by crossing acquired in terms of seed set demonstrated the suc- interspecific and intergeneric barriers. One of the cess of intraovarian pollination in P. rhoeas, as well most convincing examples is fertilization of culas in other members of Papaveraceae such as P. tured ovules of Melandrium album with pollen from somniferum, Argemone mexicana, A. ochroleuca, and M. rubrum, Silene schafta, and S. tatarica, all of Eschscholzia californica. Although reciprocal crossesCaryophyllaceae (Zenkteler 1967). Indeed, this between the diploid A. mexicana and the tetraploid appears to be a choice method for raising interspeA. ochroleuca failed due to inhibition of pollen tube cific hybrids because hybrid embryos complete growth in the style, intraovarian pollination offered their full-term development. Cultured ovules of M. the potential for eliminating this roadblock to fer- album are also found to favor pollen grains of other tilization when either species was used as the genera of Caryophyllaceae such as Cerastium female parent (Kanta and Maheshwari 1963a). An arvense and Dianthus serotinus, and fertilization extension of this method is to inject sperm directly ensues; however, the competence of zygotes to into the embryo sac; this approach holds promise form full-term embryos is found to fade after a few in plants where the sperm cells do not reach the divisions (Zenkteler, Misiura and Guzowska 1975). embryo sac after intraovarian pollination. Beginning with the works of Sladky and Havel (1976) and Gengenbach (1977a), a great effort has been made to identify the conditions for maxi(ii) Test-Tube Fertilization mizing fertilization and kernel development A step toward a more controlled encounter of under in vitro conditions in maize. These studies, the egg and sperm was achieved by culturing which basically have involved culturing pieces of unpollinated receptive ovules of P. somniferum in a the cob and pollinating the exposed silk or dustnutrient medium and dusting them with pollen ing the nucellar tissue with pollen grains, have grains. During the ensuing period of two to three examined the influence of donor plant genotype weeks, pollen germination, pollen tube growth, fer- (Gengenbach 1977b; Bajaj 1979; Higgins and tilization, embryogenesis, and endosperm develop- Petolino 1988), the ratio of ovary to cob tissue ment occur, as attested by the transformation of (Sladky and Havel 1976; Higgins and Petolino cultured ovules into mature seeds that survive on 1988), pollen quantity (Raman, Walden and the original medium. This was confirmed by histo- Greyson 1980), explant size (Higgins and Petolino logical preparations of ovules that established 1988), silk length (Dupuis and Dumas 1989), tissue beyond doubt the occurrence of double fertilization continuity of the silk (Booy, Krens and Bino 1992), in the test tube; hence, in analogy to animal sys- stage of spikelet development (Dupuis and Dumas tems, the technique is designated as "test-tube fer- 1990a), and temperature stress (Mitchell and tilization" (Kanta, Ranga Swamy and Maheshwari Petolino 1988; Dupuis and Dumas 1990b) on fertil1962). Using these general guidelines, the protocol ization frequency and seed set. Using this system, it was also successfully extended to other members has been shown that compared to the mass pollinaof Papaveraceae, as well as to members of other tion that occurs in nature, no more than a single families (Kanta and Maheshwari 1963b; Kameya, pollen grain is required to affect fertilization of an Hinata and Mizushima 1966; Zenkteler 1967; ovule, although the frequency of successful fertilZubkova and Sladky 1975). In a modified version of ization events varies greatly in different trials the protocol, whole pistils were cultured and polli- (Raman, Walden and Greyson 1980; Hauptli and nated in vitro with results similar to those obtained Williams 1988; Kranz and Lorz 1990). Since pollen with cultured ovules; this is analogous to conducttubes are able to travel a shorter distance within the ing stigmatic pollination in vitro. The technique style under in vitro conditions to affect fertilization was successful with, among others, Petunia violacea than in vivo, it appears that sperm cells do not (Shivanna 1965), Nicotiana tabacum (Dulieu 1966), undergo an extended period of maturation in the N. rustica (Rao and Rangaswamy 1972), Antirrhinum majus (Usha 1965), Dicranostigma pollen tube to become fusion competent (Dupuis franchetianum (Papaveraceae) (Rangaswamy and and Dumas 1989). Havel and Nova~k (1981) used Shivanna 1969), and Papaver nudicaule (Olson and genetic color characters of embryos and kernels to Cass 1981). In the last-mentioned example, the demonstrate double fertilization in vitro. The pollen tube enters the embryo sac by growing into development of kernels after in vitro pollination is not proof that the process of double fertilization
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has occurred or that subsequent developmental (iii) In Vitro Fertilization programs have been successful, because occasional absence of embryos and abnormalities in From the developmental point of view, in vitro endosperm development are noted in apparently fertilization with isolated, single gametes presents normal-looking kernels (Schel and Kieft 1986). In an ideal opportunity to analyze karyogamy at the this case, the lesion is not in the fertilization level of the single cell, uninfluenced by the presprocesses but in the development of the embryo and ence of other cells. From the long-range point of endosperm. view, in vitro fertilization will provide a route for One aspect of test-tube fertilization technique to direct transformation of the egg using external be emphasized here concerns its potential use in sources of DNA and for transmission of alien cytoeliminating self-incompatibility barriers. In the pro- plasm to study organelle inheritance. Using the tocol used to overcome self-incompatibility in methods available for isolation of embryo sacs and Petunia axillaris, a placenta with attached ovules, sperm cells, a beginning has been made in the together with a short length of the pedicel, was cul- study of the conditions necessary for single gamete tured and the ovules were pollinated by dusting fusion and culture of artificial zygotes in maize. them with pollen. The result was prompt germina- The initial strategy in this work is to use cleverly tion of pollen grains, rapid growth of pollen tubes, engineered microcapillary techniques to select fertilization of the egg within about two days after sperm cells individually from ruptured pollen pollination, and eventual seed set (Rangaswamy grains and to select eggs and other component cells and Shivanna 1967). Additional manipulations from enzymatically treated embryo sacs. Selection have established that cultured ovules of self-incom- is followed by the transfer of gametes to a micropatible plants do not show any preference to the drop of a fusion medium (0.55 M mannitol) placed pollen grains they encounter, as long as they belong on a coverslip; pairs of gametes are fused electrito the same species. For example, when ovules on cally. By this method, fusion has been accomone placenta in the ovary are left unpollinated and plished not only between isolated, single egg and those on another placenta in the same ovary are pol- sperm cells, but also between single sperm cells linated with self- or cross-pollen grains, the unpolli- and single synergids and central cells. The nated ovules wilt, whereas fertilization and seed set egg-sperm fusion products, when nurtured in a occur in both selfed ovules and crossed ovules. The microculture surrounded by a suspension of feeder efficacy of self- and cross-pollinations for fertiliza- cells of maize embryo origin, develop into multition in vitro was demonstrated by the absence of cellular structures (Kranz, Bautor and Lorz 1991a, differences in seed set when ovules on one placenta b). Electrofusion between isolated male and female are selfed and those on another placenta in the same gametes of wheat resulting in multicellular strucovary are crossed (Rangaswamy and Shivanna tures has also been reported (Kovacs, Barnabas and 1971a). In yet another approach, when stigmatic Kranz 1995). In an alternative method, developed pollination and placental pollination are performed without the application of electrical pulses (Figure on the same pistil in culture, the former is found to 11.5), the essential environment for the egg and lead to seed set only with cross-pollination, whereas sperm cell of maize to fuse was found to be a with the latter, both self- and cross-pollinations are medium containing a high concentration of CaCl2 found to be equally effective (Rangaswamy and at pH of 11.0 or a medium containing mannitol and Shivanna 1971b). These protocols are collectively CaCl2 (Faure, Digonnet and Dumas 1994; Kranz designated as "placental pollination." Placental pol- and Lorz 1994). Later studies showed that the first lination has also been successfully used to over- few rounds of division of the artificially created come incompatibility barriers in P. hybrida (Niimi zygote to form the multicellular mass and the 1970; Wagner and Hess 1973), Brassica campestris transformation of the mass into an embryo bear (Zenkteler, Maheswaran and Williams 1987), and obvious similarity to in vivo development of the zygote. Formation of plants occurred by germinaLilium longiflorum 0anson 1993). tion of the bipolar embryos and, occasionally, by Zenkteler (1990) has compiled a list of plants in polyembryony and by organogenesis from the which fertilization and seed development have multicellular mass (Kranz and Lorz 1993; Kranz, been achieved by the aforementioned methods. It von Wiegen and Lorz 1995). should be noted that since fertilization events occur in eggs enclosed within ovules, these experiments What is the possibility that the plants regenerfall short of a true in vitro fertilization system as ated are the products of spontaneous division of understood in animal embryology. the egg in the absence of fusion with a sperm? This
316
Pollination and Fertilization
Figure 11.5. Phase contrast micrographs of in vitro adhesion and fusion between isolated egg (EC) and sperm cells (SC) of Zea mays. Adhesion between gametes occurred 1 minute 50 seconds after transfer to a fusion medium containing 5 mM CaCI2 (a). Progress of gametic fusion is shown in sequences (b) to (i) taken at intervals over a period of 9 seconds, as indicated on the lower right corner. After fusion
(i), the sperm nucleus (SN) is still visible in the egg cell protoplast. Scale bar = 15 urn. (Reprinted with permission from J.-E. Faure, C. Digonnet and C. Dumas 1994. An in vitro system for adhesion and fusion of maize gametes. Science 263:1598-1600. © 1994. American Association for the Advancement of Science. Photograph supplied by Dr. C. Dumas.)
does not appear to be the case, as karyological, phenotypic, and segregation analysis have established that the regenerated plants are indeed hybrids of both egg and sperm parents. Electron microscopic analysis has shown that the fusion of the egg and sperm culminating in karyogamy involves diverse activities that occur in a regimented way at their own times after cell contact is established (Faure et al 1993; Kranz, von Wiegen and Lorz 1995). Of further interest are the increases in membrane-bound Ca2+, calmodulin, ER, and mitochondria in the zygote and in the deposition of cellulose around it (Tirlapur, Kranz and Cresti 1995). Suggestive of the existence of a block to polyspermy at the gamete level is the finding that the addition of extra sperm cells in the vicinity of fertilized eggs does not lead to their penetration of, or fusion with, the egg (Faure, Digonnet and Dumas 1994). A small num-
ber of isolated egg cells were found to divide in the absence of fertilization when activated by a pulse of a high concentration of 2,4-D, although the microcalluses produced did not regenerate plants. Divisions were also induced in isolated maize egg cells after fusion with sperm of other members of Poaceae, such as Coix lacryma-jobi, Hordeum vulgare, Sorghum bicolor, and Triticum aestivum (Kranz, von
Wiegen and Lorz 1995). These observations not only show that it is possible to break down species barriers in the natural system of cross-incompatibility but also pave the way for interesting experiments for the artificial control of fertilization. Alternative methods to study the intimate details of fertilization under in vitro conditions are observations of living embryo sacs isolated enzymatically from pollinated ovules, and microinjection of active sperm nuclei into the egg and central cell
Fertilization: The Beginning of Sporophytic Growth
317
of isolated embryo sacs (Zhou 1987a; MatthysRochon et al 1994). We have a long way to go before in vitro fertilization in angiosperms will become a routine procedure that can be manipulated without the aid of artificial fusion methods such as electrofusion. Considering the state of the art only a few years ago, the stepwise progress made to facilitate the encounter of egg and sperm for observation and analysis is impressive. As fertilization in angiosperms is most certainly mediated by complementary receptor sites on the sperm and the egg plasma membranes, the stage is now set for the exploration of the nature of these receptor molecules. Streamlining of the in vitro fertilization system will also lead to a generalized picture of the mechanisms that block polyspermy, an area in which the evidence is too flimsy for any firm conclusions.
that divided in the embryogenic pathway was not identified with any degree of certainty. Although it seems clear from later investigations that in some cases embryos originate by the division of the unfertilized egg, evidence for their origin from the synergids and antipodals has been circumstantial (Ferrant and Bouharmont 1994). Compared to pollen embryogenesis, gynogenesis of limited value in developmental and molecular studies due to the unpredictable nature of the response of cultured explants and the difficulty of tracing the origin of embryos to specific cells of the embryo sac. 4. GENERAL COMMENTS
It is apparent from the foregoing discussion that a multifaceted approach can be of fundamental importance in the analysis of fertilization in angiosperms. The establishment of the role of the degenerating synergid, formulation of the concept (iv) Gynogenesis of the male germ unit, isolation of relatively pure Gynogenesis refers to the activation of cells of populations of male and female gametes, and the embryo sac to form embryos in the absence of demonstration of in vitro fertilization stand as landfertilization and is a process with dual conse- marks in the progress made in this field during the quences, namely, the substitution of other stimuli last three decades. In a way that is unique to for fertilization and the construction of a complex angiosperm reproduction, the phenomenon of douembryo and plant. Gynogenesis is generally facili- ble fertilization is beginning to make sense at the tated by the in vitro culture of the flower, ovary, and molecular level when we consider the evidence, ovule before pollination. Thus, in contrast to the in albeit limited, in favor of sperm dimorphism. That vitro methods described earlier, which promote an one sperm distinguished by certain ultrastructural egg-sperm encounter, gynogenetic methods essen- features almost always seeks out the egg cell for tially seek to prevent fertilization. Embryos that are recombination indicates that cell surface molecules formed in the absence of fertilization are haploids may determine this preferential union. at the cellular level, and for this reason, gynogeneThe successful demonstration of in vitro fertilsis is often considered, along with pollen embryo- ization does not necessarily make the task of genesis or androgenesis (Chapter 17), as a means of explaining the phenomenon of gamete recognition obtaining haploid plants in quantity for breeding easy. There are few, if any, available facts concernpurposes. ing cell surface ligands and receptors that must be Culture of unpollinated ovaries of barley in a involved in gamete recognition and fusion. The mineral-salt medium led to the first reported next logical step in the development of this imporinduction of gynogenetic embryos and plantlets tant field of plant embryology is to establish in (San Noeum 1976). This work was followed by vitro conditions involving minimum experimental reports of plant regeneration from cultured unpol- manipulations for the fusion of isolated egg and linated flowers, ovaries, or ovules of wheat, rice, sperm cells in a few model systems. It is clear that barley, maize, tobacco, lily, sunflower, beet root, our ability to manipulate gametes of angiosperms Gerbera jamesonii, Lactuca saliva (lettuce; Asteraceae), has improved so rapidly that it is unrealistic to preCucurbita pepo, and Populus sp. (San and Gelebart dict the rate of progress in the biochemical and 1986; Yang et al 1986; Zhou et al 1986b). In these molecular characterization of gametes. It is likely to investigations, the origin of embryos and plantlets be determined as much by the availability of new in the absence of fertilization was attributed to the and simple techniques for manipulation of gametes haploid cells of the embryo sac, but the specific cell as by the interest and expertise of the researchers.
SECTION III
Zygotic Embryogenesis
Chapter 12
Developmental Biology of the Endosperm 1. Histology and Cytology of the Endosperm (i) Structure and Development of the Endosperm (ii) Cellularization of the Endosperm (iii) DNA Amplification 2. Nutritional Role of the Endosperm (i) Endosperm Haustoria (ii) Chemical Composition of the Endosperm (iii) Embryo Abortion and Endosperm Development 3. Gene Action during Endosperm Ontogeny (i) Endosperm Mutants (ii) Synthesis of Housekeeping Proteins 4. Synthesis and Accumulation of Endosperm Storage Reserves (i) Mutants Defective in Starch Synthesis (ii) Storage Protein Synthesis 5. Analysis of Storage Protein Gene Expression (i) Maize Storage Protein Gene Expression (ii) Barley Storage Protein Gene Expression (iii) Wheat Storage Protein Gene Expression (iv) Storage Protein Gene Expression in Rice and Oat 6. Cloning and Characterization of Endosperm Storage Protein Genes (i) Zein Genes (ii) Hordeins and Other Genes from Barley (iii) Gliadin and Glutenin Genes (iv) Rice Glutelin and Other Genes 7. General Comments
The primary endosperm nucleus, born out of fusion of the second sperm cell with the polar fusion nucleus during double fertilization, is the starting point for the development of the endosperm. Repeated divisions of this nucleus within the confines of the central cell generate a
cluster of free nuclei or a cellular tissue known as the endosperm. Although the endosperm is reckoned as a triploid tissue in the vast majority of angiosperms, the ploidy level of the fusion nucleus obviously determines the final nuclear constitution of this tissue. Theoretically, the ploidy level of the endosperm might be expected to vary from diploid in the Oenothera type of embryo sac, to pentaploid in the Penaea, Fritillaria, and Plumbago types, and as high as 9N in the Peperomia type. Although both diploid and pentaploid endosperms have been described, it is, however, doubtful whether a 9N endosperm is formed as a permanent tissue in any seed types investigated. The endosperm has been studied from a number of viewpoints using light and electron microscopy, tissue culture, and biochemical and molecular techniques. For many years, the patterns of division of the primary endosperm nucleus, the morphological nature of the tissue formed, and the interaction of the tissue with the developing embryo were of prime concern to investigators whose studies have been an important factor in revealing that the developing embryo is nourished by the food materials of the endosperm. During the last two decades, a large part of the research on the endosperm has been devoted to the molecular biology of this tissue, and significant information about the regulation of synthesis and the accumulation of storage materials in the endosperm has been obtained. All through these periods of activity, the endosperm of cereal grains, as a repository of reserve food materials used in human and animal nutrition, has been subject to extensive biochemical investigations. It is perhaps not surprising, therefore, that the multifaceted research to improve the quality of our cereal grains that is conducted in laboratories all over the world is 321
322 in reality concerned with the control of endosperm development. Apart from the essential interest in it as a byproduct of fertilization, the endosperm presents basic problems relevant to our understanding of progressive embryogenesis. The objective of this chapter is to consider the endosperm from such aspects that are germane to its function in the developing seed following double fertilization. First, attention will be paid to the period of division and differentiation of the primary endosperm nucleus. Second, there is a considerable amount of evidence showing that genetic characters play a major role in determining the final form of the endosperm in certain plants. The various genetic factors controlling endosperm development will be briefly considered. Third, information will be presented concerning gene expression pattern in the endosperm, with emphasis on the synthesis and accumulation of storage proteins. Much of the early work on the morphology, cytology, and genetics of the endosperm has been considered by Brink and Cooper (1947b) in an exhaustive article. Reviews of relatively recent vintage are by Bhatnagar and Sawhney (1981) and Vijayaraghavan and Prabhakar (1984) on endosperm developmental morphology; Olsen, Potter and Kalla (1992) on the cytology and molecular biology of cereal endosperms; Lopes and Larkins (1993) on gene regulation; and Shewry (1995) on biochemistry. These are immensely challenging topics addressed by a considerable accumulation of literature, and so some degree of subjective judgment has gone into the selection of material to be discussed here.
1. HISTOLOGY AND CYTOLOGY OF THE ENDOSPERM The ontogeny of the endosperm has been followed in many different species to provide a descriptive basis for future biochemical and molecular investigations. These studies have reinforced the view that the endosperm is a relatively simple and amorphous tissue marked by the presence of only a few differentiated cell types. In spite of the fact that fusion events during double fertilization are almost simultaneous, it is generally acknowledged that the division of the primary endosperm nucleus takes precedence over that of the zygote. Little is known about the signal transduction system that potentiates the division of the primary endosperm nucleus ahead of the zygote or of the factors that induce the development of the primary endosperm nucleus into a compact tissue.
Zygotic Embryogenesis
Estimates of the development of the endosperm prior to division of the zygote range from the production of a few isolated nuclei to the formation of a fully developed tissue. A quantitative analysis of mitotic activity in the primary endosperm nucleus and the zygote in Triticum aestivum is particularly revealing of the shift in the balance of competing genomes. In plants reared at 20 °C, the primary endosperm nucleus begins to divide as early as 6 hours after pollination, whereas the division of the zygote is delayed until about 22 hours. The primary endosperm nucleus completes at least four rounds of nuclear divisions by the time the zygote divides, so that by 24 hours after pollination, at least 16 free endosperm nuclei are drifting randomly in the central cell around a two-celled proembryo. By the fifth day after pollination, the endosperm has already amassed more than 5,000 cells, in contrast to little more than 96 cells in the proembryo (Bennett et al 1973b). The same general trend in the rates of nuclear or cell doubling in the developing endosperm and embryo is maintained in other species of Triticum, in Hordeum vulgare, H. bulbosum, and Secale cereale, and in several genotypes of Triticale, with differences of similar magnitude in their cell numbers resulting (Bennett, Smith and Barclay 1975). To the extent that the endosperm possesses some nutrient value even at this early stage in its ontogeny, the interval between the division of the primary endosperm nucleus and the division of the zygote has significance in the nutrition of the zygote. Although the diploid polar fusion nucleus divides spontaneously in apomictic species, autonomous division of this nucleus may also occur in cultured, unpollinated ovules of certain amphimictic angiosperms (M61, Betka and Wojciechowicz 1995). Ohad et al (1996) have isolated a female gametophyte mutation in Arabidopsis termed fertilization-independent
endosperm (fie) that allows for replication of the diploid polar fusion nucleus and endosperm development without fertilization. Continued nuclear divisions in the central cell without the trigger of fertilization argue for a general molecular explanation based on cell cycle control of the fusion nucleus.
(i) Structure and Development of the Endosperm
The patterns of division of the primary endosperm nucleus have served as the basis for a simple classification of endosperms into nuclear, cellular, and helobial types. In the nuclear type, the
Developmental Biology of the Endosperm
323
Figure 12.1. Endosperm development in Ranunculus sceleratus. (a) Longitudinal section of the ovule showing the embryo sac surrounded by two to three layers of nucellar cells and a single layer of cells of the integument. The vacuole is lined by a thin layer of endosperm cytoplasm containing free nuclei, (b) A whole mount of the embryo sac containing 128 endosperm nuclei. Arrows point to walls in
the micropylar region, (c) In this longitudinal section of the ovule, the endosperm has been completely partitioned into uninucleate, open compartments. AC, antipodal complex; AW, anticlinal wall; CV, central vacuole; Em, embryo; En, endosperm; EW, embryo sac wall; I, integument; N, nucleus; NC, nucellus; S, starch. Scale bars = 100 urn (From Chitralekha and Bhandari 1993.)
first several rounds of division of the primary endosperm nucleus are unaccompanied by cytokinesis and thus result in a few thousand free nuclei in the central cell. During the early endosperm ontogeny, the nuclei are scattered throughout the central cell, or are pushed to its periphery by a burgeoning vacuole, or stream into incipient haustorial processes. Ultrastructural observations indicate that the cytoplasm of the central cell housing the free nuclei is undistinguished and is not enriched for any special organelles, although it is considered to be metabolically active (Marinos 1970a; Schulz and Jensen 1974, 1977; Ashley 1975a; Pacini, Simoncioli and Cresti 1975; Gori 1987; Dute and Peterson 1992; Chitralekha and Bhandari 1993). One might assume that as part of the distinction that characterizes the nuclear type of endosperm, the nuclei remain free throughout the ontogeny of the tissue, but this is not always the case. Indeed, close to the final stage of endosperm ontogeny in the majority of species examined, free nuclei and the incipient cytoplasmic domain around them are sequestered with walls and become cellular (Figure 12.1). Following the initial phase of cellularization, invaginations of the plasma membrane similar to transfer cells are seen on the wall of the endosperm cells surrounding the embryo in Vigna sinensis (Hu, Zhu and Zee 1983), Zea mays (Schel, Kieft and van Lammeren 1984; Davis, Smith and Cobb 1990;
Charlton et al 1995), and Vicia faba (Johansson and Walles 1993a, 1994). To emphasize the structural specialization of the basal endosperm transfer cells of maize, it has been pointed out that transcripts and the protein product of a cDNA clone isolated from a 10-day-old endosperm are restricted to these transfer cells (Hueros et al 1995). Whereas the endosperm in the seeds of most legumes is absorbed during seed development, soybean and certain other legumes have a partial or fully developed cellular endosperm. In soybean, the outermost layer of the cellular endosperm is incorporated into the seed as a single layer of cells around the embryo (Yaklich and Herman 1995). The flexibility of the nuclear type of endosperm is illustrated in cereals in which a coenocytic stage with free nuclei precedes a cellular configuration. Later, cells of the outermost layer of the endosperm differentiate into protein-rich aleurone cells, and the inner cells become filled with starch and storage proteins (Figure 12.2). This basic histology of endosperm development has been described repeatedly in wheat (Buttrose 1963a; Evers 1970; Mares, Norstog and Stone 1975; Morrison and O'Brien 1976; Mares et al 1977; Morrison, O'Brien and Kuo 1978; Briarty, Hughes and Evers 1979; Fineran, Wild and Ingerfeld 1982; van Lammeren 1988a), barley (Forster and Dale 1983; Bosnes et al 1987; Engell 1989; Bosnes, Weideman and Olsen
324
Figure 12.2. A section of the endosperm of Hordeum vulgare showing the peripheral aleurone layers and the inner starchy cells. AL, aleurone layer; M, mitosis in the aleurone cells; NUC, nucellus; PB, protein body; S, starch granule; SAL, subaleurone layer; SE, starchy endosperm. Scale bar = 5 um. (From Bosnes, Weideman and Olsen 1992.)
1992), maize (Randolph 1936; Kiesselbach 1949; Reddy and Daynard 1983; Kowles and Phillips 1985), and rice (Terada 1928; Juliano and Aldama 1937; Hoshikawa 1976; Ellis, Gates and Boulter 1987; Ramachandran and Raghavan 1989). Although cell volume in wheat endosperm continues to increase up to 35 days after anthesis, cell divisions plateau off at 16-20 days. In this time course, cellularization is accomplished within 2 - 4 days after anthesis during which period a marked decrease in cell volume is registered (Briarty, Hughes and Evers 1979; Gao et al 1992b). Whereas the aleurone layer in wheat and other cereals is only one cell layer thick, in barley it is at least three cell layers thick; development of the barley endosperm has been claimed to show a pattern in which cells of the starchy endosperm have a shorter proliferative period than those of the aleurone cells (Cochrane and Duffus 1981; Kvaale and Olsen 1986). However, it is doubtful whether there is a relationship between the number of layers of cells in the aleurone and a prolonged mitotic activity because cell divisions have been shown to continue in the single aleurone cells of maize (Randolph 1936), wheat (Evers 1970), and rice (Ellis, Gates and Boulter 1987; Ramachandran and Raghavan 1989) even after cessation of divisions in the starchy cells. Cell division data in conjunction with anatomical studies have led to the separation of endosperm development in barley into syncytial, cellularization, differentiation, and maturation phases. Most of the interesting changes occur during the differentiation phase, when the peripheral endosperm cells become electron dense and highly vacuolate
Zygotic Embryogenesis
and form the first aleurone cells at the placental region known as the crease or groove. At the same time, the large cells in the central endosperm are partitioned into smaller cells. In the maturing grain, aleurone cells adjacent to the crease area become thick walled in comparison to the rest of the aleurone tissue. In the mature endosperm are recognized, according to their disposition, three types of starchy cells, namely, irregular cells at the wings, prismatic cells that fan out from the crease area, and subaleurone cells (Bosnes, Weideman and Olsen 1992). During grain maturation, the contents of the crease aleurone cells are transformed into membrane-enclosed blobs of cytoplasm containing ribosomes, plastids, mitochondria, vacuoles, and a generous supply of dictyosomes. There seems little doubt that if the membrane surrounding the islands of cytoplasm is a plasma membrane, the crease aleurone cells might function as transfer cells (Cochrane and Duffus 1980). Aleurone cells in the crease region of the endosperms of Setaria lutescens (Rost and Lersten 1970), Echinochloa utilis (Zee and O'Brien 1971), and Sorghum bicolor (Giles, Bassett and Eastin 1975) (all of Poaceae) have structural features of typical transfer cells with wall invaginations. In developing wheat grain, wall ingrowths are confined to the crease aleurone and subaleurone cells that lie immediately below the aleurone layer. Moreover, the development of the crease aleurone cells is accompanied by a massive proliferation of the nuclear envelope that results in the accumulation of large amounts of rough ER (Mares, Jeffery and Norstog 1976; H. L. Wang et al 1995). In many cereals there are, besides the aleurone transfer cells, and often bordering the central cell containing the developing endosperm, elongate nucellar cells known as nucellar projections, also decorated with wall ingrowths (Cochrane and Duffus 1980; Smart and O'Brien 1983; Wang, Offler and Patrick 1995). A realistic evaluation of the crease aleurone cells and nucellar projections is that they amplify severalfold the plasma membrane surface area available for solute transport from the mother plant to the endosperm. In cereals, the interface between the embryo and the endosperm is a single layer of aleurone cells known as the germ aleurone. The cells of the germ aleurone layer are free of the aleurone grains that fill the normal aleurone cells of the rest of the endosperm (Bechtel and Pomeranz 1977; Cochrane 1994). Besides in Poaceae, a nuclear endosperm is found in Cucurbitaceae, Fabaceae, Arecaceae, and Proteaceae. Maheshwari (1950) has listed isolated
Developmental Biology of the Endosperm
genera showing the nuclear type of endosperm in additional families of monocots and dicots. In members of certain families, such as Acanthaceae, Cyrillaceae, Gesneriaceae, Icacinaceae, Loasaceae, Loranthaceae, Santalaceae, and Scrophulariaceae, in which the endosperm is of the cellular type, the first and subsequent divisions of the primary endosperm nucleus are followed by cytokinesis. This results in the formation of a population of cells that fills the entire central cell and may even extend beyond its boundary. The helobial type is a less common form of endosperm development and is found only in the monocots. The definitive feature of the helobial type of endosperm is the asymmetrical partitioning of the central cell into a large micropylar cell and a small chalazal cell following the first division of the primary endosperm nucleus. Subsequently, the micropylar cell executes a brief free nuclear phase followed by cellularization, whereas the chalazal cell remains undivided, or is partitioned into small cells, or, more often, divides in a limited way unaccompanied by cytokinesis to become coenocytic (Vijayaraghavan and Prabhakar 1984). As part of overall accounts of embryogenesis and seed formation, descriptions of endosperm development in an increasing number of plants continue to appear in the botanical literature, but they have not introduced any new concepts on the structure and morphology of this tissue. (ii) Cellularization of the Endosperm
As the endosperm is the major storage tissue of developing seeds of many plants, the importance of the cellularization lies in the fact that the accumulation of storage products and their precursors occurs in whole cells and not in isolated nuclei. Apart from this interest, the formation of cell walls in the endosperm presents basic problems relevant to the cytological basis underlying delayed formation of walls between isolated nuclei dispersed in an amorphous cytoplasm. Three questions relating to wall formation in the endosperm have attracted a great deal of interest and discussion: Does cytokinesis take place by the formation of a phragmoplast? What is the role of microtubules in the assembly of walls? What influence does the embryo sac wall exert in the cellularization of the endosperm? Observations based on light and electron microscopy form the basis for most descriptions of wall formation during endosperm development. Whether a phragmoplast is formed during cytokinesis between free nuclei of the endosperm is an issue
325
that has vexed some cytologists. Based on studies of free nuclear stages of endosperms of Helianthus annuus (Newcomb 1973b; Yan, Yang and Jensen 1991b), Stellaria media (Newcomb and Fowke 1973), Triticum aestivum (Morrison and O'Brien 1976; Mares et al 1977), Quercus gambelii (Singh and Mogensen 1976), Gossypium hirsutum (Jensen, Schulz and Ashton 1977), Papaver somniferum (Bhandari, Bhargava and Chitralekha 1986), Phaseolus vulgaris (Yeung and Cavey 1988), Glycine max (Dute and Peterson 1992), and Ranunculus sceleratus (Chitralekha and Bhandari 1993), and of helobial endosperms of Haemanthus katherinae (Newcomb 1978) and Asphodelus tenuifolius (Liliaceae) (Chitralekha and Bhandari 1992), the likely sequence seems to be the centripetal growth of the embryo sac wall toward the central vacuole, dividing the peripheral layer of cytoplasm into uninucleate open compartments independent of phragmoplasts and cell plates; the fusion of the walls with each other laterally at their origin; and nuclear divisions accompanied by the formation of periclinal phragmoplasts, cell plates and walls that close the open ends of the cells formed. It has also been frequently reported that wall formation begins with free nuclei at the micropylar end of the embryo sac and extends to the chalazal end. Dictyosomes and their vesicles, as well as ER and occasional arrays of microtubules, are seen in conjunction with the freely growing anticlinal walls during endosperm cellularization (Newcomb and Fowke 1973; Singh and Mogensen 1976; Jensen, Schulz and Ashton 1977; Newcomb 1978; Fineran, Wild and Ingerfeld 1982; Gori 1987; van Lammeren 1988a; Webb and Gunning 1991; Yan, Yang and Jensen 1991b; Dute and Peterson 1992; Chitralekha and Bhandari 1993; XuHan and van Lammeren 1993). It is important to note that even when it has been claimed that phragmoplasts form between free nuclei, their resemblance to typical phragmoplasts leaves much to be desired (Ponzi and Pizzolongo 1984). A recent study has shown that the placement of initial walls during cellularization of the barley endosperm syncytium is controlled by microtubules. Although the cytokinetic apparatus lacks a preprophase band of microtubules, a radial system of microtubules emanates from the interphase nuclei and organizes the cytoplasm into discrete units surrounding each nucleus. The walls grow at the boundaries of the nucleo-cytoplasmic domains defined by the microtubules and partition the peripheral cytoplasm into an alveolate system (Brown, Lemmon and Olsen 1994). A somewhat similar configuration of microtubules appears dur-
326
Zygotic Embryogenesis
ing cellularization of the endosperm in Phaseolus vul- the number of nuclear divisions completed after garis (XuHan and van Lammeren 1994). The consis- the formation of the primary endosperm nucleus. tent occurrence of microtubules delimiting the Nuclear activity is reflected in the extent to which nucleo-cytoplasmic units has inspired the notion endosperm haustoria with a limited number of that rather than being a passive support system, the nuclei, as compared to the multicellular tissue, microtubular cytoskeleton undergoes a striking undergo repeated endoreduplication. A striking reorganization during the initial cellularization of example is the uninucleate chalazal endosperm haustorium of Arum maculatum, which exhibits a the endosperm. After the inception of the anticlinal wall from ploidy level of 24,576N, corresponding to 13 the embryo sac wall, the pattern of subsequent wall endoreduplication cycles, whereas cells of the formation involves the building up of an incipient endosperm proper remain triploid (Erbrich 1965). phragmoplast. Indeed, the formation of phragmoEndosperms of various cereals offer considerplasts and cell plates is easier to observe during this able advantage for studying nuclear cytology due phase of endosperm development than during cel- to the prolonged cellular phase when progressive lularization of the free nuclei. Some of the early changes in DNA content occur. The extent of DNA studies using the wall-free liquid endosperm cells changes in the developing endosperm of maize is of of Haemanthus katherinae have provided a dynamic special interest since a number of reports have charpicture of mitosis in the endosperm; a beautiful cin- acterized the nuclei extensively. The increase in emicrographic record of the process, produced by nuclear volume that is often seen during polyBajer (1965), continues to be used even to this day ploidization and endoreduplication has been cited to illustrate mitosis in living plant cells. From the as indicative of an increase in the DNA content of model of cell wall formation that incorporates data the endosperm ranging from 6C to 384C (Punnett from polarized light, phase contrast, differential 1953; Tschermak-Woess and Enzenberg-Kunz 1965; interference contrast, immunofluorescence, and Cavallini et al 1995). Following upon an earlier electron microscopy, there seems little doubt that the observation that the DNA content of the endosperm phragmoplast is initially formed at late anaphase by of young maize grains detected by Feulgen the consolidation of small droplets rich in elements microspectrophotometry increased up to 24C (Swift of the dictyosomes or ER, and that it coexists with a 1950), later studies showed that coincident with the massive accumulation of microtubules. Not unlike decline in mitotic activity, there was a variable the situation described in other plant cells, these increase in the DNA content of the endosperm droplets coalesce to form the cell plate (Hepler and nucleus, followed by a decrease. In studies using Jackson 1968; Lambert and Bajer 1972; de Mey et al flow cytometry of nuclear preparations of the entire 1982). Schmit and Lambert (1987,1990) have shown endosperm, only a modest increase of about 12C in that endosperm cells of H. katherinae have a perma- the average DNA content per nucleus was observed nent system of microfilaments, and it is possible, at the peak period (Kowles and Phillips 1985; and even likely, that actin may participate in the Kowles, Srienc and Phillips 1990; Lur and Setter microtubule-guided transport of vesicles during 1993). A tentative conclusion from these data is that phragmoplast formation. preferential amplification or underreplication of part of the genome during endoreduplication is not a way of life for maize endosperm nuclei.
(iii) DNA Amplification Odd as it may seem, cellularization of the endosperm is not generally associated with a normal cycle of DNA synthesis and mitosis. Two conventional routes followed by the cells of the endosperm that escape a normal cell cycle are endoreduplication and polyploidy. D'Amato (1984) has reviewed the literature on the occurrence of these phenomena in the endosperms of diverse angiosperms; as tabulated therein, the range of endoreduplication and polyploidy displayed by endosperms may be considerable, depending upon the location of cells, the presence of haustoria, and
Studies of developing endosperms of hexaploid triticales, oat (Herz and Brunori 1985), wheat (Herz and Brunori 1985; Chojecki, Bayliss and Gale 1986; Brunori, Forino and Frediani 1989), rice (Ramachandran and Raghavan 1989), and barley (Giese 1992) by microspectrophotometry or autoradiography have also shown that some form of endoreduplication occurs in cell populations and causes a gradual rise in DNA content from 3C to 24C in triticales, oat, and wheat, and from 3C to more than 30C in rice (Figure 12.3). In the context of a general increase in DNA content of endosperm cells with time, there is a marked decrease in the
Developmental Biology of the Endosperm
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however, that aleurone cells belong to the same league as starchy cells, because only a simple doubling of DNA content, with some nongeometric fluctuations, has been observed in the examples studied (Maherchandani and Naylor 1971; Keown, Taiz and Jones 1977). The mechanism of endoreduplication in the endosperm poses a fundamental problem, since it brings into focus the control systems for induction of the S phase and inactivation of the M phase of the cell cycle. Two key observations from a work on maize endosperm deserve to be mentioned in this connection. One is that DNA synthesis during the early stages of endosperm development is maintained by an increase in the amount and activity of S phase related protein kinases, which are implicated in the onset of the S phase in animal cells. Secondly, endoreduplicated cells contain an inhibitor that suppresses the activity of an M phase promoting factor (Grafi and Larkins 1995). The biological and functional significance of endoreduplication in the endosperm cells is still a matter of debate; according to a popular view, as in the cells of legume cotyledons, an increase in the DNA content of endosperm cells may be important for the synthesis and accumulation of storage products. Unfortunately, amplification of specific DNA sequences related to the function of the developing endosperm has not been demonstrated, because the evidence has turned out to be negative (Giese 1992). 0 6
12 IS 24 30 36 42 48 54 60 66 72 78 84 ONA per nucleus (C units)
Figure 12.3. Distribution pattern of DNA content of nuclei in the developing endosperm of Oryza sativa. DA, days after anthesis. (From C. Ramachandran and V. Raghavan 1989. Changes in nuclear DNA content of endosperm cells during grain development in rice [Oryza sativa]. Ann. Bot. 64:459-468, by permission of Academic Press Ltd., London.)
number of nuclei with lower DNA values and a proportionate increase in nuclei with higher DNA values. A similar relationship exists during endosperm development in some noncereal monocotyledons (Marciniak 1993). The basic changes in DNA values just described were elucidated mostly for the starchy cells or for both starchy and aleurone cells of cereal endosperms. Although assumed to remain unaltered during cytodifferentiation, the nuclear DNA content of developing aleurone cells reportedly varies when the cells are analyzed in the absence of contaminating starchy cells. It seems improbable,
2. NUTRITIONAL ROLE OF THE ENDOSPERM Within the confines of the embryo sac, the free nuclei of the developing endosperm surround the embryo like a loose string of beads and eventually form a compact tissue around it. It is therefore obvious that some kind of interaction must take place between the endosperm and embryo. The best documented claims of interaction invoke a role for the endosperm in providing nurture and nutrients to the developing embryo. These claims, along with the structural modifications of developing embryos for nutrient uptake, will be discussed in Chapter 13. This section considers the outgrowth of haustorial structures, the chemical composition of the endosperm, and the relationship between embryo abortion and endosperm activity in wide crosses all of which constitute additional circumstantial evidence that relates the endosperm with embryo nutrition.
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Zygotic Embryogenesis
(i) Endosperm Haustoria
A general feature of endosperm organization in certain angiosperms is the production of outgrowths known as haustoria. These have been described in exquisite detail by whole mount preparations of the entire endosperm, which facilitate isolation of the parts intact. The haustoria arise at either the micropylar or the chalazal end or at both ends of the developing endosperm and appear as branched tubular processes that invade adjacent parts of the ovule such as the nucellus, integuments, and chalazal tissues. They are found mostly in the nuclear and cellular types of endosperms and occasionally in the helobial type. One arbitrarily selected representative of each of the three categories will be described here to emphasize the infinite complexity of the endosperm haustoria. It might appear unnecessary to obtain evidence of the actual transfer of metabolites to the haustorium in order to recognize that at least in some species, haustorial action might supplement the nutritional resources of the endosperm. An example of the micropylar haustorium is provided by Impatiens roylei. In the most common pattern noted, the first division of the primary endosperm nucleus separates a micropylar and a chalazal chamber. Subsequently, the micropylar chamber is partitioned into three small cells, the uppermost of which is transformed into a tubular haustorium. The haustorium retains its original form for a variable period of time, then it branches extensively in the micropylar region of the embryo sac, extending even into the funiculus (Dahlgren 1934). A view of the chalazal haustorium comes from the account of endosperm development in Nothapodytes foetida (Icacinaceae). The haustorium has its origin in the chalazal endosperm cell formed by the division of the primary endosperm nucleus. This cell enlarges and sends out narrow tubes that extend toward the chalaza. The continued ramification of the haustorium is accomplished by contact between the haustorium initial and an adjacent cell in the chalazal tissue of the ovule. The outcome of this union is the transfer of the cytoplasm and nucleus of the chalazal cell into the haustorium. At the same time as these events occur, the nucleus of the haustorium initial becomes distorted in shape as it streams into the haustorial branches (Swamy and Ganapathy 1957). These observations show that at some point in its development, the haustorium is enriched with protoplasm from the adjacent chalazal cell and thus its function is deeply entwined with the integration of materials from neighboring cells.
Figure 12.4. Development of the chalazal and micropylar haustoria in the endosperm of Rhinanthus minor. A section of the fully developed seed is shown. CH, chalazal haustorium; E, endosperm; EM, embryo; I, integument; M, micropyle; MH, micropylar haustorium; P, placenta; V, vascular connection; W, winglike part of the seed. Scale bar = 200 urn. (From Nagl 1992.)
The endosperm of Rhinanthus minor (Scrophulariaceae) is described as developing both chalazal and micropylar haustoria. These are formed when the products of the first division of the primary endosperm nucleus become polytenic and penetrate the chalazal and micropylar cells of the ovule (Figure 12.4). From the functional perspective, the most significant observation is the appearance of numerous wall labyrinths in the outer region of the chalazal haustorium where it interdigitates with adjacent cells (Nagl 1992). The convergence of numerous mitochondria alongside the wall projections and the lysis of chalazal cells in contact with the haustorium strongly suggest an active transport of the lysed material through the plasma membrane. Histochemical studies of the haustorium of Linaria bipartita (Scrophulariaceae) have shown that the cytoplasm is enriched for RNA, proteins, histones, and starch (Bhatnagar and Kallarackal 1980). These are hallmarks of a metabolically active cell, but whether they are related to its presumed function in the absorption of metabolites is not known. It is within the range of current cytochemical and cytological techniques to provide additional information to place on a sound footing such an important assumption as the role of the endosperm haustorium in nutrient absorption.
Developmental Biology of the Endosperm (ii) Chemical Composition of the Endosperm
The nutrient function of the endosperm is determined not only by the tissue's morphology and its intimacy to the embryo, but also by its chemical composition. Moreover, several observations point to the view that a knowledge of the chemical composition of the nuclear and cellular types of endosperms is necessary to obtain a general picture of the effect of this tissue on the nutrition of the embryo beginning with the zygote stage. In view of the difficulty of obtaining large quantities of nuclear endosperm, biochemical analysis has been limited to the liquid endosperm of Cocos nudfera (coconut; Arecaceae). Popularly known as coconut milk or coconut water, the liquid endosperm of coconut is the morphological equivalent of a nuclear endosperm and is widely used as a growth adjuvant in plant tissue culture media. As described in the next chapter, the first successful culture of proembryos was accomplished by supplementing the mineral-salt medium with coconut milk. What are the chemical factors present in coconut milk that dramatically induce the growth of embryos and other explanted organs in culture? Results from chemical fractionation have shown that coconut milk is a conglomeration of several inorganic ions, amino acids, organic acids, vitamins, plant hormones, sugars, sugar alcohols, and other substances in concentrations that are used in tissue culture media. Indeed, some of the substances may not be of any real value in the promotion of growth of cultured plant organs and, thus far, no single compound or group of compounds has come to be accepted as being able to duplicate the major growth promoting activity of coconut milk in its entirety (Raghavan 1976b). Limited information is available on the chemical composition of the cellular endosperms of some plants and of their effects on the growth of cultured organs. Because this work has been summarized previously, only a few general comments will be made here (Raghavan 1976b). Maize endosperm at the milky stage contains several amino acids and hormones, including the cytokinin zeatin. Among other cereals, wheat, rye, barley, and rice have endosperms that are known to contain auxins. By far the most potent source of endosperm as a growth promoter of cultured tissues is the liquid content of the vesicular embryo sac of Aesculus woerlitzensis (horse chestnut; Hippocastanaceae), which contains both free nuclei and cellular endosperm. Chlorogenic acid, myo-inositol, and auxin are some of the compounds with potential
329 growth promoting properties identified in the endosperm extract of horse chestnut. Based on the information from chemical analyses in conjunction with tissue culture data, it is believed that the innate in vivo function of endosperm tissues of Datura stramonium, Juglans regia (walnut; Juglandaceae), Allanblackia parviflora (Guttiferae), Cucumis sativus, Prunus persica and related species, Hyphaene natalensis (Arecaceae), and Lupinus luteus
is concerned with the supply of nutrients to the growing embryo. Similarly, the identification of substances such as GA and sugars in the endosperm tissues of Aesculus californicus, Echinocystis macrocarpa (Cucurbitaceae), and apple, among others, reflects the potential utilization of these tissues by the embryo. Monitoring the fate of GA during embryo growth might permit alternative explanations, and so these examples need further study. (iii) Embryo Abortion and Endosperm Development
In horticultural and breeding practices, embryo abortion is commonly encountered in unsuccessful crosses that fail to set seeds. Although fertilization occurs normally in such crosses and embryos begin to grow in a relatively healthy way, lethality is rampant during the subsequent growth of embryos. Hybrid embryos display a wide range of irregularities, some of which are described in Chapter 14; the concern here is to examine evidence suggesting the relationship of embryo abortion to a lack of endosperm activity. Renner (1914) first called attention to the fact that the collapse of embryos in unsuccessful crosses between Oenothera biennis X O. muricata and O. biennis X O. lamarckiana was pre-
ceded by the disintegration of the endosperm beginning soon after fertilization, which thus deprives the embryo of an immediate source of food. As shown by detailed cytological studies of endosperm development in aborted seeds in an extensive range of interspecific hybrids, endosperm failure is more frequent than is generally believed and might take place in discrete ways during the division of the zygote. Comparison of ovule development in reciprocal crosses involving diploid and tetraploid races of Lycopersicon pimpinellifolium and in crosses of these races with L. peruvianum provides a fine example of the diversion of the primary endosperm nucleus into an alternative pathway during its early ontogeny. In terms of cell number, the initial rate of development of the endosperm in inviable crosses does not differ appreciably from that observed in successful
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crosses, but afterward, the endosperm of the latter outstrips that of the former in its growth. The structural changes seen in the disintegrating endosperm cells are more dramatic than the decrease in cell number. In the hybrid ovule as early as 144 hours after pollination, cells of the endosperm at the chalazal end become vacuolate and less dense than those at the micropylar end. Although the endosperm survives for some time more, it undergoes no further divisions. Instead, as a result of the dissolution of cell walls and the fusion of the protoplasm and of the nuclei with those of contiguous cells, the endosperm is reduced to a few giant cells surrounding the embryo (Cooper and Brink 1945).
Zygotic Embryogenesis
An important factor governing the continued growth of the embryo and endosperm in a fertilized ovule is the nature of their interaction with the neighboring cells; therefore, some attention has been paid to the behavior of the nucellus and integuments in ovules of wide crosses. Several investigators, beginning with Renner (1914), have noted that the development of a weak endosperm and embryo abortion in inviable crosses are associated with the proliferative growth of a tissue from the nucellus. In a comparative histological examination of ovules from crosses between Nicotiana species, seed collapse was found to be associated with a retarded growth of the endosperm, which In Trifolium ambiguum X T. repens cross, division of attained only a fraction of the growth attained in the primary endosperm nucleus ceases at different the selfed parent, and with a pronounced hyperstages of development of the hybrid ovules, but in plastic growth of the nucellus. The term "somatono case does the endosperm proceed beyond the plastic sterility" is used to describe the hyperplasia 128-nucleate stage; in many hybrids, endosperm of the maternal tissue. It has been found that development is stymied at even earlier stages. In embryo failure is also associated with a deficiency contrast, selfed T. ambiguum has a complete in the development of the conducting elements endosperm including a well-developed chalazal within the seed, foreshadowing a suppression of haustorium (Williams and White 1976). The ultra- nutrient transport to the endosperm; hyperplastic structural changes in the hybrid endosperm cells growth of the nucellus has been ascribed to the are also impressive and seem to impinge on the syn- consequent abnormal distribution of nutrients thetic activities of the cells and on nutrient transport (Brink and Cooper 1941). into them. For example, in the endosperm cells of A gene dosage effect leading to a change in the Hibiscus costatus X H. aculeatus and H. costatus X H. ratios of ploidy levels among the embryo, furcellatus hybrids, the rough ER appears as short endosperm, and maternal tissues is seen in the irregular segments lacking ribosomes, whereas the ovules of several infertile crosses that show tumor nuclei contain multiple nucleoli (Ashley 1975a, b). formation from the inner epidermis of the integuIndubitable proof that endosperm abnormalities are ment (endothelium). In reciprocal crosses between related to the transport of nutrients is the demon- 4N L. pimpinellifolium and 2N L. peruvianum, the stration that wall ingrowths characteristic of the endothelium assumes a densely cytoplasmic connormal endosperm are almost completely sup- tour and surrounds the endosperm except for a pressed in the hybrid endosperm generated in small gap at the chalazal end. Tumor formation is crosses between normal diploid and autotetraploid initiated from the dorsal region of the endothelium maize (Charlton et al 1995). In the various examples near the chalazal end. As the tumor assumes form, studied, cells of the unhealthy endosperm have it begins to grow without restraint and extends into been found to show uninformative types of mitotic the embryo sac as a burgeoning mass of cells. The abnormalities such as lagging chromosomes, fused cells of the endosperm surrounding the embryo nuclei, and nuclei of high or low chromosome num- deteriorate progressively to the point that the bers. However, an understanding of the extent to embryo is engulfed by the overgrown endothelial which disturbances in the endosperm in inviable cells (Cooper and Brink 1945). It is no wonder that crosses may cause starvation of the embryo is cer- abortion of the already undernourished embryo is tainly lacking at the present time. It must be con- hastened by these changes. cluded on logical grounds that the hybrid embryos An imbalance in the chromosome number are starved to death due to the inability of the phys- between embryo and endosperm is postulated to iologically disturbed endosperm to manufacture cause abnormalities in the endosperm and consethe exacting nutrients required for embryo growth quent embryo abortion in crosses between diploid or to absorb them from the surrounding maternal and tetraploid varieties of Citrus (Esen and Soost tissues and transmit them to the embryo. This 1973). Considerations of the requirement for a balmeans that the deleterious genes exert their primary ance between maternal and paternal genomes in effect on the endosperm rather than on the embryo. the endosperm for seed set have led to a number of
Developmental Biology of the Endosperm
hypotheses to account for endosperm failure and embryo abortion in wide crosses (Kowles and Phillips 1988). A most useful observation based on data from a wide variety of intraspecific- and interspecific-intraploidy crosses is that the endosperm develops abnormally when the maternal-paternal ratio in the tissue deviates from 2:1. This has been incorporated into the notion of endosperm balance number (EBN) to explain endosperm development in crosses with different levels of ploidy. According to this hypothesis, the genome of each species is assigned an EBN on the basis of its crossing behavior with a standard species and, for crosses to be successful, the EBN has to conform to a 2:1 maternal-paternal ratio (Johnston et al 1980; Johnston and Hanneman 1982; Camadro and Masuelli 1995; Ehlenfeldt and Ortiz 1995). From the practical side, the attractiveness of this hypothesis is that the EBN can be manipulated to break the crossing barriers and to create new hybrids; from the theoretical side, it is unclear how the ratio of the maternal-paternal genomes is detected and how a deviant ratio causes endosperm malfunction. Using a mutant indeterminate gametophyte (ig) to cre-
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methods have been widely used to obtain transplantable seedlings from aborted seeds of interspecific and intergeneric crosses that are traditionally condemned as being incapable of further growth. In certain cases, continued growth of the hybrid embryo is secured by implanting it on a normal endosperm, which is then cultured, thereby initiating a nurse culture. In a limited number of cases, efforts directed toward rescuing hybrid embryos by culturing ovules and ovaries have proved rewarding. Examples of hybrid embryo rescue by these different tissue culture protocols have been described or tabulated in published reviews (Raghavan 1977b, 1985, 1986b, 1994; Collins and Grosser 1984; Williams 1987) or books (Raghavan 1976b, 1986c), and the reader is referred to these sources for further information. The appearance of haustorial structures on the endosperm, the presence in this tissue of chemical substances with potential growth promoting functions, and the rescue of hybrid embryos from ovules lacking a normal endosperm, present evidence of the potential role of the endosperm in the nutrition of the embryo. These observations, considered in conjunction with the various ultrastructural modifications of the embryo and suspensor cells for the absorption and translocation of metabolites from the surrounding cells (Chapter 13), make a compelling case for the nutrition of the embryo by endosperm metabolites.
ate various ploidy level combinations in crosses in maize, Lin (1984) concluded that development of the endosperm is affected by the parental source of its chromosomes and that the 2:1 maternal-paternal chromosome number is critical in the development of a healthy endosperm. Although this hypothesis can readily explain why 2x X 4x or 4x X 2x crosses with 2:2 and 4:1 maternal-paternal chromosome ratios, respectively, in the endosperms will fail, it is 3. GENE ACTION DURING ENDOSPERM not applicable to crosses in which endosperm ONTOGENY development is not governed by a ratio of chromoIt is natural to think of the development of the some numbers. endosperm in terms of the synthesis of storage The histology of the failure of endosperm devel- compounds and proteins necessary for the fundaopment and its side effects on the maternal tissues mental processes of cellular metabolism and houseof ovules of unsuccessful crosses are obviously in keeping functions. Three major experimental line with the view that gene action deprives hybrid approaches have been pursued to gain an underembryos of a continuing supply of nutrients to standing of how genes control the complex process complete development. The postulated nutritional of endosperm development in cereals. Historically, relationship between the embryo and endosperm spontaneous kernel mutations in maize and barley tends to suggest that the cryptic potentiality of the have provided a tractable genetic system for monihybrid embryo to complete development may be toring the effects of marker genes on the developrealized if it is allowed to grow in an artificial ment and differentiation of the endosperm. A large medium supplied with nutrient substances that are body of work has centered on the correlations of normally identified with the endosperm. Early in RNA and protein synthesis with endosperm develthe development of embryo culture as a research opment, and on the isolation of cDNA clones for tool, Laibach (1925) demonstrated that progenies specific proteins and the analysis of their exprescan be obtained from nonviable seeds of Linum sion in the developing endosperm. In recent years, perenne X L. austriacum hybrid by excision and cul- extensive efforts have gone into the sequencing of ture of embryos before the embryos begin to disin- genes regulating endosperm storage product syntegrate. Since this pioneering work, embryo culture thesis and into the search for conserved nucleotide
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sequences of developmentally regulated genes and for the expression of promoter sequences of genes in transgenic plants. This section will deal with the genetic control of endosperm development, the regulation of RNA and protein synthesis, and the expression of genes for housekeeping proteins in the endosperms of cereals, in which they have been studied intensively. Due to the sheer volume of information, synthesis and accumulation of storage products in the endosperm will be considered in a separate section.
Zygotic Embryogenesis
mutant kernels have an extremely low level of invertase, impairment of the movement of photosynthates between the pedicel and endosperm, and the consequent osmotic imbalance, offer a plausible explanation of the effects of the mutant gene on the endosperm (Lowe and Nelson 1946; Miller and Chourey 1992; Cheng, Taliercio and Chourey 1996). The basic histological manifestation of mutation in the de-17 locus is the failure of endosperm cells facing the placenta to form a specialized absorbing tissue for transmitting nutrients from the maternal plant to the endosperm (Brink and Cooper 1947a). (i) Endosperm Mutants A large number of dek mutants have been The singular strength of maize as a genetic sys- induced by pollinating maize ears with pollen tem among higher eukaryotes is the occurrence of grains mutagenized with ethyl methanesulfonate morphologically identifiable mutants that affect (EMS). The Ml plants raised from mature grains virtually all stages of its life cycle. In considering were self-pollinated and the harvested ears generthe regulatory intervention of genes in kernel ated the stock of dek mutants for subsequent genetic development in maize, most analyses have been analysis. The endosperms of the mutants varied conperformed to date with mutants that affect both the siderably in their phenotypic appearance, with endosperm and the embryo and with those that are many having a collapsed or shrunken appearance; specific for endosperm development. Those in others were fully formed but were distinguished which growth of both embryo and endosperm is from the wild type by a floury or opaque appearimpaired are known as defective kernel (dek) mutants; ance and a soft or fluid consistency. It was not included in the group that affects only endosperm unusual to find mutant kernels that were lighter in development are shrunken (sh), wx, brittle (bt), sugarycolor than the normal due to blockage of carotenoid (su),floury(fl), and opaque (op) mutants. Since the lat- synthesis. Generally, the mutant phenotype is ter group of mutations impinges on the nature of revealed within two weeks after pollination, at the storage reserves in the endosperm, it is appro- which time the growth of the defective endosperm priately considered in the next section. slows down. Effects of the mutant gene on the embryo are seen even earlier than on the endosperm The first dek mutants described turned out to be (Neuffer and Sheridan 1980; Sheridan and Neuffer those in which the endosperm was normal but the grain did not have a viable embryo (Jones 1920). 1980). Compared to the wild type, mutations impose True dek mutants with defective endosperm were serious restraints on the accumulation of dry matter, described by Mangelsdorf (1923), who found total proteins, and storage proteins in the grains in which ovule development was arrested endosperm. From these analyses, an informal recogalmost immediately after fertilization or after a nition of three broad classes of mutants has small amount of endosperm had been formed. emerged: (a) those showing a reduced accumulation Additional defc-like mutants are designated as of dry matter and proteins during the entire reduced endosperm (re-1, re-2) (Eyster 1931), minia- endosperm ontogeny; (b) those showing an initial ture seed (mn) (Lowe and Nelson 1946), and defec- normal rate of accumulation, followed by an early tive seed (de-17) (Brink and Cooper 1947a). In decline; and (c) those in which an initial lag in the general, mutant grains attain only a fraction of the rate of accumulation is coupled with an early termiweight of normal grains while maintaining a regu- nation of accumulation (Manzocchi, Daminati and lar initial pattern of growth of the endosperm and Gentinetta 1980; Manzocchi et al 1980). In almost all embryo. Closer studies of mn mutant kernels have cases examined, the average DNA content of the shown that subsequent disintegration of the vas- nucleus of mutant endosperm cells was found to be cular elements at the chalazal end of the ovary lower than that of the wild type; two idealized posseemingly interrupts nutrient supply from the sibilities - a low proportion of nuclei in the higher maternal tissues through the pedicel. Provision of DNA classes or the occurrence of fewer rounds of the MN gene in the wild-type grain serves to endoreduplication - can obviously tilt the balance in encode an endosperm-specific cell wall invertase favor of a low DNA value (Kowles et al 1992). necessary for the mobilization of sucrose. Since the An array of mutants equally diverse as those
Developmental Biology of the Endosperm
just described has been obtained by the insertion of mutator (mu) transposons into the maize genome. This is accomplished by crossing standard lines with pollen from mu stock; the Fl plants grown from the resulting caryopses are then self-pollinated. Defective kernels are identified on the ears and are used for subsequent genetic dissection. Among the range of phenotypes observed, the most common are those containing a reduced endosperm and a small embryo, and others that have an empty pericarp and are almost always embryoless. Several mutants identified by transposon tagging have been found to be allelic to the dek mutants obtained by EMS mutagenesis (Scanlon et al 1994). These observations lead to the inference that following fusion of the sperm cell with the diploid polar fusion nucleus, many different genes must function in an orderly manner to effect the development of the endosperm into a functional nutritive tissue. The potential to gain deep insight into the genetic program governing endosperm development in maize by molecular cloning has been enhanced by the isolation of mutants from mu stocks.
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(ii) Synthesis of Housekeeping Proteins
As a rapidly growing tissue, the endosperm must synthesize an enormous range of proteins necessary for cellular metabolism. However, in contrast to the extensive literature on storage protein synthesis, little information is available on the synthesis of metabolic proteins during endosperm development. The main reason for this is that synthetic activities of the widely investigated endosperm tissues are inextricably linked to the accumulation of storage products. Most importantly, the analysis of housekeeping proteins includes changes in the activities of enzymes for the metabolism of starch, proteins, and lipids, and in the levels of amino acids and nucleotides. Of necessity, much of the evidence from such investigations that relates to the synthesis of housekeeping proteins is correlative, and hence its significance is not fully understood. A few studies that have analyzed gene expression patterns in the endosperm dissociated from the synthesis of storage proteins will be described later.
Further evidence for the action of a multiplicity Following fertilization, a readily available supply of genes during endosperm development has of substrates for the division of the endosperm come from analyses of barley mutants with a nucleus must be present. This may be provided by defective endosperm. Deserving recognition for transcription of the genome of the maternal tissues of their importance are spontaneous endosperm the ovule or of the endosperm nucleus. Evidence mutants designated as shrunken endosperm (se and based on autoradiography of 3H-uridine incorporaseg), in which shrunken kernels are caused by the tion shows that active transcription is initiated in the expression of the mutant genes in the maternal tis- endosperm of barley as early as the free nuclear stage sues of the plant. Mutant seg endosperm types are (Bosnes and Olsen 1992). The question of how soon distinguished from the wild type by a premature after fertilization the genome of the endosperm termination of grain filling caused by necrosis of nuclei begins transcription has been investigated in the chalaza, or by upsets in the endosperm growth maize by crossing two strains with different elecpattern that lead to characteristic abnormalities trophoretic isozymes for a particular locus and by but permit normal development of the chalazal tis- screening the endosperm at different stages of develsues (Jarvi and Eslick 1975; Felker, Peterson and opment for expression of the variant enzymes. It was Nelson 1984, 1985, 1987). Another group of found that several enzymes of the maternal allele mutants with a defective endosperm includes the become active two or three days earlier than the gene chemically induced shrunken endosperm expressing products of the paternal allele. The earlier expression xenia (sex) mutants, in which the mutant genes of enzymes from the maternal side has been attribexert their effect on tissues of the entire grain. As uted to their origin in the nucellus. However, other in the case of maize, the ramifications of sex muta- experiments showed that transcription of some tion on the endosperm of barley are extensive, isozyme genes in the endosperm nucleus begins relranging from the total lack of aleurone cells to the atively soon after fertilization but translation of the complete loss of starchy endosperm traits, and message is delayed until the endosperm becomes celeven to the presence of new cell types not found in lular (Chandlee and Scandalios 1983). the wild-type endosperm; the embryo may remain A study of catalase activity profiles has shown viable or nonviable. These effects are so profound that continued development of the maize as to suggest that the mutations cover genes par- endosperm necessitates fundamental changes in ticipating in all phases of development of the the control of gene expression for the three endosperm (Bosnes et al 1987; Bosnes, Weideman unlinked genes of the enzyme (CAT-1, CAT-2, and and Olsen 1992). CAT-3) in the different parts of the kernel. CAT-1 is
334
present throughout the 30-day developmental period analyzed, whereas CAT-2 is expressed toward the period of kernel maturity and CAT-3 appears only during the initial period of development. The appearance of isozymes correlates well with the expression of transcripts for the respective genes in RN A blots, indicating that the synthesis of the enzyme is controlled at the transcriptional level. The presence of CAT-1 mRNA in the pericarp as well as in the starchy and aleurone cells of the endosperm, presence of CAT-2 mRNA only in the aleurone layer, and presence of CAT-3 mRNA only in the pericarp are evidence of strong tissue-specific regulation of the synthesis of these isozymes (Tsaftaris and Scandalios 1986; Wadsworth and Scandalios 1989). The reason for the unique, developmentally controlled pattern of expression of catalase genes is not at present understood, and investigations on the endosperms of other cereals will be valuable for comparative analysis. Like catalase, pyruvate orthophosphate dikinase, a key enzyme in photosynthetic carbon dioxide fixation in C4 plants, illustrates a tissue-specific and developmentally regulated pattern of gene expression during endosperm development in wheat. The changes in the enzyme titer in the endosperm, with a peak at mid-maturation followed by a decrease, are correlated with changes in mRNA activity. At the tissue level, activity of the enzyme confined to the cells of the aleurone layer is paralleled by the appearance of mRNA in these cells (Aoyagi, Bassham and Greene 1984; Aoyagi and Chua 1988). Genes for glyceraldehyde-3-phosphate dehydrogenase and alcohol dehydrogenase (enzymes of the glucose-phosphate metabolism) show an increased expression correlated with progressive endosperm development in maize up to about 15 days after pollination, followed by a decrease (Russell and Sachs 1991). Trans-acting factors required for developmental expression of the maize ADH-1 gene appear to be present in the endosperm of rice (Kyozuka et al 1994). As will become evident in the following section, the pattern of accumulation of storage proteins and their mRNAs in developing endosperms is not much different from that displayed by the photosynthetic and cytosolic enzymes. 4. SYNTHESIS A N D ACCUMULATION OF ENDOSPERM STORAGE RESERVES
Because of the role of the endosperm as a major dietary source, the importance of the analysis of gene expression concerned with the synthesis of storage reserves in this tissue cannot be overemphasized. An
Zygotic Embryogenesis
understanding of the mechanism by which genes for the synthesis of the three principal storage products, namely, starch, proteins, and lipids, are regulated in the endosperm would be of great importance to the genetic engineering of cereal grains with improved nutritional qualities. A good deal of work, aided mostly by mutants in which the coordination and differential expression of endosperm-specific genes are disturbed, has been done to characterize the synthesis and deposition of storage products during endosperm development in our major cereal crops. This section will review the basic data on the nature of the endosperm storage products, mutants that have led to the identification of key steps in the biosynthetic pathway of storage products in the endosperm tissues of various plants, and the cell biology and biochemistry of the accumulation of proteins in the endosperm. A nutritional role for the storage proteins is supported by their presence in large quantities in the seed and by their rapid degradation during the early stages of germination as the embryo is transformed into a seedling. In seeds of some members of Fabaceae and Arecaceae, extensive deposition of polysaccharides like galactomannans and xyloglucans occurs on the inner cell walls of the endosperm and virtually occludes the cell contents. In addition, the cells contain small amounts of storage lipids and proteins (Meier and Grant Reid 1977; DeMason, Sexton and Grant Reid 1983; DeMason 1986; DeMason, Chandra Sekhar and Harris 1989; Edwards et al 1992). Although it is believed that these cell wall polymers serve as food materials, they are not further considered here.
(i) Mutants Defective in Starch Synthesis
This account will be prefaced by a brief reference to conventional biochemical studies that have identified the enzymes that link the synthesis of starch to its main substrate, sucrose. An important initial reaction in the sucrose-starch conversion is catalyzed by sucrose synthase, which splits sucrose to fructose and UDP-glucose. Another set of reactions involving hexokinase and hexose phosphate isomerase results in the transformation of fructose to glucose-6-phosphate. Formation of glucoses-phosphate is also provoked from UDP-glucose by the action of pyrophosphorylase followed by a reaction mediated by a mutase. These changes, which occur in the cytoplasm, are followed by the conversion of glucose-6-phosphate to starch inside the amyloplast. This step is also catalyzed by a pyrophosphorylase and involves the formation of ADP-glucose, which donates its glucose to an increasing chain of
Developmental Biology of the Endosperm
glucose molecules. Starch exists as straight-chain amylose and as the branched-chain amylopectin, the latter being formed by the action of a branching enzyme. Although different biosynthetic pathways that lead to the same end products occur in maize and in some other plants, the basic steps described here could be rightfully considered as the most prevalent. Several genes affect the type and quantity of carbohydrates in the mature endosperm of maize and, consequently, genetic approaches have been used extensively to determine the biochemical lesions caused by mutations. The sh-1 mutation leads to a shrunken or collapsed endosperm and is attributed to the presence of a reduced amount of starch. At the protein level, the mutant endosperm is highly deficient in sucrose synthase; this in turn results in the accumulation of elevated levels of sucrose in the endosperm (Creech 1965; Chourey and Schwartz 1971; Chourey and Nelson 1976; Cobb and Hannah 1983). A special aspect of all sh-1 mutants showing a substantial loss of sucrose synthase is that they retain a residual level of enzyme activity. The arguments about the normal enzyme and the residual enzyme have hinged around the question of whether the two enzymes are coded by the same or different genes; it is now resolved that the residual activity is due to a minor enzyme, similar in catalytic properties to the bulk enzyme, but encoded by a second gene, SUCROSE SYNTHASE (SUS) (Chourey 1981). A model of different in vivo functions for the two isozymes has gained currency with the demonstration that the isozymes are located in different parts of the endosperm. The enzyme encoded by SH-1 generally accumulates in the bulk of starch-containing cells, whereas the SLZS-encoded enzyme is restricted to the aleurone layer and the basal endosperm transfer cells (Chen and Chourey 1989; Heinlein and Starlinger 1989). Finally, transcripts of SH-1 and SUS genes have been localized in the starchy endosperm and the aleurone layer, respectively (Springer et al 1986; Rowland and Chourey 1990). This finding indicates that the rate of transcription or message stability may be a controlling factor in the accumulation of the two isozymes. At the molecular level, both the SH-1 and SUS genes have been cloned and characterized (Werr et al 1985; McCarty, Shaw and Hannah 1986; Gupta et al 1988). Another mutant resulting from upsets in the carbohydrate metabolism of the endosperm is vox, which forms exclusively amylopectin, instead of a mixture of amylose and amylopectin as in the normal endosperm. The enzymatic basis for this differ-
335
ence has been attributed to the absence in the mutant endosperm of starch-granule-bound nucleoside diphosphate sugar transferase enzymes, which transfer glucose from ADP-glucose or UDP-glucose to starch (Nelson and Rines 1962; Nelson and Tsai 1964). As in the case of the sh mutant, the wx endosperm maintains some residual enzyme activity possibly coded by a different gene (Nelson, Chourey and Chang 1978). Cloning of the WX locus and characterization of its gene product have shown that some mutant phenotypes result from the production of a structurally altered protein; several other stable mutant phenotypes are caused by insertions or deletions within the WX gene transcription unit (Echt and Schwartz 1981; Shure, Wessler and Federoff 1983; Wessler and Varagona 1985). Greatly reduced starch synthetic capability is associated with sh-2 and bt-2 mutants. The mutant block has been identified with a reduced activity of the enzyme ADP-glucose pyrophosphorylase in the endosperm; this enzyme catalyzes the formation of ADP-glucose from glucose-1-phosphate and ATP (Tsai and Nelson 1966). However, the presence of detectable levels of enzyme activity in several alleles of both mutants suggested that they may possess a pyrophosphorylase independent of SH-2 and BT-2 gene functions; this proved to be the case when it was found that 95% of the activity in the wild type exists in the heat-labile form and 5% in the heat-stable form. Both mutations reduce the heat-labile form appreciably and the heat-stable activity only marginally (Hannah and Nelson 1976; Hannah, Tuschall and Mans 1980). Although these observations point to the genes' preference for functioning in a nonstrucrural role in some posttranslational steps, two later studies have strengthened the argument that SH-2 is a structural gene. In one strategy, using the transposable element dissociation {ds), which has the ability to inhibit the function of SH-2, it was found that upon removal of the ds element, gene function is restored in the endosperm. Moreover, revertants that restore wildtype enzyme levels possess enzyme molecules with altered allosteric and kinetic parameters (Tuschall and Hannah 1982). Confirming that SH-2 is indeed a structural gene for the maize endosperm ADPglucose pyrophosphorylase, the deduced amino acid sequence of a SH-2 cDNA clone has revealed much similarity to that of the bacterial enzyme (Bhave et al 1990). Using rice ADP-glucose pyrophosphorylase cDNA as a probe, a BT-2 cDNA clone was isolated from maize endosperm and partially sequenced. When RNA isolated from various bt-2 mutants was probed with the maize cDNA
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clone, some mutants had wild-type levels of transcripts and others had transcripts of altered size, suggesting that BT-2 is a structural gene for the transcript (Bae, Giroux and Hannah 1990). Not much is known about the bt-1 mutant, which also contains extremely reduced amounts of starch in the endosperm. The deduced amino acid sequence of a cDNA clone of the BT-1 gene and the copurification of the protein with plastid membranes have indicated a possible role for the gene product in amyloplast membrane transport, especially in the uptake of adenylate molecules (Sullivan et al 1991; Sullivan and Kaneko 1995). It thus appears that significant differences exist in the molecular mechanism of the inhibition of starch deposition in the mutant endosperms, despite their phenotypic similarities. The su mutant is interesting because the principal carbohydrate storage product in the endosperm is the high-molecular-mass compound phytoglycogen. Since accumulation of phytoglycogen occurs at the expense of amylose and amylopectin, it is not surprising that the su endosperm is deficient in starch-debranching enzyme. Chromatography on hydroxyapatite column showed that su endosperm extract lacks one of the three peaks of enzyme activity associated with the nonsugary endosperm and has much reduced activity in the other two peaks. In the multistep pathway of sucrose-to-starch conversion, it is not clear how a deficiency in the debranching enzyme would increase sucrose levels. A related mutant is ae, whose wild-type gene encodes a major starch-branching enzyme. Effects of the mutant gene are manifest in a reduction in amylopectin, a small increase in amylose, and a minor reduction in starch content (Pan and Nelson 1984). Consistent with their endosperm-specific nature, the mutations that abolish the activity of enzymes in the endosperm do not interfere with enzyme synthesis in the embryo. This is true for sucrose synthase at the SH-1 locus (Chourey and Nelson 1976), starch-granule-bound transferase at the WX locus (Akatsuka and Nelson 1966), ADPglucose pyrophosphorylase at the SH-2 locus (Preiss, Lammel and Sabraw 1971), and debranching enzyme at the SU locus (Pan and Nelson 1984). Some of the genes involved in starch metabolism of the endosperm of maize that have been cloned are characterized by a highly variable number of introns. The number of introns ranges from 2 in the BT-1 gene (Sullivan et al 1991), to 4 in the SU gene (James, Robertson and Myers 1995), 13 in the WX gene (Klosgen et al 1986) and 15 in the SH gene (Werr et al 1985).
Zygotic Embryogenesis
Mutants regulating WX gene expression in the endosperm have been well documented in rice (Sano 1984). As in the case of maize, as many as 13 introns have been reported in the WX gene of rice (Z.-Y. Wang et al 1990; Hirano and Sano 1991). In the grains of normal and transgenic rice, the WX gene is expressed in a tissue-specific manner exclusively in the endosperm. The expression of this gene is also developmentally regulated, transcripts being present at maximum levels during the early to middle stages of endosperm development (Hirano and Sano 1991; Hirano et al 1995; Z.-Y. Wang et al 1995). Spontaneous mutants affecting carbohydrate metabolism of the endosperm are rare in other cereals, although various kinds of endosperm mutants similar to the su and sh have been obtained by chemical mutagenesis in rice (Satoh and Omura 1981; Yano et al 1984, 1985; Kumamaru et al 1988). (ii) Storage Protein Synthesis
The section on this topic will begin with reference to some terminology currently used to describe seed storage proteins. On the basis of their solubilities, seed storage proteins are classified into albumins, globulins, prolamins, and glutelins. The albumins are soluble in water and in dilute aqueous buffer at a neutral pH, whereas the globulins are characterized by their solubility in salt solutions and insolubility in water. Prolamins are soluble in alcohol, and glutelins belong to a class of proteins that are soluble in dilute acid or alkali solutions. What is known about the distribution of the different kinds of endosperm storage proteins indicates that some of the major cereal grains can be identified by the type of reserve proteins of their endosperms. The major storage proteins in the endosperms of maize, barley, and sorghum are of the prolamin type. Glutelins account for nearly half of the storage proteins of the wheat endosperm, whereas about 40% is constituted of prolamins. Glutelins were thought to be present in the largest amount in rice, accounting for at least 80% of the total protein of the starchy endosperm. However, it now appears that the relative contribution of glutelins to the total proteins of the rice endosperm has been overestimated and that prolamins and globulins also contribute significantly to the total protein content (Krishnan and White 1995). Although prolamins of maize continue to receive a great deal of attention, it is to be noted that glutelins, which constitute about 35%-45% of the storage proteins of the endosperm, have hardly
Developmental Biology of the Endosperm
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been studied. Globulins, which are the predominant the rough ER in the synthesis of wheat prolamins, storage proteins in the embryos of angiosperms and but the exact mode of transport of the protein from gymnosperms, are the major reserve proteins in the the ER to the protein body in the vacuole has been endosperm of oat. Albumin is also of wide-spread interpreted in different ways (Campbell et al 1981; occurrence in embryos; there are only a few reports Miflin, Burgess and Shewry 1981; Bechtel, Gaines of their occurrence in endosperms. Endosperm stor- and Pomeranz 1982; Parker 1982). One telling age proteins are often designated by a common piece of evidence was the identification of the name derived mostly from the generic name of the wheat homologue of an ER-resident protein, plant, such as zein (prolamin from Zea mays), immunoglobulin heavy-chain-binding protein hordein and hordenin (prolamin and glutelin, (BiP), in the ER and protein bodies of the developrespectively, from Hordeum vulgare), avenin (pro- ing endosperm of T. aestivum; the label was scarcely lamin from Avena sativa), gliadin, glutenin, and trit- present in the Golgi apparatus. The most generalicin (prolamin, glutelin, and globulin, respectively, ized views of these observations are that storage from Triticum aestivum and other species of culti- protein molecules aggregate within the ER along vated wheat), secalin (prolamin from Secale cereale), with the resident protein to form the protein body kafirins (prolamin from Sorghum bicolor), coixin (pro- and that intact protein bodies are transported to the lamin from Coix lacryma-jobi), and oryzin and vacuoles by a novel route that does not utilize the oryzenin (prolamin and glutelin, respectively, from Golgi apparatus (Levanony et al 1992; Rubin, Oryza sativa). Levanony and Galili 1992). An apparent absence of the Golgi apparatus during the active phase of proOrigin of Protein Bodies. The usual concomi- tein accumulation in the cells of the endosperm of tants of endosperm development in cereal grains T. aestivum suggested that this organelle is not are the synthesis of storage proteins and their involved in the transport of the protein product to deposition in the vacuoles as protein bodies. A the vacuole (Briarty, Hughes and Evers 1979). The well-defined aspect of storage protein accumula- picture that began to emerge from later investigation is the appearance of protein bodies in the vac- tions is that Golgi bodies are present abundantly in uoles shortly after the endosperm becomes cellular the developing endosperm adjacent to the proteins and an increase in the size and the number of pro- concentrating within the ER. At this critical period, tein bodies thereafter. Synthesis of storage proteins, an enlargement of the cloud of Golgi vesicles, contheir transformation into protein bodies in a taining material resembling storage proteins, and translocation-competent conformation, and, finally, connections between sheets of rough ER and eletheir transport to the site of storage occur in com- ments of the Golgi vesicles are also frequently plicated ways, and even in the best studied systems observed in suitable preparations (Bechtel, Gaines and Pomeranz 1982; Parker 1982; Parker and some uncertainty remains about the processes. Hawes 1982; Bechtel and Barnett 1986). The weight Some of the most influential generalizations on of evidence is thus suggestive, but not complete the mechanism of protein body formation and its enough to say, that the wheat storage proteins are transport have come from investigations on the transported to protein bodies by a mechanism endosperms of different species of Triticum. The involving the Golgi bodies. It is also conceivable usual parameter used to classify gliadins and that as a first step in protein body formation, the glutenins of wheat endosperm is electrophoretic proteins might begin to condense within the Golgi mobility. The subunits of gliadin thus identified are vesicles, for subsequent transport to the vacuole. a-, (3-, y-, andft>-gliadins,whereas the glutenins are Support for this view has come from immunocytoconstituted of a low- and a high-molecular-mass chemical observations showing that in developing class. Early ultrastructural studies of T. vulgare and endosperm cells of T. aestivum, the ER lumen is T. durum showed the presence of electron-dense bypassed in favor of Golgi vesicles for the packagprotein deposits surrounded by tubular mem- ing and transport of gliadin (Kim et al 1988). branes probably originating from the ER in the Another immunocytochemical study, though developing endosperm. Despite the low resolution implicating the Golgi complex as the site of proof the electron micrographs, these observations led cessing and transport of the proteins, does not, to the suggestion that storage proteins are synthe- however, rule out the lumen of the ER as the site of sized on the rough ER and are deposited in protein their synthesis (Stenram, Heneen and Skerritt bodies by a process of internal secretion (Graham et 1991). From these differences in interpretation, one al 1962; Morton, Palk and Raison 1964). Subsequent is left with a less-than-satisfactory view that the investigators have persistently argued for a role for
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synthesis and transport of storage proteins in the wheat endosperm revolve around the ER and Golgi vesicles, at least transiently, in a complex way. It is unlikely that electron microscopy and immunolocalization can truly reflect the dynamics of the synthesis and transport of storage proteins in the endosperm, especially when the proteins are constituted of different subunits. Some new information about the role of the Golgi apparatus in protein transport has come from the ability of Xenopus laevis oocytes to sponsor the formation of -a and ysubunits of gliadin when injected with the respective mRNAs. Labeling with radioactive precursor combined with electrophoresis was used to follow the accumulation of proteins in the injected oocytes and in the outside medium. The striking observation made in this experiment was that whereas agliadin accumulates inside the oocyte, y-gliadin is secreted into the medium. Since oocytes of X. laevis do not have vacuoles, y-gliadin is believed to be exported from the ER to the Golgi apparatus and secreted into the medium. This view was also supported by the fact that disruption of the Golgi apparatus by monensin halts the secretion of ygliadin from the oocyte (Simon et al 1990). Evidently, some activation event determined by the structural differences between a- and y-gliadins must allow the gliadins to be distinguished for their assignment to markedly different subcellular transport routes. One final point in this context is that the expression of deletion mutants of y-gliadin in the oocytes showed that the C-terminal region is responsible for secretion of the protein (Altschuler et al 1993). Immunocytochemical evidence shows that during the transport of y-secalins, which are synthesized in the ER, to the site of their accumulation, protein body formation in rye endosperm cells is analogous to that in wheat and involves the Golgi apparatus (Krishnan, White and Pueppke 1990). The biochemical similarity of endosperm prolamins of the wild grass Haynaldia villosa (Poaceae) to wheat gliadins has made it of interest to localize the protein using appropriate antibodies. Absence of label in the inclusions of the ER and the specific labeling of protein bodies found in the vacuoles and vesicles associated with the Golgi complexes provide strong support for a role for the Golgi apparatus in the transport of storage proteins, if not in their synthesis (Krishnan, White and Pueppke 1988). The major mobilization of zein protein bodies in the maize endosperm commences when the cells
Zygotic Embryogenesis
are still undergoing division. The first change in the endosperm cell auguring its altered function is the appearance of small, membrane-enclosed, spherical deposits. The deposits begin to accumulate into dense protein bodies in vesicles produced by localized dilations of the ER that occur along the cisternae or at their enlarged ends. It is difficult to decide from the electron micrographs alone whether the protein is synthesized outside or inside the membranes (Khoo and Wolf 1970). Additional understanding of the role of ER in zein synthesis was provided by studies showing that membrane-bound polysomes and the mRNA population present in the rough ER of the endosperm direct the synthesis of virtually the same zein protein in vitro as they do in vivo (Larkins and Dalby 1975; Larkins, Bracker and Tsai 1976; Larkins, Jones and Tsai 1976; Larkins and Hurkman 1978; Melcher, 1979; Torrent et al 1986). Consistent with the view that fully formed protein bodies are surrounded by ER, a cell-free system that synthesizes zein using polysomes and mRNA derived from protein bodies as templates has also been described (Burr and Burr 1976; Burr et al 1978; Viotti et al 1978). A finding with implications for the role of ER in zein synthesis is that injection of zein mRNA into X. laevis oocytes potentiates the synthesis of proteins compartmentalized within ER vesicles (Larkins et al 1979; Hurkman et al 1981). Any disadvantage in the use of endosperm from intact grains of maize for biochemical studies has been partially ameliorated by the successful culture of the endosperm in liquid suspension or on agar medium. The cultured cells maintain their endosperm phenotype and synthesize zeins that accumulate in protein bodies as in the endosperm cells of developing kernels (Shimamoto, Ackermann and Dierks-Vending 1983; Felker 1987). Although the classification and nomenclature of zeins have remained a contentious issue for a long time, a consensus seems to have emerged in recent years. Accordingly, these proteins are classified into five individual zeins of molecular masses of about 10,15,19, 22, and 28 kDa. These are also known as a-zeins (19 and 22 kDa), fi-zein (15 kDa), y-zein (28 kDa), and 5-zein (10 kDa). Immunolocalization has shown that the different classes of zeins are distributed heterogeneously in the protein bodies, with further variation dependent upon the location of the organelles within the endosperm cells. Generally, the protein bodies of the subaleurone cells are small and contain almost exclusively |3and y-zeins, whereas those confined to the more central cells of the endosperm contain a-zeins also.
Developmental Biology of the Endosperm
An interesting feature of the protein bodies containing, a-, (3-, and y-zeins is that the oc-zein is confined to the central region of the organelles and the p"- and y-zeins become peripheral (Lending et al 1988; Lending and Larkins 1989). Cells of the rice endosperm are unusual in harboring different types of protein bodies that differ in their morphology, the nature of accumulated proteins, and their subcellular origin. A model that has gained some acceptance recognizes two types of predominantly prolamin-containing spherical protein bodies and one type of glutelinenriched, irregularly shaped crystalline body. Electron microscopic observations have supported the view that the three types of protein bodies share a common site of synthesis in the rough ER (Harris and Juliano 1977; Bechtel and Juliano 1980; Oparka and Harris 1982; Yamagata and Tanaka 1986). It has been claimed that different domains of the rough ER, designated as the protein body ER and cisternal type ER, are the site of the synthesis of prolamins and glutelins, respectively. The bases for this view are the clear identification of prolamin-derived protein bodies delimited by the rough ER and of glutelin-derived bodies delimited by single-lamellar membranes, and the differences in the content of prolamin and glutelin mRNAs in the two domains of the rough ER. Whereas the cisternal type of rough ER is enriched for glutelin mRNA, the protein body ER harbors mostly prolamin mRNA (Li, Franceschi and Okita 1993). Evidence has also appeared indicating that a special protein located in the ER lumen might bind to the nascent prolamins, promoting their folding and assembly into protein bodies and thus retaining them in the ER (X. Li et al 1993). Some studies have implicated the Golgi vesicles in a formative way in the final configuration of the prolamins and glutelins (Bechtel and Juliano 1980; Oparka and Harris 1982; Yamagata and Tanaka 1986). The definitive view of the role of the Golgi vesicles now appears to be that they mediate the transport of only the glutelin protein bodies synthesized in the cisternal type ER to the site of their deposition (Krishnan, Franceschi and Okita 1986; Li, Franceschi and Okita 1993). There is little evidence that can be marked out as suggestive of a mechanism that preferentially segregates glutelin mRNA and the protein product to the cisternal type of ER and the Golgi vesicles, respectively. Much of the interest in protein body formation in endosperms of other plants hinges on the involvement of ER in the synthesis of the proteins
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and of Golgi bodies in their transport. In the developing endosperm of oat, globulins and avenin are deposited in spatially distinct subcellular domains. The main aggregation of globulin takes place within the vacuole, whereas most of the avenin appears to assemble directly within the rough ER, whence it migrates to the vacuole (Saigo, Peterson and Holy 1983; Lending et al 1989). Additional support for the synthesis of storage proteins within the rough ER comes from observations on endosperm development in barley (Cameron-Mills and von Wettstein 1980) and Sorghum bicolor (Taylor, Schiissler and Liebenberg 1985; Krishnan, White and Pueppke 1989). In barley, the proteins, following their synthesis in the ER, are transported to the vacuoles, where they form protein bodies; in sorghum, the proteins aggregate within the site of their synthesis. There is no evidence for the involvement of Golgi vesicles in the synthesis or transport of proteins in oat, barley, and sorghum endosperms. Other abundant proteins such as thionins, which are synthesized in the endosperm of wheat and barley, are detected around storage protein bodies as electron-dense spheroids; a role for ER in their synthesis has not been demonstrated (Carmona et al 1993). Although the endosperm of Ricinus communis (castor bean; Euphorbiaceae) stores abundant oil, there seems to be no constraints on the accumulation of proteins in the same cells. The protein body of the castor bean endosperm has a relatively complex structure, containing phytin globoids and protein crystalloids embedded in an amorphous proteinaceous matrix surrounded by a single membrane. The origin of protein bodies has been traced to small vacuoles that become filled with the protein (Tully and Beevers 1976; Gifford, Greenwood and Bewley 1982). Vacuolelike inclusions containing a particulate phase are also believed to be the site of oil body synthesis in the castor bean endosperm (Harwood et al 1971). Protein Bodies of the Aleurone Cells. Unlike the cells of the starchy endosperm, the aleurone layer is made up of living cells. Although the aleurone layer is not a storage tissue like the starchy endosperm, the cells are filled with aleurone grains - spherosomes and protein granules of a chemical composition different from that of the storage proteins already considered. In wheat (Morrison, Kuo and O'Brien 1975), rice (Bechtel and Pomeranz 1977), barley (Jacobsen, Knox and Pyliotis 1971; CameronMills and von Wettstein 1980; Olsen, Potter and Kalla 1992), and oat (Bechtel and Pomeranz 1981;
Zygotic Embryogenesis
340
15 DAYS
20 DAYS
25 DAYS
35 DAYS
Figure 12.5. Diagrammatic representation of the formation of protein bodies in aleurone cells of the endosperm of Zea mays. Protein precipitates are first detected (15 days) in the vacuole, where they continue to accumulate (20 days). Later the proteins condense, forming a large, electron-
transparent core (25 days). Spherosomes are apposed to the surface of the fully developed protein body (35 days). (From Kyle and Styles 1977. Planta 137:185-193. © Springer-Verlag, Berlin.)
Peterson, Saigo and Holy 1985), the aleurone grains are packaged as discrete particles of phytin, protein, and carbohydrate enclosed within a unit membrane. In electron micrographs, the phytincontaining area of the aleurone grain is recognized as one or more electron-dense globoids; another electron-dense area is constituted of a protein-carbohydrate complex that exists as a matrix or as inclusions. Lipid droplets or spherosomes surrounding the enclosing membrane like a ring complete the profile of the aleurone grain (Jones 1969; Jacobsen, Knox and Pyliotis 1971; Morrison, Kuo and O'Brien 1975). Buttrose (1963b) showed that aleurone grains of wheat originate in vacuoles as deposits of moderate electron density, with protein accumulation progressing with the maturity of the caryopsis. The assembly of aleurone grain in the oat endosperm differs significantly from that described in wheat. Although the vacuole remains as the site of its assembly, the aleurone is initiated by a sequential and progressive deposition of phytin and protein into the vacuole, with phytin deposition preceding that of protein. Compelling evidence in support of the view that the two types of deposits are targeted to different compartments is the early subdivision of the vacuole by internal membranes (Peterson, Saigo and Holy 1985). A similar vacuolar origin of the aleurone grain in the form of dense deposits (Figure 12.5) is the rule in endosperms of barley (Cameron-Mills and von Wettstein 1980) and maize (Kyle and Styles 1977).
the work done with barley has shown that cc-amylase concerned with the digestion of starch in the endosperm is synthesized in the aleurone cells in response to GA coming from the embryo during the early hours of imbibition of the grain. Although the level of regulation of oc-amylase by GA is attributable to changes in gene expression, it is not known what keeps the aleurone cells from expressing the genes in the endosperm of developing and mature grains. Synthesis of carotenes and anthocyanins in the aleurone cells of maize endosperm has come under intense investigation from a developmental perspective, and several genes encoding enzymes for the production of these pigments have been cloned. The anthocyanin biosynthetic pathway in the aleurone cells has served as a model for studies in the genetic interactions between regulatory and structural genes. A discovery of great interest for investigations into the genetic and tissue-specific regulation of expression of genes involved in anthocyanin production is that such genes, introduced by microprojectile bombardment into aleurone cells of a mutant endosperm unable to synthesize the pigment, can faithfully program synthesis of the correct pigment exclusively in the target cells (Klein, Roth and Fromm 1989).
The aleurone cells have provoked a great deal of research in connection with the mobilization of starch during germination of cereal grains. Much of
Metabolism of Storage Proteins. The changes in the pattern of accumulation of total storage proteins and their fractions during endosperm development have been followed in wheat, maize, barley, rice, rye, sorghum, and castor bean. In wheat, the production of most storage proteins
Developmental Biology of the Endosperm
occurs throughout the period of grain maturation; toward the latter phase of maturation, however, accumulation of low-molecular-mass proteins overtakes that of high-molecular-mass proteins (Flint, Ayers and Ries 1975; Pavlov et al 1975; Mecham, Fullington and Greene 1981; Singh and Shepherd 1987). Grain filling in maize is due to an increasing accumulation of globulins and zein from two weeks after pollination up to the mature stage, whereas albumins show a two-phase pattern, reaching a peak midway through endosperm development and declining further with time (Jones, Larkins and Tsai 1977; Dierks-Ventling 1981). In all essential respects, the pattern of accumulation of glutelin in the developing maize endosperm parallels that for zein (Ludevid et al 1984). In terms of their increasing molecular mass and other properties, the prolamins of barley endosperm are designated as B-, C-, and Dhordeins; B-hordeins are sulfur rich; C-hordeins are poor in sulfur; and high-molecular-mass proteins are D-hordeins. Based on genetic studies and protein characterization, three subfamilies, B1-, B3-, and y-hordeins, have been recognized in the Bhordein group. During development, the relative amount of C-hordein is greatest in the young endosperm; it then decreases, remaining constant through grain maturity. In the B-hordein group, Blhordein is found to increase throughout endosperm development whereas B3- and yhordeins behave like C-hordein. D-hordein appears to accumulate toward the final phase of endosperm development (Shewry et al 1979; Rahman, Shewry and Miflin 1982). Another protein of potential nutritional value found in the barley endosperm is thionin; accumulation of this protein follows a course almost parallel to that of Dhordein (Ponz et al 1983). The changes in the proportions of a 75-kDa y-secalin and &>-secalin as the major and minor components, respectively, of the developing rye endosperm mirror to a large extent those for Bl- and C-hordeins of barley (Shewry et al 1983). In rice, two groups of polypeptides (22-23 kDa and 37-39 kDa) composed of glutelin continue to increase coordinately during development, whereas the levels of the 57-kDa and 76-kDa polypeptides remain fairly constant throughout endosperm development (Figure 12.6). The relative proportion of glutelin to prolamins accumulated is high in 10-day-old grains but decreases in 25-dayold grains due to an increased accumulation of prolamins (Yamagata et al 1982; Li and Okita 1993). Total proteins of castor bean endosperm (Gifford, Greenwood and Bewley 1982) and prolamins of
341
7-39k(
26k2-23k( 18k13k-
4 5 6 7 8 9 10 11 12 45 9 11 13 15 17 19 21 23 25 28 I
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Figure 12.6. Protein composition of the endosperm of Oryza sativa. (a) SDS-PAGE of the extant proteins of the developing endosperm. Numbers at the bottom refer to the fresh weight of a grain (mg) at the indicated stage, days after flowering (DAF). The apparent molecular masses of the proteins are given in kDa on the left, (b) Changes in the relative amounts of the proteins given as peak heights of densitometric scanning of the gel. The peak height of the 76-kDa protein is designated as 100%. (From Yamagata et al 1982. PlantPhysiol. 70:1094-1100. ©American Society of Plant Physiologists.)
sorghum endosperm (Taylor, Schiissler and Liebenberg 1985) show the same patterns of accumulation characterized by an almost linear increase throughout development. Assay of the major storage albumins and oleosins of the endosperm of Coricmdrum sativum (coriander; Apiaceae) using antibodies raised against similar proteins from carrot seeds showed that the albumins accumulate at a slightly earlier stage of development than the oleosins (Ross and Murphy 1992).
342 The dependence of continued accumulation of storage materials on protein synthesis has been demonstrated by the incorporation of labeled precursors in developing endosperms of rice (Luthe 1983; Zhu, Shen and Tang 1989; Li and Okita 1993), barley (Shewry et al 1979; Giese, Andersen and Doll 1983; Giese and Hejgaard 1984), wheat (Greene 1983), and oat (Luthe 1987). Because of the difficulty of harvesting endosperms at very early stages of development, these experiments have not generated a set of well-documented data on the pattern of synthesis of individual proteins during the developmental time span of the endosperm. From the foregoing account it is not clear whether the inability of endosperm cells at early stages of development to accumulate storage reserves is due to a high turnover or due to lack of the components necessary for their synthesis. Along the same lines, it is also not determined whether differences exist in the turnover rates of the different storage proteins in the species studied.
Zygotic Embryogenesis
products a few days before their detection in the endosperm cells in vivo (Figure. 12.7). Results of these studies in principle correlate well with those on protein metabolism in showing a coincidence between the periods of increased in vitro translational capacity of mRNA and increased protein synthesis or accumulation. (i) Maize Storage Protein Gene Expression
As the major class of storage proteins of the maize endosperm, zein has been studied intensively with the focus on gene expression. Following the first detection of zein mRNAs 12 days after pollination, the accumulation of these mRNAs in the developing maize endosperm reaches a maximum between 18 and 22 days, and thereafter it declines somewhat until maturation. Using groups of cDNA clones corresponding to 10-, 15-, 19-, and 22-kDa zeins as probes, it was estimated that the efficiency of expression of members of different zein families varies somewhat; generally, transcription of the gene encoding 15-kDa zein is initiated earlier and is 5. ANALYSIS OF STORAGE PROTEIN GENE more active than that of 19- and 22-kDa zeins EXPRESSION (Marks, Lindell and Larkins 1985a; Kirihara et al 1988). Patterns of accumulation of transcripts of Studies on the expression of storage protein individual zein genes also depend upon the growth genes during endosperm ontogeny have considered in vitro protein synthesis by mRNA and conditions and genotypes of the plant (Langridge, polysomes, changes in mRNA accumulation and Pintor-Toro and Feix 1982a, b; Kriz, Boston and synthesis, and the expression of cloned genes. Since Larkins 1987). At the cellular level, the noticeable these topics are of vital importance in understand- characteristic of zein genes is their exclusive expresing the structure of endosperm storage protein sion in the nucleus and cytoplasm of the endosperm genes and their expression in transgenic systems, cells, with the exception of the cells of the aleurone layer. As shown in Figure 12.8, a meaningful comthey will be briefly reviewed here. It was mentioned earlier that in vitro synthesis parison of transcript localization allows one to recof zein is programmed by polysomes and mRNA ognize specific territories of expression for each zein isolated from the developing endosperm of maize; mRNA within the developing endosperm (Dolfini et similar studies have reported the cell-free synthesis al 1992). Zein gene transcripts are also expressed in of globulins primed by polysomes of oat suspension cultures derived from the endosperm of endosperm (Luthe and Peterson 1977; Fabijanski maize, but expression occurs at a reduced level as and Altosaar 1985; Fabijanski, Matlashewski and compared to that of the endosperm of developing Altosaar 1985), hordeins and thionins primed by grains (Manzocchi, Bianchi and Viotti 1989; Ueda polysomes of barley endosperm (Brandt and and Messing 1991). Transcripts of the much less Ingversen 1976; Matthews and Miflin 1980; Ponz et investigated glutelin genes show a rate of accumulaal 1983; Rahman et al 1984), prolamins primed by tion in the developing endosperm similar to that of polysomes of sorghum endosperm 0ohari, Mehta 19- and 22-kDa zeins (Perez-Grau et al 1986). and Naik 1977), gliadin primed by polysomes and The apparent lack of coordination in the expresmRNA of wheat endosperm (Greene 1981; Okita sion of the individual zein genes raises the question and Greene 1982; Reeves, Krishnan and Okita of their control during endosperm development. 1986), secalin primed by mRNA of rye endosperm Determination of the levels of the different zein (Shewry et al 1983), and prolamins and glutelins mRNAs in run-off transcription assays from isoprimed by mRNA of rice endosperm (Yamagata lated endosperm nuclei of different ages has proand Tanaka 1986; Yamagata et al 1986). The storage vided evidence that the levels of the two oc-zein proteins are generally detected in the translation transcripts are generally higher than those of genes
343
Developmental Biology of the Endosperm
Time after anthesis (days) 18 22 26 34
14
18
22 26 30
(ft)
B1 60
f / / /
40
»
83 —»
N
20
C1
0 n.
"14 18 22 26 30
Time after anthesis (days) Figure 12.7. In vitro synthesis of hordeins by RNA isolated from the endosperm of Hordeum vulgare. Top panel: SDS-PACE of proteins translated from membrane-bound polysomes (a), polysomal poly(A)-RNA (b), polysomal RNA (c), and total RNA (d) at different days after anthesis.
Bottom panel: Densitometric determination of the relative incorporation of the labeled amino acid into proteins; (e-h) correspond, respectively, to (a-d). (From Rahman et al 1984.)
encoding p*-, y-, and 8-zeins during all developmental periods studied. Among the two a-zeins, the one encoding the 22-kDa a-zein was transcribed more actively than that encoding the 19kDa protein. Since the high transcriptional activity of the 22-kDa a-zein gene is not predicted by the low levels of its mRNA accumulation on polysomes, both transcriptional and posttranscriptional regulation of the zein genes appear to be
operating during endosperm development (Kodrzycki, Boston and Larkins 1989). Preferential accumulation of 10-kDa zein associated with the presence of a high level of its transcripts occurs in the endosperm of an inbred line of maize, BSSS-53, whereas an underexpression of this protein and its transcripts is seen in another inbred line, Mo-17 (Kirihara et al 1988; Schickler, Benner and Messing 1993). Evidence from analyses
344
Zygotic Embryogenesis
cation that they function to modulate transcript stability in some ways. PROBES In considering the relationship between the transcriptional activity of zein genes and the accuM1 mulation of protein products in the endosperm, the occurrence of mutations credited with reducing zein synthesis needs to be taken into account. Two of these mutations characterized initially were op-2 and fl-2, both of which cause a profound reduction in the synthesis of zein polypeptides while permitE19 ting an increase in the high-lysine-containing, nutritionally superior nonzein proteins (Mertz, Bates and Nelson 1964; Nelson, Mertz and Bates 1965). In the op-2 mutant, depending upon the particular M6 maize strain background, there is a partial to an almost complete inhibition of synthesis of 22-kDa zein at the same time that the levels of the lysinerich protein and elongation factor-la (EF-loc) are significantly increased (Lee et al 1976; Habben, G1 Kirleis and Larkins 1993; Habben et al 1995). The endosperm of this mutant begins to synthesize zein several days later and ceases activity earlier than the wild-type endosperm (Murphy and Dalby Figure 12.8. A diagrammatic representation of the pattern of 1971). The impact of EF-lct is that, as a lysine-rich accumulation of transcripts of zein and glutelin genes durprotein that binds aminoacyl-tRNAs to ribosomes, ing the development of the endosperm in Zea mays. The it could be expected to contribute significantly to diagram is based on in situ hybridization using probes for the increased lysine content. Kernels of the defective zein (Ml, M6, and E19) and glutelin (G1) sequences. DAP, days after pollination. (From S. F. Dolfini et al 1992. Spatial endosperm-B30 (de*-B30) mutant also show a preferregulation in the expression of structural and regulatory ential reduction in the accumulation of 22-kDa zein, storage-protein genes in Zea mays endosperm. Devel. whereas another mutation, op-7, causes a greater Genet. 13:264-276. © 1992. Reprinted by permission of reduction in the synthesis of 19-kDa than in the synWiley-Liss Inc., a subsidiary of John Wiley & Sons Inc.) thesis of 22-kDa zein. Other mutations, such as op-6, fl-2, and mucronate (me), suppress the synthesis of all of linkage relationships between the high mRNA zeins to the same extent (Lee et al 1976; Jones 1978; levels and the BSSS-53 allele for the 10-kDa zein di Fonzo et al 1979; Salamini et al 1983; Soave and structural gene locus seems to indicate that a postSalamini 1984). A newly characterized mutant, optranscriptional regulation of the mRNA level by a 15, shows a large reduction in the accumulation of trans-acting factor accounts for the increased accu27-kDa zein, but contains high concentrations of 19mulation of 10-kDa zein in the inbred line BSSS-53; and 22-kDa zeins (Dannenhoffer et al 1995). Effects a mechanism imposed by the lack of one or more of the mutation extend to the level of the structure frans-acting factors might account for the underexof the protein bodies and the distribution of the varpression phenotype in Mo-17 (Cruz-Alvarez, ious classes of zeins in them. For example, the disKirihara and Messing 1991; Schickler, Benner and organized protein bodies with clumps of P- and Messing 1993; Chaudhuri and Messing 1994). y-zeins interspersed within a mass of a-zein found Tissue-specific frans-acting factors also seem to in the endosperm cells of the/Z-2 mutant provide a determine the transcription of the 19-kDa and 15striking visual contrast to the organized organelle kDa zein genes and the 28-kDa glutelin gene by structure in the cells of the wild-type endosperm binding to their 5'-flanking regions. Five sites in the (Lending and Larkins 1992). promoter region of the 19-kDa zein gene capable of binding to a crude protein fraction of maize The complete mechanism by which the mutaendosperm nuclei include a 22-bp region contain- tions cause a reduction in zein synthesis is yet to be ing a 15-bp-long consensus sequence conserved in determined. Convincing evidence that the mutavirtually all zein genes investigated (Maier et al tions affect the regulatory functions of genes rather 1987, 1988; So and Larkins 1991; Ponte et al 1994). than their organization was provided by restriction The action of frans-acting factors carries the impli- analysis of genomic DNA of op-2, op-7, and fl-2 DAP
10
13
17
20
Developmental Biology of the Endosperm
mutants with cloned cDNA probes for zein genes; this work showed that the coding sequences of the mutant genes had no deletions or structural alterations that might affect zein accumulation (Burr and Burr 1982). The patterns of accumulation of specific zein mRNAs in the mutant endosperms are relatively well described in several studies that show that the reduction of zein proteins is correlated with reduced amounts of the corresponding mRNAs (Pedersen et al 1980; Burr and Burr 1982; Langridge, Pintor-Toro and Feix 1982a; Marks, Lindell and Larkins 1985a; Kodrzycki, Boston and Larkins 1989; Lopes et al 1994; Dannenhoffer et al 1995; Neto et al 1995). In particular, during development of the op-2 endosperm, none of the zein mRNAs can be detected at 12 days, as they would be in the normal endosperm. The mRNAs for 19kDa zeins attain relatively normal levels by 28 days. Since the mRNAs arrive at that level slowly, it is clear that the mutation delays the expression of the 19-kDa zein gene (Figure 12.9). The appearance of 15-kDa zein mRNAs is also delayed and the transcript level is greatly reduced in the mature endosperm. Although the level of 22-kDa zein polypeptides is substantially reduced by the op-2 mutation, deficiency in certain subgroups of 22kDa zein transcripts, rather than in the whole range of transcripts that encode this protein, appears to cause the reduction (Burr and Burr 1982; Marks, Lindell and Larkins 1985a). This explanation of the effects of the op-2 mutation on zein protein mRNA synthesis does not, however, preclude additional levels of regulation. In vitro culture studies have shown that the accumulation of 22-kDa zein and its mRNAs in the endosperm of the op-2 mutant is favored when an excess of nitrogen is supplied in the medium (Balconi et al 1993). Thus, there appears to be a correlation between the level of nitrogen supply and the phenotypic effect of the mutation. Two groups of investigators have cloned the OP-2 gene by transposon tagging with the help of the mobile elements suppressor mutator (spm) and activator (ac). As expected, the gene is expressed in the wild-type endosperm but not in nonendospermic tissues or in the endosperm of the op-2 mutant (Schmidt, Burr and Burr 1987; Motto et al 1988; Gallusci, Salamini and Thompson 1994). The theoretical translation of cDNA clones suggests that the gene encodes a transcription factor that contains a leucine zipper DNA-binding protein known as the bZIP domain (Hartings et al 1989; Schmidt et al 1990). The endosperm-specific expression of the OP-2 gene correlates well with the expression of
345
DAP 16 27.000
16.000 14.000 10.000
18
20 27.000
10,000
Figure 12.9. SDS-PACE of zeins extracted from wild-type (a) and op-2 mutant (b) of Zea mays to show the developmental regulation of zein gene expression. Developmental stages are indicated at the top as days after pollination (DAP). Molecular masses of the zein fractions are shown on the right. (From Kodrzycki, Boston and Larkins 1989. Plant Cell 1:105-114. © American Society of Plant Physiologists. Photograph supplied by Dr. B. A. Larkins.)
the OP-2 protein in the nuclei of maize endosperm cells (Varagona, Schmidt and Raikhel 1991). A view advanced to explain the molecular basis for zein gene expression is that OP-2 protein activates zein gene transcription by binding to promoters. Some support for this idea has come from the demonstration that the protein product of the OP-2 gene expressed in Escherichia coli can bind in vitro to upstream regions of a zein gene. Similarly, the OP-2 protein transiently expressed in the protoplasts of cultured maize endosperm cells or in the cells of the intact maize endosperm can transactivate the 22-kDa zein gene promoter (Schmidt et al 1990; Ueda et al 1992; Unger et al 1993). Conversely, the protein of an op-2 mutant causing a disruption in zein gene expression is unable to bind to the same zein gene promoter (Aukerman et al 1991). To account for these complementary results, a 20nucleotide target site has been identified in the promoter of the 22-kDa zein gene (Schmidt et al 1992). Because this sequence is not present in the promoters of other zein genes, this is prima facie evidence to explain the differential effect of the op-2 mutation on the 22-kDa zein. Unraveling the ramifications of the mutation on zein gene expression is likely to assume complex proportions as other tran-
346
scriptional regulatory proteins are identified (Yunes et al 1994; Holdsworth et al 1995). Another bZIP protein that might play a role in the regulation of the 22-kDa zein gene is the product of a maize endosperm cDNA clone (OHP-1) that binds to the OP-2 target site in the promoter region of the 22-kDa zein gene (Pysh, Aukerman and Schmidt 1993). Considering the fact that the OP-2 gene affects the expression of all subunits of zein, a finding with regulatory implications for the role of OP2 protein is that this protein transactivates the promoters of P-zein of maize and a P-zein homologue from the related member of Poaceae, Coix lacryma-jobi (Neto et al 1995). Another relevant observation is that mutation in the OP-2 gene down-regulates the accumulation of cytoplasmic pyruvate orthophosphate dikinase (PPDK) in the endosperm. Since PPDK is an enzyme involved in C4-photosynthesis, this finding might provide a meeting point for the roles of the OP-2 gene in coordinating the expression of zein genes and genes for carbon partitioning (Maddaloni et al 1996). The tide of reports of the isolation of transcription factors from the endosperm has continued to rise. Characterization of a salt-soluble 32-kDa protein (B-32) from the endosperm of wild-type maize and its absence in the op-2 and op-6 endosperms have led to the highly interesting idea that B-32 protein plays a role in the control of zein synthesis, perhaps as a transcription factor. The protein is coordinately expressed with zein during the development of the endosperm but is localized in the soluble cytoplasm, rather than in protein bodies. It has been speculated that B-32 protein is responsible for regulating zein gene expression in an indirect way. This view assumes that B-32 protein is encoded by the OP-6 gene, which in turn regulates zein synthesis, and that the OP-2 gene activates the OP-6 gene (Soave et al 1981; di Fonzo et al 1986). It is, however, doubtful that B-32 protein is a product of the OP-6 locus, and its abundance and location in the cytoplasm would argue against a direct interaction with zein or zein genes as a regulatory protein. The structure of the gene encoding B-32 protein and the deduced amino acid sequence have not given any indication of the presumed relationship of the protein to the OP-6 gene or its translation product (di Fonzo et al 1988; Hartings et al 1990; Bass et al 1992). According to Lohmer et al (1991), the B-32 gene is transactivated in the presence of the OP-2 gene product, indicating that the interaction between the OP-2 and B-32 genes is at the transcriptional level and is not indirect through the activation of the OP-6 gene. Admittedly, B-32
Zygotic Embryogenesis
protein has received a good deal of attention as a regulator of zein gene expression, but its precise function remains unresolved. Another contribution to our understanding of the action of fl-2, de*-B30, and me mutations in maize showed that endosperms carrying these mutations overproduce a 70-kDa water-soluble protein (B-70) associated with both protein bodies and the rough ER (Galante et al 1983). Dramatic increases in the levels of both B-70 protein and its corresponding mRNA occur in the endosperms of these mutants between 14 and 20 days after pollination. This is followed by a decline in mRNA levels to the vanishing point, whereas B-70 protein levels remain high in the morphologically perturbed protein bodies (Boston et al 1991; Zhang and Boston, 1992). It appears that B-70 is a cytosolic chaperonelike protein with affinities to BiP and HSP-70 (Fontes et al 1991; Marocco et al 1991). Another candidate for a regulatory function in zein gene expression is an unusual 24-kDa protein associated with the zein fraction of the endosperm of the fl-2 mutant (Lopes et al 1994). Results from the cloning of the gene for the 24-kDa protein and from its deduced amino acid sequence indicate that the gene is expressed as a 22-kDa ct-zein with a defective signal peptide (Coleman et al 1995). Irrespective of the nature of the B-70 and 24-kDa proteins, there is circumstantial evidence to suggest they are involved in some ways in the disruption of zein processing and deposition in the endosperms of the mutants. The phenotypic effect of the op-2 mutation typically results in the formation of a soft, floury endosperm. A significant development affecting the phenotype of the op-2 mutant is the modification of all or parts of the endosperm to a vitreous or glassy consistency. Genes that condition this phenotype are known as OP-2 MODIFIERS; the same genes are also active in changing the phenotype of the endosperm in other mutants such as fl-2. Wallace et al (1990) showed that the principal change in the endosperm storage protein of the modified op-2 mutant is a two- to fourfold increase in the y-zein content without any appreciable effect on the other classes of zeins. A central question in analyzing the effects of the various modifier genes is whether there are differences in the fundamental mechanisms of gene transcription or posttranslational events affecting protein synthesis. Experiments designed to answer this question have shown that opaque modifiers confer an increase in the level of y-zein mRNAs; since this occurs in the absence of any increase in the number of gene copies, it appears that the increased gene expression is due to
Developmental Biology of the Endosperm
enhanced mRNA transcription or stability (Geetha et al 1991; Lopes and Larkins 1991). However, there is only a marginal difference between the transcription rates of the "y-zein gene in the op-2 mutant and in the modified op-2 genotypes. This has ruled out a direct role for the modifier genes in the transcription of the y-zein gene and suggests a posttranscriptional role for them (Or, Boyer and Larkins 1993). (ii) Barley Storage Protein Gene Expression
Some observations support the view that the amounts of different hordeins synthesized by the developing barley endosperm are determined by the abundance of corresponding mRNAs present, with little room for translational and posttranslational controls. Quantification of the relative amounts of B1-, B3-, and C-hordeins synthesized in vitro by mRNA preparations from developing endosperm showed that the amount of Bl-hordein increased, and that of B3- and C-hordeins decreased, with age. The population of the corresponding mRNA species determined by dot-blot hybridization using cDNA clones also showed similar changes during endosperm development (Rahman et al 1984). It must be kept in mind that this kind of correlative evidence can never be considered complete because it is difficult to ascertain whether the amount of mRNA present at any given time is due to gene transcription or mRNA turnover. However, in vitro transcription rates of all hordein genes are more or less the same despite the fact that the steady-state levels of some hordein mRNAs are low; this indicates that the high levels of hordein mRNAs are due to posttranscriptional regulations (Sorensen, CameronMills and Brandt 1989).
347
Aalen et al 1994; Kalla et al 1994; Aalen 1995). Transcripts of another aleurone-specific recombinant (LTP-1) also accumulate characteristically in the aleurone cells during late grain development (Skriver et al 1992). Steady-state levels of mRNAs that encode a protein with some sequence homology to LEA proteins begin to accumulate in the aleurone and in the embryo midway through development and remain high in the aleurone cells of the mature endosperm (Hong, Barg and Ho 1992). Genes for chitinase isolated from barley endosperm display either aleurone-specific or both aleuroneand starchy endosperm-specific patterns of expression. This protein is generally reckoned as part of the grain's defense arsenal against invading pathogens (Swegle, Kramer and Muthukrishnan 1992; Leah et al 1994). Mutants impaired in the synthesis and deposition of hordeins have been identified in cultivated varieties of barley. The mutant Rise-56 suffers a 30% reduction in overall hordein accumulation and, in particular, the amount of B-hordein is reduced by about 75%. This reduction has been traced to the absence of B-hordein mRNA in the mutant endosperm due to a deletion covering most of the hordein-2 locus (Kreis et al 1983). In another mutant, Ris0-15O8, the amounts of B- and Chordeins are reduced to about 20% and 7%, respectively, of those present in the parent line at the same time as the amount of D-hordein is increased fourfold. The changed amounts of B-, C-, and Dhordeins in this mutant are accompanied by reduced levels of B- and C-hordein mRNAs and a twofold increase in the amount of D-hordein mRNA, and by a dramatic reduction in the in vitro transcription activity of the genes encoding B- and C-hordeins (Thompson and Bartels 1983; Kreis et al 1984; Cameron-Mills and Brandt 1988; S0rensen, Cameron-Mills and Brandt 1989). Thus, the primary lesion caused by the mutations appears to be at the transcriptional level. The catastrophic effects of the mutation in Rise-1508 have been further traced to transcriptional inactivation of the genes by methylation, whereas hypomethylation of the genes is correlated with their specific expression in the endosperm of the parent (Sarensen 1992).
The use of cDNA clones isolated from aleurone cells of the barley endosperm has yielded vital information on the time course and pattern of accumulation of aleurone mRNAs. Transcripts of an aleurone-specific cDNA clone (B11E) and its genomic clone (LTP-2) show a nearly 200-fold increase in abundance exclusively in the aleurone cells beginning in the early-stage endosperm and extending to the stage of peak accumulation. During desiccation of the grain, transcript levels decrease; they disappear completely from the aleurone cells of the mature grain (Figure 12.10). In con(iii) Wheat Storage Protein Gene trast, mRNAs complementary to other cDNA clones Expression expressed in both the aleurone and the embryo are present more or less uniformly in the aleurone Storage protein mRNAs of the developing throughout grain development and decrease coinci- wheat endosperm are attached to both membranedentally with those of B11E (Jakobsen et al 1989; bound and free polysomes, with the former makOlsen, Jakobsen and Schmelzer 1990; Liu et al 1992; ing up the major component of the storage protein
348
Zygotic Embryogenesis
DPA
0
2 4
OPI
6 8 10 12 16 20 25 30 35 40 45 50 2
kb
Figure 12.10. Temporal and spatial patterns of accumulation of transcripts of aleurone layer-specific genes during the development of the endosperm in Hordeum vulgare. (a) Northern blot hybridization showing steady-state levels of B11E mRNA in unfertilized ovules (0 days postanthesis, DPA), deembryonated grains (2-10 DPA), isolated aleurone-pericarp (12-45 DPA), mature aleurone (50 DPA), and mature aleurone imbibed for two days (2 days postimbibition, DPI). (From Olsen, jakobsen and Schmelzer
1990. Planta 181:462-466. © Springer-Verlag, Berlin.) (b, c) In situ hybridization of a section of the grain with labeled LTP-2 antisense strand. Dark-field micrographs showing hybridization of the probe to the peripheral aleurone cells, but not to the starchy cells of the endosperm. A portion boxed in (b) is magnified in (c). AL, aleurone layer; CC, cross-cell layer; MA, modified aleurone cells; NE, nucellar epidermis; PE, pericarp; SE, starchy endosperm. Scale bars = 100 urn (b) and 250 urn (c). (From Kalla et al 1994.)
messages. Based on hybridization kinetics, it has been estimated that both types of polysomes are constituted only of superabundant and intermediate abundant classes of mRNAs. Although there is no evidence for selective amplification of gliadin and glutenin genes in the developing wheat endosperm, it appears that more than half of the translated mRNAs direct the synthesis of the gliadins (Pernollet and Vaillant 1984). The results of changing patterns in RNA synthesis and in levels of the transcripts of different subunits of gliadins and glutenins have been taken to indicate that the subunit genes are expressed coordinately and that the amounts of mRNAs for the proteins increase sharply until the mid-maturation phase of endosperm development and then decline. The major RNA transcripts that accumulate during the peak period of endosperm development encode the a/P- and y-gliadins. The pattern of accumulation of mRNAs of triticin, a globulinlike storage protein that constitutes only about 5% of the total wheat endosperm protein, is very similar to that of
gliadins. As no protein is detected in the developing endosperm before its homologous mRNA is synthesized, a transcriptional control mechanism appears to coordinate storage protein synthesis in the wheat endosperm (Terce-Laforgue and Pernollet 1982; Okita 1984; Bartels and Thompson 1986; Reeves, Krishnan and Okita 1986; Singh et al 1993). These data are in general accord with those described for maize and barley endosperms. In attempts to characterize the protein factors that regulate transcription of gliadin genes, Vellanoweth and Okita (1993) have identified six sequences in the promoter of a oc/p-gliadin gene that interact with different nuclear proteins from the developing endosperm (Figure 12.11). When nuclear extracts from endosperms of different stages of development were assayed, activities of four of the nuclear factors were found to appear at early stage to midstage, reach a maximum at midstage, and then decline later during endosperm ontogeny. The temporal changes in the activities of nuclear factors are coincident with the periods of
Developmental Biology of the Endosperm
349 -163 Spel
-265 Xbal
-46 Ncol
I
CHD
CAAT
xs
TATA
SN
SN
c5 c4 c3 c2 d
1 2
3
4
7
8
9
10
11 12 13 14
Figure 12.11. Interaction of nuclear proteins of Triticum aestivum with the cx/p-gliadin promoter, (a) A schematic representation of the promoter region of the o/P-gliadin gene. Numbers refer to positions upstream of the transcriptional start site. TATA and CAAT elements are shown in boxes. The arrow indicates the direction of transcription. The probe and competitor fragments used in the experiment shown in (b) are indicated, (b) Electrophoretic mobility shift assay using restriction fragments SN (-163 to -46 bp) in lanes 1-7 and XS (-265 to -163 bp) in lanes 8-14.
The amounts of crude nuclear proteins used were (ug): 0.5 (lanes 1, 8); 1 (lanes 2, 9); 2 (lanes 3, 10); 4 (lanes 4, 6, 7, 11, 13, 14); and 8 (lanes 5, 12). Lanes 6 and 13 and lanes 7 and 14 contained a 100-fold molar excess of unlabeled XS and SN DNA fragments, respectively. The five complexes formed with SN are indicated on the left as d to c5; those formed with XS are marked on the right as d ' to c4'. A sixth, low-mobility complex formed with SN is not seen in the figure. (From Vellanoweth and Okita 1993; photograph supplied by Dr. T. W. Okita.)
maximum accumulation and decrease of gliadin mRNA in the endosperm. Activity of a fifth nuclear factor was found at high levels only at mid-development; a sixth nuclear factor displayed a constant level of activity throughout endosperm development. Because the last-mentioned nuclear factor remains active even after transcription for gliadins has stopped, it has been surmised that some binding proteins may regulate gliadin expression by a negative frans-acting effect. An interaction between the putative trans-acting factors and sequences of the endosperm box (see p. 350) and GCN4-like (for General Control Nonderepressible) motifs
that bind yeast regulatory proteins found in the low-molecular-mass glutenin has also been demonstrated by in vivo dimethylsulfate (DMS) footprinting. Not only is the binding highly specific to endosperm nuclear proteins, but the two types of proteins are found to bind sequentially, whereby the endosperm box motif becomes occupied during the early stages of development of the endosperm and the GCN4 sequence is coupled during the later stages (Hammond-Kosack, Holdsworth and Bevan 1993). From these results, a good case can be made that specific sequences in the promoter regions of the gliadin and glutenin
350 genes combine with endosperm nuclear factors to regulate their temporal and spatial expression during development. (iv) Storage Protein Gene Expression in Rice and Oat
Most of the evidence linking storage protein gene expression with endosperm development in rice comes from studies of glutelin and prolamin genes, the former representing a significantly higher proportion of the total proteins than prolamins. What is the nature of the control imposed on the relative expression of these proteins? In the absence of significant differences in the steady-state levels and transcriptional rates of glutelin and prolamin genes in the developing endosperm, it has been argued that differential recruitment of the respective mRNAs into polysomes might explain the differences in protein accumulation (Kim, Li and Okita 1993). In a similar way, there is a predominance of globulin over avenin in the endosperm of oat; yet, the amount of avenin mRNA is equal to or greater than the amount of globulin mRNA present during most periods of endosperm development (Chesnut et al 1989). Given the uncertainties about the involvement of nontranscriptional processes in the synthesis of proteins, the reasons for the translational efficiencies of these genes deserve further study. The possibility that expression of glutelin and prolamin genes during rice endosperm development is noncoordinately regulated arises out of experiments on the transcriptional activities and transcript levels of several cDNA clones of these gene families. Comparative patterns of expression of 15.2-kDa prolamin messages belonging to two different homology classes showed that although both mRNAs increase during the 10 to 25 days following anthesis, there is a three- to fourfold increase in the abundance of one message during the early phase of endosperm development (Kim and Okita 1988b; Kim, Li and Okita 1993). Like the 15.2-kDa prolamin genes, the gene for a 13-kDa prolamin continues to be expressed at a high level up to the late stage of endosperm development (Masumura et al 1990). The expression of glutelin genes belonging to three subfamilies (GT-1, GT-2, and GT-3) shows that although transcripts of the homologous GT-1/GT-2 genes steadily accumulate during endosperm development beginning 5 days after anthesis, those of GT-3 accumulate maximally during the 5 to 10 days after anthesis, followed by a steady decline to imperceptible levels (Okita
Zygotic Embryogenesis
et al 1989; Takaiwa and Oono 1991; Takaiwa et al 1991; Kim, Li and Okita 1993). At different times during rice grain development, the glutelin gene is expressed not only in the endosperm but also in the embryo and pericarp (Ramachandran and Raghavan 1990). Izawa et al (1994) have characterized a bZIP transcription factor from the rice endosperm (RITA1) that shows an affinity to ACGT motifs. The expression of the RITA-1 gene in the aleurone and starchy endosperm cells of transgenic rice grain implies that this transcription factor might play a major role in the control of storage protein gene expression in the endosperm. 6. CLONING AND CHARACTERIZATION OF ENDOSPERM STORAGE PROTEIN GENES
Isolation and characterization of the relevant genes are necessary to gain insight into the molecular mechanisms of storage protein synthesis in the endosperm. The structure of the genes encoding many endosperm storage proteins is now known. Analyses have defined and delineated the differences among the different genes and have led to the recognition of a 7-nucleotide sequence (TGTAAAG), termed the -300 bp element, in the endosperm storage protein genes of several cereals. This sequence is frequently referred to as the "endosperm box" and is believed to program the endosperm-specific expression of the genes (Figure 12.12). Later progress in this area came with the dissection of promoter sequences of the genes, construction of deletion mutants, and introduction of chimeric genes into transgenic plants to follow the genes' spatial and temporal expression. This section presents an overview of recent findings pertaining to the various levels of characterization of endosperm storage protein genes. (i) Zein Genes
Based on classical genetic approaches, nucleic acid hybridization analysis, and Southern blot hybridization, there have been several revisions of the estimated number of genes encoding zein. The consensus of the prevailing opinion is that a-zein is encoded by a large multigene family of 50-100 gene members, whereas (3-, y-, and 8-zeins are encoded by genes present in a few copies. Several cDNA and genomic clones encoding zeins have been characterized to provide a clear picture of the molecular biology of the gene. Genes characterized during the early period of zein gene cloning
Developmental Biology of the Endosperm
PBHR184:
•563
351 -302 WCAT6TAAAGTGMTAAGGT
pW8233:
GTAAAGTGAMTAAGATGAGlrCATGCAT
pMLl: ZG99: -333 Figure 12.12. Conserved distal sequences of the -300 bp endosperm box in hordein (pBHR184), a-gliadin (pW8233), 21-kDa zein (pML1), and 19-kDa zein (ZG99) genes. The sequence of the repeat of the -300 bp element in the B1 hordein gene is incomplete because the repeat contains one of the two EcoR1 sites that define the boundaries of the
sequenced fragment. Asterisks indicate identical residues in each pair of aligned sequences; those parts of the element that are common to the prolamin and zein genes are boxed. Numbering is relative to the ATC codon. (From Forde et al 1985a. Nucl. Acids Res. 13:7327-7339, by permission of Oxford University Press.)
belonged to the two major classes of oc-zeins (19 kDa and 22 kDa). A list of cDNA or genomic clones of the two a-zeins includes those isolated by Weinand, Briischke and Feix (1979), Geraghty et al (1981), Wienand, Langridge and Feix (1981), Burr et al (1982), Geraghty, Messing and Rubenstein (1982), Marks and Larkins (1982), Hu et al (1982), Spena, Viotti and Pirrotta (1982), Pedersen et al (1982), Heidecker and Messing (1983), Marks, Lindell and Larkins (1985b), Kriz, Boston and Larkins (1987), Wandelt and Feix (1989), and Liu and Rubenstein (1993). Clones encoding 15-kDa zein (Marks and Larkins 1982; Marks, Lindell and Larkins 1985b; Pedersen et al 1986), 27-kDa zein (Wang and Esen 1986; Das and Messing 1987), and 10-kDa zein (Kirihara, Petri and Messing 1988; Kirihara et al 1988), were isolated subsequently, completing the characterization of clones representing all zein subclasses. cDNA clones corresponding to some storage proteins of maize endosperm previously considered as zeins have been assigned to 28-kDa and 16-kDa glutelins because of their close homology in both nucleotide and amino acid sequences to gliadin and hordein (Prat et al 1985; Boronat et al 1986; Prat, Perez-Grau and Puigdomenech 1987; Gallardo et al 1988; Reina et al 1990a, b).
genes have consensus CAAT and TATA sequences in their 5'-noncoding region and one or more polyadenylation sites in the 3'-noncoding part. Differences between the different zein genes, which probably account for their operation, can be traced to the duplication of an original gene, the presence of repeated sequences, the insertion of sequences by duplication of small segments, and nucleotide changes. Based on the structure of individual genes, there is some interesting speculation that members of the zein multigene family have evolved from a common ancestral sequence. Protein structures of zeins deduced from cDNA clones show no homology to major storage proteins of other cereals. Among the different zeins, there is close similarity between the 19-kDa and 22-kDa zeins, as both contain a sequence of 20 amino acids repeated nine times. The amino acid repeats are not part of the sequence of the 15-kDa zein, which thus does not share homology with the 19-kDa and 22kDa zeins. The mature 10-kDa has no homology to the other zeins characterized (Wilson and Larkins 1984; Heidecker and Messing 1986). Although the sequences and structure of the 28-kDa glutelins are different from those of zeins, the 5'- and 3'-flanking regions of the two groups of genes show considerable homology (Boronat et al 1986).
Comments about the structure of the zein genes will be limited to general features that are important in their functioning. The zein genes belong to a rare class of eukaryotic genes that do not contain any introns, thus making their coding regions very long. All zein genes have repeated sequences, transcriptional regulatory sequences characterized by the presence of two or more promoters, as well as the —300 bp element and transcription initiation sites. The unusual promoter system may account for the high level of zein expression in the endosperm. Like typical eukaryotic genes, zene
The transcriptional regulation of zein genes has been monitored by introducing individual genomic clones into heterologous tissues and examining their expression in the absence of other members of the multigene families. Transformed yeast cells used as a transcription system are able to recognize the promoter regions of 19- and 22-kDa zein genes and synthesize the appropriate transcripts (Langridge et al 1984). By integrating the gene into Agrobacterium tumefaciens, it was shown that the 5'flanking regions of 15-kDa, 19-kDa, and 22-kDa zein genes contain sufficient information to accu-
352
Zygotic Embryogenesis
rately direct transcription in transformed sun(ii) Hordeins and Other Genes flower tissues or in transgenic Petunia hybrida from Barley (Matzke et al 1984; Goldsbrough, Gelvin and Larkins 1986; Ueng et al 1988). In addition, the proThe first characterized hordein genes included moters of some zein genes have been shown to those for B1-, B3-, and C-hordeins (Forde et al direct expression of fused CAT, GUS, or luciferase 1985a, b). Comparison of the 5'-flanking regions of genes in protoplasts of carrot somatic cells, tobacco barley, wheat, and maize prolamin genes showed mesophyll cells, and maize endosperm trans- the presence of short, highly conserved sequences formed by electroporation or by polyethylene gly- within 600 bp of the translation initiation site. col (Boston et al 1987; Schwall and Feix 1988; Besides a CAAT sequence and a TATA-box, a Quayle, Hetz and Feix 1991; Giovinazzo et al 1992). sequence with potential significance in the control The effect of deletion of DNA sequences flanking of gene expression was conserved around -300 bp the gene encoding a 19-kDa zein on zein mRNA in all three classes of prolamin genes. Sequence expression in sunflower tissues and CAT expres- homologies were also found between B- and Csion in carrot protoplasts has helped to identify hordeins. Other investigators have subsequently functional upstream noncoding sequences that are cloned and characterized additional Bl- (Brandt et required for optimum transcription. This region is al 1985; Chernyshev et al 1989), y- (Cameron-Mills unusual in sharing minor homology with a virus and Brandt 1988), and C-hordein (Rasmussen and enhancer core sequence (Roussell et al 1988; Brandt 1986; Entwistle 1988) genes. The aleuroneThompson et al 1990). specific genes LTP-1 and LTP-2 from barley encode Initial experiments using zein genes under the a putative nonspecific lipid transfer protein; both control of homologous regulatory elements did not genes contain regulatory motifs such as the MYB lead to gene expression in a tissue-specific manner (identified with myeloblastosis-associated viruses) and did not always reveal the synthesis of and MYC (identified with myelocytomatosis-assodetectable protein in the cells. In later experiments, ciated viruses) recognition sites (Linnestad et al transcriptional fusions composed of 15-, 19-, and 1991; Skriver et al 1992; Kalla et al 1994). These con28-kDa zein genes placed under the control of the sensus binding sites might interact in the regulaflanking regions of the gene for (3-phaseolin (the tion of the LTP-1 and LTP-2 genes, whose protein globulin storage protein of the embryos of products may have a protective function in the Phaseolus vulgaris), CaMV 35S promoter, or the GUS aleurone layer. A gene designated as B22E, which is reporter gene were found to be expressed in the expressed in the endosperm and in the embryo of protein bodies of the endosperm and embryo of barley, encodes two unidentified proteins charactransformed Petunia hybrida, tobacco, and terized by high cysteine contents (Klemsdal et al Arabidopsis (Hoffman et al 1987; Williamson et al 1991). 1988; Quattrocchio et al 1990; Geli, Torrent and In the first report of a regulated expression of a Ludevid 1994; Bagga et al 1995). However, reports barley hordein gene in transgenic plants, Marris et of endosperm-specific activity in transgenic al (1988) fused the 5'-flanking region of Bl-hordein tobacco plants of a 22-kDa zein gene promoter gene to the CAT reporter gene. Transfer of the placed upstream of the GUS reporter gene have chimeric gene into tobacco plants showed that the proved to be unreliable (Schernthaner, Matzke and hordein gene regulatory sequences direct the synMatzke 1988; Matzke et al 1990; Matzke, Stoger and thesis of CAT only in the endosperm in a developMatzke 1993). Both 19-kDa and 22-kDa zeins are mentally regulated manner. However, attempts to synthesized in the cytoplasm of unicellular, uninu- transform plants with other constructs of the cleate green alga Acetabularia mediterranea follow- hordein gene have given variable results. For ing microinjection of the genes into the nuclei and example, a chimeric B22E promoter-GlTS gene the transplantation of the nuclei into enucleate introduced into barley grains by particle bombardalgal cells (Langridge et al 1985; Brown, Wandelt ment is functional in directing transcription in the and Feix 1986). In this system, and in the Hela cell aleurone cells of the endosperm, but it is not active and Xenopus germinal vesicle transcription sys- in the seeds of transformed tobacco (Klemsdal et al tems, the 19- and 22-kDa zein genes are transcribed 1991). The expression of the promoter of the Blwith unequal efficiency from two widely placed hordein gene linked to the GUS gene delivered promoters (Langridge, Pintor-Toro and Feix 1982b; into barley endosperm is confined to mature aleuLangridge and Feix 1983; Brown, Wandelt and Feix rone cells and excludes the developing aleurone 1986). cells (Knudsen and Miiller 1991). Use of a similar
Developmental Biology of the Endosperm
type of transient assay for deletion analysis of the promoter of chitinase gene CHI-26 has made it possible to identify a 20-bp sequence that contains two hexameric direct repeats and confers developmental and aleurone-specific expression (Leah et al 1994). In the transient assay, the promoters of the LTP-1 and LTP-2 genes also display activity in the aleurone cells (Skriver et al 1992; Kalla et al 1994). It appears that barley endosperm storage protein genes contain sequences specific to conferring expression separately in monocot and dicot tissues and in the aleurone and starchy cells of the endosperm. (iii) Gliadin and Glutenin Genes A large multigene family encodes the gliadins, which have probably evolved by repeated divergence from an ancestral gene. The various gliadins are related and are coded by three gene subfamilies, namely, ct/p-, y-, and o-gliadins, the oc/p-type being the largest subfamily. The basic structural features of the coding regions and promoter elements of the oc/ P-gliadins have been deduced from cDNA and genomic clones (Bartels and Thompson 1983; Forde et al 1983; Anderson et al 1984; Kasarda et al 1984; Rafalski et al 1984; Okita, Cheesbrough and Reeves 1985; Sumner-Smith et al 1985; Reeves and Okita 1987). Deletions, duplications, and single base substitutions are believed to generate the sequence differences between the genes in each subfamily. Genomic sequences have revealed typical 5'-promoter elements, two polyadenylation signals, and a lack of introns. At the protein level, the deduced amino acid sequences share a primary structure of six domains consisting of a signal peptide, a region of repeated sequences arranged in tandem, and two polyglutamine stretches interrupted by two regions having distinctive amino acid compositions. There is a close correspondence between the coding sequences of oc/p-gliadin and y-gliadin genes, especially in the lack of introns. The amino acid sequences derived from the ygliadin gene reveal a signal peptide, a region high in glutamine and proline, and a C-terminus (Okita 1984; Okita, Cheesbrough and Reeves 1985; Bartels et al 1986; Rafalski 1986; Sugiyama, Rafalski and Soil 1986; Scheets and Hedgcoth 1988). The main focus of nucleotide sequence analysis of the glutenin genes has been on the high-molecular-mass subunits. They are encoded by 6-18 genes and belong to a small multigene family. The coding sequences of the genes are uninterrupted by introns, and the promoter regions contain a
353
sequence that somewhat resembles the conserved -300 bp element of the endosperm box. The proteins encoded by the glutenin subunit genes possess a structure composed of three linked domains, namely, an N-terminal domain of about 100 amino acids, a repetitive region of over 500 amino acids, and a C-terminus of about 40 residues (Thompson et al 1983; J. Forde et al 1985; Sugiyama et al 1985; Thompson, Bartels and Harberd 1985; Halford et al 1987; O. D. Anderson et al 1989). Several ris-acting motifs that include the endosperm box and the DNA sequences that bind the GCN4-JUN (for JUNNA) class of mammalian and yeast regulatory proteins form a conspicuous part of the structure of a low-molecular-mass glutenin gene. At the protein level, the sequence has a signal peptide, a central repetitive region enriched for glutamine and proline residues, and N- and C-terminals (Colot et al 1989). Although the nucleotide sequence of a triticin cDNA clone shares some features of gliadin and glutenin genes, its predicted amino acid sequence shows strong homology to US proteins of legume embryos (Singh et al 1993). The sequences of the gliadin and glutenin genes that regulate their expression have been sought by transient assays and by introduction into heterologous hosts. Promoters of a/P-gliadins were weak in transient expression assays in tobacco protoplasts and in protoplasts prepared from maize endosperm suspension culture (Aryan, An and Okita 1991; Blechl et al 1994). However, parts of the gliadin promoter placed upstream of either a nonfunctional or a functional version of the NOS promoter can restore the transcriptional activity of the nonfunctional element or enhance the activity of the functional sequence in tobacco protoplasts (Aryan, An and Okita 1991). Approaches to identify cfs-acting elements important in the spatial and tissue-specific transcription of glutenin genes have been more rewarding than with gliadin genes. Chimeric genes consisting of low-molecular- and high-molecularmass glutenin gene promoters linked to the CAT gene display endosperm-specific expression in the seeds of transgenic tobacco. The results of deletion series of low- and high-molecular-mass glutenin genes showed that sufficient information to confer endosperm-specific expression lies between -360 to -160 bp and -433 bp upstream of the respective promoters (Colot et al 1987; Robert, Thompson and Flavell 1989). When transcription of truncated high-molecular-mass glutenin gene fragments was enhanced by CaMV 35S promoter, only sequences spanning -375 to -45 bp in the upstream region
354
Zygotic Embryogenesis Endosperm-specific expression
pGUS45T
3.35 ± 2.05 (13) •45
POUS45R
-375
-*—
1
pGUS45
-45
1
•148
I
- * • - » -
1 -375
pousise
1
pQUSZM
1 "I-
1.51 ± 1.00(10)
-ise |
—^-
-375
3.56 ± 2.25 (14)
1 -1S3 |
•378 POUS163
2.74 ± 1.59 (14)
I
- • « •
-375 pOUS148
2.36 ± 1.76 (10)
I
-37S
0.51 ± 0.31 (15)
-234
0.26 ± 0.22 (18)
I
POUS283
0.34 ± 0.16(9)
pen 21 .s
0.32 ± 0.12 (7)
POUS45O
J
0.66 ± 0.58 (9)
Figure 12.13. Deletion analysis of the promoter of a highmolecular-mass wheat glutenin gene in transgenic tobacco. The acceptor plasmid contained CaMV 35S promoter, the CUS gene, and the 3'-end of the NOS gene. The coordinates on the glutenin gene indicate the distance in bp of the end points from the initiation site for glutenin transcription. Polarity of the promoter fragments is shown by arrows.
Data on the right indicate GUS activity (picomoles ± standard error) in the seeds of transgenic tobacco; numbers in parentheses are the number of plants used for CUS assay. Endosperm-specific expression is indicated by + or - . (From Thomas and Flavell 1990. Plant Cell 2:1171 -1180. © American Society of Plant Physiologists.)
were found essential to potentiate endosperm-specific GUS gene expression in transgenic tobacco seeds (Figure 12.13) (Thomas and Flavell 1990). A functional analysis of the 5'-flanking region of a silent high-molecular-mass glutenin gene in transgenic tobacco has led to the proposal that single base changes in the -280 bp sequence immediately upstream of the transcription start site might result in the inactivation of the promoter. Sequences required for tissue and temporal specificity of the expressed high-molecular-mass glutenin gene in transgenic tobacco are also incorporated in the -280 bp region; since this includes only two core sequences of the conserved -300 bp element, there is some question about the essentiality of the latter for tissue-specific expression of the gene (Halford et al 1989). In transient expression assays using protoplasts
the high-molecular-mass glutenin gene. This is apparently mediated by sequences found in the endosperm box of the glutenin promoter and suggests that such binding sites could act as cis-acting elements in regulating gene expression (Hilson et al 1990). The ability of Escherichia coli and yeast cells to express biologically active proteins has made possible an important advance in analyzing wheat storage protein gene expression (Bartels, Thompson and Rothstein 1985; Neill et al 1987; Galili 1989). Galili (1989) showed that the DNA sequence encoding the glutenin high-molecular-mass subunit cloned into a bacterial expression vector directs the synthesis of large amounts of the corresponding protein. Confirmation that the protein synthesized by the bacteria is identical to the native wheat protein subunit was provided by solubility characteristics. A practical consequence of this discovery is that it makes possible the analysis of the structure of the protein and the identification of its sequences that affect the quality of the dough.
of Nicotiana plumbaginifolia, mammalian transcrip-
tion factors of the leucine zipper class are found to stimulate transcriptional activity of promoters of
Developmental Biology of the Endosperm
355
moters effective in directing transcription in isolated tobacco protoplasts and in transgenic tobacco. As indicated in the previous section, three sub- Three regions of the GT-3 promoter, -945 to -726 families comprise the rice glutelin genes; each sub- bp, -346 to -263 bp, and -181 to +7 bp from the family contains five to eight copies. Several cDNA transcription start site, are required for the clones and genomic clones of glutelin genes have endosperm-specific expression and temporal regubeen isolated and characterized (Takaiwa, Kikuchi lation of chimeric genes in transgenic plants (Leisy and Oono 1986, 1987, 1989; Takaiwa et al 1987; et al 1989; Okita et al 1989; Zhao, Leisy and Okita Masumura et al 1989a; Okita et al 1989; Takaiwa 1994; Croissant-Sych and Okita 1996). A major difand Oono 1991). Genes of all three classes contain ference between glutelin genes of the GT-3 and GTthree short introns precisely at the same positions 1 classes is that in the 5'-flanking region of the in the coding regions. A comparison of genomic latter, information required for both tissue speciclones showed that the 5'-flanking regions of GT-1 ficity and temporal regulation is located in a single and GT-2 are closely related, whereas GT-3 has lit- domain (between -441 and -237 bp) (Takaiwa, tle or no homology upstream of -267 bp to either Oono and Kato 1991). A difference between the GT-1 or GT-2. The derived sequence of the protein location of the promoter elements of the GT-1 gene shows an amino terminus with features typical of a in heterologous and homologous hosts was also signal peptide. From molecular analyses of glutelin revealed when the gene was expressed in transcDNA clones, sequence homology of the primary genic rice, suggesting limitations in the recognition sequence of the gene to the US globulin gene of of critical sequences by transcription factors in the heterologous host (Zheng et al 1993). The legume embryos has become apparent. Analysis of rice prolamin cDNA clones encod- endosperm-specific expression of a rice prolamin ing a 15.2-kDa protein revealed that they contain gene in transgenic tobacco appears to be controlled relatively long 5'-untranslated sequences of about by the upstream region from -680 to -18 bp (Zhou 150-200 bp followed by coding regions of about and Fan 1993). As detected by transient assay, 450 bp. The encoded proteins have a signal peptide endosperm-specific expression of the y-kafirin gene but conspicuously lack repeated sequences (Kim is conserved in the grains of maize, sorghum, and and Okita 1988a, b). Prolamin cDNAs that encode C. lacryma-jobi and is caused by a -285 bp proximal 10-kDa and 13-kDa proteins show close similarity region of the promoter (de Freitas et al 1994). to the structure of cDNA clones that code for the An aleurone-specific gene, LTP-2, from barley 15.2-kDa protein (Masumura et al 1989b, 1990). endosperm that encodes a nonspecific lipid transfer The oat globulin gene is a member of a small protein has been discussed earlier. Kalla et al (1994) multigene family with a nucleotide sequence char- have shown that expression of this gene is under acterized by the presence of three introns and the strict temporal and spatial control in transgenic rice, lack of a -300 bp element. The protein product directing transcription exclusively in the aleurone shares structural features with the 11S globulin of layer from shortly after the onset of aleurone cell legume embryos (Walburg and Larkins 1986; differentiation to mid-maturity of the grain. At preShotwell et al 1988, 1990; Schubert et al 1990). The sent, to account for the differential expression of this nucleotide sequence of avenin, a member of a gene in the developing endosperm, it is assumed multigene family of about 25 genes, shows signifi- that the active promoter sequences contain cis-regucant homology to the gliadins of wheat and the B- latory elements that bind to nuclear proteins. hordein of barley; the coding sequence lacks Although transcripts of the 12S oat globulin gene introns, and the promoter region has a -300 bp ele- are found in both the embryo and the endosperm ment (Chesnut et al 1989; Shotwell et al 1990). of transgenic tobacco, the protein product is localGenes for prolamins of rye (y- and a)-secalin) (Kreis ized exclusively in the endosperm. This suggests et al 1985; Hull et al 1991), sorghum (y-kafirin) that posttranscriptional regulatory processes are (DeRose et al 1989; de Freitas et al 1994), and Coix involved in the expression of this gene in the hetlacryma-jobi (coixin) (Ottoboni et al 1993) are also erologous host (Manteuffel and Panitz 1993; characterized by the lack of introns but they con- Schubert et al 1994). tain the -300 bp element. Both y-kafirin and coixin As can be discerned from this account, there is a show varying degrees of homology to different considerable industry for the isolation of various subunits of zein (de Barros et al 1991; Leite et al endosperm storage protein genes and their intro1992; Ottoboni et al 1993). duction into heterologous hosts. This work is Glutelin genes of rice contain functional pro- fueled by the potential practical benefits that will (iv) Rice Glutelin and Other Genes
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accrue from the ability of the genes to code for useful proteins in seeds of plants of otherwise limited use. As of now, the main stumbling block is a level of expression of these proteins in the heterologous hosts that is too low to be economically useful. 7. GENERAL COMMENTS
As a product of double fertilization, the endosperm has attracted the attention of plant embryologists for the variety of ways this tissue has evolved from the primary endosperm nucleus. The first few rounds of division of the endosperm nucleus follow divergent patterns in different species, but eventually the mature tissue ends up as a formless mass of cells surrounding the embryo. The conspicuous events in the formation of the endosperm are marked by stages in which cell divisions predominate first and then cell enlargement takes over. In the endosperms of many species, the cells compensate for the lack of divisions in the later part of their ontogeny by abnormal nuclear phenomena such as polyploidy and endoreduplication. In the early period of investigations of the endosperm, the importance attached to the ontogeny and cytology of the tissue tended to be overshadowed by views about the tissue's possible functions. The endosperm is considered a potent source of chemical substances that promote the growth of the early stage embryos. Although direct evidence for
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the absorption of endosperm nutrients by the developing embryo is lacking, in attempts to relate embryo growth to endosperm nutrients, the endosperm has been analyzed morphologically, biochemically, and genetically. Persuasive though the results of these analyses are, they fall short of evidence with definitive functional implications. Throughout the history of the study of endosperms, most investigators have focused on the cereal grains. From the physiological point of view, the endosperm of cereal grains supplies the nurture and nutrients for the embryo during its transformation into a seedling plant. Although the topic is outside the purview of this chapter, the biochemical ramifications of the endosperm in germinating seeds and grains have provided fundamental information about the metabolic and catabolic pathways of the major seed storage products, namely, starch, proteins, and lipids. The modern era of molecular biology and genetic engineering is beginning to bridge the gap in our knowledge of the gene expression program leading to the accumulation of storage products during the development of the endosperm. Information currently available about the structure and expression of the major storage protein genes of the endosperms of some of the economically important cereal crops will continue to have a profound influence on our future attempts to improve the quality and quantity of the food crops and to genetically engineer the endosperm storage proteins of other plants.
Chapter 13
Embryogenesis and Physiology of Growth of Embryos The Zygote (i) Volume Changes and Cell Wall Synthesis (ii) Infrastructure of the Zygote The Multicellular Embryo (i) Classification of Embryo Types (ii) Comparative Embryogenesis in Monocots and Dicots (iii) Reduced Embryo Types (iv) Cell Lineage and Cell Fate during Embryogenesis Cellular Aspects of Embryogenesis (i) Ultrastructural and Histochemical Characterization (ii) Cell Division and Cell Elongation (iii) DNA Synthesis Tissue and Meristem Differentiation during Embryogenesis (i) Shoot and Root Apical Meristems (ii) Origin of the Quiescent Center The Suspensor (i) Ultrastructure of the Suspensor Cells (ii) Nuclear Cytology of the Suspensor Cells (iii) Physiology and Biochemistry of the Suspensor Nutrition of the Embryo (i) Nutrient Supply to the Zygote (ii) Embryo-Endosperm Relations (iii) Nutrient Supply from the.Mother Plant Embryo Culture (i) Seed Embryo Culture (ii) Culture of Immature Embryos and Precocious Germination (iii) Culture of Proembryos 8. General Comments
After fertilization, the egg is transformed into the zygote, which embarks upon one of the most critical periods in its development as it is partitioned
into cells that ultimately make up the body of the embryo. What makes the zygote so unique a cell is that it is the product of fusion of two gametes, the sperm contributing the paternal genome and the egg providing the maternal counterpart. The point in time when the egg and sperm fuse together to be woven into a new sporophyte marks the beginning of the ontogeny of the species. The phase of ontogeny concerned with progressive division of the zygote to form the embryo is known as embryogenesis, or as zygotic embryogenesis (to avoid semantic confusion with somatic embryogenesis and pollen embryogenesis). Classical histological analyses of embryo development in a large number of plants have generated an invaluable guide to the division patterns of the zygote and its immediate derivatives, and contemporary studies are beginning to provide some much needed insight into the associated cellular and molecular changes. The history of the study of the development and physiology of embryos can be considered to have evolved in three distinct phases. Much of the early work was focused on the first few rounds of division of the zygote and on the subsequent morphogenesis of cells to give rise to the embryo. This early work, published in scores of scientific papers over the years, gave rise to the thesis that each division plane of the fertilized egg and its descendants is accomplished according to a genetic blueprint characteristic of the species. In later years, a large number of studies were undertaken on the structural characteristics of embryos and their relationship to morphological development and function. One aspect of these studies was a concern with the ultrastructural changes that accompany the transformation of the zygote into a full-term embryo. A third focus has been on the much debated but little 357
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understood area of nutrition of the embryo. also play a role in the evolution of the zygote. For Experimental approaches to gaining insight into example, in Gossypium hirsutum (Jensen 1968), the nutritional interactions during embryogenesis Hibiscus costatus, H. furcellatus (Ashley 1972), have relied on perturbations of the growth of Lagerstroemia speciosa (Lythraceae) (Raghavan and embryos that involve their isolation and culture at Philip 1982), and Brassica campestris (Sumner 1992), there is a dramatic decrease in the size of the fertilvarious stages of development. The account in this chapter will begin with ized egg to about half the size it was before fertiloverviews of the zygote and various aspects of its ization; the zygote of Nicotiana tabacum also division that generates the embryo and the subse- experiences a shrinkage, but it is less striking quent tissue and meristem differentiation patterns in (Mogensen and Suthar 1979). In some plants the the embryo. Part of the account will also be con- egg actually enlarges after fertilization as it is cerned with the cell biology and cytology of transformed into the zygote. This is the case in embryos as they complete their genetically permissi- Datura stramonium (Satina and Rietsema 1959), ble development. Later sections will deal with the Cypripedium insigne (Poddubnaya-Arnoldi 1967), physiology of embryogenesis, emphasizing the Jasione montana (Erdelska and Klasova 1978), Oryza nutrition of embryos of different ages. Three reviews sativa (Jones and Rost 1989b), Hordeum vulgare (Raghavan 1991, 1993; Raghavan and Sharma 1995) (Engell 1989), Monotropa uniflora (Olson 1991), and have discussed the topics covered here from slightly Arabidopsis (Mansfield and Briarty 1991). In different perspectives and the reader is referred to Arabidopsis, a change in the distribution of the microtubules, from a random orientation to a prethem for additional information. dominantly transverse orientation in a subapical band, coincides with the growth in length of the zygote (Webb and Gunning 1991). 1. THE ZYGOTE Within the zygote lies the ability to form an entire plant, a feat that the zygote accomplishes by extensive changes in form in defined and dramatic ways and by a progressive change from the undifferentiated to the differentiated state. Thus, the zygote can truly be considered as the unicellular stage of a multicellular organism, possessing all essential properties of interaction of the adult. Common to the changes that occur during the progressive transformation of the zygote into an embryo are such processes as cell division, cell expansion, cell maturation, cell differentiation, and formation of meristems. In a very young embryo, all cells are involved in producing a new generation of daughter cells. As development continues, cell divisions, which lead to the formation of specialized tissues and organs, are segregated to certain parts of the embryo predictable by their position in the cell lineage. Very little is known about the mechanisms that bring about the restriction of functional activities in the developing embryo. (i) Volume Changes and Cell Wall Synthesis
Once fertilization has been accomplished, many properties of the egg change during the first few hours of its life as the zygote. Even though the postfertilization changes occur in the cytoplasm, physical changes occurring outside the cytoplasm
Special attention has been focused on the chalazal part of the egg, which in many species, as seen in Chapter 6, lacks a cell wall in the unfertilized state. The laying down and the consolidation of a new wall at the chalazal end are important milestones in the evolution of the zygote in G. hirsutum (Jensen 1968), Capsella bursa-pastoris (Schulz and Jensen, 1968c), Quercus gambelii (Singh and Mogensen 1975), N. tabacum (Mogensen and Suthar 1979), Glycine max (Folsom and Cass 1992), Helianthus annuus (Yan, Yang and Jensen 1991), and B. campestris (Sumner 1992); increased activity of dictyosome vesicles and cortical microtubules coincident with cell wall regeneration seems to validate these observations (Jensen 1968; Schulz and Jensen 1968c; Yan, Yang and Jensen 1991b; Sumner 1992). In several species of Rhododendron and in Ledum groenlandicum (Ericaceae), a callose wall is laid down around the zygote during the first two days after fertilization (Williams et al 1984). The fact that the wall essentially insulates the newly formed zygote from the influence of cells of a different genotype in the immediate neighborhood probably has some significance in the subsequent induction of the sporophytic divisions. (ii) Ultrastructure of the Zygote
No generalizations can be made about the ultrastructural upheavals that occur in the egg after fertilization except that the simplicity of the zygote as
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revealed in the electron microscope almost betrays size and number of vacuoles increase markedly its potential to form a whole new plant. In Gossypium toward the micropylar end of the zygote (Figure hirsutum, polarity is already established in the egg 13.1). With minor variations, these changes are conby the presence of a large micropylar vacuole. The sistently found to occur in the fertilized egg cells of position of the vacuole remains unchanged as the Capsella bursa-pastoris (Schulz and Jensen 1968c), egg is transformed into the zygote. However, cyto- Linum usitatissimum (D'Alascio-Deschamps 1972), plasmic organelles like plastids, ER, mitochondria, and jasione montana (Erdelska and Klasova 1978). In and ribosomes begin to float in the egg within hours Papaver nudicaule and Zea mays, whose unfertilized after fertilization and take up new positions at the egg cells have a micropylar nucleus and a large chachalazal end, where they form an investment lazal vacuole, fertilization is accompanied by a around the nucleus. This shift in organelle positions reversal of cytoplasmic polarity as the zygote accentuates the polarity already present in the unfer- acquires a chalazal nucleus and a micropylar vactilized egg. Observations of the egg at different times uole (Olson and Cass 1981; van Lammeren 1986). after fertilization leave no doubt that the increase in Other ultrastructural observations, such as the total amount of ER as well as in the tube-con- increase in the number and complexity of the taining ER, the increase in size and number of starch organelles, indicate that the zygote is a metaboligrains in the plastids, the elaboration of additional cally active cell. The standard changes suggestive of cristae in the mitochondria, the generation of new altered cell functions observed include alterations ribosomes, and the formation of polysomes from the in the form of the ER, a marked increase in the numexisting ribosomes, are all part of the basic response ber of starch grains in the plastids, the elaboration of the egg to fertilization (Jensen 1968). In of additional cristae in the mitochondria, the Arabidopsis, the large micropylar vacuole of the egg appearance of a new generation of ribosomes and undergoes a transient reorganization upon fertiliza- their transformation into polysomes, and increased tion as it is replaced by numerous, small, vesiclelike dictyosome activity; these changes are described in vacuoles. This is followed by the elongation of the great detail in the zygotes of Z. mays (Diboll 1968; zygote, a decrease in the number of vacuoles, and van Lammeren 1986), C. bursa-pastoris (Schulz and the eventual formation of an extremely large vac- Jensen 1968c), Epidendrum scutella (Cocucci and uole occupying approximately two-thirds of the Jensen 1969c), Linum usitatissimum (Deschamps total length of the cell at the micropylar end. Nuclear 1969), L. catharticum (D'Alascio-Deschamps 1981), migration from the basal part of the zygote to the Q. gambelii (Singh and Mogensen 1975), H. annuus chalazal end completes the program of organelle (Yan, Yang and Jensen 1991b), Arabidopsis redistribution that results in a distinct apico-basal (Mansfield and Briarty 1991), and Brassica campestris polarity (Mansfield and Briarty 1991). The target of (Sumner 1992). In the eggs of Petunia hybrida fertilization in Glycine max also appears to be the (van Went 1970c), L. usitatissimum (D'Alasciolarge micropylar vacuole of the egg, which is frag- Deschamps 1972), and Q. gambelii (Mogensen 1972), mented into a bunch of small vacuoles distributed an increase in the number of lipid bodies after ferthroughout the zygote; additionally, there is an tilization is a feature not observed in other plants. increase in the number and complexity of the amy- The presence of starch-containing plastids and lipid loplasts in the zygote (Folsom and Cass 1992). The bodies is considered an adaptive adjustment of the polar distribution of cytoplasmic organelles of zygote to budget energy-related materials needed mature egg cells is also maintained unchanged in during its early division. The abundance of increasthe zygotes of Petunia hybrida (van Went 1970c; ingly complex mitochondria and the large quantity Vallade 1980), Quercus gambelii (Singh and of ribosomes might indicate a high metabolic activMogensen 1975), Monotropa uniflora (Olson 1991), M.ity of the egg upon fertilization. That fertilization hypopitys (Olson 1993), and Helianthus annuus (Yan, signals the production of mRNAs and their engageYang and Jensen 1991b). In plants in which the egg ment by ribosomes, is reflected in the abundance of cell does not possess an incipient polarity, present polysomes in the zygote. Consistent with this obserindications are that the zygote displays a polarized vation, it has been found that in C. bursa-pastoris distribution of organelles, most strikingly of the ER, (Schulz and Jensen 1968c) and Nicotiana rustica ribosomes, plastids, and mitochondria, which clus- (Sehgal and Gifford 1979), fertilization is accompater around the nucleus at the chalazal end. nied by an increase in the stainable RNA content of Accentuating the polarity of the zygote is a gradient the egg; in situ hybridization of ovules of C. bursaof decreasing density of plastids and mitochondria pastoris with 3H-poly(U) has revealed not only an toward the micropylar end. Conversely, both the increase in the mRNA content of the zygote but
Figure 13.1. Ultrastructural profile of zygotes. (a) The zygote (z) of Quercus gambelii. ds, degenerating synergid; f, filiform apparatus; I, lipid bodies; n, nucleus; ps, persistent synergid. Scale bar = 10 urn. (From Mogensen 1972; photograph supplied by Dr. H. L. Mogensen.) (b) The zygote of Zea mays, en, endosperm; m, mitochondria; n, nucleus; s, starch granule. Scale bar = 10 um (From van Lammeren 1986; photograph supplied by Dr. A. A. M. van Lammeren.)
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also a gradient in its accumulation, with a high concentration in the chalazal part (Raghavan 1990b). Eggs of other plants, such as N. tabacum (Mogensen and Suthar 1979) and Triticum aestivum (You and Jensen 1985), show variable ultrastructural details but display little change after fertilization. In Z. mays, postfertilization increases in the activity of oxidative enzymes and the accumulation of reserve substances bypass the zygote and are generally confined to the nucellus and integuments (Singh and Malik 1976). It is clear that several concomitant changes coordinated by many assembly lines must be taking place during the formation of the zygote, but they differ among different species. One consequence of these changes is that the zygote essentially becomes a repository of developmental information that can play a role in morphogenesis. To a large extent, this is reflected in the subsequent division of the zygote to form the embryo. 2. THE MULTICELLULAR EMBRYO
Changes in the ultrastrucrure of the zygote are preparatory to its first division. However, this division might not occur immediately after fertilization or immediately after the reorganization of the cytoplasm, since the zygote remains quiescent for a period of time, ranging from a few hours to several months, before it divides. At the time of its division, the zygote appears as a highly polarized structure. Polarity is attributed to two factors. First, the orientation of the zygote is polar in the sense that its one end is cemented to the embryo sac wall at the micropylar end, and the other end projects into the central cell facing the chalazal end. Second, polarity is also bestowed on the zygote by its ultrastructural profile previously described. The division of the zygote is the next milestone in the establishment of polarity. This division is almost invariably asymmetric and transverse, cutting off a large, vacuolate, basal cell toward the micropylar end and a small, densely cytoplasmic, organellerich terminal cell. In Arabidopsis, the occurrence of zygote division is presaged by the presence of a preprophase band of microtubules corresponding to the position of the future cell plate (Webb and Gunning 1991). In mutants that are unable to form cortical microtubules, cell divisions are random due to irregular cell expansion and the failure of division plane alignment. A line of reasoning based on the observation that mutant embryos differentiate normally has led to the suggestion that the basic differentiation pattern in the developing embryo
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does not depend upon the ordered arrangement of the cytoskeletal elements (Traas et al 1995). A mathematical analysis of the relative areas of the apical and basal cells seems to contradict the conventional view of the universality of the size relationships of these cells, since in a significant number of cases, the apical cell may be large and the basal cell small, or both cells may be of the same size (Sivaramakrishna 1978). A consistent, unequal division is morphogenetically significant, because it points to some underlying cytoplasmic program that determines the position of the mitotic spindle and the orientation of the cell wall. Ultrastructural and histochemical studies have shown that the marked degree of asymmetry displayed by the two-celled embryo is complemented by subtle variations in the distribution of organelles and in the concentration of macromolecules. For instance, in Gossypium hirsutum (Jensen 1963), Capsella bursa-pastoris (Schulz and Jensen 1968c), and Quercus gambelii (Singh and Mogensen 1975), the terminal cell of the two-celled embryo is similar to the chalazal part of the zygote in that it has a dense cytoplasm enriched with organelles, whereas the basal cell is relatively impoverished of organelles. In the two-celled embryo of G. hirsutum, the terminal cell is also characterized by the presence of a large number of plastids and large and variably shaped mitochondria, whereas the basal cell has a preponderance of vacuoles of various sizes and shapes (Jensen 1963). In G. hirsutum (Jensen 1963) and Vanda sp. (Alvarez and Sagawa 1965b), a functional differentiation between the terminal and basal cells is signaled by a higher concentration of RNA in the former. By contrast, in Stellaria media (Pritchard 1964b), C. bursa-pastoris
(Schulz and Jensen 1968c), and Limnophyton obtusifolium (Alismaceae) (Shah and Pandey 1978), RNA concentration is higher in the basal cell than in the terminal cell. Apparently an increased concentration of macromolecules does not infallibly predict in all cases the fate of the cell in a particular morphogenetic pathway. (i) Classification of Embryo Types
Partitioning of the terminal and basal cells leads to the formation of the mature embryo and, for this reason, the plane of division of one or both these cells and the fate of the daughter cells formed have an important bearing on the classification of embryo types in angiosperms. Generally, the organogenetic part of the embryo is derived from the terminal cell with little or no contribution from
362 the basal cell; during embryo formation, the first division of the terminal cell may be transverse, longitudinal, or, rarely, oblique. These facts have served as a basis for classifying different types of embryos in angiosperms. A widely used system of classification has identified two major groups, one in which the division of the terminal cell of the twocelled embryo is longitudinal, and the other in which the division wall is oriented in a transverse plane (Johansen 1950). Within these major groups, different embryo segmentation types are recognized and identified by the name of the family in which many examples of the type are found. However, it should be noted that in some families more than one type of embryo development, represented by one or two genera in each case, has also been observed. Accounts of the ontogeny of representatives of each segmentation type are given by Johansen (1950) and Maheshwari (1950), and so in this account ontogenetic information is passed over. In describing embryogenesis, the term "proembryo" is used to designate the early stages of development of the embryo; the initiation of cotyledons is considered as a good cut-off point for the end of the proembryo stage.
Zygotic Embryogenesis cotyledons and plumule, and the latter to the root meristem, root cap, and hypocotyl; a small suspensor is also derived from the basal cell (Jones 1927). The Asterad type of embryo development or its variations have been described in members of Asteraceae, Geraniaceae, Lamiaceae, Oxalidaceae, Polygonaceae, Rosaceae, and Urticaceae (dicots), and Liliaceae and Poaceae (monocots) (Natesh and Rau 1984).
Transverse Division of the Terminal Cell. Three embryo developmental patterns (Solanad, Caryophyllad, and Chenopodiad) are recognized on the basis of the transverse division of the terminal cell and the participation of the division products of the terminal and basal cells of the two-celled proembryo in the formation of the mature embryo. The Solanad type, found in several species of Nicotiana, is characterized by the formation of the embryo from the terminal cell and by division of the basal cell to form a suspensor. Besides members of Solanaceae, members of Hydnoraceae, Linaceae, Papaveraceae, and Rubiaceae display the Solanad type of embryo formation. In the Caryophyllad type, the basal cell does not divide after it is cut off, and the organogenetic part of the embryo is derived exclusively from the division of the terminal cell. These characLongitudinal Division of the Terminal Cell. teristics are clearly evident during the embryogeneDepending upon the extent of the contribution of sis of Sagina procumbens (Caryophyllaceae) (Soueges the basal cell of the two-celled proembryo to the 1924). A suspensor is not a regular feature of the formation of the organogenetic part of the mature Caryophyllad type of embryos; if one is present, it is embryo, Crucifer (or Onagrad) and Asterad types also derived from the terminal cell with contribuof embryo development have been recognized in tions from the basal cell, as seen in Saxifraga granulata the group exhibiting longitudinal division of the (Saxifragaceae) (Soueges 1936). The Caryophyllad terminal cell. In the Crucifer type, exemplified by type of embryo development has been described in Capsella bursa-pastoris and Arabidopsis, the basal cell plants included in Crassulaceae, Droseraceae, contributes very few derivatives to the formation Fumariaceae, Holoragaceae, Portulacaceae, and of the organogenetic part of the embryo, which is Pyrolaceae (dicots), and Alismataceae, Araceae, thus generated almost entirely by the terminal cell. Ruppiaceae, and Zannuchelliaceae (monocots). The cells derived from the basal cell form a shortAnother addition to the list is Monotropa uniflora, in lived, filamentous structure known as the suspenwhich the basal cell collapses after it is cut off (Olson sor; more will be said about the suspensor in a later 1991). The Chenopodiad type has the distinction of section of this chapter. This type of embryogeny having the division products of both the terminal has also been described in plants belonging to and basal cells integrated into the organogenetic part Onagraceae, Bignoniaceae, Lamiaceae, Lythraceae, of the embryo. Generally, the hypocotyl of the Papilionaceae, Ranunculaceae, Rutaceae, and mature embryo has its origin in the basal cell, which Scrophulariaceae among the dicotyledons, and in also forms a short suspensor. In addition to being Juncaceae and Liliaceae among the monocotylefound in members of Chenopodiaceae, this type of dons. In contrast, the embryo in the Asterad type is embryo development is typical of plants included in generated by the division of the terminal and basal Amaranthaceae and Polymoniaceae (Natesh and cells of the two-celled proembryo. The description Rau 1984). of embryogenesis in Lactuca sativa reveals a nice division of labor between the derivatives of the terA type of embryo development in which the minal cell and those of the basal cell in the forma- first division of the zygote itself is oblique or lontion of the embryo, the former giving rise to the gitudinal (Piperad type) was slow to be recog-
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nized and was thought to be found only in a few generally transcend the boundaries of dicots and families of dicotyledons, such as Piperaceae, monocots. Balanophoraceae, Dipsacaceae, and Loranthaceae. To understand how the zygote progresses from a The genera and species assigned to this type are not single cell to a complex multicellular embryo in many, and only occasionally has it been identified which fundamental tissues and organs are demarin monocots (Tohda 1974; Natesh and Rau 1984). cated, embryogenesis of representatives of dicots We see that descendants of the basal cell of the and monocots will be considered. Any one of the two-celled proembryo defy a neat packaging, many dicot and monocot embryos that have been because in some cases, part of this cell population studied over the years will illustrate the general becomes enmeshed in the organogenetic part of the principles of embryogenesis easily, but some plants embryo, the rest forming a suspensor, whereas in have been favorites for this purpose. Embryogenesis other cases, the basal cell population wholly forms in Capsella bursa-pastoris is almost a textbook examthe suspensor or simply wilts away. The general ple of the Crucifer type of embryogenesis in which conclusion from these observations is that although the first division of the terminal cell of the two-celled the terminal cell is already determined as the prog- proembryo is longitudinal (Figure 13.2). Each of the enitor of the embryo, the determinative state of the two cells thus formed again divides by longitudinal walls to yield a quadrant, followed by an octant basal cell is at best described as being labile. Embryo development in Paeonia anomala, P. stage by division of the four cells by transverse moutan, and P. wittmanniana, first described by walls. Although the embryo is populated at this Yakovlev and Yoffe (1957), does not fit into any of stage by a relatively small number of homogeneous the types just described, because the failure of wall cells, there are functional differences between the formation during the first few rounds of division of upper and lower tiers of cells. For example, the stem the zygote results in embryogenesis that starts with tip and cotyledons in the adult embryo are believed numerous free nuclei suspended in a cellular bag. to be formed from the upper tier of cells, and the Later, the nuclei migrate toward the periphery of hypocotyl generated from the lower tier. In situ the cell, where wall formation is initiated to form a hybridization of embryos of Brassica napus with multicellular proembryo. A further remarkable cloned genes for the storage proteins, cruciferin and change occurs as the marginal cells of the proem- napin, has shown that the determinative event bryo differentiate as embryo primordia. Although established by the transverse division of the quadseveral primordia are born out of a proembryonal rant is reflected in the gene expression patterns in cluster, only one outlives the others and differenti- later stage embryos. In these embryos, cells derived ates into a viable embryo. Independent investiga- from the lower tier accumulate high levels of trantions by other workers (Cave, Arnott and Cook scripts whereas, except in the cotyledons, cells 1961; Carniel 1967a) on additional species have derived from the upper tier are devoid of transcripts essentially confirmed the unusual features of (Fernandez, Turner and Crouch 1991). The point is embryogenesis in this genus. A free nuclear phase important because the pattern of transcript accumuis typical of embryogenesis in gymnosperms, and lation does not appear to reflect tissue or cell types its occurrence in Paeonia appears to be the but follows the boundary set up by an earlier division. However, the developmental fate of a group of angiosperm equivalent of a gymnosperm feature. three or four layers of cells derived from the upper tier of the quadrant (designated as the epiphysis) and of the root apical meristem derived from the ter(ii) Comparative Embryogenesis in minal cell of the suspensor (designated as the Monocots and Dicots hypophysis) is expressed by their failure to accumuThe essence of embryogenesis is the establish- late storage protein transcripts. It appears that gene ment of tissue systems and primordial organs in expression patterns in these groups of cells are set, the embryo. Obviously, a precise sequence of divi- not by the division boundaries, but by their biosions of the zygote and proembryo, as well as a chemical and physiological characteristics. high degree of inductive interactions between the growing parts, bears heavily on the final form The next round of division of cells of the octant attained by the embryo. It is important to note that embryo is periclinal and results in the formation of although mature embryos of dicots and monocots a globular embryo of 16 cells. That some covert appear morphologically different, their early divi- changes may foreshadow the differential potencies sion sequences are basically similar. It is, therefore, of cells of the embryo at this early stage is shown by expected that the modes of embryo development localization of transcripts of a Kunitz trypsin
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f Figure 13.2. Stages in the division of the zygote and early development of the embryo and suspensor in Capsella bursa-pastoris. (a) Zygote. (b) Asymmetric division of the zygote. (c) Transverse division of the basal cell and longitudinal division of the terminal cell, (d) Quadrant stage
embryo, (e) Octant stage embryo, (f) Early globular embryo, (g-i) Three stages of the globular embryo, showing the division of the hypophysis (h). Suspensor is outlined in the square bracket. The micropylar end is toward the bottom of the page. (From Soueges 1919.)
inhibitor gene {KTI-3) in the cells at the micropylar of Triticum aestivum in the presence of auxin and pole of globular embryos of soybean (Perez-Grau inhibitors of auxin transport have supported a role and Goldberg 1989). The expression of the KTI-3 for auxin in the establishment of bilateral symmetry gene is one of the earliest indicators of polarity in in monocot embryos (Fischer and Neuhaus 1996). the globular embryo. The globular embryo is The role of polar auxin transport in the maintephased into the heart-shaped stage when it attains nance of embryonic form was the focus of reports bilateral symmetry due to a lateral expansion of the using the more easily accessible somatic embryos distal pole that gives rise to a pair of cotyledons. of carrot. Not only do polar auxin transport The elements of the future shoot apex, which inhibitors block transitions in embryo development remains essentially in a dormant state throughout beginning as early as the globular stage, they also the life of the embryo, are carved out from a few cause the formation of giant globular and oblong cells lying between the cotyledons. Based on the embryos (Schiavone and Cooke 1987). action of inhibitors of auxin transport on cultured Microsurgical experiments on heart-shaped and embryos of B. juncea resulting in the formation of torpedo-shaped somatic embryos have shed furcotyledons as a fused collarlike structure around ther light on the role of polar auxin supply and disthe apex, rather than as two discrete lateral out- tribution by showing that the basal root half of the growths, a model proposed to account for the embryo grows more rapidly when cultured alone attainment of bilateral symmetry in embryos grow- than when it is grafted back to the apical shoot half ing in vivo has invoked a requirement for polar and cultured. However, when grafted somatic auxin transport (Liu, Xu and Chua 1993b). Indeed, a embryos are treated with a polar auxin transport small percentage of embryos of the pin-formed inhibitor, the basal halves behave as if they have flower mutant (pin-1.1) of Arabidopsis, which has a been cultured separately. Not surprisingly, treatdefect in polar auxin transport, strikingly resemble ment of isolated basal sections with IAA forced treated embryos of B. juncea (Okada et al 1991). The them to grow as if they were part of a graft union patterns of growth displayed by isolated embryos with the apical part (Schiavone 1988). These obser-
Embryogenesis and Physiology of Growth of Embryos
Figure T3.3. Late stages in the development of the embryo and suspensor of Capsella bursa-pastoris. (a) Heart-shaped embryo, (b) Torpedo-shaped embryo, (c) Walkingstick-shaped embryo, (d) Mature embryo, c, cotyledon; ra, root apex; s, shoot apex. Suspensor is outlined in the square bracket. The micropylar end is toward the bottom of the page. (From Schaffner 1906.)
vations also attest to a remarkable dependence of the embryonic root, rather than the shoot, on polar auxin transport. After the cotyledons are initiated, division and differentiation of cells in the basal tier of the embryo give rise to the hypocotyl. There is considerable elongation of the cotyledons and hypocotyl at this stage, which gives rise to the torpedoshaped embryo. A root apex delimited in the torpedo-shaped embryo is the progenitor of the embryonic radicle (Figure 13.3). During progressive embryogenesis, continued growth of the embryonic organs, compounded by spatial restrictions within the ovule, causes the embryo to curve at the tip to assume the walking-stick-shaped stage
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and finally to take on the shape of a horseshoe (Schaffner 1906; Soueges 1919). Although organs characteristic of the adult embryo differentiate as early as the heart-shaped or torpedo-shaped stage, subtle physiological and biochemical changes continue in the cells of the embryo and in the surrounding tissues before the maturation program sets in. It was discussed earlier that a hallmark of the Crucifer type of embryo development is the ambiguity about the role of the descendants of the basal cell of the two-celled proembryo. That some of these cells are ultimately incorporated into the embryo of C. bursa-pastoris has been established by careful cell lineage studies (Schaffner 1906; Soueges 1919). Initially, the basal cell of the two-celled proembryo is partitioned transversely. Further divisions in this newly formed pair of cells are,
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Figure 13.4. Stages in the development of the embryo of Triticum aestivum. (a) Division of the zygote. (b) Division of the terminal and basal cells, (c) Globular embryo (d) Club-shaped embryo, (e) Formation of the scutellum and coleoptile. (f) Diagrammatic section of the mature embryo. The micropylar end is toward the bottom of the page, cl, coleoptile; co, coleorhiza; ep, epiblast; r, root; s, suspensor; sa, shoot apex; sc, scutellum; v, first leaf of the plumule. (From Batygina 1969.)
however, restricted to the terminal cell, which produces, by transverse walls, 8-10 cells held together as a filament constituting the suspensor. The suspensor is terminated at the chalazal end by the organogenetic part of the embryo, and at the micropylar end by the basal cell, which has now become disproportionately large (Figures 13.2, 13.3). All the cells of the suspensor, except one, are eventually crushed and obliterated by the growing embryo; this fortunate cell (hypophysis) is in contact with the embryo and plays an important role in the formation of the root apex. The hypophysis divides transversely to form two cells, and the descendants of these cells contribute to the formation of the root cortex, the root cap, and the root epidermis of the mature embryo. It is as if a single cell from a group of about 10 cells born out of simple mitotic divisions of a mother cell is directed into a specialized channel of continued differentiation, while others perish along the way. Practically nothing is known about the mechanisms that cause changes in the developmental options of the cells of the suspensor. In their morphological appearance, mature
Zygotic Embryogenesis
embryos of monocots are strikingly different from those of dicots because of the presence of only one cotyledon. Studies on the comparative ontogeny of monocot and dicot embryos were bedeviled by controversy for some time, but there is now general agreement that the development of the embryo up to the octant stage is almost identical in dicots and monocots. The main point of contention is with regard to the origin of the shoot apex and cotyledon in the monocots. For a period of time, the dominant view was that in monocots the shoot apex is lateral, and the cotyledon is terminal, in origin. Extensive ontogenetic studies (Swamy and Lakshmanan 1962a, b; Ba et al 1978; Swamy 1979; Guignard 1984), however, seem to support the view that both the shoot apex and the cotyledon share a common origin from the terminal cell of a three-celled proembryo. Accordingly, what is perceived as a lateral shoot apex is one that is displaced from its terminal position by the aggressive growth of the single cotyledon. Sagittaria sagittaefolia (Alismaceae) exhibits a pattern of embryogenesis that is typical of many monocots. Here the differentiative potencies of the cells are established after the first transverse division of the zygote. The terminal cell gives rise to the cotyledon and shoot apex, whereas the hypocotyl-shoot axis and the entire suspensor are derived from the
Embryogenesis and Physiology of Growth of Embryos
basal cell. The cells destined to form the shoot apex are initially sluggish, so that the rapidly growing cotyledon dwarfs the shoot apex. As the hypocotyl is organized in the lower part of the proembryo, there is a concomitant differentiation of the procambium and formation of root initials (Swamy 1980). In their mature structure and ontogeny, embryos of Poaceae do not have much in common with other monocots. The main features of the grass embryo are the development of an absorptive organ known as the scutellum; the presence of caplike structures covering the radicle and plumule, known respectively as the coleorhiza and the coleoptile; and a flaplike outgrowth, called the epiblast, on one side of the coleorhiza. Embryo development in Triticum aestivum will be considered as an example (Figure 13.4). Here, following a transverse-oblique division of the zygote, both cells divide by oblique walls to form a four-celled proembryo. Repeated divisions of these cells yield the mature embryo. Following a globular stage of 16-32 cells, the embryo becomes club shaped. It is at this stage that the scutellum appears as a vague elevation in the apical-lateral region of the embryo. At the same time, the opposite side of the embryo begins to enlarge, marking the differentiation of the shoot apex and leaf primordia. The final major event of embryogenesis is the differentiation of the radicle, which has an endogenous origin in the central zone of the embryo (Batygina 1969). In some other members of Poaceae, such as Zea mays, the pattern of cell divisions in the proembryo becomes unpredictable even as early as the second round (Randolph 1936). The use of the transcription factor-encoding homeobox gene KNOTTED-1 (KN-1) has provided a promising experimental design to show that the first histologically recognizable sign of the shoot meristem in developing Z. mays embryos is foreshadowed by the expression of this marker gene. The scutellum, which is formed almost simultaneously with the shoot apical meristem, does not express the KN-1 gene. Since this gene is known to be active throughout the postembryonic shoot ontogeny of maize, its transcription probably connotes a consistent feature of the shoot meristem from the time it is cut off in the embryo (Smith, Jackson and Hake 1995). A different result is obtained when the expression of a rice homeobox gene is followed in developing rice embryos. The gene is expressed in wild-type globular embryos well before organs are initiated, as well as in mutant embryos that lack detectable embryonic organs. On the face of it, this observation does not
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reveal that homeodomain proteins directly determine shoot development during embryogenesis (Sato et al 1996). (iii) Reduced Embryo Types It is clear that embryogenesis in angiosperms is a closely orchestrated process involving a great many cells whose activities are interlaced. As a result, the mature embryo attains the basic organization consisting of a bipolar axis with one or two cotyledons. The embryo is terminated at each pole by an apical meristem, and the point of attachment of the cotyledons separates the embryonic axis into a hypocoryl-root region and an epicotyl-plumule region. The former region has, at its lower end, the primordial root (radicle), whereas the primordial shoot (plumule) is attached to the stem part called the epicotyl. From the earliest investigations of embryogenesis, it has become apparent that embryo development does not proceed to completion in certain angiosperms. These plants thus shed seeds that harbor underdeveloped or reduced embryos (Figure 13.5). Members of Scrophulariaceae, Balanophoraceae, Ranunculaceae, Orobanchaceae, and Pyrolaceae among dicots, and Orchidaceae, Burmanniaceae, and Pandanaceae among monocots, are noteworthy in this respect, as are additional species belonging to at least 13 more families (Rangaswamy 1967; Natesh and Rau 1984). A paradigm of a seed with a reduced embryo is Monotropa uniflora. Here the embryo is generally embedded in the endosperm and consists of no more than two cells separated by a transverse wall (Olson 1980, 1991). Equally striking for its reduced embryo is the seed of Burmannia pusilla (Burmanniaceae), in which embryogenesis terminates at the quadrant stage of the apical cell (Arekal and Ramaswamy 1973). A higher level of embryo differentiation is found in the seeds of various orchids (Arditti 1979) and of root parasites like Orobanche aegyptiaca and Cistanche tubulosa (both of Orobanchaceae) (Rangaswamy 1967); at the time of shedding, seeds harbor globular embryos that do not show any differentiation into radicle, hypocotyl, cotyledons, epicotyl, and plumule. Mature seeds of AlectA36S ra
vogelii
and
Striga
gesnerioides
(both
of
Scrophulariaceae) contain heart-shaped embryos with rudimentary cotyledons and a radicular pole (Okonkwo and Raghavan 1982). Despite the lack of morphological differentiation, it has become apparent that undifferentiated embryos of certain plants display subtle differences between shoot and root poles. For example, in both O. aegyptiaca and C.
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Zygotic Embryogenesis
sc
Figure 13.5. Seeds with reduced embryos, (a) Longitudinal section of the seed of Monotropa uniflora. E, embryo; En, endosperm; H, haustorium. Scale bar = 10 urn. (From Olson 1980.) (b) Longitudinal section of the seed of Burmannia pusilla. e, embryo; em, endosperm; np, nucellar pad. Scale bar = 10 um. (From Arekal and Ramaswamy
1973.) (c) Longitudinal section of the embryo of Spathoglottis plicata. d, distal end; p, proximal end. Scale bar = 50 |jm. (From Raghavan and Goh 1994.) (d) Longitudinal section of the seed of Striga gesnerioides. em, embryo; en, endosperm; re, root cap; sc, seed coat. Scale bar = 10 um. (From Okonkwo and Raghavan 1982.)
tubulosa, cells at one end of the embryo, identified as tigation; by identifying when and where embryogethe radicular pole, are small, whereas those at the nesis is blocked, one could determine the causal facopposite, plumular pole are large and vacuolate tors involved in the process. (Rangaswamy 1967). Although embryos of Eriocaulon robusto-brownianum and E. xeranthemum
(iv) Cell Lineage and Cell Fate during (Eriocaulaceae) are typically considered as reduced Embryogenesis types, they show some differential cell division activity resulting in a relatively quiescent region Because of the precise regularity of the cleavage and an actively dividing sector. The former region patterns during embryogenesis that lead to the represents the locus of differentiation of the epi- establishment of histologically recognizable tiers of cotyl; the cotyledons arise from the latter region. cells, there are persuasive arguments for the view The embryos, however, do not differentiate a radi- that cell lineages have highly specific developmental cle or a hypocotyl (Ramaswamy, Swamy and fates. This has no doubt formed the basis of the clasGovindappa 1981; Ramaswamy and Arekal 1982). sification of embryo types wherein embryogenesis is In the embryos of the vast majority of orchids, the considered to adhere to certain laws concerned with proximal, chalazal region is occupied by a group of the origin of cells at a given stage, the number of cells slightly smaller in size than those at the distal, cells produced in each generation, the role of cells micropylar region. As shown in Vanda sp. (Alvarez formed, and the determinative fate of the early and Sagawa 1965b) and Spathoglottis plicata (both formed cells (Crete 1963). However, because of the Orchidaceae) (Raghavan and Goh 1994), total RNA difficulty of establishing the precise location of cells and poly(A)-RNA, respectively, are concentrated in descended from the terminal or basal cells of the equal density in all cells of the embryo; however, proembryo beyond a few rounds of divisions, it has the lack of biochemical differentiation between the not been determined whether the early regularity of chalazal and micropylar parts does not preclude the the cell division pattern is continued late in embryocells at the chalazal region from initiating divisions genesis. Consequently, conclusions about cell linto form the shoot apical meristem, whereas the cells eages during embryogenesis based on studies thus of the micropylar part continue to enlarge. These far reviewed reflect more what can be followed than examples serve to illustrate the occurrence of devel- what should be followed. opmental blocks at programmed stages of embryoA way out of this dilemma is to screen chimeric genesis. What causes the arrest of embryo seedlings endowed with genetic markers such as development in these plants requires further invesanthocyanin or chlorophyll deficiency and to follow
Embryogenesis and Physiology of Growth of Embryos
the distribution of groups of marker cells. From an analysis of sectorial chimeras of Gossypium hirsutum, Christianson (1986) concluded that the cotyledons and the first and second seedling leaves, rather than arising from the embryonic apex, are formed from cellular compartments earmarked in the globular stage embryo. Compartments thus identified are two sets of eight cells for the cotyledons, two cells for the first leaf, and a single cell for the second leaf. In another study on the role of cell lineage during embryogenesis, embryos of Zea mays were screened for the production of somatic sectors from genetic mosaics generated by acute X-irradiation of ears heterozygous for cell marker mutations at different times after pollination and fertilization. Several conclusions from this work on the developmental fate of cells of the embryo and on the origin of the embryonic shoot apex deserve mention here. The occurrence of sectors in more than one-half of the scutellum following irradiations carried out after the first longitudinal division of the terminal cell of the two-celled proembryo provided evidence that the products of the first division do not contribute equally to the growth of the embryo. Based on the observation that irradiation after the first division produces mainly whole plant sectors, it may be valid to argue that the shoot meristem is produced from the progeny of only one of the cells of this division. In many mutants, sectors were observed in both the embryo axis and the scutellum, indicating that cell lineages of these two organs do not become separate until some time after the first or second longitudinal division of the two-celled proembryo. The distribution of sectors following germination led to the view that primordial cells of the shoot meristem delineated peripherally during the early stage of embryogenesis are destined to produce the first few nodes of the adult plant, whereas the later nodes are descended from the more internal cells. That the determination of the shoot meristem is a gradual process that proceeds acropetally is evident from the fact that the number of bifurcated shoots (twin shoots) produced by irradiation declines from the multicellular proembryo stage (8-10 days after pollination) and, as later-stage embryos are irradiated, there is an apical progression in the level at which splitting occurs. Later-stage embryos also become progressively insensitive to irradiation, indicating that the shoot system is fully established in the proembryo. The orientation of the sectors from one side of the shoot to the other (saddle sectors) has demonstrated that the shoot meristem arises
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from the region of the embryo containing longitudinally oriented files of cells (Poethig, Coe and Johri 1986). To determine cell lineages giving rise to the root and hypocotyl, Scheres et al (1994) analyzed bluestaining sectors in transgenic plants of Arabidopsis containing the maize transposable element ac interrupting a GUS gene-CaMV 35S promoter construct. The ac element randomly excises from the construct to activate GUS gene expression in the cells in which the event occurs. Assuming that blue sectors in the seedling represent descendants of cells in which transposon excision restores GUS expression, it is possible to relate specific cells in the heart-shaped embryo to these embryonic organs from an analysis of sectors spanning a part or all of the root and hypocotyl regions. Without going into the details of the complex analysis involved in this work, the results have led to the conclusion that distinct cell lineages are not specified for the root and hypocotyl, although the derivatives of the hypophysis are restricted to the columella and the central region of the root. However, it was possible to trace the hypocotyl, an intermediate zone near the upper boundary of the root, the root and root meristem back to compartments in the four tiers of cells derived from the lower tier of cells of the globular embryo (Figure 13.6). Since the sector boundaries are not as sharp as the morphological boundaries, on the whole it appears that cell lineages play a relatively minor role during embryogenesis and that there is no overt, determinate pattern of cell divisions giving rise to the major embryonic organs. Clonal analysis of embryogenesis in additional species using critical markers should make it possible to reevaluate the different types of embryo development based on histological analysis. Whether the cell fate is specified in the young embryo or not, cells of the postglobular embryo initiate a program of continued divisions and morphogenesis to form the apical meristems. Mutations that target specific meristems in the seed embryo are useful for determining the fate of the cells present in the meristem during postembryonic stages of ontogeny. Some of the investigations undertaken with meristem mutants relate to the clonal analysis of the development of the tassel and ear shoot in maize (Johri and Coe 1983), the cell lineage patterns in the shoot meristem of maize and sunflower embryos (McDaniel and Poethig 1988; Jegla and Sussex 1989), the alteration in the shoot apical meristem associated with reproductive transition (Sung et al 1992; Bai and Sung 1995), and the production of abnormal
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Zygotic Embryogenesis
Figure 13.6. Mapping of the end points of colored sectors formed on the seedling of transgenic Arabidopsis. The end points of the large sectors demarcate the earliest divisions, and those of progressively smaller sectors mark later divisions. The sectors are indicated in relation to the seedling diagrammed at left. Black bars indicate the exact location of the sector terminus as determined; bars with white ends indicate estimated sector ends. (A) Sector demarcating the division of the upper and lower tiers of the globular embryo. (B) Root and hypocotyl sectors. (C) Cotyledon shoulder sec-
tors. (D) Hypocotyl sectors. (E) Root sectors with apical ends overlapping basal ends of hypocotyl sectors. (F) Root sectors with apical ends disconnected from basal ends of hypocotyl sectors. (G) Intermediate zone sectors. On the vertical axis, the bars represent cell walls of adjacent epidermal cells. 0, lower wall of the first epidermal cell with root hair; 3, third epidermal cell below the junction of cotyledon vascular strands. (From Scheres et al 1994. Development 120:2475-2487. © Company of Biologists Ltd.)
roots in Arabidopsis (Benfey et al 1993). Findings from these investigations have to some extent altered previously held concepts about the relationship between meristems and contiguous regions; however, since these studies do not materially contribute to our understanding of the developmental fate of early division phase embryos, they will not be further considered. Results from the culture of seed embryo segments have shown that the ability to produce working parts of the missing structures is generally restricted to segments containing the shoot apical meristem. Thus, whereas shoot segments develop into complete seedlings, root and hypocotyl segments retain their typical organization without regenerating shoots (Raghavan 1976b). As shown in an investigation using torpedo-shaped somatic embryos of carrot, region specification seems to operate even in the early stages of embryogenesis in a fashion similar to that seen in seed embryos. It appears that, following extirpation of the lower half of the embryo, virtually any part of the shoot consisting of portions of the hypocotyl or of the hypocotyl and root tissues was able to regenerate a new root, but remnants of the shoot with just the cotyledons and the apical notch did not possess root-forming potential. In contrast, restoration of a normal shoot from a root portion was a rare event and occurred only from root segments consisting of the extreme end of the root pole. During root regeneration from the shoot portion, re-formation of the
missing regions of the embryo begins in the same order and polarity as in an uncut embryo, with the root meristem following the hypocotyl-root tissues (Schiavaone and Racusen 1990, 1991). From this it seems that genetic information required for directing the patterning and timing of events of later stages of embryogenesis is present in the cells of the shoot pole fragment. The extent to which the undifferentiated cells of proembryos and the differentiated regions of slightly more advanced embryos are assigned to a particular fate probably depends upon a succession of events and upon changes in a variety of components. In the proembryo, the events are sustained until the cells are determined as the shoot or root regions. The position developed here with respect to the fate of the territories of early torpedo-shaped embryos is that the events continue until the meristems are fully established. As more cells are produced in the developing embryo, cellular interactions become increasingly complex due to unknown changes in the component processes and their effects on the differentiated cells. For example, based on the capacity of shoot and root segments to form the missing parts, one can conclude that different regulatory states exist in the fragments destined to regenerate. Since global statements such as these do not explain the mechanism of cell lineage and cell fate in developing embryos, one may hope that they will stimulate the formulation of cogent questions and experiments about the systems.
Embryogenesis and Physiology of Growth of Embryos
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developmental fate of the hypophysis is apparently determined after it is cut off, because ribosome density and concentrations of nucleic acids and The foregoing account has been entirely con- proteins in the cells of the root cap and root epidercerned with embryo growth resulting in recogniz- mis cut off by the hypophysis appear intermediate able changes in embryo size and shape. However, between those of the embryo cells and suspensor size and shape changes, as indicators of the growth cells. The earliest stage of embryogenesis in which of a multicellular structure, leave much to be cytological differences between cells are noted in desired, since the tacit assumption remains that the anticipation of tissue and organ differentiation is embryo attains its form by spatial and temporal the heart-shaped embryo. Here the cells of the promodulations in cell division, cell elongation, and cambium and ground meristem appear more cell differentiation. Histological diversity attained highly vacuolate than those of the protoderm in the embryo might be presaged by detectable (Schulz and Jensen 1968a). ultrastructural and cytochemical changes in critical Ultrastructural studies of embryos of other cells and tissues. What patterns might these cellu- angiosperms are less complete than those lar activities follow in a growing embryo? described in C. bursa-pastoris and contribute little to our understanding of cell specialization during embryogenesis. Early embryogenesis in Hordeum (i) Ultrastructural and Histochemical vulgare involves an increase in the population of Characterization ribosomes and mitochondria in all cells. As the It is hard to imagine that, following the distinct embryo completes two or three rounds of divisions, ultrastructural and histochemical differences that there is a progressive decrease in the distribution of characterize the terminal and basal cells of the two- vacuoles in the cells of the suspensor (Norstog celled proembryo, further differentiation of the 1972a). Accumulation of vacuoles and multivesicuproembryo can occur without similar accompany- lar bodies in the cells of the apical part of the ing changes. There is no question that there are proembryo of Secale cereale is considered indicative some subcellular differences in the distribution of of early stages of cell differentiation to form the organelles as the proembryo is repeatedly parti- scutellum (Stefaniak 1984). In Helianthus annuus, tioned, but whether these changes are symptomatic cells of the embryo have more ribosomes than of cell differentiation in the embryo is a debatable those of the suspensor, giving these cells a more issue. In Capsella bursa-pastoris, as the basal cell ofelectron-dense appearance (Newcomb 1973b). the two-celled proembryo divides transversely to According to Singh and Mogensen (1975), cells of form the suspensor, the terminal cell acquires more the embryo and suspensor of Quercus gambelii seem ribosomes and it stains more intensely for proteins to amplify the ultrastructural features of the termiand nucleic acids than do the suspensor cells. On nal and basal cells, respectively, of the two-celled the other hand, ER and dictyosomes are more proembryo. As the embryo is phased out into the abundant in the basal cell than in the terminal cell globular stage, suggestive of increased metabolic (Schulz and Jensen 1968c). As the terminal cell activity, the organelle population undergoes a comforms the globular embryo and the suspensor cell plete reorganization. Increases in plastid number is partitioned transversely into a filament, ribo- and complexity, mitochondrial number, dictyososome density is distinctly greater in the cells of the mal activity, ribosomal aggregation, and the embryo than in those of the suspensor. The pres- amount of ER are a conspicuous part of the ence of starch and lipid bodies in the terminal cell processes that peak in the late globular stage of the three-celled proembryo and their disappear- embryo. By contrast, the number and distribution ance during the division of this cell have suggested of these organelles in the suspensor cells remain that these storage bodies probably function as a unchanged. Besides the conventional subcellular source of energy for mitosis (Schulz and Jensen structure, some unique changes that have been 1968a). When the hypophysis is cut off from the ter- described are the disappearance of starch and lipid minal cell of the suspensor in the globular embryo droplets and an increase in ribosome density and in of C. bursa-pastoris, it acquires a basic subcellular the number of autophagic vacuoles in globular profile different from that of cells of the embryo; embryos of Petunia hybrida (Vallade 1980); the presthis is evident in the highly vacuolate cytoplasm, ence of extensive nuclear evaginations and vesicles the low ribosome density, and the generous in five- to eight-celled embryos of Oryza sativa amounts of ER, starch, and lipids in the cell. The (Jones and Rost 1989b); the appearance of large 3. CELLULAR ASPECTS OF EMBRYOGENESIS
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mitochondria in the cells of the globular embryo of Arabidopsis (Mansfield and Briarty 1991); and the presence of vacuoles with flocculent materials, membrane fragments, and weak electron-scattering bodies in the cells of the globular embryo of Viciafaba (Johansson and Walles 1993b). Cells of the heart-shaped and older embryos have the normal complement of organelles typical of meristematic cells, and no generally characteristic ultrastructural changes augur the onset of tissue and organ differentiation in them. Clearly, in a situation in which newly formed cells are constituted into tissues in a developing organ, it is unrealistic to expect major upheavals in cell structure. Some exceptions are noteworthy, such as a 4- to 16-fold proliferation of the rough ER found in the cotyledons of Phaseolus vulgaris during the cell elongation phase, coincident with storage protein accumulation (Briarty 1980). At the histochemical level, tests on embryos of various plants have encountered higher concentrations of RNA in the cells of the presumptive cotyledons than in the rest of the embryo, though it is not surprising that transcriptional activity of the cells increases preparatory to differentiation (Rondet 1962; Pritchard 1964b; Schulz and Jensen 1968c; Vallade 1970; Norreel 1972; Syamasundar and Panchaksharappa 1976; Shah and Pandey 1978). Another measure of activity of developing embryos is the distribution of enzymes of respiratory metabolism. Determinations made in the embryos of Gossypium hirsutum show a striking correlation between tissue and organ differentiation and the activity of succinic dehydrogenase, a key enzyme in aerobic respiration. A high enzyme titer is found in all recognizable organs of the embryo, such as the suspensor, cotyledons, hypocotyl, shoot apex, subapical region, and the elongating radicle, during the period of their active growth (Forman and Jensen 1965). Alcohol dehydrogenase is involved in anaerobic metabolism, and multiple forms of the enzyme are generally present in cells. Differentiation of the axis and cotyledons of embryos of Phaseolus vulgaris is associated with the appearance of a fastmoving isozyme of alcohol dehydrogenase in these organs (Boyle and Yeung 1983). Since the embryo grows in an environment insulated from atmospheric oxygen, perhaps a case can be made for a functional role in normal cellular activities for organ-specific isozymes of the relevant enzymes of anaerobic metabolism. These observations not only give meaning to the process of embryogenesis, but also suggest that a cytochemical examination of embryogenesis with more refined techniques should prove instructive.
Zygotic Embryogenesis
(ii) Cell Division and Cell Elongation A generally accepted notion is that cell divisions predominate during the early period of embryogenesis and that cell enlargement prevails during the later period. A variation of this theme is observed in the embryos of Capsella bursa-pastoris and Gossypium hirsutum, in which the increase in cell number is interrupted by a lag period during cotyledon initiation; this results in a conventional S-shaped curve for cell number from zygote to the globular embryo, and another S-shaped curve from the heart-shaped to the mature embryo (Pollock and Jensen 1964). In the globular embryos of Hordeum vulgare, Oryza saliva, and Triticum aes-
tivum, an exponential increase in cell number is maintained only up to the 20-40-celled stage, after which the rate of increase declines up to the 100-200-celled stage and again up to the 1,000celled stage. This has been attributed to subtle changes in cell metabolism preparatory to changes in form in the embryos (Nagato 1978). Changes in cell size are tightly linked to embryogenic divisions and, notwithstanding the lack of extensive quantitative studies, it is clear that a reduction in cell size accompanies the first several rounds of division of the zygote. In G. hirsutum and C. bursa-pastoris, the zygote remains the largest cell in the life of the embryo, and the average cell size is reduced to about one-fifth that of the zygote even as early as the first division. Cell size drops even further up to the 100-celled embryo stage in G. hirsutum and up to the young globular embryo stage in C. bursapastoris (Pollock and Jensen 1964). A distinctive reduction in cell size up to the midglobular stage of the embryo also occurs in Arabidopsis. The reduction in cell size is accompanied by progressive decreases in cell-to-nuclear volume ranging from 365 in the zygote to about 13 in the midglobular embryo (Mansfield and Briarty 1991). These changes probably help to establish a favorable balance between the nuclear size and cytoplasmic volume for rapid divisions. The distribution of mitotic figures is uniform in very young embryos of G. hirsutum and C. bursapastoris and continually changes to reflect the onset of tissue and organ differentiation, assuming a Ushaped pattern in heart-shaped embryos and ending up in a Y-shaped pattern in torpedo-shaped embryos. Apparently, the onset of specific histogenic patterns during embryo development depends upon the attainment of a critical cell number before the shape change occurs, and is followed by asymmetric distribution of cell divisions. Thus,
Embryogenesis and Physiology of Growth of Embryos
the transition of the globular embryo to the heartshaped form seems to result from an increased mitotic activity in the distal part of the embryo (Ushaped pattern) that gives rise to the cotyledons. A different pattern of distribution of cell divisions (Yshaped pattern), accompanied by cell elongation, might account for the transition of the heartshaped embryo to the torpedo-shaped stage (Pollock and Jensen 1964). The key facts that may provide clues to a more explicit understanding of the factors that control differential cell divisions in a homogeneous tissue are, however, not available. One might suspect that some cytoplasmic gradient is being set up by cell division factors diffusing in the direction of the embryo where potential divisions occur, but as yet we have no evidence for this.
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1967), Vicia faba (Brunori 1967), and Brassica napus (Silcock et al 1990) in the middle or later periods of embryogenesis indicate that the DNA content of the cells does not deviate from the 2C level even as dehydration of embryos preparatory to quiescence of seeds sets in. It is possible that with progressive desiccation of the embryo, other factors, such as available water supply, might limit DNA synthesis. A great deal of our present knowledge of the dynamics of DNA synthesis and accumulation in developing embryos has been acquired from analyses of cotyledons of legumes and other plants that accumulate storage proteins (Figure 13.7). After an initial period of growth by cell divisions, the subsequent growth of cotyledons is marked by cell expansion accompanied by the accumulation of enormous quantities of storage products, mostly proteins. Scharpe and van Parijs (1973) showed that (iii) DNA Synthesis cells of developing cotyledons of Pisum sativum The concept of constancy of DNA in a cell pre- continue to synthesize DNA after cell divisions dicts that DNA synthesis must keep pace with the have ceased; this leads to a progressive increase in production of cells so that each new cell will have a the DNA amount in the cells of full-grown cotylefixed amount of DNA. One might hope that the dons to a level as high as 64C. The general pattern precise division of the zygote to produce a multi- of changes in DNA levels of cotyledonary cells of P. cellular embryo will depend upon an efficient arvense closely resembles that described in P. DNA duplication mechanism, followed by cytoki- sativum (D. L. Smith 1973). These two studies have nesis. This picture is obscured in some studies been augmented by reports of a continued increase because of complications introduced by endoredu- in the nuclear DNA content of the parenchymatous plication. The DNA content of the zygote has been cells of cotyledons of Vicia faba (Millerd and shown to range from the expected 2C-4C amounts Whitfeld 1973), Gossypium hirsutum (Walbot and in Tradescantia paludosa (Woodard 1956), Hordeum Dure 1976), Glycine max (Dhillon and Miksche vulgare, and H. vulgare X H. bulbosum (Bennett and 1983), Phaseolus vulgaris (Briarty 1980; Johnson and Smith 1976; Mogensen and Holm 1995), to attain a Sussex 1990), and Sinapis alba (Fischer et al 1988). In 3C-6C level in Petunia hybrida (Vallade et al 1978), the embryos of V. faba (Millerd and Whitfeld 1973; or to become as high as 16C in H. distichum (Mericle Borisjuk et al 1995) and G. hirsutum (Walbot and and Mericle 1970). DNA amounts in the nuclei of Dure 1976), it has been established that the increase two- to four-celled embryos of these species show a in DNA content of cells following completion of similar range, from a 2C amount in T. paludosa to 8C division is by endoreduplication; that endoredupliin H. distichum. In the globular stage embryo of cation occurs at the same time that cell elongation Vanda sanderiana, there is a gradient of increasing and storage protein accumulation proceed adds DNA values from 2C to above 8C in cells from the interest to the possible relationship among the distal meristematic tip to cells in the proximal three events. parenchymatous region (Alvarez 1968). With furThe question as to whether the increase in DNA ther development, as in heart-shaped and torpedo- content of cotyledonary cells represents endoredushaped embryos of Glyine max and Vitis vinifera, plication of the entire genome or selective amplifithere is a stabilization of DNA content in the cells at cation of certain sequences has engendered some the 2C-4C levels (Chamberlin, Homer and Palmer discussion. According to Walbot and Dure (1976), 1993b; Faure and Nougarede 1993). In Gossypium the presence of identical reassociation kinetics of hirsutum, the amount of DNA per cell remains nonrepetitive DNA from cotton cotyledons before fairly constant from the early heart-shaped to the and after endoreduplication, the absence of satelcotyledonary stage embryos, although there is lites after analytical ultracentrifugation of the two nearly a 30-fold increase in cell number during this DNA samples, and the constancy in the number of transition (Fisher and Jensen 1972). Analyses of rRNA cistrons per unit of the two DNA samples embryos of Lactuca sativa (Brunori and D'Amato argue against selective amplification. Results in
374
Zygotic Embryogenesis
Cell Num ber
Root Tip
•1
2C
4C
8C
16C
32C
64C
128C 256C
1 2C
f 4C
1 8C
1 16C
32C
64C
128C
256C
32C
64C
128C
2S6C
Stage V
,J 2C
•
| ^ _ 4C
8C
tec
Stage VI
4-
r- • - 1 4C 2C 1
•
- • 1 • i • F • 32C 8C
16C
i_ ^—
- ^ 128C
—I 2S6C
64C
Stage VII -
2C
sc
4C
tec
MI\i u I
•_
I 64C
1 128C
1 256C
1 32C
1 64C
1 128C
1 256C
32C
64C
128C 2S6C
32C
12"
••
8'
Stage VIII
4"
Ju 2C
6 4 2 "
4C
.---F-J
8C
16C
Stage VIII Axis
2C
4C
• 8C
'6C
Figure 13.7. Distribution of nuclear DNA in the root tip cells, cells of developing cotyledons (stages IV to VIII), and stage VIII embryo axis of Phaseolus vulgaris. Stage IV is the youngest and stage VIM is the oldest embryo analyzed. Nuclear DNA contents are given in C values. (From Johnson and Sussex 1990. Chromosoma 99:223-230. © Springer-Verlag, Berlin.)
accord with this finding have come from a comparison of the hybridization kinetics of cloned soybean embryo storage protein gene with an excess of DNA isolated from embryo and leaf. The presence of identical smooth curves resulting from the annealing of the two DNA samples attests to the absence of selective gene amplification during continued DNA synthesis in the embryo (Goldberg et al 1981a). Additionally, Southern blot hybridization of DNA from the embryos and leaves of soybean with probes for glycinin, the major soybean storage protein, and for a 15-kDa protein showed similar patterns, confirming that selective gene amplification is not a factor to be reckoned with during
endoreduplication (Fischer and Goldberg 1982). Results from the use of a wide selection of molecular probes in hybridization experiments have also discounted gene amplification and random synthesis of DNA as factors in DNA accumulation in the cells of cotyledons of Phaseolus vulgaris Qohnson and Sussex 1990). If storage protein genes or genes for other essential proteins are not selectively amplified in the cotyledons, it is a mystery why the cells contain a lot more genome than all the proteins it can code for. Conceivably, the extra DNA copies may be of restricted but strategic significance as a storage form of nucleotides during germination. 4. TISSUE AND MERISTEM DIFFERENTIATION DURING EMBRYOGENESIS
To give legitimacy to the embryo as the future sporophyte, the three primary meristems - namely, the protoderm, ground meristem, and procambium and the shoot and root apical meristems are blocked out sequentially at different stages of embryogenesis. In the embryo of Capsella bursa-pastoris, the first meristem to differentiate is the protoderm, which is formed by a periclinal division of cells of the octant embryo. Because of the nature of this division, the packet of cells of the globular embryo can be neatly separated into eight external cells of the protoderm, sheltering an internal group of eight cells. After they are cut off, cells of the protoderm continue to divide anticlinally to keep pace with the increasing volume of the embryo and they gradually begin to acquire characters of the epidermis (Schulz and Jensen 1968a). That early during embryogenesis the peripheral layer Of cells becomes superficially similar to epidermal cells is attested by the presence of cutin on globular and heart-shaped embryos (Rodkiewicz, Fyk and Szczuka 1994). The expression of a gene that encodes a candidate protein involved in cutin biosynthesis on the protoderm of early stage carrot embryos is consistent with this observation (Sterk et al 1991). At the heart of the timing of initiation of the protoderm lies the question whether all cells in the embryo are undetermined until the protoderm is carved out or whether genes for protoderm formation are turned on at an earlier stage. In Citrus jambhiri, the commitment to differentiate an epidermis is made by the zygote itself through the development of a cuticle around its outer surface; this trait is faithfully transmitted to the external layer of cells of the embryo. Failure of epidermal re-forma-
375
Embryogenesis and Physiology of Growth of Embryos
tion in embryos whose epidermis had been surgically removed has shown that both epidermal and subepidermal cell layers are already locked into their developmental fate along divergent pathways early during embryogenesis (Bruck and Walker 1985a, b). The ground meristem and the procambium are derived from the inner core of cells of the globular embryo. The cells of the ground meristem may differentiate into a cortex or into both cortex and pith. In either case, the peripheral layer of cells of the central core of the proembryo is the progenitor of the cortex of the mature embryo. During the elongation of the embryo that begins around the torpedo-shaped stage, the cells of the peripheral layer divide periclinally to form three or four concentric layers of cells of the embryonic cortex. Formation of the pith from the ground meristem is foreshadowed by repeated divisions of the cells of the central region of the globular embryo, which continue as the pith of the hypocotyl part of the mature embryo. It was mentioned earlier that the presence of transcripts of the KTI-3 gene is a molecular marker of the micropylar end of the soybean globular embryo. Because the transcripts persist in the cells of the ground meristem of heart-shaped and cotyledon stage embryos, it is conceivable that these cells have their origin in cells of the globular embryo harboring KTI-3 transcripts (Perez-Grau and Goldberg 1989).
the analysis of tissue differentiation in the embryos of several other plants, but limitations of space do not permit us to take up these examples. On the whole, the pattern of tissue formation is a crucial process of embryogenesis and one of enduring histogenetic interest, but one about whose mechanisms we know very little. (i) Shoot and Root Apical Meristems
Differentiation of the shoot and root apical meristems in the embryo makes the preexisting, but cryptic, bipolarity of the embryo more obvious. Investigations on the histogenesis of embryos of Nerium oleander (Mahlberg 1960), Downingia baci-
galupi, and D. pulchella (Campanulaceae) (Kaplan 1969) have shown that a precise pattern of cell divisions and differentiation distinguishes the formation of the shoot and root apical meristems. In N. oleander, the earliest indication of the shoot and root apices is the appearance of pockets of partially differentiated meristematic cells at both poles of the globular embryo. These cells are interpreted as direct descendants of the octant stage embryo rather than as newly generated cells with meristematic potential. In all three species, differentiation of the shoot apex is also augured by a change from the random distribution of cell divisions to their concentration in the apical half and by cytohistological differentiation into a peripheral and a Procambium initiation is a critical feature of his- central zone. It is to be noted that these changes togenesis of the embryo and occurs in the cells of occur before the appearance of the cotyledons, sugthe cortex or pith of globular embryos before the gesting that the cotyledons are the first formative primordia of the cotyledons are formed. As shown organs produced by the shoot apex. The visible difin Nerium oleander (Apocynaceae), elements of the ferentiation of the shoot apex generally coincides procambium are first seen in the innermost part of with the change in the shape of the embryo from the cortex of the early heart-shaped embryo at the globular to heart shaped and is attributed to the level of the presumptive cotyledons. More initials continued meristematic activity of the protoderm are laid down on either side of the original file and and the underlying layer of cells in the apical part this leads to the formation of a complete ring of pro- of the embryo. The formation of the central zone of cambial cells. After the procambium is established the shoot apex is prefigured by the cytological difin the hypocotyl of the developing embryo, it ferentiation of three or four layers of cells below the extends acropetally into the cotyledons and shoot protoderm. Differentiation of the shoot apex is apex, and basipetally into the root apex. Thus, by complete when it becomes dome shaped and the time the embryo becomes torpedo shaped, it has lapses into a state of quiescence or dormancy in the a solid cylinder of procambium extending through mature embryo. its entire length and into the cotyledons (Mahlberg Depending upon the types of embryo develop1960). In Rhizophora mangle (Rhizophoraceae), files ment (described in a previous section), the root of procambium linking the shoot and root apices are meristem and the root cap have their origin in the apparently initiated independently, because they terminal cell or basal cell of the two-celled proemappear discontinuous for a brief period until they bryo. For illustrative purposes, an example of the are connected to the hypocotyl procambium by embryo in which the root apex is derived wholly cotyledonary strands (Juncosa 1982). Besides these from the terminal cell of a two-celled proembryo is selected examples, contributions have been made to that of D. pulchella. Limited transverse and longitu-
Zygotic Embryogenesis
376
dinal divisions of the terminal cell lead to the formation of a large proembryo consisting of four superimposed tiers of one to four cells in each tier. During the organization of the root apex, cells of the most basal tier divide periclinally to give rise to the root cap. Protodermal cells of the next tier, toward the apical part of the proembryo, also divide periclinally to contribute to the root cap. In addition, cells of this second tier generate derivatives that form the root cortex and meristem (Kaplan 1969). In C. bursa-pastoris, in which the embryonic root apex is derived from the hypophysis, the initial division of this cell is in the transverse plane. By further transverse and longitudinal divisions, the daughter cell close to the suspensor gives rise to the root cap and root epidermis, and the one close to the organogenetic part of the embryo generates the rest of the root apex, including the root meristem (Schaffner 1906; Soueges 1919). Whatever factors determine the extent to which the terminal and basal cells of the two-celled proembryo are involved in the formation of the root apex, the embryonic root apex - compared to the shoot apex - displays a remarkable degree of architectural plasticity that allows it to generate closely integrated final products. (ii) Origin of the Quiescent Center A feature of the angiosperm root meristem is the presence of a group of cells that divide rather infrequently or not at all. This packet of cells, known as the quiescent center, is characterized by low DNA, RNA, and protein synthetic activities; the combination of cellular properties has led to the suggestion that the quiescent center originates as a result of the decreasing metabolic activity of certain cells in the embryonic root meristem. Despite the merit of this suggestion, and the use of a variety of methods to monitor the formation of the quiescent center in developing embryos, the evidence in favor of it is at best ambiguous. Relevant data typically invoked to characterize the appearance and disappearance of the quiescent center in developing embryos include the tempo in mitotic activity, staining reactions, and the pattern of 3H-thymidine incorporation (Sterling 1955; Rondet 1962; Vallade 1972; Jones 1977; Clowes 1978a, b). Localization of mRNA in developing embryos of C. bursa-pastoris using 3H-poly(U) as a probe has shown that autoradiographic silver grains caused by annealing of the label are plentiful in the cells of the developing embryo, except in the hypophysis and its derivatives, which constitute the quiescent center in the root apex of the mature
embryo. In seedling roots, the same cells that fail to anneal 3H-poly(U) also fail to incorporate 3H-thymidine, indicating their identity as the quiescent center. These correlations support the conclusion that the quiescent center in the root of C. bursa-pastoris has its origin in the hypophysis of the embryonic suspensor (Raghavan 1990a). 5. THE SUSPENSOR
In the study of angiosperm embryogenesis, interest at the morphological to molecular levels has been bestowed largely on the organogenetic part of the embryo, with little or no attention beyond anatomical descriptions given to the suspensor. This is mainly due to the well-grounded notion that the role of the suspensor is purely of a mechanical nature, namely, to push the growing embryo deep into the chalazal end of the embryo sac, close to the source of nutrients. During the past three decades, a number of ultrastructural and cytological studies, along with a few physiological and biochemical investigations, have led to a revival of interest in the structure and physiology of the suspensor and to a refreshing departure from the classical views about its function. The current view of the structure-function relations of the suspensor is that the suspensor is a dynamic part of the embryo complex, functioning in the absorption and short-distance translocation and exchange of metabolites necessary for the growth of the embryo. Although a filament of 8-10 cells attached to the organogenetic part of the embryo, as seen in Capsella bursa-pastoris or Arabidopsis, is one of the
most familiar images of the suspensor, comparative studies of embryogenesis in angiosperms have shown that suspensors come in various sizes and shapes. A few of these interesting variations in suspensor morphology have been collated, illustrated, and described (Natesh and Rau 1984; Raghavan 1986c). In many plants, the morphological complexity of the suspensor is attributed to the extension growth of its cells, which reach the extraembryonal tissues of the ovule and even the ovary. In embryology literature, these outgrowths are frequently referred to as haustoria, implying that they are specialized structural adaptations for the absorption of nutrient substances. In some families, such as Orchidaceae, Podostemaceae, and Trapaceae, in which complex suspensors with haustoria have been described, the ovules are characterized by the complete absence of endosperm or by the presence of a reduced amount of endosperm. This indicates that nutrients absorbed
Embryogenesis and Physiology of Growth of Embryos
by the suspensor haustoria may provide an important source of energy for the growth of the embryo. However, direct evidence has been difficult to obtain, as most observations are based on the intimate association of the haustoria with the extraembryonal tissues and do not bear directly on the actual transfer of nutrients or on the ultrastructural modifications of the haustoria for nutrient uptake and transfer. As will be described, ultrastructural modifications suggest such a function for the regular, nonhaustorial suspensor cells of several plants.
377
proteins in the heart-shaped embryo. Gradually, the middle lamella of cells begins to loosen, and eventually the suspensor is obliterated by the burgeoning embryo. The basal cell generally remains active for some time after the collapse of the other cells (Schulz and Jensen 1969). The presence of a series of invaginations on the walls of cells of the suspensor of C. bursa-pastoris has been a focal point of ultrastructural study. The invaginations arise from the outer lateral walls of the mid-suspensor cells of the globular embryo and increase in number and complexity at the heartshaped stage. The wall projections are a feature mat (i) Ultrastructure of the Suspensor Cells the cells of the suspensor share with the basal cell. Despite the fact that the suspensor is an The inner wall of the micropylar and lateral parts ephemeral organ that does not become a part of the of the basal cell are also thrown up into an elabomature embryo, structural adaptations for specific rate network of invaginations projecting into the functions find their highest expression in the cells cytoplasm. In fact, the projections first appear in of the suspensor. With regard to the alignment of the wall at the micropylar end of the zygote and the organelles and the density of their distribution, increase in number, peaking in the basal suspensor suspensors of different species thus far investi- cell of the heart-shaped embryo (Schulz and Jensen gated, as well as the different cells of a suspensor, 1968c, 1969). are astonishingly diverse. Other aspects of the susAs will be seen later, Phaseolus is a paradigm for pensor, such as the number of cells present and defining and interpreting the nuclear cytology of their life span, also vary in the different species. suspensor cells in an angiosperm. In P. coccineus, The first detailed account of the ultrastructural the suspensor is polarized from the proembryo cytology of the suspensor was provided in Capsella stage onwards with respect to mitotic activity. bursa-pastoris. For purposes of discussion, three Although divisions mostly cease in the more basal stages have been identified in the life of the suspen- cells of the suspensor by the proembryo stage, they sor: (a) the octant embryo, whose suspensor con- continue in the cells toward the chalazal region. sists of 6 cells; (b) the globular embryo, whose The suspensor attains its maximum number of suspensor attains the maximum cell number and about 200 cells in the heart-shaped embryo; at the appears as a filament of 10 cells interposed between cotyledon stage, it reaches its maximum size. The a large, vacuolate basal cell and the embryo; and (c) presence of a large number of cells in the suspenthe heart-shaped embryo, whose suspensor attains sor, however, does not enhance its longevity the maximum length and lives out its genetically (Yeung and Clutter 1978). permissible life span. Throughout its development, Cell specialization in the suspensor cells of P. the suspensor is functionally connected with both coccineus begins with the appearance of wall infoldthe embryo and the basal cell by numerous ER-con- ings as early as the proembryo stage. Soon after, the taining plasmodesmata. Although cells of the sus- cells become committed to a developmental pathpensor harbor the usual array of organelles, way characterized by a disproportionate increase ribosomes appear to be specially concerned with in the number of ribosomes, plastids, mitochoncellular interactions of this organ. In line with this dria, dictyosomes, and smooth ER, which continview of the seemingly special role of ribosomes, it ues until the cotyledon stage. In considering the has been observed that at the octant stage of the ultrastructure of the wall of developing suspensor embryo, the suspensor cells have a respectable cells, it is significant that an increase in the number number of ribosomes, which show an increasing of invaginations parallels the increase in organelle gradient toward the chalazal end. In the globular density and that a raft of organelles, particularly embryo, ribosomes gradually dissipate in the cyto- ER, dictyosomes, and mitochondria, lie in close plasm; they completely disappear from the suspen- proximity to the wall ingrowths. These observasor cells of the heart-shaped embryo. Symptomatic tions further strengthen a role for the wall projecof the inevitable failure of the cellular machinery of tions in nutrient exchange (Yeung and Clutter the suspensor is the onset of cytoplasmic degener- 1978). ation accompanied by the loss of nucleic acids and Although the suspensor in Stellaria media is
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Figure 13.8. Morphology and infrastructure of the suspensor of Medicago scutellata. (a) A globular embryo showing vacuolation in the chalazal and basal suspensor cells, (b) Electron micrograph of a portion of the basal suspensor cell of (a), showing wall ingrowths, (c) A heart-shaped embryo with suspensor. (d) Electron micrograph of a portion of the basal suspensor cell of (c), showing mitochondria associ-
Zygotic Embryogenesis
ated with wall ingrowths. BS, basal cell; BSC, basal suspensor cell; END, endosperm; M, mitochondria; S, starchcontaining integumentary cells; Wl, wall ingrowth. Scale bar = 10 urn (a), 1um (b, d), 100 um (c). (From Sangduen, Kreitner and Sorensen 1983; photographs supplied by Dr. E. L. Sorensen.)
formed from the terminal cell of the two-celled Kiihner 1976), Medicago sativa, M. scutellata proembryo (Caryophyllad type), the undivided (Sangduen, Kreitner and Sorensen 1983a), Alisma basal cell presents a profile similar to, and no less lanceolatum, and A. plantago-aquatica (Alismaceae) complex than, that of the cells of the suspensor. (Bohdanowicz 1987) give every appearance of havOne manifestation of the complexity portrayed in ing undergone some degree of specialization the basal cell is the appearance of an array of mas- unique to each species. sive wall projections on the inner surface, along Wall labyrinths of the type just described appear with a preponderance of mitochondria near them. to be a widespread phenomenon (Figure 13.8) and Microbodies, plastids, and extensive whorls of have been documented in suspensor cells of P. vultubular ER, which are generally absent from the garis (Schnepf and Nagl 1970), Diplotaxis erucoides cells of the embryo, become a conspicuous part of (Simoncioli 1974), T. majus (Nagl 1976b), Brassica the organelle complement of the basal cell and the napus (Tykarska 1979), Vigna sinensis (Hu, Zhu and cells of the suspensor. The plastids of the suspensor Zee 1983), M. sativa, M. scutellata (Sangduen, cells have an unusual morphology with numerous Kreitner and Sorensen 1983a), Alyssum maritimum tubules and electron-translucent inclusions. There (Brassicaceae) (Prabhakar and Vijayaraghavan is also a decreasing gradation in the complexity of 1983), Alisma lanceolatum, A. plantago-aquatica plastids and in the number of microbodies from the (Bohdanowicz 1987), Glycine max (Dute, Peterson basal cell to the chalazal suspensor cells (Newcomb and Rushing 1989; Chamberlin, Homer and Palmer and Fowke 1974). Plastids found in the suspensor 1993a), and Viciafaba (Johansson and Walles 1993b). cells of Pisum sativum (Marinos 1970b), Phaseolus There is now considerable and varied evidence vulgaris, P. coccineus (Schnepf and Nagl 1970; Yeungbased on the transfer cell morphology of the susand Clutter 1979), Ipomoea purpurea (Ponzi and pensor cells to support the solute absorption theory Pizzolongo 1973), Tropaeolum majus (Nagl and of suspensor function first articulated by Schulz
Embryogenesis and Physiology of Growth of Embryos
and Jensen (1969). According to this view, metabolites from the endosperm or from the surrounding diploid cells of the ovule are delivered to the embryo through the suspensor cells, with the wall imaginations facilitating the transfer by increasing the surface area of absorption. The plasmodesmata perforating the embryo-suspensor boundary may serve to maintain an open channel of communication for the flow of solutes between the suspensor and the embryo. That the path of nutrients to the developing heart-shaped embryos of P. vulgaris and P. coccineus is through the suspensor has become evident from the work of Yeung (1980). The results showed that if 14C-sucrose is administered through pods or to isolated embryos, much of the radioactivity appears in the suspensor and in the suspensor pole of the embryo. Somewhat similar results were obtained in the transport of a labeled polyamine, putrescine, in the ovules of P. coccineus (Nagl 1990). Although the new knowledge of the wall architecture of the cells of the suspensor of angiosperm embryos has pushed the frontiers of our understanding of the function of this organ to a higher level, until more is known about the gradient properties of the suspensor and its surrounding cells, the mechanism of nutrient incorporation into the embryo through the suspensor will remain unclear.
(ii) Nuclear Cytology of the Suspensor Cells
379
gradient of DNA amounts, with low values in the cells close to the embryo, intermediate values in the neck region of the suspensor, and very high values in the cells of the basal region. Endoreduplication in the suspensors of Alisma lanceolatum and A. plantago-aquatica is nonconventional since it affects only the large basal cell. The nucleus of the cell rhythmically bloats in size and, based on the increase in nuclear volume, it has been estimated that in the latter species the nucleus attains a ploidy level as high as l,024N (Bohdanowicz 1973, 1987). Although the chromosomal constitution of endoreduplicated suspensor cells of several species has revealed polyteny, the phenomenon has been most extensively studied in P. coccineus (Nagl 1967) and P. vulgaris (Nagl 1969a, b). Like polytene chromosomes in the salivary glands of dipteran larvae, those of the suspensor cells of the two species of Phaseolus also display puffs, which represent sites of DNA synthesis and transcription. However, some DNA-synthesizing puffs are found on chromosomes only at specific stages of embryogenesis, indicating the possibility of a stage-specific variation in DNA replication in the same polytene chromosome (Tagliasacchi et al 1984). A gene identified in the polytene chromosome of P. coccineus by in situ hybridization using a cloned probe encodes a protein that inhibits fungal endopolygalacturonase (Frediani et al 1993). Since this is a stress-related protein, it is too early to assess the significance of this observation.
As shown by the molecular hybridization of Investigations on the nuclear cytology of suspensor cells of various angiosperms have shown 25S, 18S, and 5S 3H-labeled rRNA fractions to DNA that endoreduplication coupled with the presence of suspensor cells of P. coccineus, genes for rRNA of polytene chromosomes distinguishes their are clustered at the nucleolus-organizing regions of development. A recurrent theme that has since polytene chromosomes (Durante et al 1977). emerged from these studies is that despite its short However, only the ribosomal cistrons embedded in life span, the suspensor is made up of transcrip- random nucleolus-organizing centers are functiontionally active cells that may play a regulatory role ally active and synthesize RNA in a given chromoin the development of the embryo. Research on the some. In general, compared to the heterochromatic cytology of suspensor cells has been reviewed regions of the chromosomes, the bands engaged in before and will not be considered here again RNA puffing are undermethylated; consequently, (Raghavan 1986c). transcriptional activity of polytene chromosomes On the basis of nuclear volume or quantitative may rely on specific molecular components of Feulgen microspectrophotometry, values for the DNA (Frediani et al 1986a, b). When the activities DNA content of suspensor cells so far reported of the same chromosome puffs at different stages of are analyzed by autoradiography range from 16C in Sophora flavescens (Fabaceae) embryogenesis 3 (Nagl 1978) to 8,192C in Phaseolus coccineus (Brady of H-uridine incorporation, they are found to 1973). In P. coccineus (Brady 1973; Nagl 1974) and engage in RNA synthesis with different amplitudes Eruca sativa (Corsi, Renzoni and Viegi 1973), the (Forino et al 1992). Another intriguing observation outcome of endoreduplication in cells in the differ- is that differing labeling patterns appear in the ent regions of the suspensor is somewhat different, polycistronic transcription units for 25S, 18S, and but its consequences in terms of gene regulation are 5S rRNA genes when polytene chromosomes are probably the same. In both these species, there is a annealed with fractions of DNA banded in the ana-
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lytical ultracentrifuge, showing that rRNA cistrons of the same class are interspersed with different DNA sequences (Durante et al 1987). Given the nottoo-low number of the chromosomes in the suspensor cells and given the complex, convoluted nature of the chromosomes, these cytological observations are illuminating and provide a good beginning toward the goal of achieving a complete understanding of the transcriptional apparatus of the polytene chromosomes. The question as to whether polytene chromosomes undergo selective DNA amplification has been subject to intensive study. Autoradiography of 3H-thymidine incorporation into suspensor cells of P. coccineus showed that in most cells the label is found exclusively in the heterochromatic region of the polytene chromosomes although in a small number of cells the label decorates the entire nucleus. These results have been interpreted to indicate that certain genes of the heterochromatic regions undergo DNA amplification, in addition to the scheduled DNA synthesis, due to endoreduplication (Avanzi, Cionini and D'Amato 1970). Consistent with the observation that cells in the different regions of the suspensor display varying degrees of endoreduplication, selective DNA amplification is found to prevail in the puffs of the highly polytene chromosomes of cells in the micropylar region of the suspensor, whereas cells produced during the early stages of suspensor development show uniform labeling over the nuclei due to endoreduplication (Cremonini and Cionini 1977). The occurrence of selective DNA amplification has been confirmed in the analytical ultracentrifuge from the density shifts in DNA isolated from suspensor cells of P. coccineus (Lima-deFaria et al 1975). This work showed the presence in the suspensor cells of a class of DNA with buoyant density that has no parallel with DNA isolated from other parts of the plant. As shown in the same work by a lower saturation value in hybridization experiments with total suspensor DNA and rRNA, certain DNA sequences, particularly the genes for rRNA, are underreplicated in the genome of the suspensor cells. Thus, a seemingly fragile balance between amplification of certain genes and underreplication of other genes governs the nuclear activities of cells of the suspensor during a major part of its life. Although these observations stress a regulatory role for the polytene chromosomes of the suspensor cells, precise determination of the function of their gene products, given the small size and transient nature of the suspensor, requires a protracted study.
Zygotic Embryogenesis (iii) Physiology and Biochemistry of the Suspensor
Based on cytochemical, physiological, and biochemical information, the suspensor cells are considered to be metabolically active throughout their life span; the activities may be geared to the absorption of nutrients or for self-autolysis. In Brassica campestris, several enzymes of oxidative metabolism are found to be maximally active in the suspensor cells between the globular and torpedo-shaped embryos and to diminish thereafter (Malik, Singh and Thapar 1976; Malik, Vermani and Bhatia 1976; Bhalla, Singh and Malik 1980a, 1981b). The preponderance of enzymes in the actively growing suspensors presumably indicates the need for a high metabolism connected with nutrient uptake. However, the cessation of growth of the suspensor promptly initiates hydrolytic enzyme activities beginning in the cells at the micropylar end. The presence of specialized plastids in the suspensor cells of Phaseolus coccineus and P. vulgaris (Nagl 1977;
Gartner and Nagl 1980) has led to the view that the acid phosphatase of these organelles may possibly play a role in the autolysis of the suspensor. In Tropaeolum majus, whose suspensor produces haustoria, self-digestion of the suspensor is initiated by degenerative changes in the mitochondria coupled with the presence of high acid phosphatase activity in the haustorium (Nagl 1976c; Gartner and Nagl 1980). Other enzymes, such as alkaline phosphatase and acetylesterase, are also active in the cells of the suspensor and in the haustorium, becoming maximally effective at the early cotyledon stage of the embryo when suspensor autolysis is well under way (Singh, Bhalla and Malik 1980). Comparative analyses of RNA and protein metabolism of the suspensor and embryo of P. coccineus have provided evidence of higher transcriptional and translational activities in the suspensor cells than in the cells of the embryo. Expressed on a per cell basis, RNA content and the rate of RNA synthesis are low during the early development of the suspensor, increasing to a maximum at the late heart-shaped or early cotyledon embryo stage, and then declining. Protein content and the rate of protein synthesis also register low values in the suspensor cells of early stage embryos but increase substantially in the suspensors of late stage embryos. Characteristically, at most stages analyzed, cells of the suspensor contain more RNA and proteins and synthesize them more efficiently than do the cells of embryos of comparable age (Walbot et al 1972; Sussex et al 1973). Somewhat similar
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One prevalent notion about the role of hormones results have been obtained for the comparative biosynthetic activities of the embryo and suspensor in the suspensor is that in the early stages of develcells of T. majus (Bhalla, Singh and Malik 1981a). opment, the embryo does not bear the burden of Calculations based on the DNA content and rates of synthesizing the hormones necessary for its growth, RNA synthesis in suspensor cells of P. coccineus at but that this function is taken up by the suspensor different stages of embryogeny have shown that the (Cionini et al 1976; Bennici and Cionini 1979; Yeung template activity of these cells in the late heart- and Sussex 1979). A role attributed to gibberellins is shaped or early cotyledon stage embryos is higher that, in the suspensor of P. vulgaris, the hormone than is expected of the diploid gene copies present increases the translational activity and maintains in them (Clutter et al 1974). A clear expression of the the protein level in the embryo. The role of the susprotein synthetic activity of the suspensor geared to pensor in enhancing translational activity was embryo nutrition is seen in the transient accumula- shown by a decrease in protein synthesis in tion of the storage proteins, legumin and vicilin, in embryos detached from the suspensor and cultured the suspensor cells of globular embryos of Viciafaba (Brady and Walthall 1985; Walthall and Brady 1986). even before these proteins begin their massive, A convincing demonstration of the effect of gibberellins in enhancing transcriptional activity of embryo-specific accumulation (Panitz et al 1995). The focus on the synthesis of growth hormones suspensor cells is the hormone-induced formation by the suspensor and their relevance for the func- of RNA puffs and the increased RNA synthesis in tion of this organ represents a major and essentially certain regions of the polytene chromosomes of P. novel addition to our knowledge of its physiology coccineus (Nagl 1974; Forino et al 1992). and biochemistry during the last 20 years. Auxins The significance of the presence of other growth (Alpi, Tognoni and D'Amato 1975; Przybyllok and hormones in the suspensor remains largely Nagl 1977), gibberellins (Alpi, Tognoni and unknown. In Tropaeolum majus, changes in the conD'Amato 1975; Alpi et al 1979; Picciarelli et al 1984; centrations of peroxidase in the suspensor and the Picciarelli and Alpi 1987; Piaggesi et al 1989; embryo are believed to ensure the regulation of Picciarelli, Piaggesi and Alpi 1991), cytokinins endogenous auxin diffusing from the suspensor (Lorenzi et al 1978), and ABA (Perata, Picciarelli into the embryo. This correlation is somewhat typand Alpi 1990) have been identified in the suspen- ical of organs that synthesize or utilize IAA for sors of P. coccineus and other plants. The suspensor growth (Singh, Bhalla and Malik 1979). of P. coccineus has 10 times more gibberellin activity In conclusion, it is clear that the suspensor is an than the heart-shaped embryo and contains as early embryonic organ that is programmed for termany as six different gibberellins (Picciarelli and minal differentiation and, except in a few cases, it Alpi 1986; Piaggesi et al 1989). The high concentra- does not contribute any cells to the sporophytic tion of gibberellins is reflected in the ability of a generation of the plant. Yet, it cannot be naively cell-free extract prepared from homogenized sus- assumed that the primary function of the suspenpensors to synthesize precursors of GA, as well as sor is to orient the embryo in close proximity to the GAj, GAy and GA8, from radioactively labeled sub- source of nutrition in the embryo sac or in the strates (Ceccarelli, Lorenzi and Alpi 1979,1981a, b). ovule. Important findings about the structure of In the heart-shaped embryo of P. coccineus, the walls of suspensor cells implicate these cells in cytokinins are also present in higher concentrations the absorption of metabolites from the surroundin the suspensor cells than in the cells of the ing tissues and in the transport of metabolites to embryo proper; a complete reversal of the status of the embryo. The cells of the suspensor, and espeboth gibberellins and cytokinins occurs in the sus- cially their polytene chromosomes, are clearly pensor and embryo at the cotyledon stage (Alpi, involved in transcriptional activity, although the Tognoni and D'Amato 1975; Lorenzi et al 1978). physiological role of the gene products syntheChanges in the auxin content of the suspensor have sized is not understood. Finally, the accumulation drawn little attention, although it appears that the of growth hormones by the suspensor has genersuspensor contains higher amounts of auxin than ated great interest in elucidating the possible funcdoes the embryo (Alpi, Tognoni and D'Amato 1975; tion of hormones in promoting embryo growth. Przybyllok and Nagl 1977). Very little ABA is pre- Despite these advances in our understanding of sent in the suspensor of P. coccineus during the ini- the suspensor, further studies are needed to gain tial stages of its development, but some increase, new insights into the physiological and molecular with two peaks, is noted subsequently (Perata, bases for its terminally differentiated state and for Picciarelli and Alpi 1990). its interaction with the embryo. The future direc-
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tions in the study of the suspensor are briefly considered by Yeung and Meinke (1993) in a thoughtful essay. 6. NUTRITION OF THE EMBRYO To a large extent, continued development of the embryo depends upon its interaction with the milieu of the embryo sac, the endosperm, and the cells of the ovule surrounding the embryo sac. Thus, in contrast to the static image conjured by pictures of embryos in isolation, it is reasonable to assume that developing embryos maintain a channel of communication with the external cellular environment. This traffic has in turn generated the reprise of a cherished objective of experimental plant embryologists, namely, to identify the substances that provide the nurture and nutriments to developing embryos, but this goal remains largely unattained. (i) Nutrient Supply to the Zygote It is noted that the primary endosperm nucleus generally begins to divide ahead of the zygote, thus ensuring that a mass of cells charged with storage products is in place when the zygote begins to divide. However, few would agree that the nutrition of a specialized cell like the zygote is, in the final analysis, grounded on the advance performance of another cell of a different genotype. This has led to a renewed interest in the ultrastructural analysis of the embryo sac, because an elucidation of the mechanism of zygote nutrition logically requires a consideration of the structure of the embryo sac before fertilization. It now appears that in a broad sense, the structural modifications of the embryo sac devoted to the nutrition of the egg persist after fertilization or find their counterpart in the embryo sac of the fertilized egg. It is likely that several other modifications of the embryo sac and its surrounding tissues for the nutrition of the female gametophyte described in Chapter 6 are carried forward unchanged after fertilization and have the same primary consequences for the zygote.
Zygotic Embryogenesis
and Palmer 1984; Chamberlin, Horner and Palmer 1993a), Arabidopsis (Mansfield and Briarty 1991), and Butomus umbellatus (Fernando and Cass 1996), or adjacent to the degenerating nucellus in wheat (Morrison, O'Brien and Kuo 1978). In Vicia faba, invaginations from the embryo sac wall are formed close to the zygote, as well as at the chalazal end opposite the surviving cells of the nucellus, referred to as the nucellar cap (Johansson and Walles 1993b). In an investigation to trace autoradiographically the fate of 14C-labeled photosynthates in the ovules of soybean, it was found that at the zygotic stage the label was concentrated over the wall projections of the embryo sac and in the hypostase at the chalazal end. The interesting conclusion drawn from this study - that two pathways of solute transport, a primary one from the outer integument to the base of the zygote and to the central cell, and a secondary one through the hypostase, operate to supply nutrients to the zygote - obviously bolsters a role for the wall invaginations in the process of nutrient supply (Chamberlin, Horner and Palmer 1993a). Zygote nutrition is a problem with wide ramifications and of considerable interest; the stimuli and nutrients that this cell normally receives from the rest of the plant body deserve to be characterized.
(ii) Embryo-Endosperm Relations From the time embryologists first began to understand the relevance of the endosperm in seed formation, the function of nurturing the embryo has traditionally been assigned to this tissue. Although evidence in favor of the role of the endosperm as a nurse tissue for the developing embryo is more circumstantial than experimental, the idea has gained great credibility over the years. The absence of the endosperm in mature seeds of some plants known to produce this tissue at early stages of ovule development seems to indicate that the endosperm is not a permanent source of nutrition but is apparently consumed by the growing embryo. In seeking structural adaptations in the proembryos that mediate the transfer of nutrients from the endosperm, as discussed in the previous section, much attention has centered on wall proliferations from the suspensor cells. The invaginations from the outer wall of the suspensor cells in contact with the endosperm, as seen in Capsella bursa-pastoris (Schulz and Jensen 1969, 1974),
The number of investigations that encompass the structure of the embryo sac wall after fertilization is comparatively small, and invariably all refer to the presence of wall projections akin to transfer cells. The projections originate from the inner wall of the embryo sac at the micropylar end close to the Diplotaxis erucoides (Simoncioli 1974), Phaseolus coczygote in Euphorbia helioscopia (Gori 1977), cottoncineus (Yeung and Clutter 1979), Vigna sinensis (Hu, (Schulz and Jensen 1977), soybean (Tilton, Wilcox Zhu and Zee 1983), and Glycine max (Chamberlin,
Embryogenesis and Physiology of Growth of Embryos
Homer and Palmer 1993a), and from the inner wall of the embryo sac at the micropylar or chalazal pole, as seen in pea (Marinos 1970a; Harris and Chaffey 1986), Haemanthus katherinae (Newcomb 1978), Medicago sativa (Sangduen, Kreitner and Sorensen 1983a), and Vigna sinensis (Hu, Zhu and Zee 1983), seem in particular to invite considerable speculation about the possible functions of the imaginations in nutrient transport into or from the endosperm. One investigation has shown that as the zygote of G. max is phased into a multicellular embryo, there is a sequential development of invaginations in at least four different sites in the embryo sac wall in addition to some wall ingrowths that appear in the cells of the inner integument and nucellus. This observation has drawn attention to the possibility that the wall of the embryo sac functions both as a sink and a source of apoplastic transport to nourish the growing embryo (Folsom and Cass 1986). In view of the postulated linkage between wall ingrowths and short-distance transport of solutes, it does not seem unreasonable to find evidence for the presence of ingrowths on the walls of proembryos (Dute, Peterson and Rushing 1989) or the walls of the embryonic cotyledons (Johansson and Walles 1993a). It is relevant to point out that a plasma membrane almost invariably follows the contour of the wall projections; this may not be coincidental, since the membrane greatly increases the surface area of the invaginations. Occasionally, concentrations of various organelles in the vicinity of the wall proliferations have been described with the implication that they serve to increase the active transport of solutes across the plasma membrane. However, the possible direction of the flow of solutes remains unclear because there is nothing to preclude the absorption of nutrients from the endosperm into the suspensor or the transport of metabolites from the degenerating suspensor into the endosperm. A sense of uncertainty prevails also about the direction of solute flow through the invaginations that arise from the inner wall of the suspensor cells. The timing of the direct absorption of endosperm nutrients by the embryo may also be worth emphasizing at this point. Direct absorption generally happens only after the suspensor has degenerated to the point that individual cells are separate from one another. The direct utilization of the endosperm by the growing embryo is indicated by the appearance of a conspicuous zone of destroyed endosperm cells around the cotyledons, as in dicots (Erdelska 1980,1985,1986), or ahead of
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the tip of the scutellum and coleoptile, as in cereals (Smart and O'Brien 1983; Jones and Rost 1989b). In a similar way, the accumulation of legumin and vicilin in the endosperm cells surrounding the embryo of Vicia faba and their subsequent decay may be looked upon as processes involved in the supply of nutrients to the growing embryo (Panitz et al 1995). Significant increases in the cationdependent phosphatase activity during tobacco seed development as detected biochemically and by ultracytochemical localization methods have been attributed to the active transport of assimilates to the endosperm and embryo (Mogensen and Pollak 1983; Mogensen 1985). To the extent that the cytoplasm of lysed cells can serve as a source of metabolites, the expanding growth of the embryo sac after fertilization has profound effects on the nutrition of the embryo. Expansion of the embryo sac is evident mostly at its micropylar or chalazal end when cells of the nucellus or integument begin to disintegrate and release free cytoplasm and nuclei; some of the newly liberated organelles are even incorporated into the endosperm (Pacini, Cresti and Sarfatti 1972; Norstog 1974; Schulz and Jensen 1974; Pacini, Simoncioli and Cresti 1975; Hardham 1976). In Capsella bursa-pastoris, cells of the residual intact chalazal nucellus enlarge to form a tissue (chalazal proliferating tissue) that protrudes into the embryo sac. The cells of this tissue actively incorporate precursors of both DNA and RNA and accumulate poly(A)-RNA during the period of their active growth. However, by the time the embryo attains the globular stage, degeneration in the chalazal proliferating tissue is initiated in the cells close to the embryo sac and continues until all the cells have broken down (Pollock and Jensen 1967; Schulz and Jensen 1971; Raghavan 1990b). The general interpretation of these various observations is that proliferations on the embryo sac wall aid in the passage of cytoplasmic nutrients from the lysed ovular cells into the central cell. (iii) Nutrient Supply from the Mother Plant
It is difficult to find structural evidence for the supply of nutrients from the vegetative parts of the plant to the growing embryo. From physiological experiments it is known that seeds that accumulate large quantities of storage reserves in the embryo or in the endosperm act as powerful sinks for metabolites from other parts of the plant. With legume seeds, the use of 14CO2, which is incorporated into photosynthates, has given the most con-
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vincing demonstration of the accumulation in the embryo of sugars manufactured in the leaf. Considering the amount of storage reserves synthesized by developing seeds, the vascular system of the ovule might be expected to play a role in translocating precursors to the embryo or to the endosperm. In developing pea seeds, it has been found that by the time storage protein synthesis is initiated in the cotyledons, most of the endosperm has been consumed. Storage protein synthesis coincides with a marked increase in the amount of vascular tissues in the ovule and funiculus, as well as a marked increase in the number of phloem transfer cells in the latter (Hardham 1976). However, it is uncertain whether nutrients transported through the vascular tissues from the vegetative parts of the mother plant are involved in the nutrition of the embryo or are transformed into storage products. An investigation that traced the fate of 14C-photosynthate into the embryo sac of soybean at the zygote stage was referred to earlier. Extension of this study has provided evidence for the flow of labeled photosynthate through the vascular system into the micropylar, chalazal, and lateral poles of the embryo sac for the nurture of developing globular to heart-shaped embryos (Chamberlin, Horner and Palmer 1993a). Yaklich et al (1992) have shown that the outermost layer of cells of the endosperm of soybean (considered analogous to aleurone cells) is reinforced by the generation of a significant number of Golgi vesicles and an abundance of ER coincident with the period of storage protein accumulation in the embryo. This observation has suggested the possibility of the translocation of photosynthates from the seed coat to the aleurone cells and their secretion into the milieu of the embryo for purposes of nutrition and storage protein synthesis. The main ideas and results that have flowed from several laboratories have been presented here to show that embryos developing in vivo employ different strategies for their nurture. These strategies can be related to specific stages of embryogenesis, with the wall ingrowths' having a prominent role at the zygote or proembryo stage. An immunochemical localization of high levels of a GTP-binding protein (G-protein) in developing embryos of Arabidopsis has fostered the argument that nutritional relations of embryos involve a source-sink interaction (Weiss, Huang and Ma 1993). Because G-proteins are involved in a variety of signal transduction pathways, growth hormones that pass into embryos from various sources can be considered as good candidates for signals transduced by these proteins. The nutrition of embryos is a subject that
Zygotic Embryogenesis
inevitably involves whole-plant physiology, ultrastructure, and molecular biology, with an array of interacting variables impinging on the problem, so only general conclusions are possible at this stage. As described in the following section, it seems easier to obtain information about the chemical nature of the regulatory substances necessary for embryo growth from in vitro culture than from an assault on embryos growing in vivo. 7. EMBRYO CULTURE
The cultivation of embryos excised from ovules and seeds under aseptic conditions in a medium of known chemical composition constitutes the basic methodology of embryo culture. By growing embryos outside the environment of the ovule it is possible to identify the special nutrient or hormonal requirements essential for their continued growth, differentiation, and morphogenesis. Ideally, one would hope to follow progressive embryogenesis in vitro in a defined medium, starting with a zygote, but in a true sense this aim is still a distant goal. Most of the work has been done on the culture of relatively mature and differentiated embryos excised from quiescent seeds. The logic behind this was the hope that a knowledge of the nutrient requirements for the growth of excised seed embryos under controlled conditions would provide clues about the nature of the storage materials utilized during seed germination. Another trend seen in this area was toward the culture of progressively younger embryos with a view to determining their changing nutritional requirements. From a theoretical angle, embryo culture draws attention to the degree to which growth and behavior of embryos of different ages may be regulated exogenously by nutritional or hormonal substances or by physical modifications of the culture environment; from an applied perspective, the technique offers a means of obtaining seedlings from inviable hybrids and overcoming dormancy of seeds.
(i) Seed Embryo Culture
Embryo culture has a long history dating back to the work of Hannig (1904), who obtained transplantable seedlings from embryos excised from the seeds of Raphanus sativus, R. landra, R. caudatus, and Cochlearia danica (Brassicaceae) grown in a medium containing macronutrient salts and cane sugar. Since the publication of this work, the subject has developed rapidly, generating a long list of plants whose embryos have been successfully cultured
Embryogenesis and Physiology of Growth of Embryos
under aseptic conditions in defined media as a means of providing useful information on the physiology of embryo growth. Rather than deal encyclopedically with these studies, the account given here is in large part based on the work done on a few selected species to elucidate the requirements for growth and differentiation of embryos in culture. Carbon and Nitrogen Nutrition. Although supplementation of the culture medium with a carbon energy source is not essential for inducing the growth of isolated seed embryos, it is the general experience that the growth and survival of embryos are markedly enhanced by its presence. A caveat should be added here: Even with the ideal sugar in the medium, the growth of embryos may be influenced by other factors, such as the genotype of the donor plant (Dunwell 1981). At the morphogenetic level, addition of sucrose to the medium has been shown to promote root growth or the growth of both shoot and root primordia of cultured embryos (Rietsema, Satina and Blakeslee 1953; Buffard-Morel 1968). Embryos of Datura stramonium (Rietsema, Satina and Blakeslee 1953) and pea (Davis 1983) require a higher sucrose concentration for optimum root growth than that required for optimum shoot growth, and that may partly explain the reason why a concentration of sucrose effective in enhancing root growth inhibits shoot growth. The favorable effect of dextrose on the root growth of embryos of Cocos nucifera is dependent upon the composition of the mineral-salt medium, because a high nitrogen-containing medium like Murashige-Skoog medium is particularly effective in this respect (Del Rosario and de Guzman 1976). When one realizes that shoot and root primordia are already laid down in the embryo at the time of culture, it becomes clear that the role of sugar in the morphogenesis of embryonic organs has to be viewed in a limited context. To the extent that supplementation of the medium with various nitrogen compounds tells us something about the amino acid metabolism of cultured embryos, a few generalizations have emerged. One is that embryos are able to grow moderately well in a medium containing nitrates or ammonium salts as the sole source of nitrogen. Secondly, the amide glutamine has proved to be an efficient source of nitrogen for the growth of embryos of several plants (Paris et al 1953; Rijven 1956; Matsubara 1964). Thirdly, mutual antagonism and synergism exist between different amino acids in the growth of cultured embryos. For example, a mixture of 20 amino acids put together in the proportions present in casein hydrolyzate is as effective
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as casein hydrolyzate in promoting growth of preheart-shaped and early heart-shaped embryos of D. innoxia and D. stramonium. This occurs despite the fact that the mixture contains readily utilized, partially utilized, and even relatively toxic compounds. Moreover, favorable effects of the complete amalgam are not reproduced by incomplete mixtures of amino acids or by single amino acids (Sanders and Burkholder 1948). The growth of embryos of cereals such as wheat, barley, and maize is inhibited by the simultaneous addition of lysine and threonine to the medium; the inhibition is alleviated by methionine, homoserine, or homocysteine (Bright, Wood and Miflin 1978; Green and Donovan 1980). Since these observations indicate that embryos are starved for methionine and its metabolic intermediates by feedback inhibition of enzymes of the aspartate pathway, they are of interest for possible selection of feedback-sensitive mutants. Hormonal Effects. Following the discovery of plant hormones, especially auxins, gibberellins, and cytokinins, concerted efforts have been made to study their effects on the growth and morphogenesis of cultured embryos. The general conclusion is that although the addition of hormones modifies the growth of cultured embryos, there is no evidence to indicate the need for a specific hormone for the growth of embryos of a wide range of plants. As far as the effects of hormones on the growth of embryonic organs are concerned, they are quite similar to those displayed by the corresponding organs of a seedling plant. As seen in cultured embryos of Capsella bursa-pastoris, the effect of IAA is manifest in the promotion of growth of the radicle at low concentrations; the inhibition of growth of the radicle, hypocotyl, and shoot at intermediate concentrations; and callus initiation at high concentrations (Raghavan and Torrey 1964). However, it appears that globally the most effective auxin for inducing callus growth in cultured embryos is 2,4-D and, as will be seen in Chapter 16, in many instances calluses thus produced provide the starting material for the induction of somatic embryos. Among the various parameters of auxininduced callus growth on cultured embryos, the genotypic effect of the donor plant has come under intense scrutiny. In oat (Cummings, Green and Stuthman 1976), wheat (Maddock et al 1983), and Poa pratensis (McDonnell and Conger 1984), generic differences account for the ability of cultured embryos of more or less the same age to produce differentiating cultures; in maize, the response of embryos of the same and different ages to 2,4-D is
386 dependent on the particular genotype (Green and Phillips 1975). Varietal differences, as well as the growing conditions of the plant from which embryos are excised, seem to determine the course of auxin-induced regeneration of callus from cultured barley embryos (Bayliss and Dunn 1979). The organ that differentiates the callus in response to auxin application appears to vary in embryos of different species, as well as in embryos of different ages of the same species. In mature oat embryos, the radicle seems to be the most responsive organ and the scutellum undergoes necrosis, whereas whole young embryos are transformed into a callus without any necrosis of tissues (Cummings, Green and Stuthman 1976). In maize, irrespective of the age of embryos at excision, calluses are most successfully induced from the scutellum (Green and Phillips 1975). This organ is also the most proliferative in immature embryos of barley, whereas growth in the more mature embryos occurs from the mesocotyl-radicle region (Bayliss and Dunn 1979; Dale and Deambrogio 1979). The ease of callus regeneration from specific organs of cultured embryos does not mean that other organs of the embryo are incapable of regeneration; apparently, in embryos of different plants, cells of certain organs with an increased potential for uncontrolled growth possess a selective advantage and tend to overgrow cells of other organs. Contrary to the general experience that roots are relatively insensitive to GA, the hormone promotes root elongation and the production of lateral roots in cultured embryos of C. bursa-pastoris. If one considers the effects of GA on embryos of different ages, older embryos are seen to produce roots with abundant laterals whereas younger embryos form roots with fewer laterals (Veen 1963; Raghavan and Torrey 1964). Cultured embryos of two different ages of cotton also display differences in GA sensitivity, such as the enhancement of cotyledon maturation and cell elongation in immature embryos, and the acceleration of cell elongation, cell division, and axis growth in older embryos (Dure and Jensen 1957). These observations are in accordance with a speculative view that GA effectively induces growth in embryos in which the meristems are sufficiently well developed. The limited information available on the morphogenetic effects of cytokinins concerns kinetininduced inhibition of root growth in cultured embryos of C. bursa-pastoris. The degree of inhibition depends on the age of embryos and the conditions of culture. In cultures grown in complete darkness, relatively low concentrations of kinetin,
Zygotic Embryogenesis
which inhibit root elongation in heart-shaped embryos, are marginally inhibitory in slightly older embryos and are without effect in torpedo-shaped and mature embryos. In light-grown cultures, kinetin inhibits root elongation in embryos of all ages but promotes precocious leaf expansion and callus growth (Veen 1963; Raghavan and Torrey 1964). An indirect role for an auxin-cytokinin mixture in inducing the growth of embryos is seen in the observation that seed embryos of Colocasia esculenta (taro; Araceae) develop into plantlets only when they are cultured in the presence of the endosperm. Indicative of a role for hormonal substances of endospermic origin in the growth of the embryo, it was found that NAA and 6-methylaminopurine can replace the endosperm effect on cultured embryos (Nyman et al 1986,1987). From this brief survey, it appears that despite the different growth modifications induced by hormones in cultured seed embryos, there is no evidence to demonstrate or exclude the involvement of a specific hormone in the morphogenesis of embryos. This indicates that mature embryos excised from quiescent seeds have a sufficient supply of endogenous hormones for the initiation of growth when they encounter a relatively simple nutrient medium, a carbon energy source, the normal composition of the atmosphere, and a favorable temperature. As will be seen, a role for hormones in the growth of immature embryos and proembryos is more explicit than their role for mature embryos. (ii) Culture of Immature Embryos and Precocious Germination
During the early period of embryo culture investigations, Dieterich (1924) discovered that whereas mature embryos grown in culture generally produce seedlings that look normal in most respects, cultured immature embryos inexplicably skip the later part of the embryogenesis program and grow into rudimentary, weak seedlings. These malformed seedlings were referred to as "kiinstliche Fruhgeburt" and the process whereby the embryo becomes competent to respond to germination cues before it has fully played out its maturation program is now well known as precocious germination. This apparent dichotomy in the growth patterns of mature and immature embryos has repeatedly been noted in subsequent years by various investigators and has invited frequent comment. In developmental terms, during precocious germination, the normal process of embryogenesis that would lead to a period of arrested
Embryogenesis and Physiology of Growth of Embryos
growth of the embryo in the seed resulting in quiescence or dormancy, is replaced by an alternate program that leads to germinative growth. In most plants there is a vulnerable period during embryogenesis when embryos germinate precociously upon transfer to a culture medium; those cultured earlier than this period fail to grow, whereas older embryos germinate normally. One approach employed to investigate the causes of precocious germination and its control is by manipulation of the medium composition. This is based on the view that immature embryos are already predisposed to evolve into full-term embryos when they complete their development within the ovule, but upon excision and culture, they are prevented from doing so by the limitations imposed by the nutrient medium. The first noteworthy attempt at manipulation of the culture medium to control precocious germination utilized barley embryos as the test material. Normal growth of mature barley embryos cultured in an agar medium containing mineral salts and sucrose is characterized by the enlargement of the scutellum and the development of the first leaf and root within their respective sheathing structures, whereas precocious germination of immature embryos involves the formation of short, spindly seedlings. If the medium is additionally fortified with casein hydrolyzate, tomato juice, or other natural plant extracts, precocious germination is forestalled in the immature embryos and an embryogenic program is reinstated (Kent and Brink 1947). The effectiveness of casein hydrolyzate in preventing precocious germination of barley embryos suggested a role in the process for one or more components of this nutrient mixture, such as the inorganic phosphate, sodium chloride, or amino acids. When the growth of immature embryos of barley in media to which the three groups of components of casein hydrolyzate were added singly or in combination was monitored, it was found that amino acids and sodium chloride, mixed in the same proportion in which they are present in casein hydrolyzate, effectively duplicate the activity of casein hydrolyzate in suppressing germination. Surprisingly, immature embryos completed the embryogenic program in a medium containing sucrose or mannitol at concentrations providing an osmotic pressure equal to that of casein hydrolyzate; this supports the view that inhibition of precocious germination results from the high osmotic pressure of the medium and is not due to a specific nutrient requirement satisfied by casein hydrolyzate (Ziebur et al 1950). The tendency of immature barley embryos to germinate
387
precociously is diminished to a considerable degree by high intensities of light, moderately high temperatures, and reduced oxygen tension in the culture milieu (Norstog and Klein 1972). An arresting observation by Norstog (1972b) that precocious germination of cultured immature barley embryos is inhibited by ABA is relevant to this discussion. This result, which pointed to a role for endogenous ABA as an inhibitor of precocious germination during normal seed development, has been confirmed in experiments with embryos of Brassica napus (Crouch and Sussex 1981; Wilen et al 1991), Gossypium hirsutum (Choinski, Trelease and Doman 1981), Triticum aestivum (Triplett and Quatrano 1982; Morris et al 1985; Hess and Carman 1993), Glycine max (Ackerson 1984a; Eisenberg and Mascarenhas 1985), Oryza sativa (Stinissen, Peumans and de Langhe 1984), Hordeum vulgare (Morris et al 1985), Medicago sativa (Xu, Coulter and Bewley 1990), Zea mays (Rivin and Grudt 1991), and Linum usitatissimum (Wilen et al 1991). Results of a typical experiment are shown in Figure 13.9. More definitive evidence favoring a role for ABA in the control of precocious germination of embryos has come from other investigations, lhle and Dure (1970,1972) examined the activity of certain proteases involved in the mobilization of stored reserves of cotyledons of cotton embryos as an event connected with germination and showed that culture of immature embryos triggers precocious germination as well the development of enzyme activity by premature transcription of mRNAs. Because removal of the embryo from the ovule promotes precocious germination, it appeared that the ovule tissues might supply to the embryo some regulatory molecules that prevent germination. This was found to be the case with the discovery that addition of an aqueous extract of the ovule or ABA to the medium halts precocious germination of immature embryos and the development of enzyme activity in them. In attempts to correlate the levels of ABA in the reproductive tissues to the patterns of growth of cultured embryos, it was found that embryo axes excised from mature soybean seeds containing low levels of ABA germinate soon after culture, whereas those from immature seeds with high levels of endogenous hormone do not germinate or require a long lag period before germination. When the endogenous ABA is removed from immature embryos by washing them in water or by drying them in the pod, embryos germinate precociously (Ackerson 1984a, b). According to King (1976), embryos excised from immature wheat grains containing high concentra-
388
Zygotic Embryogenesis
100-
10' 3
ABA concentration (mol-m ) in culture medium Figure 13.9. Germination index of developing embryos of Medicago sativa (stages III, youngest, to IX, oldest) on a medium containing 1% sucrose and different concentrations of ABA. Germination of embryos of stages Ill-V was inhibited by low concentrations of the hormone. (From Xu and Bewley 1991. J. Expt. Bot. 42:821-826, by permission of Oxford University Press.)
tions of ABA germinate infrequently as compared to the high germination percentages of mature embryos excised from dry grains containing low levels of the hormone. High concentrations of ABA are found in the embryos of Phaseolus vulgaris when they are just phased out of the stage vulnerable to precocious germination, and again at seed maturity as they begin to desiccate (Morris 1978; Hsu 1979). We see here a curious situation in which a sustained supply of ABA from the ovule tissue checks the germination instincts of the developing embryo and commits it to a pathway of maturation and dehydration inside the seed. At this time, the embryo is liberated from the chemical influence of the ovule tissue and germinates in a normal fashion when confronted with favorable conditions. Investigations on the protein metabolism of germinating embryos also cast some light on the role of ABA in the regulation of precocious germination. As reviewed in Chapters 14 and 15, synthesis of a variety of proteins, including LEA proteins, storage proteins, and defense-related proteins, is slowed down or inhibited in precociously germinating embryos, whereas in the presence of a concentration of ABA in the medium appropriate to suppress precocious germination, cultured embryos accumulate these pro-
teins to the same extent as those growing in situ. Other investigations have, however, questioned a role for ABA in preventing precocious germination. In addition to ABA, osmotically active agents such as mannitol and sucrose can also inhibit precocious germination and, in some cases, a high osmotic pressure is more effective in this role than exogenous ABA. Moreover, the results of experiments to determine the levels of endogenous ABA in embryos cultured in high osmoticum are consistent in showing that inhibition of precocious germination is not necessarily accompanied by a rise in ABA levels (Finkelstein and Crouch 1986; Morris et al 1988, 1990; Barratt et al 1989). Whereas both ABA and osmoticum effectively prevent precocious germination of immature embryos of M. sativa, proteins that are the hallmarks of embryos developing in situ in the pod are synthesized only in the presence of the osmoticum (Xu, Coulter and Bewley 1990). Another work showed that ABA and osmoticum are differently effective at different stages of embryogenesis and that when both agents are present, the combined inhibitory effect is greater than the sum of the individual effects (Xu and Bewley 1991). These various observations are in line with the view that ABA is not the universal regulator of precocious germination of embryos; even when a distinct effect of the hormone is documented, rather than being the primary effector of the process, ABA might inhibit precocious germination secondarily by blocking water uptake by the embryos. One of the points to be considered in assessing the control of precocious germination is the importance of the constraint imposed by the seed environment and the associated seminal tissues. Because two potential regulatory factors, namely, ABA and osmoticum, have been studied with inconclusive results, it seems likely that other specific signals supplied by the ovular tissues may predispose the embryo to complete its maturation program rather than opt for the germinative mode of development. Release of the embryo from the influence of the maternal environment has been achieved by premature desiccation of the seed; subsequent hydration of the seed switches the embryo from a developmental to a germination mode. As in precocious germination, embryos of prematurely desiccated seeds are most prone to germinate during a narrow developmental window restricted to the period before they naturally desiccate and pass into the quiescent or dormant state (Kermode and Bewley 1985,1988). At the biochemical level, premature drying suppresses developmental protein synthesis in the embryo and
Embryogenesis and Physiology of Growth of Embryos
induces germination protein synthesis; a switch in the synthesis of mRNAs from those coding for developmental proteins to those encoding germination proteins also occurs during premature drying and germination (Dasgupta and Bewley 1982; Misra and Bewley 1985a; Oishi and Bewley 1992; Xu and Bewley 1995a). Precocious germination and germination after premature drying are two faces of the same coin, reflecting on the need for completion of the embryo maturation program for successful seedling development. Furthermore, one may assume that cultured immature embryos germinate precociously because they have lost access to the pool of endogenous signals from the maternal tissues and premature drying depletes the maternal tissues of the signals necessary for inducing embryo maturation. (iii) Culture of Proembryos
During their growth within the ovule, proembryos, including the zygote, are heterotrophic in nature and are dependent not only upon the metabolites present in their cells, but also upon those diffusing from the endosperm and the surrounding maternal tissues. From the embryo culture perspective, this suggests that the nutritional requirements for the culture of proembryos are bound to be more exacting than those for immature embryos, and mature, differentiated embryos. In addition, the technical difficulties involved in isolating uninjured zygotes and proembryos have made it impossible to study the initiation and progress of cell divisions in culture in all but a few species. To follow the course of development in this field, the topic will be reviewed here under the following headings: (a) effect of nutrients of endospermic origin; (b) effect of the physical parameters of culture; (c) role of hormones; (d) role of the suspensor; (e) zygote culture; and (f) culture of fertilized ovules and embryo sacs.
389
pantothenic acid) and an assortment of organic substances (adenine, glycine, and succinic acid). Although still smaller embryos (200-500 [im long) failed to grow, or grew feebly in this medium before turning into undifferentiated callus, a dramatic recrudescence of growth occurred in these embryos when the medium containing vitamins and organic addenda was further supplemented with nonautoclaved coconut milk. This was attributed to the presence of a hormonal factor, designated as the "embryo factor," which was later obtained in a form sufficiently pure enough to promote growth of embryos in a dilution as low as 1:19,000 on a dry weight basis as compared to 1:110 for the crude coconut milk (van Overbeek, Siu and Haagen-Smit 1944). In later years, these studies spawned several successful attempts to culture proembryos of other plants by the use of plant extracts of endospermic or nonendospermic origin, most notably of maize by the use of water extracts of date, banana, wheat gluten hydrolyzate, and tomato juice (Kent and Brink 1947); D. tatula by diffusates from young seeds of Lupinus luteus and old seeds of Sechium edule (Cucurbitaceae) (Matsubara 1962); Cucumis sativus, Cucurbita maxima, and C. moschata by extracts of their respective endosperms (Nakajima 1962); and Secale cereale by use of its own endosperm as a nurse tissue (Stefaniak 1987).
Nutrient media containing undefined natural plant extracts cannot be regarded as chemically defined until the essential functions of the latter can be duplicated by, and restricted to, known compounds. An important step toward the formulation of a completely synthetic medium for the culture of proembryos of barley is noteworthy in this context. Norstog (1961) showed that undifferentiated barley embryos, which generally fail to survive in White's mineral-salt medium, can be successfully cultured by supplementing the medium with coconut milk. The smallest embryo thus cultured had probably no more than 100 cells (about 60 fim long), and its Effect of Nutrients of Endospermic Origin. An survival and subsequent growth in culture were early lead in the successful culture of proembryos enhanced by the addition to the medium of glutacame from the work of van Overbeek, Conklin and mine or a mixture of amino acids besides coconut Blakeslee (1942) on the physiology of growth of milk. In a further modification of this medium, a embryos of Datura stramonium. From previous rigorous definition of growth requirements for the studies of these investigators, there was some gen- culture of barley embryos as small as 90 ^m was eral agreement that it was possible to grow heart- achieved with the demonstration that a phosphateshaped and torpedo-shaped embryos of D. enriched White's medium at an optimum pH of 4.9 stramonium to the plantlet stage in a mineral-salt fortified with glutamine and alanine (400 mg/1 medium enriched with a mixture of vitamins (nico- each) as major nitrogen sources and lesser amounts tinic acid, thiamine, pyridoxine, ascorbic acid, and of leucine (20.0 mg/1), tyrosine, phenylalanine, cys-
390 teine, and tryptophan (all at 10.0 mg/1) is an effective substitute for coconut milk (Norstog and Smith 1963). The effectiveness of endosperm extracts of Cucumis sativus, Cucurbita maxima, and C. moschata
in promoting the growth of their respective embryos, mentioned earlier, is similarly matched by manipulation of the hormonal and organic additives to the medium, because a mixture of IAA, 1,3diphenylurea, and casein hydrolyzate supports growth of embryos to the same extent as does a medium containing the embryo factor (Nakajima 1962). The conclusion from these results is that growth induction in cultured proembryos by endosperm extracts is a measure of the effect of specific chemical components of the endosperm. Effect of the Physical Parameters of Culture.
Interest in the effect of the physical conditions of culture on the continued growth of proembryos was stimulated by the common observation that the amorphous liquid endosperm in which the proembryos are constantly bathed has a high osmolarity. Measurements have shown that the ovular sap surrounding proembryos has a very low osmotic potential value (negative), which substantially decreases (becomes more negative) with embryo maturity (Ryczkowski 1960; J. G. Smith 1973; Yeung and Brown 1982). This has led to the view that the osmotic pressure of the medium might play a role in promoting the growth of proembryos in vitro. The importance of this parameter of the culture medium in the control of the growth of proembryos has not been critically tested, although the balance of evidence is in favor of adjustment of the medium to be isotonic with the ovular sap for this purpose. As indicated earlier, the embryo factor from coconut milk fully met the requirements for continued growth of pre-heartshaped embryos of D. stramonium. However, another work showed that whereas mature embryos of this species grow in the complete absence of sucrose in the medium, earlier-stage embryos require progressively higher concentrations of sucrose. For example, a medium containing 8%-12% sucrose is found optimal for continued growth and differentiation of globular and pre-heart-shaped embryos. Interestingly enough, these embryos grow to the same extent in a medium containing 2% sucrose plus enough mannitol to be isotonic with 8%-12% sucrose (Rietsema, Satina and Blakeslee 1953). From this it appears that the essentiality of sucrose in embryo culture medium is to be accounted for wholly or in part as an osmotic stabilizer rather than as a carbon energy
Zygotic Embryogenesis
source. The successful culture of proembryos of Capsella bursa-pastoris (Rijven 1952; Veen 1963), D. tatula (Matsubara 1964), Linum usitatissimum (Pretova 1974), and Triticum aestivum (Fischer and Neuhaus 1995) has also been linked to the provision of a high osmolarity in the medium. Although success has been sporadic, a new small-size-limit in proembryo culture in a medium of high osmolarity has been attained with 25^45-^m-diameter proembryos of Arabidopsis (Wu et al 1992). The need to gradually reduce the osmolarity of the medium during the growth of embryos without sequential transfer from one medium to another has resulted in two technical modifications of the culture system. For the culture of embryos of C. bursa-pastoris as small as 50 ium in diameter, a continual change in the osmolarity of the medium was obtained by using two media of different compositions, solidified in juxtaposition in a Petri dish. During growth in the medium of high osmolarity, the embryo was subjected to continually changing osmotic conditions due to diffusion from the medium of low osmolarity (Monnier 1976a). A culture system comprised of two agar layers, with the top layer having a higher osmolarity than the bottom layer, was used to obtain continued growth and differentiation of 8-36-celled proembryos of Brassica juncea. Proembryos were embedded in the top layer, whose osmolarity decreased during culture (Liu, Xu and Chua 1993a). Although no illuminating ideas as to how osmolarity of the medium regulates the cells of the tiny embryos to engage their intrinsic ability for division and growth have been generated by these investigations, an effective control of the flow of metabolites and inorganic ions into the cells of the embryo by the osmoticum appears a distinct possibility. Role of Hormones. Because of the widespread use of hormones as adjuvants in tissue culture media for growth induction in organ, tissue, and cell cultures of plants, it is not unexpected that proembryo culture has come under the spell of hormones. Embryos of C. bursa-pastoris have figured prominently in the investigations using hormones. Although some workers (Rijven 1952; Veen 1963) had successfully cultured heart-shaped embryos in an inorganic liquid medium of high osmolarity secured by the addition of 12%-18% sucrose, a later work showed that it is possible to culture these same embryos in an agar-solidified mineral-salt medium supplemented with 2% sucrose. Growth of still smaller embryos (up to about 55 \xm long) was secured by fortifying this medium with IAA,
Embryogenesis and Physiology of Growth of Embryos
kinetin, and adenine sulfate (Raghavan and Torrey 1963). The addition of the hormone mixture eliminated the need for a medium of high osmolarity, suggesting that the picture of osmotic regulation of the growth of proembryos is somewhat misleading. Nonetheless, the need for a properly balanced mixture of hormones for the growth of these embryos is in a large part met by an osmoticum or by an increase in the concentration of major salts by a factor of 10. The more the mineral salts of the medium became fully competent to support the growth of proembryos, the more they appeared likely to be the governing factor in regulating proembryo growth. From a comparative analysis of different inorganic salt media, Monnier (1976a) found that the high-salt medium of MurashigeSkoog is superior for the culture of proembryos of C. bursa-pastoris. A concurrent experiment also revealed the interesting point that excellent growth of proembryos ensued when they were cultured in a modification of this medium with increased levels of Ca+ and K+ and a decreased concentration of NH4NO3. In other approaches, enhanced growth of proembryos of C. bursa-pastoris was obtained by supporting them on a bed of polyacrylamide instead of agar (Monnier 1975) or by increasing the partial pressure of O2 in the medium (Monnier 1976b). Although C. bursa-pastoris has dominated the field of proembryo culture involving manipulation of hormonal and other additives to the medium, a successful report of culture of proembryos of Linum usitatissimum by kinetin (Pretova', 1986) is noteworthy. As experiments in the culture of early division phase proembryos and of the single-celled zygotes of these plants are contemplated, simultaneous changes in the chemical and physical conditions of culture offer a means of channeling the work in new directions. Role of the Suspensor. As discussed earlier, in the division of labor of the whole embryo, the suspensor with its wall projections for solute absorption and transport is presumed to channel exogenous stimuli for the growth of the organogenetic part of the embryo. This is especially true at the proembryo stage, when the suspensor is at the prime of its life before degradative changes set in. This has raised the question as to whether it is possible to demonstrate a causal relationship between the presence of the suspensor and the growth of proembryos in culture. The simple approach to interfering with the influence of the suspensor is to sever its connection with the embryo and follow the growth of the embryo in culture. This is no
391
doubt an exacting task; it has been performed with only a few species, perhaps involving a limited number of embryos. From experiments using this approach, it was found that continued growth of proembryos of Eruca sativa (Corsi 1972), Phaseolus coccineus (Nagl 1974; Yeung and Sussex 1979), and C. bursa-pastoris (Monnier 1984) is more enhanced in the presence of an attached suspensor than in its absence. Growth of embryos of P. coccineus is promoted even by the presence of a detached suspensor kept in close proximity in the medium (Yeung and Sussex 1979). A string of experiments with embryos of P. coccineus involving supplementation of the medium with growth hormones and determination of the growth hormone levels separately in the embiyo and suspensor has provided indirect evidence showing that the alleged suspensor effect is due to the production of GA and cytokinins. Whereas intact pre-cotyledonary stage embryos grow in a mineral-salt medium, the chance of survival of embryos of the same age deprived of the suspensor is very remote unless GA is present in the medium. GA is, however, inimical to the growth of suspensorless post-cotyledonary stage embryos as compared to intact embryos of the same age grown in a hormone-free medium (Cionini et al 1976; Yeung and Sussex 1979). These responses of embryos to GA are correlated with a falling level of endogenous GA in the organogenetic part and a rising level in the suspensor of the pre-cotyledonary stage embryos. As the suspensor begins to undergo senescence in the post-cotyledonary stage embryos, there is a sharp decline in the concentration of GA in the suspensor and a corresponding increase in its titer in the embryo proper (Alpi, Tognoni and D'Amato 1975). Likewise, addition of zeatin or zeatin riboside fosters the growth of suspensorless pre-cotyledonary stage embryos of P. coccineus, but post-cotyledonary stage embryos scarcely respond to the cytokinins (Bennici and Cionini 1979). As prototypes of hormones that promote cell divisions, cytokinins like zeatin and its riboside might be expected to be present in the organogenetic part of the post-cotyledonary stage embryos in a quantity sufficient to promote embryo growth. Indeed, this has been found to be the case, in a manner that is comparable to the presence of the same cytokinins in the suspensor part of the pre-cotyledonary stage embryos (Lorenzi et al 1978). These results provide a strong argument for a role for hormonal gradients from the suspensor in promoting growth of proembryos. This concept might serve as baseline infor-
392
Zygotic Embryogenesis
Figure 13.10. Culture of isolated zygotes of Zea mays, (a) Section of a polarized zygote 3 hours after culture. (b)Zygote showing first division 19 hours after culture; arrowheads point to the newly formed cell wall, (c) Section through an
embryo formed from a zygote 11 days after culture, showing the embryo proper and suspensorlike part. N, nucleus. Scale bars = 10 urn (a, b) and 50 urn (c). (From Leduc et al 1996; photographs supplied by Dr. C. Dumas.)
mation for formulating a medium for growing in vitro still smaller embryos, and even the zygote,
favorable effect of the nurse tissue may conceivably depend upon the release of undefined nutrients into the medium, these results indicate that, analogous to the culture of isolated proembryos, requirements for regenerating plants from zygotes are likely to be exacting and varied. The potential of the maize system for the transfer of foreign genes has been underscored by preliminary experiments showing transient expression of microinjected reporter genes.
of P. coccineus.
Zygote Culture. Up to the present time, only two successful attempts at zygote culture have been reported. Zygotes extruded from embryo sacs of barley developed into embryolike structures that yielded fully fertile plants when cocultured with embryogenic microspores of barley (Holm et al 1994). Through the use enzyme digestion and microdissection, zygotes of maize excised from embryo sacs were similarly induced to form fertile plants by simulating typical in vivo embryonic development (Leduc et al 1996). The unexpected finding was that the same nurse tissue of embryogenic microspores of barley was very effective in inducing continued growth and divisions of explanted maize zygotes (Figure 13.10). As the
Culture of Fertilized Ovules and Embryo Sacs. Since embryos must be isolated in order to be cultured, the difficulty of extracting them from the inner sanctum of the ovule could well be one of the reasons why there are so few reports of successful culture of proembryos. From this perspective, the culture of ovary, ovule, and embryo sac has proved to be particularly rewarding for the analysis of the
Embryogenesis and Physiology of Growth of Embryos
393
growth requirements of proembryos and zygotes of transfer of embryos to a medium of a different certain plants that have hitherto defied attempts at composition is a relatively simple protocol for excision and culture. As in embryo culture studies, regenerating plants in vitro from the zygote (M61, much effort in ovule culture has gone to devising an Matthys-Rochon and Dumas 1993,1995). appropriate medium. Some early investigations reported only limited success in the culture of isolated ovules of Papaver somniferum containing the 8. GENERAL COMMENTS zygote or two-celled proembryo in a mineral-salt Embryogenesis in angiosperms, in whatever conmedium containing 5% sucrose; rapid growth of the text of study, leads ultimately to the analysis of celluenclosed nascent sporophyte was obtained in a lar mechanisms using histological, cytological, medium supplemented with casein hydrolyzate, genetic, biochemical, molecular, and tissue culture yeast extract, or kinetin (Maheshwari 1958; techniques. The distinction between dicots and Maheshwari and Lai 1961). Other examples, such as monocots offers little difficulty because differences in Zephyranthes sp. (Kapoor 1959), Capsella bursa-pas-embryogenic development between the two classes toris (Lagriffol and Monnier 1985), Hordeum vulgare relate mainly to the division patterns that herald his(Topfer and Steinbiss 1985; Holm et al 1995), togenesis and organogenesis. Although different patTriticum aestivum (Zenkteler and Nitzsche 1985; terns of early embryogenesis have been described, Comeau et al 1992), and Zea mays (Schel and Kleft there is little or no indication of the functional mean1986), can also be cited to illustrate the successful ing of these division pathways. Admittedly, many application of ovary and ovule culture to promoting materials that are needed by the zygote to commence the growth of the enclosed zygote or proembryo. divisions are stockpiled or synthesized by this cell Accelerated growth of the ovular tissues in rela- during a period of growth preceding division; howtion to the growth of the enclosed embryo or vice ever, there is no consistent pattern of distribution of versa may also determine the success or failure of these materials in the zygote that foreshadows the ovule culture. This problem has been analyzed in a major parts of the embryo. The zygote shows maxigeneral consideration of the need to formulate a mum dependence for its growth on the endosperm, medium for the culture of cotton ovules at the both for organic nutrients and for cell division and zygote stage. Working from the experience of pre- morphogenetic stimuli. vious investigators, who obtained perfectly During embryogenesis, a variety of important respectable seeds with underdeveloped embryos changes occur, commonly including activation of from cultured proembryo stage ovules of cotton, many metabolic pathways associated with the proStewart and Hsu (1977) found that postfertilization duction of new tissues and organs. This is reflected ovules of cotton can be cultured with a reasonable in the progressive transition from heterotrophy of degree of success in a medium containing low con- the proembryos to autotrophy of mature and difcentrations of IAA, kinetin, and GA along with 15 ferentiated embryos. Interaction between the mM NH4+. The key to the combined use of hor- embryo and surrounding endosperm has a role in mones and ammonium ions was the observation determining the acquisition of metabolic compethat the addition of hormones alone enables the tence. Taken as a whole, the physiological and cytoovules to attain a normal size, whereas NH4+ pro- logical properties of the suspensor serve to motes growth and division of the zygote. illustrate one of the few possible ways hetIntermediate systems between embryo culture erotrophic nutrition of the embryo is sustained. and ovule culture have been developed for some Several technical innovations suggest that methplants to obtain growth in vitro of zygotes and ods are at hand to monitor the requirements for proembryos. Such systems include the culture of growth of the fertilized egg and proembryos outzygote-containing embryo sacs of Z. mays sur- side the confines of the embryo sac. In successful rounded by the nucellus with or without a block of experiments designed to nurture the zygote and the endosperm (van Lammeren 1988b; Campenot proembryo in defined media, it might even be poset al 1992; M61, Matthys-Rochon and Dumas 1993, sible to identify the nutrient substances provided 1995; Leduc, Matthys-Rochon and Dumas 1995) by the endosperm and those present in the zygote and the culture of embryo sacs of Cucumis sp. con- and proembryo. Availability of this information taining proembryos and free nuclear endosperm will permit more specific conclusions to be drawn (Custers and Bergervoet 1990). Initial culture of about the biosynthetic pathways activated during embryo sacs of Z. mays enclosing zygotes or two- progressive embryogenesis in angiosperms. celled proembryos combined with a subsequent
Chapter 14
Genetic and Molecular Analysis of Embryogenesis 1. Genetic Control of Embryogenesis (i) Embryo Abortion in Wide Hybrids (ii) Radiation-induced Mutations (iii) Spontaneous Embryo Mutants 2. Gene Expression Patterns during Embryogenesis (i) Diversity and Changes of mRNA Population during Embryogenesis (ii) Gene Activity during Fertilization and Early Zygote Divisions (iii) RNA Metabolism in Developing Embryos (iv) Protein Metabolism in Developing Embryos 3. Characterization of Genes Essential for Embryogenesis (i) Embryo-lethal Mutants (ii) Genetic Control of Embryonic Pattern Formation 4. Expression of Genes for Housekeeping and Structural Proteins (i) Genes for Glyoxylate Cycle and Related Enzymes (ii) Heat-Shock Protein Genes (iii) Expression of Other Genes 5. Expression of Genes Related to Desiccation Tolerance and Dormancy (i) LEA Genes from Cotton and Other Dicots (ii) LEA Genes from Wheat and Other Monocots (iii) Is Embryo Desiccation Developmentally Regulated by ABA? (iv) Genes Involved in Dormancy Regulation and Seedling Growth 6. Regulation of Expression of Defense-related Genes (i) Lectin Gene Expression in Embryos of Cereals and Grasses (ii) Expression of Defense-related Genes in Embryos of Other Plants 7. General Comments
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Completion of embryogenesis in angiosperms results in the formation of a full-term embryo, which is an assemblage of organs whose cells and tissues are specialized for functions essential for the initiation and continuation of the sporophytic life of the plant. As seen in Chapter 13, our current understanding of embryogenesis owes much to extensive descriptive accounts gained from light and electron microscopic studies and experimental work relating to the culture of embryos. Based on these accounts, it is convenient to consider embryogenesis in terms of a series of developmental processes aimed at creating a recognizable morphological structure. Most important among these processes are the establishment of the precise spatial organization of cells derived from the first few rounds of division of the zygote, the differentiation of cells of common origin to create diversity in the resulting embryo, and biochemical preparations for embryo maturation, desiccation, and dormancy. As part of the mature seed, the embryo unleashes another developmental program to initiate germination; however, since germination-related events are not considered in this book, the topic will not be a part of the discussion here. The developmental episodes during embryogenesis have little in common, yet there is a common background because they are at the root of many multidimensional processes involving different levels of interactions. In arbitrary terms we can conceptualize embryo development by stating that the critical aspects of morphogenesis and differentiation are completed relatively early in the time course of embryogenesis, whereas changes connected with maturation are relegated to later periods. The primary objective of this chapter is to cast the major events of embryogenesis within a genetic and molecular perspective and to explain them in
Genetic and Molecular Analysis of Embryogenesis
terms of changes in gene expression; an overview of the research efforts that reflect on the role of genomic information during maturation, desiccation, and dormancy of the embryo is also the subject matter of this chapter. Some of the questions that will be considered here are, How is the basic body plan of the embryo generated and what are the mechanisms of cell specification that occur during the initial stages of embryo development? How do the different parts of a developing seed interact in the formation of a mature embryo? What are the genes and processes that regulate embryogenesis? What are the molecular changes that direct polarity and organ differentiation in the embryo? How does the embryo acquire desiccation tolerance? It must be admitted at the outset that complete answers to these questions and to others of equal significance are not presently at hand, but investigators have begun to tackle such questions. Moreover, considering the fact that on a conservative estimate at least 20,000 diverse genes are believed to be active during embryogenesis, the problem of sorting through them to identify genes active at landmark stages should prove to be a formidable task. Although plant embryo development differs in many striking and fundamental respects from embryogenesis in animal systems, some comments about the approaches used to study the controlling mechanisms in the construction of the animal from the egg are appropriate here. The backbone of research in the early part of this century was provided by classical genetic studies using mutant genes as markers to study pattern formation, cellular determination, and the origin of diversely committed cell lineages. An alternative strategy was to isolate mutants that lead to developmental defects often reflected in anomalous patterns of morphogenesis and differentiation. The abnormalities were attributed to lesions in information flow, namely, the flow from genes to proteins. Blossoming of the field of developmental genetics led to the discovery of lethal mutants, which die after completing parts of the embryogenic program. Lethal mutants were of particular interest to embryologists because the availability of organisms whose development was arrested at relatively early stages of embryogenesis made it possible to identify genes that perform essential functions during this critical period. The advent of molecular biology has led to investigations on the expression of specific genes during embryogenesis by a multifaceted approach involving nucleic acid hybridization, construction of stagespecific cDNA libraries, analysis of the nucleotide sequence of developmentally regulated genes, identification of their protein products, and transforma-
395
tion of eggs and embryos with cloned genes and antisense RNAs. Following the lead of animal developmental biology, genetic analyses, aided mostly by mutational dissection and combined with molecular characterization at different levels of complexity, have been increasingly applied to study the fundamental genomic strategies utilized by developing angiosperm embryos. Indeed, few organs of the angiosperm plant body have recently benefited as much as embryos from the integrated genetic and molecular approach of investigators. However, identification and characterization of genes or gene products associated with embryogenesis have barely begun. Useful complements to the contents of this chapter are the reviews by Dure (1985), Meinke (1986, 1991a, b, c), Sheridan and Clark (1987), Goldberg, Barker and Perez-Grau (1989), Reinbothe, Tewes and Reinbothe (1992a), Lindsey and Topping (1993), de Jong, Schmidt and de Vries (1993), Goldberg, de Paiva and Yadegari (1994), Jurgens, Torres Ruiz and Berleth (1994), and Jurgens (1995), from which this chapter has drawn heavily to achieve a broad coverage and to present the newer developments in their proper historical perspective. 1 . GENETIC CONTROL OF EMBRYOGENESIS
A pervading role for genes in embryo development in angiosperms has been inferred in the past from analyses of embryos following wide hybridizations between plants, from analyses of the administration of ionizing radiations to developing ovules, and from investigations of spontaneous mutants with altered patterns of embryo development. Although studies described in minute detail the morphological and cytological abnormalities in developing embryos, they did not attempt to analyze gene action at the molecular level in arresting embryo growth. Moreover, many cases of cytological damage were not supported by breeding experiments designed to help understand the control of the aberrant morphology at the genetic level. Despite these limitations, results have provided important clues as to how the genotype of participating gametes determines the growth of embryos and how perturbation of the genome of certain cells affects the replicative activity and the final form of embryos. (i) Embryo Abortion in Wide Hybrids
One of the key problems in present-day horticultural and breeding practices is that crosses between distantly related species and genera of
396
plants (wide hybridizations) have not yielded many agriculturally beneficial hybrids. From a genetic point of view, this is primarily due to the fact that in attempts to transfer beneficial alien genes across interspecific and intergeneric barriers, many deleterious genes that are difficult to eliminate also find their way into the hybrid genome. The resulting disturbance in the equilibrium between growth of the maternal tissues, embryo, and endosperm leads to embryo lethality and seed collapse. In an extensive review of embryo-endosperm relations in plants, Brink and Cooper (1947b) listed six types of genetic disturbances that cause embryo abortion in hybrids. These are (a) enforced self-fertilization in a normally cross-fertilizing species; (b) crosses involving parents of the same ploidy level but belonging to different species and genera; (c) crosses between parents of the same species but of different ploidy levels; (d) crosses between parents of different species and ploidy levels; (e) unbalanced chromosome conditions, especially aneuploidy in the endosperm; and (f) maternal genotypes causing embryo arrest irrespective of the source of the male gamete. For effective hybridization, union of the egg and sperm from members of the same species with identical chromosome numbers appears a basic necessity, and even a slight asymmetry at the species level or chromosomal level can cause loss of embryos. Numerous examples of ovule development in wide hybrids have been described in previous publications (Raghavan 1976b, 1977b) and so only representative cases will be considered here. What are the kinds of disturbances that occur during wide hybridization and how do they modify the normal development of the ovule and result in embryo abortion? Of interest is the fact that the deleterious genes affect not only the growth of the embryo but also that of the endosperm. Although accounts of the ontogeny of hybrid ovules differ in details, there are some close parallels in the patterns of embryogenesis, and virtually all investigators seem to agree in identifying an early stage when developmental anomalies in the embryo begin to surface. This was first shown in reciprocal crosses between Oenothera biennis X O. muricata and O. biennis X O. lamarckiana, in which the initial
growth of embryos was not appreciably affected; however, the growth slowed down later, and eventually the embryo did not advance beyond a few cells. Development progressed further in some afflicted embryos, but survival was generally erratic and ovules enclosing underdeveloped embryos matured into the familiar, shrunken, aborted seeds (Renner 1914). Quantitative, time-
Zygotic Embryogenesis
course analysis of the growth of embryos and descriptive accounts of cellular changes associated with embryo abortion have further characterized the effects of genomic instability in hybrids. Seed development in a cross between Trifolium ambiguum and T. repens exemplifies the time-course changes associated with embryo growth in normal and hybrid plants. In selfed T. ambiguum, the zygote embarks on the embryogenic pathway soon after fertilization and proceeds through the globular, heart-shaped, torpedo-shaped, and mature stages of embryogenesis with precise regularity. Early growth of the hybrid embryo almost betrays its developmental fate, since it outgrows the selfed embryo during the first two days after pollination in the number of cells formed. Growth of the embryo subsequently slows down and, in the hybrid seeds, the embryo produces at best a tightly knit mass of about 1,000 cells and does not differentiate beyond the heart-shaped stage (Williams and White 1976). Hybrid embryos generated from T. semipilosum X T. repens cross maintain the same rate of growth as selfed T. semipilosum embryos up to four days after pollination, but their further growth bogs down by the globular stage (White and Williams 1976). Although embryos from T. repens X T. medium hybrid exhibit a regular pattern of growth up to four or five days after pollination, mitotic activity is slower in the hybrid embryos than in embryos of both parental species (Kazimierska 1978). The view that the action of the deleterious genes is initiated in the hybrid at the time of fertilization or shortly thereafter seems to be supported by this observation. In crosses involving different species of Phaseolus, a gradation is found in the stages at which deleterious genes become active and block embryo growth. In P. vulgaris X P. lunatus and the reciprocal cross, embryos cease to grow at the pre-heart-shaped and four-celled stages, respectively (Mok, Mok and Rabakoarihanta 1978; Rabakoarihanta, Mok and Mok 1979). Embryos of hybrids from P. vulgaris X P. acutifolius and P. coccineus X P. vulgaris crosses begin development in the normal way and reach the adult form, but gene action prevents embryo maturation (Mok, Mok and Rabakoarihanta 1978; Shii et al 1982). A further extreme stage of hybrid embryo development is seen in P. vulgaris X P. coccineus cross, in which embryos escape the action of deleterious genes and attain maturity (Shii et al 1982). In wide hybridization studies using various lines of Hordeum vulgare, Secale cereale, and Triticum aestivum as female par-
ents and different or closely related genera of Poaceae as pollen donors, embryos succumb to the
Genetic and Molecular Analysis of Embryogenesis
action of alien genes mostly at the globular stage (Zenkteler and Nitzsche 1984). These observations suggest that embryo abortion in wide hybrids is clearly linked in some way to postfertilization events in the embryo sac. Among the questions of general importance concerning gene action is, what are the cellular changes in the hybrid embryos that lead to their demise? In a comparative ultrastructural study of embryo development in selfed Hibiscus costatus and H. aculeatus and H. costatus X H.furcellatus hybrids,
Ashley (1972) found that the embryogenic development of the selfed parent begins with a pronounced shrinkage of the zygote and a marked segregation of organelles toward the chalazal end. In contrast, the zygote of the hybrid hardly changes physically or cytologically. However, it is with regard to the subsequent fate of the organelles that the hybrid embryo presents in a microcosm a good example of the damage inflicted by deleterious genes. Whereas the cells of the selfed embryo are endowed with a generous supply of organelles, cells of hybrid embryos are characterized by a paucity of organelles and extensive vacuolation, so that cells of the two types of embryos do not bear even the most tenuous resemblance to each other. Early degenerative changes observed in embryos of Medicago sativa X M. scutellata hybrid are con-
fined to the suspensor, which is much more reduced than suspensors of intraspecific hybrids in the number of cells formed and in the distribution of organelles in each cell. Gene action also affects the functional capacity of the suspensor, since the characteristic wall invaginations mediating in metabolic interchange at the cell surface are absent (Sangduen, Kreitner and Sorensen 1983b). These results provide a different perspective of the development of the hybrid embryo due to cellular and physiological malfunctions. As described in Chapter 12, by and large, the most general problems relating to the genetics of wide hybrids are posed by the deterioration of the endosperm and by successful embryo rescue operations. In this sense, the alien genes do not cause lethality of hybrid embryos and their effects can be considered as transient. Unfortunately, genetic analysis of hybrids to identify the individual genes that account for embryo or endosperm abnormalities has not been possible because of the many genetic changes found in wide crosses. (ii) Radiation-induced Mutations Because ionizing radiations such as X-rays and y-rays are powerful mutagenic agents, they have
397
been used in attempts to induce chromosomal and gene mutations in embryos. This approach is based on the premise that a dose of radiation that destroys the integrity of cellular DNA will cause abnormalities in the orderly development of the embryo and that these can be related to gene action. However, this objective has hardly been realized, as investigations on the effects of ionizing radiations on embryos have yielded a bag of mixed results. A large body of research on the irradiation of seeds resulted in many chromosomal mutations and in the formation of seedlings with altered morphology of the vegetative parts. Irradiation of ovules with immature embryos led to the formation of full-term embryos with abnormal organs, whereas irradiation of ovules enclosing very young embryos resulted in embryo lethality associated with abnormal cellular changes. What was lacking in these studies was a focus on specific genetic changes in the embryos so that their relationship to gene function could be understood. Impressive radiobiological contributions to embryogenesis have come from work with Hordeum vulgare. In these studies, spikes of an inbred line were exposed to acute X- or y-irradiation and subsequent growth of embryos under normal conditions was monitored. Although the level of radiation used did not lead to lethality, the histological heterogeneity of embryos made them prime targets for radiation; indeed, the relatively homogeneous early stage proembryos were less radiosensitive than the midstage proembryos and differentiating embryos. Depending upon the developmental stage of embryos at the time of irradiation, X-rays induced profound cell and tissue abnormalities that led to aberrant growth of the scutellum, root, and shoot. These disturbances can be traced to differential cell behavior in specific organs, where certain cells are killed, others begin to divide at a low amplitude, and still others continue to divide at the same rate as before. Neither the radiosensitivity of specific embryonic organs nor the drastic nature of the cellular damage seems to be anticipated during the early period of recovery of embryos because many cell generations intervene between the actual exposure and the occurrence of microscopically detectable symptoms. A particular abnormality resulting in anomalous coleoptile formation, designated as "cleft coleoptile," was extremely stage specific and appeared only following irradiation of the spike at the midproembryo stage; irradiation before or after the critical period failed to produce the cleft. In the midproembryo stage, the scutellum and shoot are most radiosensitive, whereas in the differentiating
398
embryos, these organs become insensitive and the ontogenetically younger root moves up on the list (Eunus 1955; Mericle and Mericle 1957, 1961). These observations are in line with the view that genes for the initiation of specific organs are activated at different times during embryo ontogeny, with those for coleoptile growth being very stage specific. Results similar to those obtained with H. vulgare have been reported for embryo abortion and developmental abnormalities in other plants following irradiation. In Arabidopsis, by timing the stage at which flowers or ovules are X-irradiated, it is possible to control organ formation in embryos at any developmental period between fertilization of the egg and embryonic tissue differentiation. Embryo abortion occurs when flowers harboring ovules with egg, zygote, or two-celled proembryo are irradiated. The transition of the young embryo from radial to bilateral symmetry marks a sharp turning point in embryogenesis as this is the most radiosensitive phase. Among the abnormalities resulting from irradiation at this period are changes in the number and orientation of cotyledons, which lead to syncotyly, tricotyly, anisocotyly, tetracotyly, and the formation of multiple embryos. Surprisingly, full-grown embryos escape the debilitating effects of a dose of radiation that is effective in embryos at earlier stages (Reinholz 1959). Genetic alterations thus cause the loss of growth control by cells only at a vulnerable period, and the result is abnormal morphogenesis in the developing embryos.
Zygotic Embryogenesis
establish new centers of mitosis as a prelude to forming adventive embryos. The requirement for a gene product for continued growth of embryos that was suggested by the irradiation experiments has been confirmed in recessive embryo-lethal mutants of Arabidopsis obtained by EMS mutagenesis and TDNA insertional mutagenesis following transformation with Agrobacterium tumefaciens. In a few
mutants, arrest of embryo growth is followed by abnormal divisions of the suspensor, resulting in a multi-tiered structure with as many as 150 cells in place of a normal suspensor of 6-8 cells (Marsden and Meinke 1985; Yadegari et al 1994). When the development of a mutant embryo is followed in concert with that of the suspensor, morphogenetic defects in the embryo appear to precede visible defects in the suspensor. This observation is consistent with the view that disruption of the growth of the embryo can cause anarchy in the suspensor, leading to uncontrolled growth. In these mutants, designated as suspensor (sus), defects are noted in the development of the embryo protoderm (SMS-2 and s«s-3) and in the division of cells of the inner tier of the globular embryo (sus-1, sus-2, and sws-3) (Schwartz, Yeung and Meinke 1994). Another embryo-defective mutant is twin (twn), which produces a high percentage of twin seedlings (Figure 14.1); here, secondary embryos capable of surviving seed desiccation and germinating into seedlings originate by continued division of cells of the suspensor (Vernon and Meinke 1994). Two alternative predictions made by genetic models are that SUS genes are required for normal embryo development Indirect evidence in support of the view that a and suspensor identity and that these genes are gene product is required for continued growth of involved in the production of an undefined signal the organogenetic part of the embryo but not of the for inducing the same morphogenetic changes. In suspensor has been obtained from experiments any case, it is clear that continued growth of the susinvolving exposure of immature ovules to ionizing pensor is inhibited by the organogenetic part of the radiations. In general, the suspensor cells of irradi- embryo and that when the inhibitory effect is ated embryos show a tendency to divide under con- removed by mutation or by treatments that arrest ditions that arrest the growth of the embryonal part embryo growth, the suspensor is able to express its itself. Induction of additional divisions in the sus- full developmental potential. In the case of the twn pensor cells is especially rapid when prefertilization mutation, such a simplistic explanation is probably ovules or ovules harboring proembryos are irradi- untenable because the mutation, besides promoting ated. Classic responses of the suspensor to irradia- embryogenic potential of the suspensor, disrupts the tion are seen in Nicotiana rustica (Devreux and normal development of the embryo in various ways. Scarascia Mugnozza 1962) and Capsella bursa-pas- In order to appreciate the mutagenic effects of toris (Devreux 1963), in which the abnormal sus- ionizing radiations on embryos, it is worthwhile to pensors contain a file of 8-17 cells rather than the remember that, in the experiments described, usual 4-7 cells; in Arabidopsis, with unusually long embryos were shielded during irradiation by the suspensors composed of several files of cells ovular tissues and endosperm. To what extent do (Gerlach-Cruse 1969; Akhundova, Grinikh and the abnormalities result from the direct action of Shevchenko 1978); and in Eranthis hiemalis radiation on the embryo itself and how much is (Ranunculaceae) (Haccius and Reichert 1964), in indirect? In experiments in which embryos excised which cells of the suspensor burst into growth and
399
Genetic and Molecular Analysis of Embryogenesis
Figure 14.1. Suspensor development in mutants of Arabidopsis. (a) Initiation of a supernumerary embryo from the suspensor of a mutant seed. The cell adjacent to the basal cell of the suspensor has divided to form an octant stage embryo (e2). (b) Twin embryos in a mutant seed. The embryos are not linked with the suspensor. Scale bars = 20 urn. (From Vernon and Meinke 1994; photographs supplied by Dr. D. W. Meinke.)
from unirradiated cereal grains were transplanted into irradiated endosperm, one finds that the irradiated tissue is endowed with the same mutagenic power as direct irradiation to reduce the survival period of embryos and to cause chromosomal aberrations in their cells (Avanzi et al 1967; Floris, Meletti and D'Amato 1970). Along the same lines, other experiments have shown that the use of irradiated nutrient medium or the addition of irradiated sucrose to the nutrient medium causes cytological and morphological changes reminiscent of mutagenic effects in cultured seed embryos and somatic embryos (Swaminathan, Chopra and Bhaskaran 1962; Ammirato and Steward 1969). Another approach in this research area has taken advantage of the finding that embryo growth is impaired following fertilization of a normal egg with sperm originating from irradiated pollen grains. Despite the fact that mitosis in the nucleus of the generative cell of the irradiated pollen grain is often abnormal, pollen grains germinate and produce pollen tubes, and sperm cells participate in double fertilization in the embryo sac. Cytological abnormalities are common in the cells of embryos developing from the union of one set of normal chromosomes of the egg and one set of irradiated chromosomes of the sperm (Brewbaker and Emery 1962). Irradiation of pollen grains of Lilium formosanum with a high dose of X-rays did not impair pollen germination, pollen tube growth, or production of functional sperm cells, but the treatment induced dominant lethality that results in the failure of normal zygote formation upon fertilization of an unirradiated egg. Although the endosperm
nucleus undergoes a few rounds of division and thus succumbs later than the incipient zygote, the cells generated show more cytological abnormalities than the zygote (Brown and Cave 1953, 1954; Cave and Brown 1954). Comparative studies of the effects of irradiation of the male and female gametophytes on the development of the embryo in Triticum durum showed that irradiated sperm affected embryo development more than an irradiated egg did (Donini and Hussain 1968). Mutations did not result in the immediate death of the zygote, but degeneration of the proembryo due to necrosis of cells was frequently observed after fertilization of T. durum egg with irradiated sperm. The usefulness of these lethals to the study of the developmental effects of mutations is limited because they cannot be used for progeny tests. (iii)
Spontaneous Embryo Mutants
Spontaneous embryo mutants have been identified with precise regularity in some crop plants and are thus highly suitable for studying developmental problems of embryogenesis. Moreover, compared to the lethal mutants generated by radiation treatments, most defective embryo mutants are viable when maintained as heterozygotes. Many spontaneous embryo mutations have been described, ranging from those that impair basic physiological processes of embryos, such as their lapse into quiescence or dormancy, to those that affect biochemical characteristics such as pigment or storage protein accumulation. Mutations that interfere with the quiescence of embryos induce premature embryo germination. This happens during the maturation of the seed when the embryo begins to germinate and form a seedling although the seed is still attached to the mother plant. Such mutants are known as viviparous (vp), and the phenomenon is known as vivipary. Vivipary. Due to its widespread occurrence, vivipary in maize is better understood than in other plants. Not only has the genetic control of vivipary been clearly demonstrated in maize, the system has also proved to be an attractive one for analyzing the various ramifications of gene action in regulating the maturation pathway of the embryo and endosperm in the grain. In maize, at least nine recessive genes (vp-1, vp-2, vp-5, vp-7, vp8, vp-2, w-3, y-9, and al) that can induce vivipary are known and their respective genotypes have been described in detail. Genetic and physiological studies have shown that besides causing premature ger-
400
mination, most viviparous genes interfere with other facets of grain maturation, such as ABAmediated responses, pigment synthesis, and storage protein accumulation. All except one (vp-1) of the nine viviparous genotypes are deficient in ABA; all ABA-deficient mutants except one (vp-8) share an inability to form carotene, with the result that chlorophyll synthesis is also suppressed in the seedlings. Mutation in the vp-1 locus also suppresses the accumulation of anthocyanin in the embryo and in the aleurone cells of the endosperm (Robertson 1955; Neill, Horgan and Parry 1986; McCarty and Carson 1991). Another effect of vp mutation is the failure of embryos to accumulate storage protein globulins encoded by GLB-1 and GLB-2 genes; however, globulin synthesis can be reinstated in vp-5 mutant embryos that are deficient in ABA by exogenous application of the hormone, but this treatment is ineffective in embryos of the vp-1 mutant (Rivin and Grudt 1991). In addition, use of a transposon tagging approach has led to the characterization of mutant alleles in which vp-1 function is altered in dramatic ways, such that the embryo attains a near normal state of dormancy but neither the embryo nor the endosperm develops anthocyanin (McCarty et al 1989a). It is possible that what we see here is the pleiotropic action of a single gene on germination, pigment synthesis, and the hormone level of embryos, but the nature of the metabolic blocks involved, or how blocks in the synthesis of pigments or hormones might induce vivipary, is not known.
Zygotic Embryogenesis
phenotype. Of the several physiological differences between the patterns of development of wild-type and vp-1 mutant embryos, one that may prove useful in the elucidation of the possible role of ABA in vivipary is that viviparous embryos do not desiccate at the time when wild-type embryos begin to undergo dehydration, but continue to increase in fresh weight (Wilson, Rhodes and Dickinson 1973; Neill, Horgan and Parry 1986; Neill, Horgan and Rees 1987). When the different mutants are screened for their sensitivity to ABA inhibition of germination and in vitro embryo growth, embryos of the vp-1 mutant are easily coaxed to grow in culture in the presence of ABA in the medium, indicating that the mutation in some way confers resistance to the inhibitory action of the hormone (Robichaud, Wong and Sussex 1980; Robichaud and Sussex 1986; Sturaro et al 1996). However, this does not appear to be caused by a defect in the actual mechanism of action of the hormone, as the patterns of uptake and metabolism of ABA are remarkably similar in mutant and wild-type embryos (Robichaud and Sussex 1987). As a tentative conclusion from these results, the reduced sensitivity of mutant embryos to growth inhibition by ABA may be considered to involve a stable genetic change in their signal transduction pathway concerned with ABA perception.
A different type of experiment has revealed that interfering with the ABA metabolism of wild-type maize caryopses can induce vivipary. In this experiment, developing ears are treated with fluridone, Are the effects of the VP gene mediated by an inhibitor of carotenoid biosynthesis, and the fate changes in the embryo, in the endosperm, or in of the caryopses is followed. When the treatment is both? Although most routine crosses have yielded applied between 9 and 11 days after pollination, results too ambiguous to permit the assignment of premature plumule elongation characteristic of genetic effects to either the embryo or endosperm, vivipary is found in a high percentage of the carythe use of pollen grains carrying an A-B chromo- opses. Since carotenoids are probable precursors of some interchange in crossing plants heterozygous ABA, an obvious conclusion from this work is that for a VP gene has shown that vivipary is deter- vivipary is due to a lesion in ABA synthesis (Fong, mined solely by the genotype of the embryo. This Smith and Koehler 1983). conclusion has relied on the observation that in all Some interesting insight into the processes that cases, genetically viviparous embryos germinate control premature germination and block in the prematurely whether they are surrounded by a anthocyanin pathway in the embryo has been wild-type or a mutant endosperm in the caryopsis derived from the isolation and characterization of (Robertson 1952). Anthocyanin expression in the the VP-1 gene. McCarty et al (1989b) cloned the VPaleurone, however, is independent of the embryo 1 gene by transposon tagging and showed that it is genotype and results from the coordinate control of a single-copy gene with a 2,500-nucleotide tranthe synthesis of critical enzymes in the endosperm script. RNA blot hybridizations revealed that tranby many genes, including the one that determines scripts of the gene are present in the embryo and vivipary (Dooner and Nelson 1979; Dooner 1985). endosperm tissues of the wild-type caryopses but Since vp-1 is an exceptional mutation that causes not in the caryopses of the mutant stock; consistent vivipary without altering the ABA level of with the effects of the mutation on the embryo and embryos, considerable effort has been made to endosperm, seedling tissues such as the shoot and study the biochemical and molecular basis for its root also do not express the gene. Finally, a surpris-
Genetic and Molecular Analysis of Embryogenesis
ing observation that emerged from RNA blot hybridizations using probes made to genes implicated in VT-2-regulated processes is that expression of C-l, a regulatory gene in the anthocyanin pathway, is selectively blocked in both the embryo and the endosperm of the mutant vp-1 caryopses. Reporter Plasmids:
GUS tni-oUo
i—— Em promoter
35S-Sh-GUS
GUS
jll UMVWSpremour
Effector plasmid: 35S-Sh-Vpi
.SM^I
Vpi cONA
NOS 3
CaHVKSprantour
Reporter Plasmid
Effector Plasmid
GUS Activity pmoth'ujVo'
none
4.27+0.51
35S-Sh-Vp1
587+68
Em-GUS
(137X)
none
2750+320
35S-Sh-Vp1
4170+170
35S-Sh1-GUS (1.5X)
-6 -5 log[ABA]
-4
Figure 14.2. Transactivation of EM gene by VP-1 gene in a maize protoplast assay, (a) Schematic representations of reporter and effector plasmids. Em-CUS plasmid contains 554 bp of the 5'-flanking sequence of the f/Vf gene and the nopaline synthase (NOS) gene fused to GUS gene. The 35S-Sh-GUS plasmid includes the CaMV 35S promoter, the first intron of the maize SH-1 gene, the C4Igene, and the 3'-sequence of the NOS gene fused to GUS gene. The 35S-Sh-Vp1 plasmid has the cDNA clone containing the complete protein coding sequence of VP-1 gene. Maize protoplasts were cotransformed by electroporation with
401
In an attempt to define the interaction between VP1 and C-l genes in controlling the anthocyanin pathway, Hattori et al (1992) found that overexpression of VP-1 protein in maize protoplasts of embryogenic cell origin activates a reporter gene driven by the C-l promoter. At low concentrations of ABA, VP-1 protein also strongly augments the activation of C-l; however, at high concentrations of the hormone, the protein does not stimulate reporter gene expression. Whatever may be the significance of this result in the broader role of the VP-1 gene in caryopsis development, a conclusion that can be drawn is that this gene may control the anthocyanin pathway by translational or posttranslational regulation of C-l; it is, however, a moot question whether VP-1 protein plays an essential role in the ABA-signal transduction pathway. The amino acid sequence predicted from a cDNA clone indicates that the VP-1 gene encodes a 73-kDa protein that has no detectable similarity to known proteins (McCarty et al 1991). Although this protein appears otherwise undistinguished, it is significant that its acidic domain apparently functions as a transcription factor in regulating the expression of other genes and induces strong activation in maize protoplasts of a cotransformed GUS reporter gene fused to the promoter of a presumptive target gene such as the ABA-regulated EM (for early methionine-labeled) gene from wheat embryos. Another observation pertinent to the role of ABA in the maturation program of the embryo is that addition of the hormone causes a synergistic effect on the activation of the target gene (Figure 14.2). The key implication of these results is that proteins encoded by the VP-1 gene are necessary for dormancy induction in the embryos as well as for ABA response, perhaps as transcription factors. Inhibition of precocious induction of germination-specific oc-amylase genes in the aleurone cells of the endosperm provides another setting in which the VP-1 gene might act during caryopsis development (Hoecker, Vasil and McCarty 1995). By combining both activator and repressor functions in the same protein and in cells each of the effector and reporter plasmid DNAs as shown. GUS gene activity is expressed as the mean of three readings ± standard error. Values in parentheses show times activation over basal expression, (b) Synergistic interaction of VP-1 gene with ABA. Protoplasts from the electroporation experiments were cultured in media containing different concentrations of ABA. Squares represent protoplasts electroporated with Em-CUS construct alone and triangles represent protoplasts electroporated with Em-CUS construct together with 35S-Sh-Vp1 construct. (From McCarty et al 1991; ©Cell Press.)
402 of two different genotypes, the VP-1 gene seems to integrate the control of embryogenesis and germination in an unusual way. It was mentioned earlier that the synthesis of globulins in viviparous embryos is related to the actions of the mutant gene and ABA. Northern blot analysis showed that whereas GLB-1 and GLB-2 transcripts are not detected in vp-1 mutant embryos, they are present at a slightly reduced level in embryos of certain viviparous lines deficient in ABA (for example, vp-5). Since the expression of GLB-1 and GLB-2 transcripts is enhanced in the wild-type embryos by ABA, the function of the VP1 gene product could simply be to ensure the progress and completion of an ABA-regulated process in the maturation of the embryo (Kriz, Wallace and Paiva 1989; Paiva and Kriz 1994). Synthesis and accumulation of several LEA proteins in embryos of vp mutants are modulated differently than are those of globulins. The list includes a group of 23-25-kDa LEA proteins that appear in embryos of wild-type and certain ABA-deficient mutants (vp2, vp-5, and vp-7) of maize. The proteins and their mRNAs are also precociously induced by ABA in young wild-type and mutant embryos (Pla et al 1989; Mao et al 1995). In the vp-2 mutant, the pattern of transcript expression of the gene RAB-28 (for responsive to ABA), which encodes a 27-kDa protein, is strikingly similar to that found in wild-type embryos. Although RAB-28 mRNA is not detected in the embryos of vp-1 mutant, it is inducible by low concentrations of ABA (Pla et al 1991). Another LEA protein, MLG-3, is detected in negligible amounts in vp-5 mutant embryos, but the latter accumulate the protein when they are cultured in the presence of ABA. However, culturing vp-1 mutant embryos in ABA does not result in the accumulation of MLG-3, whereas nongerminating vp-1 embryos retained on the ear or cultured in a high osmoticum can accumulate this protein (Thomann et al 1992). The most direct interpretation of these results is that MLG-3 proteins, unlike globulins, have no specific requirement for the VP-1 gene product, but other signals, such as ABA or osmotic changes, contribute to their expression in the mature embryo. On the other hand, the regulation of expression of the EM gene in vp-1 mutant embryos is rigorously dependent upon the presence of ABA in the medium and upon its perception through a mechanism mediated by the VP-1 gene, because no EM mRNA accumulates in mutant embryos of any age growing on the ear, or cultured in ABA, or subjected to an acute osmotic stress (Butler and Cuming 1993; Paiva and Kriz 1994).
Zygotic Embryogenesis
Studies on vivipary in other plants have added few new dimensions to our present understanding of this phenomenon. In mangroves such as Rhizophora mangle, in which vivipary is a natural phenomenon, embryos have a high water content and are generally insensitive to inhibition by ABA in culture (Sussex 1975). This probably facilitates transition of the embryo from a developmental mode to a germination mode while still contained within the seed. The relationship between a low ABA level in the seed and vivipary was confirmed in an ABA-deficient mutant of Arabidopsis in which the seeds were induced to germinate in the fruit in an atmosphere of high humidity (Karssen et al 1983). In other mutants in which embryos maintain a high water content, seeds often showed vivipary without any further treatment (Koornneef et al 1989). This is what one would expect of embryos growing in ovules deficient in ABA. To the extent that water stress in plants is known to cause a rise in the ABA level of cells, it is of interest to note that vivipary is associated with a high water content in the embryos. A general conclusion is that ABA might regulate the metabolism of the embryo during the transitional period between the end of embryogenic development and onset of germination by its effects on the water potential of the cells. Screening a rice genomic library with maize VP-1 gene as a probe has led to the isolation of a gene functionally homologous to the maize gene in transient expression experiments using rice protoplasts; transcripts of the gene are detected in rice embryos as early as 10 days after flowering and decrease toward maturity (Hattori, Terada and Hamasuna 1994). Isolation of genes for vivipary in rice has not hitherto been accomplished and offers great promise in broader studies. Seed Mutants Deficient in Storage Products. Growers have known for a long time that inbred lines of some cultivated crops produce seeds that are genetically defective in the expression of specific storage products in the embryo. Indeed, it was the character of the pea seeds that affects the accumulation of storage products in the cotyledons (wrinkled and green seeds) that Mendel used in the famous experiments on the inheritance pattern involving the independent assortment of more than one gene. It is appropriate that this natural collection of mutant seeds has been studied to clarify the molecular basis for the derangement in storage product accumulation. In pea, the production of wrinkled seeds is governed by two genetic loci, r (for rugosus) and rb,
Genetic and Molecular Analysis of Embryogenesis
403
and it has often been argued that these genes sepa- the embryos of rr seeds is the cloning of the r locus. rately affect starch metabolism of the embryo in a Bhattacharyya et al (1990) used an antibody to manner that results in a reduced starch content the isoform of the starch-branching enzyme to with a concomitant reduction in starch granule size clone a cDNA made to poly(A)-RNA of embryos of and an increase in the proportion of the amylose RR seeds and showed that transcripts encoding content of the starch; alleles at the rb locus also pro- the enzyme are about 10-fold less abundant in foundly affect the growth of the testa. Comparative the mutant embryos than in the normal ones. analyses of the effects of the r locus on the growth Furthermore, the absence of starch-branching and development of embryos of near-isogenic enzyme in the mutant embryos has been attributed round (RR) and wrinkled (rr) lines of peas have to an insertion in the gene encoding the enzyme revealed several physiological changes leading to a and, because of the presence of inverted repeats at derangement in starch metabolism. One invariable its termini, the inserted sequence is considered to change is the overall high fresh weight of embryos be transposonlike. of the mutant line compared to embryos homozyLectins are sugar-binding proteins and glycogous for the R-allele (Hedley et al 1986, 1994; proteins of wide occurrence in plants. Genetic Lloyd, Wang and Hedley 1996). Throughout most defects that interfere with embryo lectin contents of the developmental period studied, cells of are common among cultivated varieties of soymutant embryos also showed a notable effect of the beans. Molecular studies on a lectinless line of soylesion in possessing a higher water content and in bean have attributed the phenotypic condition to a having a higher osmotic pressure than normal virtual absence of lectin transcripts in the genome. embryos (Wang et al 1987b). A genetic effect on cell The transcription lesion in the lectinless phenotype size appeared somewhat late in embryogenesis and is apparently due to a silencing of the lectin gene was reflected in the embryos of the wrinkled line caused by the transposonlike insertion of a 3.4-kb having larger cells than those of the normal round DNA segment (Vodkin 1981; Goldberg, Hoschek line (Ambrose et al 1987). Other side effects of the and Vodkin 1983; Vodkin, Rhodes and Goldberg mutant gene are seen in the lipid content and in the 1983). It is of particular interest that seeds of transcomposition of the storage proteins of the embryos; genic tobacco plants harboring a chimeric lectin generally, mature rr embryos contain more lipid gene containing the 5'-half of the mutant gene and and less of the storage protein legumin than RR its promoter region and the 3'-half of the wild-type embryos (Davies 1980; Coxon and Davies 1982). gene synthesize both lectin mRNA and protein. Some of the parameters of wrinkled seeds, such as This shows that the control region of the mutant starch content and composition and lipid content, gene is transcriptionally competent but that insercould be reproduced in chemically induced tion of the transposonlike element blocks gene mutants of peas (T. L. Wang et al 1990). transcription by silencing the promoter (Okamuro An important insight into the basis for the dif- and Goldberg 1992). How the promoter functions ference in the osmotic potential between RR and rr are impaired by the transposon element remains embryos was gained when it was found that wrin- unknown and should prove to be an exciting area kled seeds have higher levels of free sucrose than of future research. normal seeds (Stickland and Wilson 1983). Since Mutation in a naturally occurring variety of soythe increased sucrose content will lead to a high bean that lacks the a'-subunit of the 7S seed storage osmotic pressure in the cells, questions concerning protein, p-conglycinin, has been found to be caused this phenomenon center on the defect in the con- by a deletion that extends through most of the codversion of sucrose to starch. At the biochemical ing sequences of the gene (Ladin, Doyle and level, the r mutation is believed to lead to a reduc- Beachy 1984). Another soybean seed protein mutation in the activity of the starch-branching enzyme tion includes an inversion that separates the 5'- and (1,4-oc-D-glucan, l,4-a-D-glucan-6-glycosyltrans- 3'-regions of an aberrant glycinin gene, GY-3 (Cho ferase), which results in the complete absence of et al 1989). Because the synthesis of the respective one isoform of this enzyme (Matters and Boyer transcripts predicted in the wild-type embryos is 1982; Smith 1988). This may cause a block in the inhibited in the mutants by lesions in the correpathway of starch synthesis in embryos of wrin- sponding genes, it follows that the mutations affect kled peas and may lead to a consequent accumula- gene action at the transcriptional level. tion of sucrose. A different picture emerges in considering the Among other attempts made to relate metabolic molecular basis of a null allele of a glycinin gene consequences of gene action to starch synthesis in (GY-4) and a mutation in a KTI gene in soybean
Zygotic Embryogenesis
404
KTi +
KIT
A GCTA
AG C T A
-+510
-+480
KTi3
+
KTi3"
+ 478 j | J +498 CTT GAG AGA GTT TCT GAT GAT Leu Glu Arg Val Ser Asp Asp aa!19 aal25
CTT TAG AG—T TTC TGA TGA TGA • • Leu Phe • Ser
b Figure 14.3. Nucleotide and amino acid sequence of Kunitz trypsin inhibitor genes in wild-type (KTi+) and mutant (KTi~) embryos of soybean, (a) DNA sequence ladders represent nucleotides +460 to +520 of the two genes. Nucleotides +481 to +490 of the wild type and the corresponding mutant sequences are shown to the left. Squares show the nucleotides that are conserved between the two genes; asterisks indicate nucleotides that are mutated in the KTi" gene, (b) Nucleotide and amino acid sequence comparisons of KTI genes of wild-type and mutant embryos. Only nucleotides +478 to +498 and translated amino acids 119 to 125 are shown. Arrows refer to the nucleotides mutated in the KTi" gene (shown as asterisks in [a]). The asterisks designate translational stop codons that result from the mutations. (From Jofuku, Schipper and Goldberg 1989. Plant Cell 1:427-435. © American Society of Plant Physiologists. Print supplied by Dr. R. B. Goldberg.)
embryos. The null phenotype is attributed to a point mutation that leads to the elimination of the translation start codon due to a single base change. Although this does not interfere with transcription, mRNA is not effectively retained on the polysomes like the functional mRNA (Scallon, Dickinson and Nielsen 1987). In the mutant defective in the gene encoding KTI, a frame-shift mutation due to alter-
ation of three nucleotides causes drastically reduced accumulation of KTI mRNA and KTI in the embryos (Figure 14.3); it has also been found that the KTI gene is transcribed at the same relative rate in wild-type and mutant embryos (Jofuku, Schipper and Goldberg 1989). Evidently, some posttranscriptional modifications impose restraints on the stability of mRNAs and reduce the prevalence of mRNAs or of the protein products in the mutant embryos. The lectin-nonproducing mutant seeds of soybean have a counterpart in certain genetic mutants of Phaseolus vulgaris. In cultivars like Pinto, mutations of the genes encoding the lectin, PHA, have the effect of producing embryos that are deficient in lectins. At the biochemical level, the effects of the mutation have been interpreted as the consequence of a reduced synthesis of PHA and an equally low level of PHA gene transcription (Staswick and Chrispeels 1984; Horowitz 1985; Chappel and Chrispeels 1986). A partial explanation for the molecular basis of the mutation was provided by the cloning and sequencing of the two PHA genes of Pinto called PDLEC-1 and PDLEC-2. The latter follows very closely the sequence of the genes of the wild type and of PDLEC-1 up to -240 bp relative to the transcription site. Compared to other genes, farther upstream, PDLEC-2 suffers a 100-bp deletion that contains a 60-bp tandem repeat. Since the deleted segment might function as an enhancerlike element, this deletion could account for the low level of expression of PDLEC-2 (Voelker, Staswick and Chrispeels 1986). In support of this hypothesis, it has been found that embryos of transgenic tobacco seeds harboring a PDLEC-2 allele accumulate about 50 times less PHA than embryos of plants transformed with a gene from the normal cultivar (Voelker, Strum and Chrispeels 1987). But this clearly is not the whole explanation for the effect of the mutation, given the lack of knowledge about the causes for the low expression of PDLEC-1.
2. GENE EXPRESSION PATTERNS DURING EMBRYOGENESIS
Included among the genes expressed during embryogenesis are those that encode enzymes for housekeeping activities and mobilization of food reserves, structural proteins of the cytoskeletons and membranes, and storage proteins and embryospecific genes that regulate specific aspects of embryo development. Although we are a long way from characterizing these genes and their products and analyzing the regulatory pathways of gene
Genetic and Molecular Analysis of Embryogenesis
action, current experimental approaches have led to new insights into the molecular processes controlling gene expression pattern during embryogenesis. (i) Diversity and Changes of mRNA Population during Embryogenesis
The modern global view of quantitative changes in the gene expression program during embryogenesis was developed by Goldberg et al (1981b), Galau and Dure (1981), and Morton et al (1983) from reassociation kinetics of nucleic acids of embryos of soybean, cotton, and pea, respectively. Because of the small size of the early stage embryos, which precludes their isolation in ample quantities for preparation of nucleic acids, these studies were undertaken with embryos in advanced stages of development; yet the power of this approach to the study of gene regulation is unmistakably demonstrated in these investigations. As an intermediate between the gene and the protein it encodes, mRNA populations in embryos at different stages of development should provide an overview, down to the details of changing complexity, abundance distribution, and diversity of the genes active during embryogenesis. Hybridization experiments using an excess of mRNA from cotyledon stage (30 days after flowering) and midmaturation stage (75 days after flowering) embryos of soybean with cDNA prepared from corresponding mRNAs have shown stagespecific changes in mRNA complexity, ranging from 15,000 average-sized mRNAs in the cotyledon stage embryos to more than 30,000 in midmaturation stage embryos. Considering the simple morphology of the cotyledon stage embryos, the degree of specification at the level of mRNAs is apparently enormous. In terms of the number of molecules of mRNA present per cell per sequence, it is possible to delineate superabundant (>19,000 molecules), moderately prevalent (550-800 molecules), and rare (3-17 molecules) classes of mRNAs in the soybean embryo cells. Attesting to the diversity in the distribution of the different classes of mRNA is the fact that in the cotyledon stage embryos, mRNAs belong to the moderately prevalent and rare classes, whereas in the later-stage embryos they are predominantly of the superabundant class, with small amounts of the moderately prevalent and rare classes. Interestingly, the superabundant class, which accounts for nearly 50% of the mRNA mass of midmaturation stage embryos, contains only 6-10 diverse mRNAs. Despite the fact that morphological complexity of the embryo increases with
405 development, a surprising finding is that cotyledon stage embryos and midmaturation stage embryo axes and cotyledons contain approximately the same total number (14,000-18,000) of diverse mRNAs. This signifies that in terms of the absolute number of structural genes expressed, there are no detectable changes in gene expression during morphological differentiation of the embryo and within morphologically distinct parts of the embryo (Goldberg et al 1981b). Another approach to the analysis of mRNA changes during soybean embryo development was by the hybridization of kinetic fractions of midmaturation stage embryo-abundant and rare cDNA (selected by repeated cycles of hybridization with homologous mRNA, followed by hydroxyapatite fractionation) to an excess of cotyledon stage and midmaturation stage embryo mRNA. Comparison of the mRNA sequences present in embryos of the two developmental stages showed that almost the entire sequence comprising the mass and diversity of mRNAs of the older embryos is contained in the young embryos. Although this comparison is sensitive only to changes in at least hundreds of rare transcripts, the logical conclusion is that mRNAs of embryos of different ages tend to be qualitatively similar. The interesting possibility from these results is that in soybean the entire set of structural genes that encode proteins of embryogenesis is transcribed at a very early stage, regardless of whether the genes are translated or not. Finally, an intriguing feature of the fate of embryogenic mRNA sequences is that most of them persist throughout development and are stored in the cells of the mature seed embryo (Goldberg et al 1981b). Quantitative changes in the gene expression program during embryogenesis in cotton have been followed by RNA-excess DNA-RNA hybridization studies that have identified mRNA sets from twodimensional SDS-PAGE analyses of extant proteins, proteins synthesized in vivo and in vitro, and the changing mRNA populations in embryos of different ages. As in soybean, DNA-RNA hybridization with mRNA populations from young (50 mg fresh weight), maturation phase (110 mg fresh weight), mature (from seeds), and germinated embryos implies that 15,000-20,000 genes are expressed at the mRNA level during embryogenesis. However, only a few hundred genes encode mRNAs that are abundant enough to be detected in profiles of twodimensional gels of in vitro translation products. The analysis of extant, in vivo and in vitro synthesized proteins allowed the changes in the abundant mRNAs and abundant proteins to be followed with
406
Zygotic Embryogenesis
EMBRYOGENESIS EARLY
LATE
MID
matuf,
EARLY GERMINATION
seed
SB
Figure 14.4. A diagrammatic representation of the groups of abundant mRNAs demonstrated in developing and germinating cotton embryos. (From Dure 1985. Oxford
Surv. Plant Mol. Cell Biol. 2:179-197, by permission of Oxford University Press.)
a focus on two major questions. Does the population of extant proteins in the young embryo include some proteins whose synthesis is not required for continued differentiation of the embryo? Does a mature embryo synthesize proteins that are not needed for the survival of the young seedling? When the soluble protein fractions from cotyledons of embryos of different ages are analyzed, substantial modulations are found to occur in several sets of proteins whose synthesis is developmentally regulated; gene sets expressed in embryos of the same ages are deduced from in vitro translation analysis of their mRNAs. These data are diagrammatically presented in Figure 14.4 as groups of abundant mRNAs expressed during embryogenesis. mRNAs of groups 1 and 2 are present throughout embryogenesis; whereas group 1 mRNAs are expressed during germination as well, those of group 2 are turned off after embryogenesis is completed. Compared to group 2 mRNAs, which encode for only six proteins, group 1 mRNAs encode an impressively large set of proteins, including actin and tubulin. Another point of comparison is between the products of cell-free translation of mRNAs isolated from embryos of different ages and proteins synthesized in vivo by embryos of the same age; this comparison has shown that many translatable mRNAs of group 1 are detected only when their corresponding proteins are synthesized in vivo, indicating that gene activity with regard to the synthesis of these proteins is regulated at the transcriptional level. However, proteins
encoded by group 2 mRNAs are synthesized in vitro by embryos of all ages. Expression of group 3 mRNAs occurs coincident with the burst of mitotic activity during early embryogenesis but, as cell divisions cease, their genes are turned off in the maturation phase and in seed embryos. Proteins encoded by these transcripts accumulate extensively in embryos of all developmental stages, although they are synthesized only in young and maturation stage embryos. The most extensively changing mRNAs are those that code for storage proteins (group 4). These mRNAs are found in high levels in maturation stage embryos but vanish completely in mature embryos. Two other groups of mRNAs (groups 5 and 6), which become abundant during late embryogenesis and accumulate in seed embryos, differ in one important respect. Group 5 mRNAs, which consist of two subsets and their corresponding proteins, disappear from the abundant class during the early hours of germination but can be reinstated by incubating excised immature embryos in ABA. This is just what one would expect of a group of transcripts and their protein products whose synthesis in the embryos is regulated by endogenous ABA. The other group of mRNAs (group 6) and their proteins persist during normal germination of seed embryos, as well as during precocious germination of immature embryos. There is also one final group of mRNAs (group 7), which is not expressed during embryogenesis but appears during germination (Dure,
Genetic and Molecular Analysis of Embryogenesis
Greenway and Galau 1981; Galau and Dure 1981; Chlan and Dure 1983). Comparisons of reassociation kinetics between cDNAs from cotton embryos of all three developmental stages and an excess of homologous mRNAs have identified the same groups of mRNAs whose abundant proteins are synthesized in vivo and in vitro, as well as several less abundant mRNAs. These mRNAs belong to abundance classes that show more than a twofold increase in their sequences in the seed embryos. Another type of analysis has involved hybridization of each cDNA clone to mRNAs isolated from other embryo stages. Such heterologous reassociations have shown that embryos of all ages contain most of the mRNA sequences, although sequences prevalent at one stage may be present at a reduced level in other stages. Studies of this type have also focused on some major differences in the concentrations of the more abundant mRNAs of embryos of different stages of development, the most dramatic being a stark reduction in the presumptive storage protein mRNAs in the mature embryo (Galau and Dure 1981). The hybridization kinetics of mRNAs from cotyledons of developing pea embryos to cDNAs show much that is common with, and much that is different from, those of soybean and cotton. The total complexity of mRNA sequences of different abundance classes at the early stage of cotyledon development in pea (ca. 20,000) compares favorably with that of cotton (ca. 25,000) and soybean (ca. 32,000) embryos of slightly different ages. As noted in soybean, there is an increase in the very abundant class of mRNAs during embryo development when storage protein synthesis is initiated and sustained. However, compared to soybean and cotton, in pea the rare class of mRNAs completely disappears as embryogenesis proceeds, resulting in a 10-fold decrease in the number of mRNA sequences in embryos at the late stage of cotyledon development (Morton et al 1983). The obvious adaptive value of this strategy is that genes for the production of enzymes and structural proteins are no longer required and are switched off as the embryo begins to accumulate storage proteins. Since this reduces the number of active genes in the cell to about 200, some questions have been raised about the survival capacity of such cells (Dure 1985). (ii) Gene Activity during Fertilization and Early Zygote Divisions
There have been very few investigations on the molecular biology of fertilization in gymnosperms
407
and angiosperms, and the mechanism of gene activation during the early division phase of the zygote has completely eluded us. A major impediment to these studies is that events of fertilization and partitioning of the zygote take place within the confines of the archegonium in gymnosperms and the embryo sac in angiosperms. Reference was made in Chapter 13 to studies that seemed to indicate that fertilization of the egg and subsequent divisions of the zygote are accompanied by changes in the RNA content of the cells. Unfortunately, these investigations did not address issues such as the role of the maternal genome in fertilization and early embryo development. The concept of maternal message, which has its roots in animal embryology, assumes that the mature egg contains large stores of mRNAs that become available after fertilization to code for the first proteins of the zygote. There is no hard evidence for the existence of a legacy of template information in the gymnosperm or angiosperm egg, and the source of the genetic information parceled out for the division of the zygote is unknown. One study has shown that when rice and barley florets are cultured in a medium containing 3H-uridine, no autoradiographically detectable incorporation of the label occurs in developing embryos of less than 100 cells. At the same time, 3H-leucine is readily incorporated into embryos of all ages. Because the eggs are fertilized after the florets are placed in culture, assuming that the cells of the globular embryo do not possess a large uridine pool, it has been reasoned that protein synthesis in the absence of concurrent RNA synthesis probably involves the use of stored maternal mRNA (Nagato 1979). Based on in vitro translation of poly(A)-RNA isolated from ovules of Pinus strobus before and after fertilization, evidence was obtained for the transcription of a prominent mRNA at the time of fertilization and at the beginning of embryogenesis. A cDNA clone isolated from ovules containing globular embryos was found to encode a protein homologous to the US storage protein globulin. Since transcripts of this cDNA clone are found to accumulate in the megagametophytes of fertilized, but not of unfertilized, ovules, the trigger of the fertilization event in causing transcript abundance is obvious. Although transcripts are not detected in the developing embryos, this does not preclude a possible role for fertilization-induced signals from the embryo in transcript accumulation in the megagametophyte (Whitmore and Kriebel 1987; Baker et al 1996). Analysis of genes expressed in the embryo, with the exclusion of the female gametophyte and the sporophytic tissues that constitute the bulk of the ovule, may perhaps provide a clear picture of the embryo gene activation program during fertilization.
408
Based largely on the end result, namely, activation of the egg in the absence of fertilization (parthenogenesis), many claims have been made for an independent role for the maternal genome in directing early embryo development. These claims include the spontaneous division of the egg in the absence of a stimulus of fusion with the sperm, as in apomicts (Nogler 1984; Ramachandran and Raghavan 1992a); use of physical and chemical agents to inactivate the sperm without hindering the growth and penetration of the pollen tube and thus to stimulate the division of the egg (Lacadena 1974); and the purported origin of embryos from the eggs of cultured unfertilized ovules or ovaries (Yang and Zhou 1982; Yan, Yang and Jensen 1989; Ferrant and Bouharmont 1994). Haploid plants that arise by chromosome elimination in interspecific crosses between Hordeum vulgare and H. bulbosum
also offer a basis for the assumption that maternal templates alone can support embryo development, albeit under somewhat abnormal conditions (Kasha and Kao 1970). Since the male chromosomes disappear even before they are integrated into the genome of the egg, the haploid embryo essentially develops by transcribing the stored maternal information. Although the examples considered here do not constitute a rigorous demonstration of the presence of stored mRNAs, there is probably some truth in the assertion that the angiosperm egg cell, whether it is fertilized or not, has the potential to initiate the developmental program of the embryo using maternal transcripts.
(iii) RNA Metabolism in Developing Embryos
Zygotic Embryogenesis
Rosie 1975), Oryza sativa (Zhu, Shen and Tang 1980a), Triticum vulgaris (Zhu, Shen and Tang 1980b), and Abelmoschus esculentus (Malvaceae) (Bhalla, Singh and Malik 1980b), among others, are examples that illustrate a rising level of RNA through most periods of growth of the embryos. Investigations on P. vulgaris implicate the high water content of the embryo as the stimulus for increased RNA synthesis; as the water content of the embryo begins to decrease, the rate of RNA synthesis falls to a low value and remains at this level through the period of desiccation and dormancy (Walbot 1971, 1972). Other studies have shown that the RNA metabolism of embryos of different ages and that of specific parts of the same embryo are profoundly interrelated. When just the organogenetic part of embryos of different ages of P. coccineus, consisting of rapidly dividing cells, is analyzed, RNA content per cell and the rate of RNA synthesis per cell are highest in the early heart-shaped embryo and decline thereafter. In the numerically stabilized cell population of the suspensor, these parameters are low during early embryogenesis. The decrease in the RNA synthetic activity of the embryo cells probably reflects the inability of the nucleolus to synthesize new RNA to keep up with the rapid tempo of mitosis (Walbot et al 1972; Sussex et al 1973). Although the rate of RNA synthesis declines to near zero during desiccation of the embryo, protein accumulation is unabated; the possibility exists that the proteins are encoded by mRNAs synthesized at an earlier period in embryogenesis and stored for later use (Walbot 1971).
Investigations on the RNA metabolism of cotyledons of developing embryos have generally Several early investigations have shown that supported the results obtained with whole progressive embryo development signals consider- embryos. In the cotyledons of different species able changes in the temporal and spatial patterns of there is close agreement in the time of initiation of accumulation and synthesis of bulk RNA. In the RNA synthesis, which generally follows DNA synwake of the isolation and characterization of thesis. RNA accumulates in cotyledons of near-norembryo-specific genes, these studies can be consid- mal water content in parallel with the period of ered to represent at best a gross approximation of rapid cell division activity; this is followed by a cesgene activity in developing embryos. For this rea- sation or decrease of accumulation during desiccason, only a small portion of this work will be dis- tion. The major portion, if not all, of the RNA synthesized appears to be rRNA (Galitz and cussed here. One manifestation of RNA metabolism during Howell 1965; Millerd and Whitfeld 1973; Poulson embryogenesis is the continued accumulation of and Beevers 1973; Scharpe and van Parijs 1973; D. RNA, so that by the completion of embryogenesis, L. Smith 1973; Walbot 1973; Millerd and Spencer the amount of RNA in the mature embryo will have 1974). Considered together, the RNA metabolism of increased to many times the initial value of the developing whole embryos and isolated cotyleearly stage embryo. Viciafaba (Wheeler and Boulter dons invariably indicates a changing biochemical 1967; Manteuffel et al 1976), Phaseolus vulgaris pattern with time. (Walbot 1971), Hordeum distichum (Duffus and The rise and decline in RNA accumulation and
Genetic and Molecular Analysis of Embryogenesis
synthesis during embryogenesis reflect, to some extent, the availability of sufficient mRNA. Poulson and Beevers (1973) found that although the RNA content of pea cotyledons remains fairly constant during the seed maturation period, the capacity of the cotyledons for RNA synthesis declines considerably during this period. The abruptness of this change correlates well with the observation that ribosomal preparations made from cotyledons of mature embryos show a decreasing abundance of polysomes and are practically inefficient in supporting cell-free translation. Since the full cell-free translation potential of ribosomes is realized by the addition of an artificial messenger, experimental results are consistent with a declining mRNA component in the cotyledons with age (Beevers and Poulson 1972). A genetic approach was used by Davies and Brewster (1975) to analyze RNA synthetic activity in developing pea cotyledons. The basis for this work was the observation that cotyledons of certain genotypes of pea show appreciable differences in their RNA contents. However, when plants with widely different rRNA contents are crossed, cotyledons of the Fl hybrid embryos are found to have an rRNA content identical to that of the maternal parent. Of the two models proposed to account for the essential silencing of a set of rRNA cistrons, one supposes that some aspect of gene activity, such as transcription or processing of RNA, is primarily under the control of the maternal genome. Alternatively, a selective activation of the maternal alleles might occur and result in an unusual situation in which one set of rRNA cistrons produces the same amount of RNA in the hybrid as two sets do in the parent. Clearly, both models are valid, but further work is necessary to choose between them. Chapter 13 discussed the fact that in the cotyledons of legumes, the DNA content of cells increases during the first few days of cell expansion growth. Since the large increase in RNA content occurs almost in parallel with the increase in nuclear DNA amounts, a logical question is whether there is an apparent gene dosage effect on RNA synthesis in the cotyledons. One measure of the transcriptional activity of DNA is a change in the level of RNA polymerases. Experiments by Millerd and Spencer (1974) have proved illuminating by showing that cotyledons of a variety of pea do not synthesize additional RNA polymerase in proportion to the increasing amount of nuclear DNA template available. Since this occurs even when the template activity of chromatin in the cotyledons remains constant, a role for gene dosage, in the sense of
409
loading the cytoplasm with transcription products in a quantitative way, does not appear to be valid. Investigations of Cullis (1976) with two other varieties of pea showed that an increase in the endogenous RNA polymerase activity of chromatin occurs even before the average DNA content increases in the majority of the cells and continues well into the period of DNA endoreduplication in most cells. This might suggest that some of the endoreduplicated DNA is used as a template for RNA synthesis. A new element of uncertainty was introduced into this issue when it was found in a later work that the extent of utilization of extra copies of DNA is associated with experimental conditions that affect the rate of growth of cotyledons. For example, in plants raised at 15 °C, which permits a tardy growth of cotyledons, RNA polymerase activity increases in proportion to the DNA content of cells over a wide developmental window. When cotyledons develop rapidly, as in plants grown at 30 °C, there is no increase in the level of chromatin-bound RNA polymerase activity in proportion to the DNA (Cullis 1978). The differences in the template activity of chromatin from two sets of cotyledons are interpretable in terms of differences in the number of transcriptionally active cistrons available or in the specific activity of the bound polymerase. These views are not inconsistent with the currently known mechanisms of regulation of DNA template activity. In summary, it appears that there is no firm ground for the idea that, except under special conditions, transcriptional activity in the embryonic cotyledons is related to the availability of extra copies of DNA. Endoreduplication results in a multifold increase in the informational pool of the cells of the cotyledons, but the ultimate function and disposition of this information remain to be fully understood. Gene expression pattern during embryogenesis has been deduced from changes in the most prevalent mRNA populations seen in cell-free translation profiles of embryos of different ages (Figure 14.5). The changes in the proteins synthesized and in the expression of their mRNAs during embryogenesis in cotton were described earlier. The existence of in vivo and in vitro synthesized polypeptide subsets that exhibit stage-specific expression appears to be a consistent feature of embryogenesis in maize (Sanchez-Martinez, Puigdomenech and Pages 1986; Boothe and Walden 1990; Oishi and Bewley 1992), radish (Aspart et al 1984; Laroche-Raynal et al 1984), Phaseolus vulgaris (Misra and Bewley 1985b), rice (Mundy and Chua, 1988), and soybean
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Genetic and Molecular Analysis of Embryogenesis
(Rosenberg and Rinne 1988). In both maize and radish, although the prevalent mRNAs show numerous changes during embryogenesis, two main subsets, one synthesized in young and immature embryos and the other accumulating in mature embryos, have been identified. Maturation of rice embryo is associated with the accumulation of mRNAs encoding about seven polypeptides ranging in molecular mass from 20 to 45 kDa. Most of the mRNA sequences of the mature embryos of these plants persist as stored mRNAs and, consequently, they are considered to be products of genes transcribed during embryogenesis. The concept of stored mRNAs in embryos of quiescent and dormant seeds of other angiosperms has been supported by a variety of experimental approaches. One important consideration about stored mRNAs is that they serve as templates for the synthesis of proteins of early germination, until the growing embryo begins to synthesize new mRNAs. In vitro translation products of mRNAs isolated from quiescent embryos of wheat (Helm and Abemethy 1990) and sorghum (Howarth 1990) have been shown to include transcripts of HSP genes. Additionally, cDNA libraries made to poly(A)RNA of embryos excised from dry seeds of radish (Raynal et al 1989), Vigna unguiculata (Ishibashi and Minamikawa 1990; Ishibashi, Yamauchi and Minamikawa 1990), V. radiata (Manickam et al 1996), Pinus thunbergii (Masumori, Yamamoto and Sasaki 1992), and sunflower (Almoguera and Jordano 1992), and embryos excised from caryopses of rice (Ramachandran and Raghavan 1992b) and wheat (Kawashima et a 1992) have been differentially screened, and developmentally significant cDNA clones have been described. Of special interest is a gene that encodes a Zn-containing metallothionein-like protein isolated from mature wheat embryos. The expression of this protein is gene modulated in developing embryos by ABA and not by Zn2+. In spite of much information about the structure and ubiquity of metallothioneins in animal cells, little is known about their function during plant embryogenesis.
Figure 14.5. Fluorographs showing the in vitro translation profiles of proteins synthesized by RNA from embryo axes of Phaseolus vulgaris of different stages of development. Schematic representations of the fluorographs are given on the right. E indicates the major translation products from the endogenous mRNA. Spots enclosed in boxes correspond to permanent polypeptides. (a) Embryo axes <12 days after pollination (DAP), (b) 22 DAP. (c) 32 DAP. (d) 40 DAP. (From Misra and Bewley 1985b. J. Expt. Bot. 36:1644-1652, by permission of Oxford University Press.)
411 (iv) Protein Metabolism in Developing Embryos
Investigations on the changes in protein accumulation and synthesis, polypeptide patterns, and enzyme titer have contributed in a limited way to our understanding of the utilization of genomic information during embryogenesis. Continued accumulation of proteins, principally attributable to large amounts of storage proteins, characterizes most periods of embryogenesis studied, except perhaps toward the end of seed maturity (Beevers and Poulson 1972; Scharp<§ and van Parijs 1973; Millerd and Spencer 1974; Manteuffel et al 1976; Zhu, Shen and Tang 1980a, b). In legumes such as Phaseolus coccineus, a translational control mechanism differentially regulates protein metabolism in the cells of the embryo and suspensor, the magnitude of the rate of protein accumulation in the cells of the embryo being appreciably lower than that in the suspensor cells (Walbot et al 1972; Sussex et al 1973). Toward the end of seed maturity, when protein accumulation in the cotyledons of Pisum sativum levels off, there is a concomitant decrease in the incorporation of labeled amino acids into proteins and in the amount of ribosomes present as polysomes (Beevers and Poulson 1972). The period leading up to embryo maturity in Sinapis alba (Fischer et al 1988) and Lupinus luteus (Gwozdz and Deckert 1989) is marked by low levels of polysomes that progressively increase to the midcotyledon phase and then decrease. As seen earlier, the lack of sufficient mRNAs has, with some experimental justification, dominated thoughts about the limitations on protein synthesis in the cotyledons of mature embryos. The results of experiments involving twodimensional electrophoretic separation of proteins synthesized by embryos of Zea mays indicate that morphological and physiological events of progressive embryogenesis are correlated with the synthesis of different subsets of polypeptides (Sanchez-Martinez, Puigdomenech and Pages 1986; Boothe and Walden 1990). Boothe and Walden have identified a subset of 2 polypeptides that are associated with the general maintenance of developmental functions and are synthesized throughout embryogenesis, another subset of 14 polypeptides synthesized during the period of morphological differentiation, and a third, smaller subset synthesized during the waning period of embryogenesis and concerned with the quiescence of the embryo (Figure 14.6). Although no function can be ascribed to the embryo polypeptides, studies reinforce the view that qualitative variations in gene products
412
Zygotic Embryogenesis
20
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Figure 14.6. A schematic representation of the changing pattern of synthesis of polypeptides in developing embryos of Zea mays. Solid lines and broken lines represent high and low levels of synthesis, respectively. Where synthesis was not detected over a specific period, no line is drawn. Polypeptides D1 to D25 exhibited qualitative and/or quantitative variations during embryogenesis. Polypeptide E did not show any variation during embryogenesis. Polypeptides are organized into subsets based on similarities in their pata'nd WaTdenmoT 13 ' ^ ^ P °" i n a t i ° n - ' ^ B ° ° t h e
are important in modulating embryogenesis. Gene expression at the level of protein synthesis is seen in the ability of embryos of Z. mays of different stages of development to respond to a heat shock by the enhanced synthesis of several molecular-mass classes of polypeptides (Boothe and Walden 1990). Expression of HSP-17 occurs in the nuclei of embryonic cells of Z. mays, Lycopersicon esculentum, Nicotiana tabacum, and P. sativum independent of heat stress in a developmentally regulated manner (zur Nieden et al 1995). In P. sativum, accumulation of other low-molecular-mass HSPs is targeted to the cytoplasm of the cells of embryos beginning about 18-25 days after flowering to the mature stage. When the embryo axis separated from the cotyledons was analyzed, induction of HSPs was found to be delayed and to coincide with the separation of the seed from the pericarp (DeRocher and Vierling 1994). Two small HSPs, HSP-17.6 and HSP-17.9, are expressed constitutively in developing embryos of Helianthus annuus; although HSP-17.9 begins to accumulate several
days ahead of HSP-17.6, both are found at high levels in mature embryos (Coca, Almoguera and Jordano 1994). Developmental regulation of HSPs suggests, but does not prove, that these proteins have some undetermined functions in embryogenesis not necessarily related to heat stress. The expression patterns of several enzymes are closely tied to critical periods of development of the embryo. As embryogenesis in Hordeum distichum progresses, an increase in the activity of e n z y m e s involved in sucrose cleavage (for exampie, sucrose synthetase) generally precedes carbohydrate accumulation (Duffus and Rosie 1975). A high level of expression of this enzyme is correlated with a significant degree of differentiation of the embryos of Z. mays (Chen and Chourey 1989). Glyoxylate cycle enzymes show a sequential pattern of accumulation in developing embryos of Gossypium hirsutum, with MS generally lagging behind ICL by several days (Choinski and Trelease 1978). Activities of p-oxidation e n z y m e s such as c a r a f e , m a l a t e dehydrogenase, citrate synthase, thiolase, enoyl-coenzyme-A (coA) hydratase, and hydroxyacyl-coA dehydrogenase are coordinately developed, increasing together from low levels at , *, , . . , . i , , earl y development to high levels at maturity (Choinski and Trelease 1978; Miernyk and Trelease 1981). Since translatable mRNAs for MS, ICL, and catalase appear together at an early stage of embryogenesis, the differences in the timing of their expression may reflect fundamental differences in posttranslational processing (Turley and Trelease 1990). Multiple electrophoretic forms of catalase are common, and two and three isozyme subunits of the enzyme are found in the embryos of G. hirsutum (Kunce and Trelease 1986) and Z. mays (Chandlee and Scandalios 1984; Williamson and Scandalios 1992), respectively. In Brassica napus, both ICL and MS begin to accumulate mostly during the late stages of embryogenesis, corresponding to the onset of embryo desiccation. Insofar as the enzyme molecules pile up with different efficiencies, some translational or posttranslational processes are believed to modulate them (Ettinger and Harada 1990). The highly cyanogenic nature of rosaceous stone fruits has been attributed to the presence of catabolic enzymes that degrade cyanogenic glycosides such as amygdalin and prunasin. The accumulation of amygdalin hydrolase, prunasin hydrolase, and mandelonitrile lyase in the embryos of Prunus serotina generally coincides with cotyledon development and storage of the glycosides (Swain, Li and Poulton 1992a). These enzymes are compartmentalized at the sub-
Genetic and Molecular Analysis of Embryogenesis
cellular level in the cotyledons; as shown by immunological methods, the two hydrolases are restricted to the protein bodies of the procambium, which ramifies through the cotyledons, whereas high levels of mandelonitrile lyase are associated with the protein bodies of the cells of the cotyledonary parenchyma (Swain, Li and Poulton 1992b). The compartmentalization of the enzymes apparently prevents both the premature degradation of the glycosides and the release of hydrogen cyanide. In Viciafaba, changes in the systems involved in the synthesis and utilization of ascorbic acid are closely related to the early cell division, later cell expansion, and final desiccation stages of embryogenesis. Thus, the activity of the enzyme that reduces ascorbic acid is high during the cell division and cell expansion phases of embryogenesis, whereas the embryo acquires the enzyme for ascorbic acid biosynthesis before desiccation. This ensures that the seed is ideally packaged to start synthesis of the ascorbates needed for scavenging purposes with the initiation of germination (Arrigoni et al 1992).
413
plants generated from seeds treated with chemical mutagens and ionizing radiations, and the visual screening of embryos obtained after fertilization with mutagenized pollen grains or after insertional mutagenesis with naturally occurring transposable elements, or by transformation with T-DNA from Agrobacterium tumefaciens. The power of these tech-
niques makes it possible to isolate genes involved in specific aspects of embryo development, to clone them, and to map the mutations. (i) Embryo-lethal Mutants
The widest range of interesting phenotypes defective in embryo development has been identified and characterized in Arabidopsis by a combination of the aforementioned methods. Early attempts to identify embryo-defective mutants in this species were made by Miiller (1963) and Usmanov and Miiller (1973), who isolated six different types of embryo-lethal mutants and four types of seedlinglethal mutants by X-irradiation of seeds or by pollen Despite the fact that embryos are traditionally mutagenesis. Embryo-lethal mutants that were shielded from light as they develop within the blocked at different points spanning the entire time maternal tissues, mature embryos of several plants course of embryogenesis, beginning with the globucontain chlorophyll in the differentiated chloro- lar embryo, were classified according to the stage at plasts. It is therefore not unreasonable to expect which development was stymied and according to that they contain mechanisms for the accumulation the size and color of aborted seeds and embryos, of plastid proteins in the cells. This was found to be but little insight was provided into the causes of the case in developing embryos of Phaseolus vul- lesion in the different mutants. garis, in which accumulation of Rubisco occurs The work of Meinke and Sussex (1979a, b) independent of chlorophyll accumulation and in demonstrated that recessive embryo-lethal mutants complete darkness (Medford and Sussex 1989). The can be easily isolated following EMS mutagenesis significance of this observation is that Rubisco of Arabidopsis seeds. In six mutants characterized in accumulation during embryogenesis might be reg- these investigations, embryogenesis was arrested ulated by developmental signals. during a narrow developmental window between the preglobular and heart-shaped stages. The experiments that inescapably led to the conclusion that 3. CHARACTERIZATION OF GENES these were nonallelic, recessive lethals included ESSENTIAL FOR EMBRYOCENESIS analysis of mutant sectors in chimeric Ml plants, How far can an embryo continue to develop determination of segregation and heterozygote before it must call on a specific battery of its genes ratios, and tests for temperature sensitivity and to develop further? A cornerstone in the quest for allelism. genes essential for important developmental funcEvolving out of these efforts, later investigations tions in the embryo, instead of for housekeeping isolated a great number of mutants with considerchores, is the isolation of genetic mutations. The able diversity of abnormalities in embryo developrationale behind this approach is that in the ment and a wide range of lethal phases. In a batch absence of mutations that cause developmental of 32 mutants isolated by EMS mutagenesis, develdefects, there is no way to establish the involve- opmental arrest of embryos ranged from the zygotic ment of a gene in the normal development of the to the mature cotyledon stages of development. defective organ. Of the various approaches Some mutants had lethal phases that covered a very employed to isolate mutants defective in embryo narrow range of developmental stages, whereas in development, several have had considerable suc- others, the lethal phase covered several consecutive cess, including the visual screening of embryos of stages. In their phenotypic mode, arrested embryos
474
appeared with large suspensors, distorted and fused cotyledons, or dwarfed hypocotyls, or they were devoid of hypocotyl and cotyledons. Seeds from some mutants completed embryogenesis, but germinated to form defective seedlings. The gametophytic expression of genes in several mutants at some point prior to fertilization was ascertained from the nonrandom distribution of aborted seeds in heterozygous fruits (siliques) or from the preferential location of aborted seeds toward the half of the silique close to the stigma (Meinke 1982, 1985). A library of nearly two hundred additional embryo mutants with varying degrees of abnormal development has been isolated following seed transformation of Arabidopsis with Agrobacterium tumefaciens
(Errampalli et al 1991; Castle et al 1993; Yadegari et al 1994; Vernon and Meinke 1995). By genetic analysis of progeny of selfed Fl plants and by RFLP strategy, several genes essential for embryo development have been mapped with both visible and molecular markers (Patton et al 1991). Further developmental consequences of these mutations were uncovered by cytological, biochemical, and tissue culture investigations. As reviewed in a previous section, some key mutants are characterized by abnormal growth of the suspensor at the same time that growth of the embryo proper is inhibited. In situ hybridization with several cellspecific mRNA markers, such as lipid transfer protein mRNA and storage protein mRNAs, showed that these transcripts are expressed in their normal spatial context in both mutant and wild-type embryos, as well as in the enlarged suspensor region (Yadegari et al 1994). The expression of marker genes carried under the control of promoters of a storage protein gene and a LEA gene in the embryos and suspensors of aborted seeds of some mutants also mirrors gene specificity seen in phenotypically normal seeds (Devic, Albert and Delseny 1996). These results are considered to provide molecular evidence for the view that defects in the embryo can cause the suspensor to take over the vital functions of the embryo and that the activity state of key genes is not necessarily dependent on organ formation in the embryo. Because storage protein synthesis is largely confined to the maturation phase of embryogenesis, cellular and biochemical patterns of protein accumulation are reliable markers for the developmental arrest of mutant embryos. In their ultrastructural appearance, protein bodies of some mutant embryos are identical to those of the wild-type embryos, whereas in others they resemble immature protein bodies. This picture of protein body dimorphism is
Zygotic Embryogenesis
supported with the specific evidence that mutant embryos with morphologically altered protein bodies fail to accumulate the 12S and 2S storage proteins found in the wild-type embryos. These observations emphasize that mutant genes might disrupt either morphogenesis or both morphogenesis and cellular differentiation in embryos blocked at different stages of embryogenesis (Heath et al 1986; Patton and Meinke 1990). As storage protein accumulation represents only one of a large number of biochemical activities that lead to embryo maturation, examination of other parameters of the defective embryos is necessary to identify specific cellular and molecular defects of the mutation. The culture of seeds enclosing mutant embryos in defined media has proved to be an excellent approach to recovering homozygous mutant plants for genetic and molecular studies, although the isolation of auxotrophs by this method has not been nearly as successful. In terms of general responses to identical culture conditions, it was found that mutant embryos arrested at the heart-shaped to cotyledonary stages of development can be reprogrammed in culture to produce respectable calluses that regenerate abnormal plants with defects in reproductive development, whereas embryos arrested at the globular or earlier stages fail to grow or they produce only slow-growing calluses incapable of complete plant regeneration. Embryos of one mutant (emb-30; also referred to as 112A-A) appeared to be particularly recalcitrant to rootforming activity in culture although they produced abundant callus and even leaves (Baus, Franzmann and Meinke 1986; Franzmann, Patton and Meinke 1989). The presence of abnormal developmental syndromes in mutant plants rescued in culture opens up the possibility that many genes required for embryo development also have a role in the vegetative and reproductive growth of plants. The response of seeds and embryos of one mutant (bio-1) that failed to respond to culture on a basal medium but produced normal plants in a medium enriched with biotin or desthiobiotin (a close precursor of biotin) is typical of that of a biotin auxotroph. Mutant embryos rescued in agar cultures were found to produce normal plants with viable seeds when they were transferred to soil and watered daily with either biotin or desthiobiotin (Schneider et al 1989). Further analysis showed that embryos of bio-1 contain only trace amounts of biotin, whereas arrested embryos from other mutants with similar morphology have higher concentrations of biotin than the biotin auxotroph. A sharp definition of the
Genetic and Molecular Analysis of Embryogenesis
biotin lesion was obtained by the demonstration that the bio-1 mutant of Ambidopsis can be rendered prototrophic by transformation with a chimeric bacterial 7,8-diaminopelargonic acid aminotransferase gene that catalyzes the conversion of the biotin intermediate 7-keto-8-aminopelargonic acid to desthiobiotin (Shellhammer and Meinke 1990; Patton, Volrath and Ward 1996). These findings support the conclusion that bio-1 embryos have a defective biotin synthetic pathway that disrupts the gene encoding 7,8-diaminopelargonic acid aminotransferase. As this is the first example of an embryo-lethal mutant with a defect in a defined biochemical pathway, further research on this system should assist in our understanding of the biosynthetic pathway of biotin in embryos and the role of biotin in embryogenesis. Embryo-lethal Mutants in Maize. Just as Arabidopsis is an ideal plant among dicots for studying the genetic control of embryo development, maize has proved to be tailor-made for similar investigations among monocots. Of the many features of maize that contribute to this suitability, two are particularly noteworthy, namely, the availability of good background genetic and cytogenetic information and the presence of several well-characterized transposable elements. A number of single-gene recessive mutants in maize with lesions in their embryo and endosperm development were described between 1920 and 1950. Jones (1920) and Mangelsdorf (1923) analyzed several spontaneously occurring mutants characterized by a high degree of grain abortion due to impairment in the development of the embryo and endosperm. Although this analysis did not clearly identify the stage of embryo arrest, some of the mutant kernels were able to germinate into abnormal seedlings. This is probably suggestive of some defects during later stages of embryogenesis. True embryo-lethal mutants (germless) were described by Demerec (1923), who established that the germless condition is an inherited character. Histological studies of this line showed that aborted seeds generally contained a normal endosperm, but embryo development was arrested at various stages and was followed by degeneration prior to maturity (Sass and Sprague 1950). Clearly, these mutants did not show a loss of developmental potential in the endosperm at any time, or in the embryo up to the time that led to the arrested stage. The genetic program of embryogenesis in maize was brought into sharp focus in later years by analyses of embryo-lethal mutants isolated by
415
pollinating ears with EMS-mutagenized pollen grains or by tagging with mu transposable elements. These studies involved screening several thousand plants and examining an equally staggering number of kernels showing varying degrees of defects in both embryo and endosperm development; most kernels, however, were mutants with abnormal endosperm and nonviable embryo, or with normal endosperm and nonviable embryo. Taking advantage of maize translocation stocks in the crossing of mutants with plants of B-A translocation to produce kernels that contain combinations of a mutant hypoploid embryo and a nonmutant hyperploid endosperm, or a nonmutant hyperploid embryo and a mutant hypoploid endosperm, showed that a normal endosperm is unable to rescue a mutant embryo. It was also established from a reciprocal cross that even in the presence of a mutant endosperm, a normal embryo completes its development. These results provided the principal criteria to establish that lethality in a number of mutants studied is determined by the genotype of the embryo and not of the endosperm, and that a defective endosperm, created by previous interaction, does not prevent the completion of embryo development (Neuffer and Sheridan 1980). A group of dek mutants isolated from maize are characterized by the presence of defective endosperm and of embryos arrested at different stages of development. Most of the mutant embryos develop at least to the point of differentiating one or more leaf primordia, whereas only a few are arrested at earlier stages of development and are unable to form leaf primordia. As shown in Figure 14.7, the impairment of growth in the latter group of embryos spans an array of stages, ranging from proembryo through club-shaped transition and coleoptile stages to the stage of initiation of leaf primordia (Sheridan and Neuffer 1980,1982). Some of these mutant embryos have been characterized histologically, and the unique steps in the sequence of gene action essential for normal embryogenesis have been partly defined. Thus, a requirement for normal gene functions at the transition stage and at the stage of initiation of the coleoptile-shoot axis is implied by the developmental profiles of mutant embryos dek-22 (E113A) and dek-23 (E1428), respectively. Although the initial development of dek-22 embryos is not different from that of wild-type embryos up to the transition stage, mutant embryos do not display any further morphogenetic changes necessary to pass to the coleoptile stage. Similarly, arrested embryos of dek-23 mutants do
416
"Proembryo
Zygotic Embryogenesis
Transition
Coleoptilar
Stage 1
Stage 2
Figure 14.7. Stages of developmental arrest of embryos in dek mutants of maize. Stages of development of the normal embryo are shown in the lower half of the figure. The coleoptilar stage is characterized by the presence of a coleoptile ring and a shoot apical meristem, and stage 1 is characterized by the presence of the first leaf primordium.
Besides exhibiting specific developmental defects, all mutants shown in the figure lacked leaf primordia. (From Sheridan and Neuffer 1982./ Hered. 73:318-329. © 1982 by the American Genetic Association. Print supplied by Dr. W. F. Sheridan.)
not advance beyond an abnormal coleoptile stage characterized by the lack of a shoot apex and of a coleoptile ring found in wild-type embryos poised to form a coleoptile; eventually, the embryos begin to show signs of necrosis (Clark and Sheridan 1986; Dolfini and Sparvoli 1988). Similar defects are very common in mutants bno (E747B), ptd (E1130), and cp (E1418), which lag behind wildtype embryos in their development and are blocked at specific stages of embryogenesis (Sheridan and Thorstenson 1986). One distinguishing feature of embryogenesis in mutants rgh (E1210) and/Z (E1253B) is the diversion of arrested embryos to abnormal pathways of morphogenesis that result first in the disruption of bilateral symmetry and apical meristem formation, and then in
callus-like growth (rgh) and proliferation of the embryonic axis and surrounding regions, accompanied by hypertrophy of the scutellum (fl) (Clark and Sheridan 1988). It will be recalled from Chapter 12 that dek mutations affect the endosperm, indicating that they are pleiotropic in nature. How the same genes control developmental processes in two different tissues with differing gene dosages remains to be explored. The genetically heterogeneous nature of these mutants was underscored by the growth of embryos in culture. Embryos of several mutants were rescued by the simple expedient of growing them on a mineral-salt medium with or without organic additives, whereas the developmental defect of some embryos could not be overcome even in the most enriched
Genetic and Molecular Analysis of Embryogenesis
medium tested (Sheridan and Neuffer 1980; Clark and Sheridan 1986,1988; Sheridan and Thorstenson 1986). Isolation of auxotrophs is a cherished hope of investigators in this field; similar to a spontaneously occurring mutant in maize, one of the induced mutations has been traced to a genetic block in the proline biosynthesis pathway (Racchi et al 1978; Sheridan and Neuffer 1980). The morphogenetic effects of mutations in maize generated by transposon tagging appear to be specific to embryos and result in the same array of genetic lesions described in EMS-generated mutants. A general classification has placed these mutants, designated as embryo-specific (emb), into three groups: mutants in which (a) embryogenesis is blocked during the proembryo and transition stages; (b) the block occurs during the period of differentiation of the embryo axis and scutellum and initiation of the leaf primordium; and (c) the block persists during the period of elaboration of embryonic structures beginning at the late coleoptile stage. The majority of the 51 mutants characterized belong to the second group, leaving 10-12 in each of the other two groups (Figure 14.8). Correlated genetic and developmental studies have provided proof that many genes are essential for the ordered development of the maize embryo and that the emb phenotype reflects mutations in many loci of these genes. The conclusion that the EMB loci are essential to embryo development follows from the observations that the arrest of embryo growth occurs before kernel maturity and that mature kernels harboring mutant embryos do not germinate (Clark and Sheridan 1991; Sheridan and Clark 1993). However, this is only a part of the story because no understanding of the mechanism of action of EMB genes can be gained without molecular data. Embryo-lethal Mutants in Other Plants. In broad terms, embryo-lethal mutants isolated from rice by chemical mutagenesis resemble transposontagged mutants of maize because the mutational effects are restricted primarily to the embryo and do not spread to the endosperm. In extreme cases, the mutant grain does not possess an embryo, which apparently disintegrates at the globular stage. Some mutants show defects in the differentiation of the shoot that result in a chaotic arrangement of leaf primordia at the apex and in abnormalities in their anatomy. Root initiation and development does not go awry as the shoot does, indicating that initiation of the two major embryonic organs is controlled by different loci (Nagato et al 1989; Tamura et al 1992; Hong et al 1995b). Another set of mutants has been
417
separated into groups affecting three regulatory pathways of embryogenesis, namely, organ differentiation, positional regulation of organs, and size regulation of the embryo, that occur before embryo morphogenesis. Mutations in the first pathway produce club-shaped embryos that do not possess any visible organs, as well as embryos that do not produce any shoots but form normal roots; mutations in which both shoot and root are displaced more apically belong to the second group. Mutations affecting the size of embryos can lead either to reduction or to enlargement of the embryo (Kitano et al 1993; Hong et al 1996). This close relationship between mutations affecting early embryo development, without overlap, emphasizes the point that virtually every step in the embryogenesis program is gene controlled. Both embryo and endosperm are affected in a negatively reciprocal way in a temperature-sensitive mutant of rice, embryoless-1 (eml-1). Rearing plants at day/night temperatures of 30/25 °C following pollination causes the formation of grains with no embryos or of grains with malformed embryos but with a large quantity of endosperm (Figure 14.9). Grains with large embryos and poorly developed endosperm result when plants are reared at a constant temperature of 18 °C or 20 °C (Hong et al 1995a). The subtle effects of the mutant gene in the embryo and endosperm suggest a role for the EML-1 gene product in the continued development of these parts of the grain, but the effects are mediated in some presently unknown way. Pea is another plant that has been inducted into the ranks for analysis of the genetics of embryogenesis. Mutant embryos showing defects in cotyledon development and root initiation have been isolated, and genetic studies have shown that at least three loci participate in the development of the two cotyledons in their full multicellularity and structure (Johnson et al 1994). In one mutant, designated as cytokinesis-defective (cyd), cotyledon morphology is apparently altered due to a defect in the formation of the cell wall, resulting in partially formed cell plates and multinucleate cells (Liu, Johnson and Wang 1995). It will be interesting to see whether this deficiency could be corrected by culture of embryos in a medium with appropriate supplements. (ii) Genetic Control of Embryonic Pattern Formation
The laying down of the basic body plan of the embryo, consisting of an upper shoot region,
418
Zygotic Embryogenesis Differentlatlon ot embryo proper Into scutellum and embryonic axis and differentiation of first leaf primordium
Setting apart of the embryo proper and suspensor
Elaboration of embryonic structures and Imposition of dormancy
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Figure 14.8. Summary of developmental blocks of the emb mutants in maize. Stages of wild-type embryos are shown at the top of the figure and specific developmental defects are shown immediately below. Stages (pro, proembryo; trans, transition; col, coleoptilar; and mature stages 1 to 6)
%. at which each mutant is blocked are indicated by stippling. Chr arm, chromosome arm. Each division of the scale bar equals 0.5 m m . (From Sheridan and Clark 1993; print supplied by Dr. W. F. Sheridan.)
Genetic and Molecular Analysis of Embryogenesis
419
a Figure 14.9. Sections of grains of temperature-sensitive mutants of rice, (a) An embryoless mutant, (b) A mutant containing an embryo lacking organ differentiation, (c) A mutant endospermless grain containing a giant embryo. Arrow indicates shoot and arrowhead indicates root, en, endosperm; sc, scutellum. Scale bar = 0.5 mm. (Reprinted
from S. K. Hong et al 1995a. Temperature-sensitive mutation, embryoless 1, affects both embryo and endosperm development in rice. Plant Sci. 108:165-172, with kind permission of Elsevier Scientific-NL, Sara Burgerhartstraat 25, 1055 KV Amsterdam, The Netherlands. Photographs supplied by Dr. Y. Nagato.)
a middle hypocotyl, and a lower root segment, has been the subject of considerable study and some speculation. Descriptive embryology that addresses the organizational aspects of embryo development in plants has shown that these regions can be traced back to the terminal and basal cells of the two-celled proembryo. Speculation has centered mostly on the mechanism underlying pattern formation, which reflects deeply on the question as to whether the three regions of the embryo axis are specified autonomously and develop independent of each other. A genetic approach to analyzing the mechanism of embryonic pattern formation has sought to identify mutant phenotypes in Arabidopsis that define different steps in the process and to characterize the genes involved. In saturation mutagenesis experiments using a chemical mutagen, embryonic pattern mutants were selected by large-scale screening of mutant seedlings for profound and specific defects in the apical-basal pattern, radial organization, and shape of the embryo. The rationale for this approach is that since mutants are isolated by their ability to recover from lethality during embryogenesis, any diagnostic defects detected in the seedlings are caused by patterning genes and not due to lesions in general cell metabolism. Complementation analysis has shown that nine loci selected are each represented on an aver-
age by eight mutant alleles. Included in the class of mutations with defects in the apical-basal organization of the seedling are (a) mutants that lack the apical region, including the cotyledons and the shoot meristem (gurke [gu]); (b) those that lack the hypocotyl and so have the cotyledons attached directly to the root {fackel \fk]); (c) those that lack both the hypocotyl and the root and so consist essentially of an apical piece of axis to subtend the cotyledons and shoot meristem (monopteros [mp]); and (d) those that lack both terminal pattern elements, including the cotyledons and the root, and so consist of a cellular mass with no obvious signs of apical-basal polarity ignom [gn]); these are diagrammatically represented in Figure 14.10. Mutations in two genes, KNOLLE (KN) and KEULE (KEU), disrupt the radial pattern and result in seedlings with defects in the epidermis. Grossly abnormal seedlings are produced by mutations in three genes, FASS (FS), KNOPF (KNF), and MICKEY (MIC), that affect the shape of the embryo without eliminating any pattern elements (Jtirgens et al 1991; Mayer et al 1991). Other genes implicated in the apical-basal organization of the seedling are SHOOT MERISTEMLESS (STM) and WUSCHEL (WUS), which are critical for the initiation of the shoot apical meristem, and ROOTLESS (RTL), which is critical for the initiation of the root apical meristem in the embryo (Barton and Poethig
420
Zygotic Embryogenesis
stage transition and a series of transverse divisions in the hypophysis (Figure 14.11). The defective divisions in the lower tier of cells do not interfere GURKE with the orderly pattern of divisions in the upper tier (Berleth and Jiirgens 1993). Chaotic divisions beginning as early as the octant stage of the embryo and accompanied by incomplete cross wall formation play a major role in the evolution of the kn pheFACKEL notype, but the genetic basis of the mutation also extends to the generation of multinucleate polyploid cells (Lukowitz, Mayer and Jiirgens 1996). Effects of the STM gene can be identified only at the torpedo-shaped stage, long after other pattern elements are established; a mutation in this gene results in the inability of the presumptive cells of MONOPTEROS the shoot apical meristem to initiate divisions. Thus, in the hierarchy of gene actions responsible for specifying the embryo body plan, the STM gene acts only after the GN and MP genes (Barton and GNOM Poethig 1993). The first division of the zygote of the /s mutant is asymmetric, as in the wild-type embryo, indicating that the mutation does not interfere with polarity of the embryo; however, the FS Figure 14.10. Schematic representations of the pattern gene plays a key role in the subsequent events of deletions caused by mutations in Arabidopsis and the phecell elongation and orientation of cell divisions in notypes formed. The wild-type seedling patterns with the deleted parts shaded are shown on the left, and the corre- the embryo (Figure 14.11). This is reflected in the fact that by the time the wild-type embryo has sponding mutant patterns are on the right. (From Mayer et al 1991. Reprinted with permission from Nature turned heart shaped or torpedo shaped, the mutant 353:402-407. © Macmillan Magazines Ltd.) embryo merely resembles an oversized globular embryo (Torres-Ruiz and Jiirgens 1994). Implicating a derangement of auxin metabolism in the genera1993; Laux et al 1996). Mature embryos also tion of/s mutants is the observation that wild-type respond to the effects of the gene CLAVATA3 (CLV- Arabidopsis embryos cultured in a medium contain3), which causes abnormal enlargement of the veg- ing NAA are transformed into mutant phenocopies etative and inflorescence meristems (Clark, (Fisher et al 1996). Running and Meyerowitz 1995). Souer et al (1996) have described embryos of Petunia sp. carrying a no Possible molecular mechanisms to explain the apical meristem (nam) mutation that fail to develop action of pattern-forming genes have been sought the shoot apical meristem. by the cloning and characterization of STM, NAM, Developmental profiles of embryos of GN, and KN genes. The predicted STM protein is Arabidopsis have shown that the mutant genes somewhat close to the protein encoded by the strike at the cellular level and disrupt the division maize KN-1 gene and, as in the latter, transcripts planes that are circumscribed during early embryo- are expressed in the cells of the embryo slated to genesis. The seedling phenotype of the gn mutant form the shoot apex (Long et al 1996). A novel prohas been traced back to the first division of the tein that shares a conserved N-terminal domain zygote, which results in two nearly equal-sized with several other proteins of unknown function is cells instead of two unequal cells. Subsequent divi- apparently encoded by the NAM gene (Souer et al sions of the terminal cell are anarchic in the mutant 1996). The GN gene has been found to be allelic to embryo, which also comes off without a hypoph- embryo-lethal gene EMB-30; mutation in the gene ysis (Mayer, Biittner and Jiirgens 1993). Formation produces a fused cotyledon phenotype and does of the mp mutant embryo is another case of abnor- not promote root regeneration on embryos in culmal early divisions, but abnormalities begin to ture (Meinke 1985; Baus, Franzmann and Meinke appear only after the octant stage. They include 1986; Mayer, Buttner and Jiirgens 1993). An indicarandom cell divisions in the lower tier of cells of the tion of the role of the EMB-30/GN gene in cellular octant embryo during the globular to heart-shaped functions at several stages of development of the
Genetic and Molecular Analysis of Embryogenesis
421
Figure 14.11. Embryo development in wild-type (a-d), and mp (e-h) and fs (i, j) mutants of Arabidopsis. (a, e) Octant stage embryo; asterisk marks the uppermost suspensor cell, (b, f) Early heart-shaped stage, (c, g) Late heart-shaped stage, (d, h) Torpedo-shaped stage. Arrowhead in (h) indicates the vascular system formed in the cotyledon primordium of the mutant embryo, (i) Mutant embryo at a stage comparable to the late heartshaped stage of the wild type (stages between b and c). Arrow points to irregularly arranged cells probably corresponding to the root primordium; arrowheads point to the irregular surface layer of cells, (j) Mutant embryo at a stage comparable to the heart-shaped stage of wild type shown in (c). For each stage, wild-type and mutant embryos are shown at the same magnification, co, cotyledon; cp, a central pile of cells formed from the hypophysis contiguous with the suspensor; ep. epidermis; It, lower tier of the octant; rp, root primordium with the cells of the hypophysis; su, suspensor; ut, upper tier of the octant; vp, procambium. ([a-h] from Berleth and Jurgens 1993. Development 118:575-587. © Company of Biologists Ltd. Photographs supplied by Dr. T. Berleth; [i, j] from Torres-Ruiz and Jurgens 1994. Development 120:2967-2978. © Company of Biologists Ltd. Photographs supplied by Dr. R. A. Torres-Ruiz.)
422 plant was provided, not surprisingly, by Northern blot analysis showing that gene transcripts are expressed in almost all organs of Arabidopsis tested. However, what was surprising was the discovery that the gene encodes a protein that has extensive similarity to the secretory (SEC-7) protein of yeast. SEC-7 is a cytosolic protein found in the Golgi complex and is probably involved in glycosylation, secretion, transport, and polysaccharide synthesis associated with the Golgi apparatus. In Arabidopsis, the functionally important component of the protein is restricted to a stretch of 157 amino acids, designated as the "Sec-7 domain" (Shevell et al 1994). Another proposal has assigned the GN protein a close homology to the translation product of a different yeast gene (YEC-2) with undefined functions, rather than to SEC-7 (Busch, Mayer and Jurgens 1996). The translation product of the KN gene has been found to be a member of the syntaxin family of proteins, whose main function is in directing vesicle traffic during cell division (Lukowitz, Mayer and Jurgens 1996). In any event, the principal conclusion from experimental observations is that EMB-30/GN and KN gene products may be concerned with cell division, cell expansion, and cell adhesion during embryogenesis; because of the ubiquitous nature of these proteins, they may affect related cellular events in developing embryos that are difficult or impossible to interpret in specific detail. As such, these results depict the EMB-30/GN and KN gene products as casting only a peripheral effect on pattern formation in embryos. The defective polarity in some of the mutant embryos suggests that a protein regulating cytoskeletal assembly might be sought as an implementing factor targeted by the mutation.
4. EXPRESSION OF GENES FOR HOUSEKEEPING AND STRUCTURAL PROTEINS
To understand the molecular processes that operate during embryogenesis, it is necessary to isolate a representative sample of the variety of genes active at different stages of embryo development and to study their regulation. This is easier said than done because of the sheer number of genes expressed at the mRNA level in embryos and the possibly bewildering complexity of their protein products. Presumably there are many genes that are unique to specific stages of development of embryos; a second group comprises genes for housekeeping proteins that are shared with other plant organs. Included in the first group are mainly storage protein genes and genes involved in pat-
Zygotic Embryogenesis
tern formation and morphogenesis of embryos, whereas genes encoding enzymes of general metabolism, respiration, and photosynthesis, and genes coding for proteins of cell wall biosynthesis, plant defense, desiccation, and cell division, for example, may be assigned to the second group. Following the discussion of genes involved in pattern formation and morphogenesis just completed, this section will consider genes for housekeeping proteins. Due to the large volume of information to be collated, gene action in the regulation of storage protein synthesis in the embryo will form the sole topic of Chapter 15. (i) Genes for Glyoxylate Cycle and Related Enzymes
Investigations on the regulation of gene expression during seed germination and seedling growth of Brassica napus have led to the identification of cloned mRNA sequences that accumulate at different stages of embryogenesis. Most postgermination-abundant mRNAs selected from a cDNA library made to poly(A)-RNA of darkgrown 14-hour-old seedlings were found to accumulate in the cotyledons and axes of fully germinated seedlings. When mRNAs from staged embryos were hybridized in a Northern blot with cloned seedling probes, a surprising finding was the presence of cotyledon-abundant and axisabundant gene sets at distinct stages of embryogenesis. Cotyledon-abundant mRNAs reach maximum levels in embryos poised for desiccation (35-45 days after flowering), whereas the axis-abundant mRNAs are most prevalent at 15-27 days after flowering and reach minimum levels by the time of embryo desiccation (Harada, Baden and Comai 1988). Further insight into regulation of the expression of cotyledon-abundant genes during embryogenesis has come from the work that showed that two cotyledon-abundant mRNAs encode the glyoxylate cycle enzymes ICL and MS, which participate in storage lipid mobilization (Comai, Baden and Harada 1989; Dietrich et al 1989). RNA blot hybridization showed that ICL and MS genes exhibit qualitatively similar expression patterns in the late stages of embryogenesis, but MS messages are more prevalent than ICL messages. Isolated nuclei provide a convenient system to study changes in the rates of transcription. Use of nuclei isolated from B. napus embryos of different ages to determine ICL and MS transcriptional run-off activity showed that genes for both enzymes and some other gene sets
Genetic and Molecular Analysis of Embryogenesis
remain competent to be transcribed in the embryos, with the caveat that transcription to the level necessary for germination does not occur during embryogenesis (Comai et al 1989; Comai and Harada 1990). As predicted by these observations, the genes for MS and ICL are expressed during late embryogenesis in transgenic tomato and tobacco, respectively; furthermore, the transcripts encoded by the MS gene are distributed throughout the tissues of mature embryos, including the procambium, of both B. napus and transgenic tomato (Comai et al 1992; Zhang et al 1993). Deletion analysis using the ICL gene in transgenic Arabidopsis has also identified distinct DNA sequences that are primarily, but not exclusively, expressed during embryogenesis and during seedling growth (J. Z. Zhang et al 1996). These results reaffirm the well-established tenet that embryogenesis and germination represent different facets of the early sporophytic phase and call for the operation of distinct regulatory programs to activate and repress MS, ICL, and other similarly expressed genes. Transcripts of the ICL gene detected initially in maturing embryos of Helianthus annuus are maintained at a constant low level through seed desiccation and increase disproportionately during germination (Allen, Trelease and Thomas 1988). To follow the regulation of an axis-abundant gene (AX-92) expressed at an early stage of embryogenesis, a chimeric gene construct consisting of 5'- and 3'-flanking regions of AX-92 fused to GUS gene was expressed in transgenic B. napus. GUS gene expression was first detected predominantly in the root cortex of the torpedo-shaped embryo, the activity increasing up to the cotyledonary stage embryo and no further. Expression of deletion constructs in the embryos has identified DNA sequences located in the 3'-part of the protein coding region as critical for the activation of the chimeric gene in the embryos (Dietrich, Radke and Harada 1992). Since the same minimal gene construct induces GUS expression in the root cortex of seedlings, similar mechanisms appear to underlie the expression of the gene during embryogenesis and germination.
423
expressed HSP-70 gene, but homology decreases in the transcripts produced at later stages of embryogenesis (Domoney et al 1991). Consistent with the constitutive synthesis of HSPs, the expression of transcripts of HSP-70 or small HSP genes is found to parallel protein accumulation in embryos of P. sativum (DeRocher and Vierling 1994, 1995), Helianthus annuus (Almoguera and Jordano 1992; Coca, Almoguera and Jordao 1994), Nicotiana tabacum, and Arabidopsis (Prandl, Kloske and Schoffl 1995). In describing the heat-shock effects on developing embryos of Zea mays, Marrs et al (1993) have shown that whereas transcripts of some HSP genes (HSP-81 and HSP-90) are present in embryos at normal temperatures, developmentally regulated expression of HSP82 and HSP-17.9 does not occur in uninduced embryos. However, the ability of HSP-81 and HSP-82 transcripts to respond to heat shock appears to be itself developmentally regulated, as neither transcript is thermally inducible until embryos have progressed to about 14 days after pollination. Thus, although the two genes are probably regulated by different mechanisms, both HSP-81 and HSP-82 produce the same end result, namely, enhanced heat-induced expression. (iii) Expression of Other Genes
The expression of genes that play key roles in the developmental decisions of the embryo has remained surprisingly obscure. Two homeobox genes have provided the probes for an analysis of the expression of key genes in developing maize embryos. It may be significant that transcripts of these genes, which code for transcription factors, are activated during early embryogenesis and are confined to the meristematic cells, such as the shoot and root meristems, procambium, and leaf primordia (Klinge and Werr 1995). Consistent with their widely accepted role in controlling cell division, transcripts of the maize homologue of a cell cycle control protein kinase gene (CDC-2) are also found in actively dividing maize embryo cells (Colasanti, Tyers and Sundaresan 1991). Cell wall glycoproteins, histones, and chloroplast structural proteins must be active participants in the expression of genes for structural proteins during embryo devel(ii) Heat-Shock Protein Genes opment (Figure 14.12). A variety of methods, including Northern blot, Western blot, and in situ The synthesis of HSPs in developing embryos is hybridization, have shown that both the protein the result of the accumulation of transcripts of HSP-related genes. Early stage embryos (16-28 and the mRNA of a gene for a hydroxyproline-rich days after flowering) of Pisum sativum transcribe an glycoprotein component of Z. mays cell wall accumRNA with appreciable homology to the highly mulate in the embryo axis but not in the scutellum
424
Zygotic Embryogenesis
gene are localized in the cells of the scutellum, leaf primordia, and vascular bundles of the coleoptile (Ruiz-Avila et al 1992). At different stages of development of embryos of Oryza sativa, pronounced localization of rice histone H3 gene transcripts is confined to the cells of the scutellum, coleoptile, first leaf, and root primordium, including the root cap, coleorhiza, and zone of elongation (Figure 14.13); however, transcripts are conspicuously absent from cells of the shoot apex, newly emerging leaf primordia, and the root quiescent center (Raghavan and Olmedilla 1989). As a speculative thought, the requirement for histone transcripts in DNA repair synthesis may explain their presence in the nonmeristematic cells of the embryo. Chloroplast genes are expressed constitutively in developing embryos of Gossypium hirsutum (Borroto and Dure 1986), Glycine max (Saito et al 1989; Chang and Walling 1991, 1992), and Arabidopsis (Degenhardt, Fiebig and Link 1991). During maturation of soybean embryos, chloroplasts with highly differentiated grana are transformed into amyloplasts and proplastids; this transition is correlated with a decrease in CAB gene mRNAs. The transcription of different members of the CAB gene family during embryogenesis appears to be controlled by different factors, because CABS mRNAs are found to level off more slowly than mRNAs of other gene family members during embryo maturation (Saito et al 1989; Chang and Walling 1992). Transcripts of PSB-A, RBC-L, and RBC-S (nuclear-encoded small subunit of Figure 14.12. Autoradiographic localization of transcripts Rubisco) genes detected by in situ hybridization of a cell wall hydroxyproline-rich glycoprotein gene in the are found first in the proembryos of Arabidopsis, immature embryo of Zea mays, (a-e), Dark-field microand the transcripts increase in abundance up to the scopic images of autoradiographs of sections cut at levels mature stage. The increased concentration of the shown in the diagram of the embryo on the right. C, transcripts in the cotyledons of mature embryos coleoptile; Cr, coleorhiza; le, leaf primordium; n, scutellar node; P, procambium; r, radicle; Sc, scutellum. Scale bar = would appear to correspond with the potential fate 200 urn. (From Ruiz-Avila et al 1992.) of cotyledons as the first photosynthetic organs of the seedling plant (Degenhardt, Fiebig and Link 1991). These data collectively suggest that the of immature embryos (Ruiz-Avila, Ludevid and expression of chloroplast genes during embryogenPuigdomenech 1991; Ruiz-Avila et al 1992). esis in soybean and Arabidopsis is controlled develHowever, transcripts of another gene from Z. mays opmentally, rather than by light signals. Another that encode a proline-rich, hydrophobic protein function of plastids in developing embryos is the accumulate both in the embryo axis and scutellum synthesis of amylopectin involving the starch(Jose-Estanyol, Ruiz-Avila and Puigdomenech branching enzyme. One isoform of this enzyme is 1992). Although the functional significance of these highly expressed in the young embryos of pea, and observations is not clear, a general consensus would another isoform attains maximum expression in appear to be that cells of the embryo axis and scutelold embryos. Differential expression of the enzyme lum differ in their wall protein composition. isoforms is also correlated with the nature of amyExpression of certain histone genes during lopectin synthesized in developing embryos embryogenesis appears to be both division depen- (Burton et al 1995). dent and division independent. In immature embryos of Z. mays, transcripts of maize histone H4 One of the earliest molecular markers of
Genetic and Molecular Analysis of Embryogenesis
425
Figure 14.13. Autoradiographs of sections of rice embryos hybridized in situ with 3H-histone H3 RNA probe, (a) Section of the root apex of an 8-day-old embryo. Lines are drawn on the figure to indicate the boundaries of certain parts, (b) Section of the scutellum of a 6-dayold embryo showing the labeling pattern in individual cells, cr, coleorhiza; nu, nucleus; qc, quiescent center; re,
root cap; sc, scutellum. Scale bars = 50 um (a) and 10 um (b). (From V. Raghavan and A. Olmedilla 1989. Spatial patterns of histone mRNA expression during grain development and germination in rice. Cell Diffn Devel. 27:183-196, with kind permission of Elsevier Scientific-NL, Sara Burgerhartstraat 25, 1055 KV Amsterdam, The Netherlands.)
embryogenesis in Daucus carota is a gene for an extracellular protein (EP-2) that shows homology to plant lipid transfer proteins. Transcripts of this gene are first detected in the cells of the globular embryo, including the suspensor, but there is a progressive restriction of gene expression to the protoderm of heart-shaped and older embryos until transcripts are completely absent in mature embryos (Sterk et al 1991). The time course of accumulation of transcripts of maize and Arabidopsis lipid transfer protein genes in developing embryos of the respective species is similar to that seen in carrot embryos; at the cellular level, hybridization with the labeled maize probe is observed in the epidermal cells of the coleoptile and scutellum (Sossountzov et al 1991; Vroemen et al 1996). It is speculated that lipid transfer proteins might mediate in cutin biosynthesis in the embryo epidermis by transporting the precursors to the site of synthesis. Floral MADS-box genes code for proteins that serve as transcription factors; intense expression of transcripts of the AGL-2 gene of Arabidopsis and
embryogenesis (Flanagan and Ma 1994; Heck et al 1995; Rounsley, Ditta and Yanofsky 1995). cDNA clones hybridizing to mRNAs preferentially expressed in the embryo or in both the embryo and endosperm of barley have been useful in defining developmental events in embryogenesis. Transcripts corresponding to an embryospecific clone (pZE-40) begin to appear in the club-shaped embryos of barley; with further differentiation, steady-state levels of transcripts are abundantly expressed in the scutellum, coleoptile, and coleorhiza, but the axial tissues and the scutellar epidermis are generally devoid of transcripts. This expression pattern does not change until desiccation of the grain, when the transcript levels precipitously decline (Smith et al 1992). Transcripts of some embryo- and endosperm-specific cDNA clones are also characteristically expressed in the scutellum of developing barley embryos, whereas others that encode oleosins are expressed in the scutellum or in the root-shoot axis (Olsen, Jakobsen and Schmelzer 1990; Aalen et al 1994). Identification of the protein products encoded by these genes is necessary to provide a consolidated picture of their role in the continued differentiation of the embryo. Promoter sequences of some genes are active in
AGL-15 gene of Arabidopsis and Brassica napus in the
globular and torpedo-shaped embryos and, to a lesser extent, in older embryos of the respective species might reflect the role of specific transcription factors in regulating gene expression during
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Zygotic Embryogenesis
regulating embryo-specific expression of reporter be expressed differentially in mature embryos of genes in transgenic plants. These include promot- transgenic tobacco, but the stage of embryogenesis ers of chalcone flavanone isomerase gene (CHI-A) when stable promoter activation occurs is not from Petunia hybrida in transgenic P. hybrida determined (Benfey, Ren and Chua 1989,1990). (van Tunen et al 1990); rice actin-1 gene in transgenic rice and maize (Zhang, McElroy and Wu 1991; Zhong et al 1996); maize ADH gene in trans- 5. EXPRESSION OF GENES RELATED TO DESICCATION TOLERANCE A N D genic rice (Kyozuka et al 1994); CDC-2a, mitotic DORMANCY cyclin (CYCl-At), and phytochrome genes from Arabidopsis in transgenic Arabidopsis (Hemerly et al During seed maturation, the embryo undergoes 1993; Ferreira et al 1994; Somers and Quail 1995); desiccation resulting in the loss of nearly 90% of the and asparaginase gene from Lupinus arboreus in water from the cells and then lapses into a state of transgenic tobacco (Grant and Bevan 1994). Two quiescence or dormancy. Despite its extremely low conserved regions identified in the promoters of water content, the embryo remains viable during ABA-responsive genes of many plants are a palin- this period of enforced rest, only to spring forth dromic sequence (PA, 5'-GCCACGTGGC-3') and and germinate later under appropriate environmotif I (Iwt, 5'-GTACGTGGCG-3"). When these mental conditions. Although the mechanism that sequences are inserted in appropriate constructs retains the embryo in a developmental mode durand expressed in transgenic tobacco, PA is found ing desiccation and dormancy is poorly underto confer expression in the embryonic root and Iwt stood, it is clear that as a physiological change, is found to drive developmentally regulated desiccation tolerance is a genetically programmed expression in embryos of all ages (Salinas, Oeda phenomenon. Besides the loss of water, an increase and Chua 1992). A role for PA and Iwt sequences in in the endogenous ABA level in the cells of the differentially targeting trans-acting factors to regu- embryo marks this period of development. At the late gene expression during embryogenesis is molecular level, there is a close relationship between the low embryo water potential and implied here. The expression of two of the several oncogenes expression of specific genes encoding mRNAs that usually accumulate in the dehydrated embryo. of T-DNA of root-inducing (Ri) plasmid of These mRNAs, referred to as LEA mRNAs, are Agrobacterium rhizogenes has been followed during translated in vivo during embryo maturation and embryogenesis in transgenic tobacco by histochemearly germination and rapidly disappear thereafter. ical assay for GUS gene expression. Whereas stimOne sequence family amongst these genes encodes ulation of reporter gene expression by the a set of proteins known as dehydrins. promoter of the ROL-B gene occurs uniformly in all Several aspects of the mechanism by which cells of the globular, heart-shaped, and torpedoshaped stage embryos including the suspensor, embryos sense the low water potential have been gene activity in mature embryos is restricted to the investigated, although we do not as yet have a comvascular elements and the apical meristems. Since plete picture of the cues that activate LEA genes. ROL-B gene promoter is controlled by auxin in ABA has become an obvious candidate because it plant tissues, these results invoke an association exercises a potent effect in inducing the precocious between GUS gene expression and the synthesis of accumulation of LEA mRNAs in cultured immature endogenous auxin in the embryos. By contrast, embryos. Additionally, it has been demonstrated ROL-C gene promoter activity is conspicuously that the expression of a variety of LEA genes is actiabsent in the early stage embryos but appears in vated in plant tissues subjected to osmotic or salt the phloem tissue of the hypocotyl and cotyledons stress or to desiccation. Another pertinent fact is of late heart-shaped embryos and increases in that in several plants the concentration of ABA intensity until the mature stage (Maurel et al 1990; peaks in embryos during the period of declining Chichiricco, Constantino and Spanb 1992; Aspuria, water potential, then a decrease in concentration Nagato and Uchimiya 1994). Phloem specificity of follows during the final stages of maturation (Hsu expression of the GUS gene under the control of 1979; Ackerson 1984a; Finkelstein et al 1985; Prevost ROL-C gene promoter widens the potential use of and Le Page-Degivry 1985; Galau, Bijaisoradat and the promoter in monitoring vascular tissue differ- Hughes 1987; Wang et al 1987a; Robertson et al 1989; Chang and Walling 1991). These observations entiation during embryogenesis. Two main have placed embryo desiccation, and the correlated domains and several subdomains of the CaMV 35S expression of LEA messages and proteins and promoter fused with a reporter gene are found to
Genetic and Molecular Analysis of Embryogenesis
increase in ABA content, as part of a general response of plants to a broad range of environmental stresses, of which water deficit is paramount. The concept of LEA mRNAs arose from an analysis of the changing mRNA populations during different stages of embryogenesis in cotton based on comparisons of the extant, in vivo and in vitro synthesized proteins (Dure, Greenway and Galau 1981). This work showed that a subset of 14 polypeptides that became abundant in mature embryos was a good molecular marker of the onset of embryo maturation. The disappearance of these sequences during germination of mature embryos and their premature accumulation in young embryos cultured in a medium containing ABA were also established in this study. Galau, Hughes and Dure (1986) assigned the term "LEA" to denote these mRNA sequences that are significantly more abundant in maturation stage embryos than in younger embryos, with the caveat that the term does not imply their inducibility by ABA under any experimental conditions. Using the amino acid composition of six LEA proteins of cotton as a point of reference, it was shown that LEA proteins from different species share regions of homology with three of the cotton LEA proteins (Baker, Steele and Dure 1988; Dure et al 1989). Based on this analysis, a classification of LEA proteins has recognized three groups, namely, group 1 with cotton LEA protein D-19 and wheat EM protein, group 2 with cotton LEA protein D-ll and rice RAB-21 protein, and group 3 with cotton LEA protein D-7 and LEA proteins from barley, carrot, and rapeseed. One of the most interesting developmental features of embryogenesis, and one that has received considerable attention in recent years, concerns the basis for the expression of LEA genes. It was pointed out earlier that the influence of LEA genes is felt as the embryo becomes dehydrated, and consequently, the gene products are presumed to facilitate desiccation survival of the embryo enclosed in the seed. Given the fact that desiccation stress affects the survival of every conceivable part of the plant, it is not surprising that expression of LEA genes or their protein products is commonly observed in leaves and other organs subjected to this stress. Viewed from another angle, it is significant that the promoter region of a desiccationrelated gene cloned from leaves of an extremely desiccation-tolerant plant (Craterostigma plantagineum) is activated in the embryos of transgenic tobacco as the seeds become dehydrated (Michel et al 1993). It is also of interest that the deduced amino acid sequence of a cDNA clone isolated from
427
dormant spores of the fern Onoclea sensibilis reveals some homology to LEA proteins. Typical of LEA gene expression, transcripts of this cDNA clone begin to accumulate during the maturation and desiccation phases of spores before they lapse into dormancy (Raghavan and Kamalay 1993). Structural features of LEA proteins suggested by their amino acid sequences (in particular by the presence of a tandemly repeated sequence) and their hydrophilic nature in some cases (Baker, Steele and Dure 1988; Galau, Wang and Hughes 1992b; Dure 1993), by their homology to waterstress-related proteins in other cases (Galau, Wang and Hughes, 1993), and by their uniform distribution in the cytosol of all cells of the embryo in high molar concentrations (J. K. Roberts et al 1993), are considered to be ideal to protect the embryo from desiccation damage or to repair such damage. In short, most of the speculative attention given to the function of LEA proteins has focused on desiccation survival, with ABA thrown in as a signal transducer, but further work is necessary to place these ideas in a tangible context. The intent of this section is to deal with the isolation, characterization, and expression of LEA and other genes involved in the desiccation and dormancy of embryos of different species of plants and to discuss how their expression is regulated by ABA. An unusual breadth of information into many general features of LEA gene expression arose from studies with model systems such as embryos of cotton and wheat. For this reason, these systems are treated in greater detail than others. (i) LEA Genes from Cotton and Other Dicots cDNAs for a total of 18 different LEA genes have been cloned and characterized from cotyledons of cotton. Based on sequence data, 6 of the clones have been assigned to small multigene families; each of the 6 genes contains the standard features, including TATA-box, poly(A) addition sites, and one or more introns (Baker, Steele and Dure 1988; Galau and Close 1992). As expected, mRNAs of all LEA sequences are abundant in the mature embryos and most, but not all, are modulated to varying degrees by ABA in excised and cultured embryos. However, the accumulation kinetics of LEA mRNAs during embryogenesis have placed them in two groups of coordinately regulated mRNAs (Figure 14.14). These two groups are (a) LEA genes, with 12 members whose mRNAs exhibit their maximum level of accumulation in
428
Zygotic Embryogenesis
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napus but is inducible by ABA; however, it is highly prevalent in mature embryos and disappears during the early hours of germination. In the context of stress tolerance, a further novel feature of this recombinant is that mild, nonlethal freezing of plants triggers the premature synthesis of its transcripts in embryos during subsequent growth at room temperature 0ohnson-Flanagan et al 1992). Typical of other LEA proteins, the deduced structure of the protein encoded by the gene shows extreme hydrophilicity and the presence of tandemly organized direct repeats. A somewhat similar expression pattern has been demonstrated for transcripts of a gene isolated from a cDNA library made to poly(A)-RNA of mature embryos of Raphanus sativus; the protein deduced from the nucleotide sequence shares good homology with a cotton LEA protein and wheat EM protein (Raynal et al 1989).
Investigations on precocious germination of cultured immature embryos of Pisum sativum showed that addition of ABA to the medium Figure 14.14. Gel-blot hybridization of total RNA from increases the production of two low-molecularcotyledons of cotton embryos with two groups of cDNA mass proteins, ABR-17 (molecular mass 17.2 kDa) clones encoding LEA proteins. Days indicated at the top are and ABR-18 (molecular mass 26 kDa). Although DPA, days postanthesis; DPI, days postimbibition. Stages of embryo development indicated are Cot, cotyledon; Mat, both of these proteins and their mRNAs are maturation; PA, postabscission; Grm, germination. (From detected in mature embryos, in other respects the Hughes and Galau 1989; photograph supplied by Dr. G. A. proteins differ in their spatial and temporal Galau.) expression patterns. Especially interesting are the accumulation of ABR-18 protein in the testa durembryos after ovules have severed their connection ing early seed development and the continued to the maternal plant at the funiculus (postabscis- synthesis of ABR-17 protein during germination. sion stage, an 8-day period between maturation Amino acid analysis showed that both proteins and desiccation, beginning 45-46 days postanthe- have more homology with pea disease-resistant sis); and (b) LEA-A genes, with 6 members whose response proteins than to any of the LEA proteins. mRNAs, in addition to postabscission stage expres- However, other data relating to the accumulation sion, are also modulated by ABA (Galau, Hughes of these proteins in aging tissues and in response and Dure 1986; Galau, Bijaisordat and Hughes to various stimuli unrelated to pathogenicity sug1987; Hughes and Galau 1989). Since Gossypium hir- gest that the expression of genes for these proteins sutum, used in these investigations, is a natural in the pea embryos appropriately fits the bill for allotetraploid, it may be possible that the complex desiccation response (Barratt, Domoney and kinetics of LEA and LEA-A mRNA expression are Wang 1989; Barratt and Clark 1991,1993). A strong the sum of two alloalleles encoding them. candidate for the desiccation-related protein of However, the fact that transcripts of a few ran- pea embryos is encoded by a cDNA clone isolated domly selected LEA and LEA-A alloallele pairs dis- from a pea embryo expression library using maize play similar expressions at the mRNA or protein dehydrin antiserum. Not only is this protein levels excludes this possibility (Hughes and Galau nearly homologous to dehydrins from other 1987; Galau and Hughes 1987). Several cotton LEA plants, but, typical of desiccation-related genes, and LEA-A mRNAs also accumulate in the postab- transcripts of the gene encoding it are expressed at scission stage embryos of other dicots (Jakobsen, a high level during embryo maturity (Roberton and Chandler 1992). Similar evidence for the presHughes and Galau 1994). ence of LEA proteins has come from analyses of Harada et al (1989) have identified a cDNA sunflower embryos (Almoguera and Jordano clone whose mRNA accumulates at low to unde- 1992) and embryos of Cicer arietinum (chick pea; tectable levels in immature embryos of Brassica
Genetic and Molecular Analysis of Embryogenesis
Fabaceae) (Colorado, Nicolas and Rodriguez 1995; Colorado et al 1995). A quite different category of LEA proteins is represented in sunflower embryos by ubiquitin. The characteristic abundance and up-regulation of expression patterns of transcripts of a cDNA clone encoding tetraubiquitin during embryogenesis in sunflower are very similar to those of LEA genes. The suggestion that degradation of denatured proteins produced during desiccation of embryos might involve a ubiquitin-mediated step is consistent with the known functions of this protein (Almoguera, Coca and Jordano 1995). In addition to the accumulation of a set of proteins, a feature that endows desiccation tolerance in embryos of soybean is an increase in the concentration of sugars, especially stachyose (Blackman et al 1991; Blackman, Obendorf and Leopold 1992). There is thus circumstantial evidence for the view that proteins may function in concert with oligosaccharides in the development of desiccation tolerance. Several cDNA clones, corresponding to different maturation proteins ranging in molecular mass from 18 kDa to 70 kDa, have been isolated from soybean embryos, and the expression of transcripts of these clones has been shown to increase not only in mature embryos but also in precociously matured embryos (Hsing et al 1990; Hsing and Wu 1992). The derived amino acid sequences of two of the clones analyzed show many homologies to LEA proteins (Chen et al 1992; Hsing, Chen and Chow 1992). Beyond the examples described in which LEA mRNAs and proteins have been isolated and delineated from mature embryos of dicots, some investigators have followed in developing embryos of Arabidopsis the expression of LEA transcripts isolated from other plants. These studies include localization of EM-like genes and cotton D-19 gene in mature embryos by in situ hybridization and Northern blot, respectively (Gaubier et al 1993; Finkelstein 1993) and expression of maize RAB-17 gene in the embryos of transgenic Arabidopsis (Vilardell et al 1994). Transcripts of several clones homologous to LEA genes and some novel genes isolated from developing siliques of Arabidopsis are found to be prevalent in seeds undergoing desiccation, indicating that they are LEA transcripts (Parcy et al 1994). In the case of one of the genes, a functional analysis of the promoter in a transgenic setting has indicated that putative regulatory elements similar to those of EM and storage protein (i-phaseolin genes govern the embryo specificity of its expression (Hull et al 1996).
429
(ii) LEA Genes from Wheat and Other Monocots Among the recombinants identified from a cDNA library made to poly(A)-RNA of wheat embryos was one representing about 200 bases of a message that declined rapidly during germination (Cuming 1984). The identity of this sequence, designated as EM, as a member of LEA group 1 was established in related studies that led to the isolation of a near full-length clone, the determination of its nucleotide and amino acid sequences, and demonstration of its presence in low levels early in embryogenesis, its abundance in late stage embryos, and its inducibility by ABA (Williamson, Quatrano and Cuming 1985; Litts et al 1987, 1991; Williamson and Quatrano 1988; Morris et al 1990). Another wheat LEA gene characterized by high expression in mature embryos and inducibility by ABA is PMA-2005; it encodes a protein product of molecular mass 23 kDa and is assigned to LEA group 3 (Curry, Morris and Walker-Simmons 1991). In attempts to determine the mechanism of ABA regulation of EM gene expression in developing embryos, it was found that a construct containing a 646-bp 5'-fragment of the EM gene fused with GUS gene causes a sharp increase in gene expression in near mature seeds of transgenic tobacco relative to seeds of other ages. Expression of the construct was inducible in embryos of immature seeds by incubation in ABA (Marcotte, Russell and Quatrano 1989). A protein that belongs to the leucine zipper class of transcription factors and interacts with an 8-bp core (T/CACGTGGC) consensus sequence of the ABAresponse element of the EM gene has been identified (Figure 14.15); this development and the delineation of a transcript with homology to protein kinases that increases with embryo maturity and ABA treatment are considered important steps in determining signal transduction in the ABAinduced expression of the EM gene (Guiltinan, Marcotte and Quatrano 1990; Anderberg and Walker-Simmons 1992). Lam and Chua (1991) have shown that a mutated sequence from the wheat histone H3 promoter confers a high level of expression in embryos of transgenic tobacco. Proof for the probable function of this sequence as an ABAresponsive element has come from the observation that the expression of the promoter is diminished upon germination and is reinstated by ABA. An unidentified tobacco nuclear factor has been proposed as a candidate element that interacts with the mutated sequence. Attempts to identify LEA mRNA and proteins in
430
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leucines and the single methionine are circled. Boxes indicate the residues conserved among most leucine zipper proteins. (Reprinted with permission from M. J. Cuiltinan, W. R. Marcotte, Jr. and R. S. Quatrano 1990. A plant leucine zipper protein that recognizes an abscisic acid responsive element. Science 250:267-271. © 1990. American Association for the Advancement of Science.)
barley embryos go back to experiments on the acquisition of desiccation tolerance. Transition from desiccation susceptibility in 12-day-old embryos to desiccation tolerance at 18 days is associated with the synthesis of a set of 25-30 proteins (Barrels, Singh and Salamini 1988). Isolation and characterization of a gene temporally linked with desiccation tolerance in embryos showed that it is homologous to mammalian genes encoding an NADPH-dependent aldose reductase (Bartels et al 1991). This enzyme is considered to play an important role in the synthesis of the common cell osmolyte, sorbitol, in animal cells; osmolytes generally help to balance the changes in the osmolarity of the cytoplasm caused by adverse environmental conditions. It is interesting to note that the same gene holds the key to the mobilization of defense against common traumatic effects in both plants and animals. Another gene with a potential function in desiccation tolerance that becomes transcriptionally active in developing barley embryos has a protein sequence homologous to bacterial glucose dehydrogenase and ribitol dehydrogenase (Alexander et al 1994). Three additional LEA mRNAs (B-19.1, B-19.3, and B-19.4) were identified by using a cotton LEA cDNA probe to screen a barley cDNA library made to poly(A)-RNA from ABA-treated immature embryos, and a fourth one was identified by screening the library with a B-19.3 cDNA probe. The temporal accumulation pattern of transcripts is
essentially the same for all genes and, beginning about 25 days after anthesis, it shows accumulation to high levels throughout embryogenesis. However, based on the distinct patterns of accumulation of transcripts of three members of the B-19 gene family in embryos of different ages in response to mannitol, ABA, and NaCl, it seems that different signal transduction pathways operate in their expression (Espelund et al 1992,1995). Several properties of the putative B-19 proteins, especially their strong hydrophilic nature and the internal 20-amino acid sequence repeated one to four times, support their strong similarity to the EM protein from wheat and to LEA proteins from several dicots (Hollung, Espelund and Jakobsen 1994). Studies on the regulation of gene expression during embryogenesis in maize have shown that a set of 23-25-kDa polypeptides appears during embryo maturation and disappears during germination (Sanchez-Martinez, Puigdomenech and Pages 1986). Further characterization of these proteins involved the isolation of cDNA clones from mature maize embryos that hybridize with specific mRNAs, the demonstration of the absence of transcripts of cDNA clones in young embryos and their abundance in mature embryos, and the ABA inducibility of the transcripts in immature embryos (Goday et al 1988; Gomez et al 1988). Transcripts of one of the clones (PMAH-9) are restricted in distribution to the two outer layers of cells of the scutellum and to the epidermal cells of the embryonic
Genetic and Molecular Analysis of Embryogenesis
leaves. The deduced protein of this clone has an unusually high glycine content, like other proteins with putative structural and protective functions, and based on its affinity to undine- and guanosinerich RNAs, it is considered to be an RNA-binding protein (Gomez et al 1988; Ludevid et al 1992). Another clone (MA-12) also encodes a glycine-rich protein (molecular mass 17 kDa) with conserved tracts of amino acid sequences as in other LEA proteins. Because of the extensive homology of this protein to RAB-21 from rice - a homology evidenced especially by the presence of glycine-rich domains and conserved sequences and by the Cterminal regions - it is referred to as RAB-17. The state of phosphorylation of RAB-17 protein changes during embryogenesis, because it becomes highly phosphorylated in the desiccated embryo (Vilardell et al 1990). Structural and functional analyses have demonstrated that 5'upstream sequences of the RAB-17 gene confer developmental and hormonal expression of CAT reporter gene in embryos of transgenic tobacco. Developmentally, expression of RAB-17 gene and of RAB-28, another maize stress-protein gene, is restricted to the mature embryos and, hormonally, expression is induced in immature embryos by ABA (Vilardell et al 1991; Niogret et al 1996). Moreover, the proximal promoter region of RAB-28 contains a motif homologous to the conserved ABA-responsive element (T/CACGTGGC) of wheat embryos; DNA fragments containing this motif have been shown to bind to a trans-acting factor from the embryo nuclei (Pla et al 1993). Changes in steady-state levels of mRNA of a cDNA clone, EMB-564, during embryogenesis in maize are similar to those observed with PMAH-9 and RAB-17. EMB-564 encodes a protein that shows extensive similarity to protein products of wheat EM gene and cotton and radish LEA genes (Williams and Tsang 1991). Transcriptional and translational expression of the RAB-17 gene occurs in all cells of the maize embryo, including those of the scutellum, embryo axis, and provascular tissues. With the progress of embryo maturation, gene transcripts are increasingly confined to the outermost cells of the embryo. Both the cytoplasm and nucleus have been identified as the subcellular targets of RAB17 protein accumulation. Furthermore, a specific binding of RAB-17 protein to a nuclear localization signal binding protein has been demonstrated (Asghar et al 1994; Goday et al 1994). In the larger view, this implies that nuclear protein transport relies heavily on RAB-17 protein during stress, but
431
this idea is in its infancy and requires further investigation. A pivotal feature of the maturation program of rice embryos is the progressive accumulation of transcripts of the RAB-21 gene, which is also expressed in roots, leaves, and suspension cells under stress. The gene encodes a low-molecularmass (16 kDa), basic, glycine-rich protein that accumulates only in the cytosol. The protein contains two domains with an A and a B repeat in each; these features characterize the gene and its products as a member of the LEA family (Mundy and Chua 1988). RAB-21 is a member of a small multigene family, and four members of this family that have been characterized are referred to as RAB-16A to RAB-16D. Northern blot analysis with gene-specific probes showed that mRNAs of only A, B, and C accumulate in mature embryos (Yamaguchi-Shinozaki, Mundy and Chua 1989). An indispensable region of the B genome, which confers a developmentally regulated expression in embryos of transgenic tobacco, is restricted to 482 bp of 5'-sequences (Yamaguchi-Shinozaki et al 1990). Two genes isolated from rice embryos encoding LEA proteins and a basic leucine zipper binding protein are found to display similar characteristics, such as control by ABA and activation by the hormone in transient expression assays (Hattori, Terada and Hamasuna 1995; Nakagawa, Ohmiya and Hattori 1996).
(iii) Is Embryo Desiccation Developmentally Regulated by ABA?
One of the most intriguing problems in the physiology of the growth of embryos concerns the role of ABA as a developmental regulator of embryogenesis and a mediator of the response of embryos to stress. Earlier, it was mentioned that there is an increase in the concentration of ABA in embryos during the period of their declining water content and that the addition of ABA promotes the accumulation of LEA mRNAs in cultured immature embryos. In principle, two types of models can be invoked to explain the role of ABA: (a) the hormone promotes LEA mRNA accumulation that obviously results in a cascade effect leading to desiccation, and (b) the increase in the hormone level is a secondary effect of desiccation-related events in the embryo that are potentiated by other factors. Since much of the argument in favor a specific function for ABA in embryogenesis has followed from its effects on the accumulation of LEA mRNAs
432 or their protein products in cultured immature embryos, this work will be reviewed here. Many more LEA mRNA markers have been followed in cotton embryos than in embryos of any other angiosperm to study ABA effects, and yet there is no convincing picture of how the hormone is required during maturation and subsequent events of embryogenesis. The work of Dure, Greenway and Galau (1981) initially demonstrated that a subset of proteins that are synthesized late in embryogenesis but disappear during germination can be induced prematurely in excised young embryos cultured in ABA. This study was followed by another work, which showed that out of 18 cloned LEA mRNA sequences, as many as 13 are clearly regulated by ABA in cultured immature embryos (Galau, Hughes and Dure 1986). Subsequent investigations have shown that addition of ABA to the medium regulates the accumulation of a selection of LEA mRNAs and/or proteins in cultured immature embryos of wheat (Williamson, Quatrano and Cuming 1985; Morris et al 1990), rapeseed (Harada et al 1989; Jakobsen, Hughes and Galau 1994), barley (Bartels, Singh and Salamini 1988; Espelund et al 1992; Hollung, Espelund and Jakobsen 1994), maize (SanchezMartinez, Puigdomenech and Pages 1986; Goday et al 1988; Gomez et al 1988; Williams and Tsang 1991; Thomann et al 1992; Williamson and Scandalios 1992), pea (Barratt, Domoney and Wang 1989; Barratt and Clark 1991), tobacco, soybean (Jakobsen, Hughes and Galau 1994), and rice (Hattori, Terada and Hamasuna 1995). In some cases, various types of osmotic stress and putative plant hormones such as jasmonic acid mimic the effects of ABA on the induction of LEA mRNAs and proteins (Espelund et al 1992; S. Reinbothe et al 1992; Thomann et al 1992; Barratt and Clark 1993; Hollung, Espelund and Jakobsen 1994). As described in a previous section, some suggestive evidence for the role of ABA as a regulator of embryogenesis is provided by cases of vivipary. Induction of the germination mode seen in embryos of viviparous mutants and the analysis of ABAinsensitive mutants have played a role in articulating the idea that vivipary is analogous to precocious germination in vivo. However, the few cases described neither provide absolute confirmation that vivipary is related to the failure of ABA synthesis in the embryos nor assert that the hormone regulates the accumulation of LEA mRNAs or proteins. A perspective that ABA and water stress function as environmental inducers, rather than as developmental regulators, of embryogenesis has
Zygotic Embryogenesis
been provided by Galau, Jakobsen and Hughes (1991), who have also pointed out many interpretational pitfalls in embryo culture experiments. The significant observations supporting their model are that culture of immature embryos of cotton in a mineral-salt medium under standard conditions executes a maturation program in the molecular sense and leads to an increase in LEA mRNA amounts at rates very similar to those seen in embryos in vivo up to the postabscission stage; this mRNA induction is insensitive to the addition of fluridone. Thus, evidence is tenuous that ABA is involved in the accumulation of LEA mRNAs, and the picture is complicated by the observation that the hormone or a high osmoticum does indeed cause increased accumulation of LEA messages in cultured immature embryos (Hughes and Galau 1991; Jakobsen, Hughes and Galau 1994). Finkelstein (1993) has shown that a cotton LEA gene is expressed in the seeds of an ABA-deficient mutant of Ambidopsis at levels found in the wildtype seeds; this observation does not point to the necessity of endogenous ABA as an essential control signal for LEA gene expression. A hypothesis proposed to explain the postabscission program of embryogenesis includes ovule abscission that prepares the embryo for desiccation and an environmentally responsive mechanism resulting in water stress. Obviously, this may not seem to be an attractive hypothesis to those with a bent toward hormonal explanations, as this discounts a role for endogenous ABA in embryo desiccation. The discovery of LEA genes and the focus on the meaning of their proteins in the desiccation tolerance of embryos represent a major breakthrough in the molecular embryology of angiosperms in the last 10 years. The next areas of impact will be the determination of how desiccation leads to gene expression and an understanding of the pathways involved in desiccation tolerance of the embryos. (iv) Genes Involved in Dormancy Regulation and Seedling Growth
In a great many plants, desiccation of the embryo is a prelude to the seed's lapse into dormancy. Several critical metabolic events are elaborated in the embryo during the final stages of embryogenesis, which lead to the dormant state. Some of these events include changes in water content, acquisition of desiccation tolerance, changes in pigmentation, synthesis of ABA, and accumulation of LEA and storage proteins. Mutations in genes that regulate these events produce embryos
Genetic and Molecular Analysis of Embryogenesis
433
with defects in the following areas: storage product and produced seeds that were green, desiccation accumulation, pigmentation, maturation program, intolerant, and nondormant. The mutation reduced and seed morphology; the embryos also have mul- the amount of, but did not eliminate, the principal tiple lesions in the regulation of the final stages of seed storage proteins (Nambara, Naito and development. Analysis of these mutations has McCourt 1992). Most of these features were also made feasible a genetic approach to the identifica- noted in the seed phenotypes of two additional tion of the genes involved in the dormancy of mutants that are characterized as severely ABA insensitive (Ooms et al 1993). Hence, the data sugembryos. Since ABA affects many aspects of late embryo gest that some products of the ABI-3 gene regulate development, including dormancy, water relations, an ABA-dependent stage of embryogenesis that germination, and stress tolerance, mutational prevents a germinative type of development and approaches to the identification of genes regulating induces dormancy. In support of this view is the late embryogenesis have logically related the observation that the ABI-3 gene encodes a protein defects to disturbance in the ABA metabolism of that shares regions of sequence homology with VPembryos. Using ABA-deficient (aba) mutants of 1 gene of maize (Giraudat et al 1992). As noted earArabidopsis, it was initially found that ABA plays an lier (p. 401), VP-1 protein acts as a transcription important part in the induction of seed dormancy, factor and, by implication, the ABI-3 gene product because the mutant seeds germinate even without can be considered to regulate embryo dormancy at undergoing a period of dormancy; in contrast, the transcriptional level. seeds of the wild type that contain ABA remain An understanding of the precise roles of ABI-1, dormant (Karssen et al 1983). In a later work, which ABI-2, and ABI-3 genes in inducing seed dormancy screened a large number of mutants for seed ger- is complicated by the fact that seeds of all three mination in the presence of inhibitory concentra- mutants skip the usual period of dormancy and tions of ABA, three ABA-insensitive (abi) loci, germinate, yet embryos of abi-1 and abi-2 mutants designated as abi-1, abi-2, and abi-3, were identified. continue to synthesize storage proteins. Do these These mutants differed in their vegetative growth genes regulate the same pathways leading to dorand seed development; abi-1 and abi-2 were charac- mancy or do the genes constitute independent regterized by wilting due to disturbed water relations ulatory networks with and without the involvement but had wild-type levels of storage proteins and of ABA? An answer to this question may lie in the corresponding mRNAs, whereas the abi-3 mutants complete molecular characterization of the ABI had reduced amounts of these macromolecules. In locus and the proteins it encodes. The ABI-1 gene severe mutant alleles of abi-3, the embryos are has been cloned independently by two groups of found to lapse directly into the germination mode investigators and shown to code for a Ca2+-moduwhile they are still enclosed within the seed coats lated phosphatase having a carboxy-terminal (Koornneef, Reuling and Karssen 1984; Finkelstein domain with high affinity to serine-threonine proand Somerville 1990; Nambara et al 1995). When tein phosphatases. The involvement of serine or ABA deficiency and hormone insensitivity are threonine phosphatases in signaling processes combined in double mutants, aba/abi-3 recombi- makes the ABI-1 gene product a prime candidate to nants are found to generate seeds that do not des- integrate ABA- and Ca2+-dependent stimuli with iccate normally, that remain green, and that the phosphorylation status of the target memgerminate without lapsing into dormancy; more- branes of embryos (Leung et al 1994; Meyer, Leube over, seeds of the double mutant are deficient in and Grill 1994). Since seeds of abi-1 mutants germisome of the major storage proteins and other low- nate and produce long roots in the presence of molecular-mass polypeptides (Koornneef et al inhibitory concentrations of ABA, it has been sug1989; Meurs et al 1992). gested that ABI-1 protein might neutralize the varThe foregoing observations suggest that seed ious phosphorylation events necessary for cell development in Arabidopsis is much more sensitive division in the root meristem, ensuring that the to ABA action than is induction of dormancy. To embryo remains dormant (Leung et al 1994). Thus, further explore the role of the ABI-3 gene in regu- it seems that ABI-1 and ABI-3 gene products, which lating the late embryogenesis program, uniconazol, provide the link with the genome, prepare the an inhibitor of GA biosynthesis, was used to isolate embryo for dormancy by regulating two strategimutants defective in the normal germination pro- cally different pathways. gram. One of the lines isolated was insensitive to the inhibitor due to a mutation in the ABI-3 gene
The genetic approach to the study of the embryo maturation program has been augmented
434
Zygotic Embryogenesis
by the isolation and characterization of mutant desiccation phases, such as the temporal and spaphenotypes defective in maturation-associated tial expression of cruciferin, oleosin, and LEA embryo morphology. The leafy cotyledon (lec-1) phe- genes, are not affected by the mutation. The implinotype isolated by screening seeds of transgenic cation is that a battery of genes controlling indeArabidopsis plants produced by T-DNA insertion is pendent regulatory pathways may operate a paradigm of such a mutant with striking defects concurrently to control the embryo maturation proin the maturation-specific events of embryogene- gram and germination and that a large majority of sis. Besides being desiccation intolerant and non- these genes remain unidentified. dormant, the mutant embryos are defective in Examination of other mutants has confirmed storage product accumulation and lack the protein that activation of a set of genes during late embryoand lipid bodies characteristic of wild-type genesis initiates strategies for dealing with the embryos. However, the distinguishing feature of growth of seedlings after germination. Several this mutant is the reversion of cotyledons to a newly isolated fus mutants fall into the group showleaflike state by the development of trichomes and ing defects in the embryo and the seedling; embryos stomata on their adaxial surface, by a complex of these mutants go through a normal program of pattern of vascularization associated with the for- development but have the peculiar virtue of overmation of differentiated tissues, and by the resem- production of anthocyanins in their cotyledons blance of cotyledonary parenchymatous cells to (Baumlein et al 1994; Castle and Meinke 1994; mesophyll (Meinke 1992; Meinke et al 1994). Three Misera et al 1994). In addition, seedling lethality other mutants with related phenotypes that pro- resulting in limited seedling development is an duce trichomes on cotyledons but that generally important pervasive factor throughout the early differ from each other in certain other aspects of stages of germination of seeds of fus mutants. Since embryo maturation have allowed delineation of a several fus seedlings respond to sugars and horgroup of regulatory genes that function during late mones differently than do the wild-type seedlings stages of embryogenesis. Mutant phenotypes of lec- and since they do not become etiolated in the dark, 1, lec-1.2 (an allele of lec-1), lec-2, and fusca (fus-3) are it has been suggested that gene products needed for the most closely related because these mutations normal responses to carbohydrates, hormones, and affect similar features of the late embryogenesis light are synthesized during embryogenesis. program. Differences between lec-1 and fus-3 Characterization of one of the FUS gene products embryos are restricted mostly to the frequency of showed that it belongs to a new class of proteins that stomata and the position of anthocyanin accumula- is hydrophilic, a-helical, with several consensus protion. In contrast, lec-2 embryos are desiccation tol- tein kinase-C phosphorylation sites. Since some fus erant, do not germinate precociously, and show a mutants are alleles of other Arabidopsis mutants pattern of storage product accumulation closer to defective in light-regulated responses, FUS proteins the wild-type embryos than to lec-1 or fus-3 have been implicated in regulating signal transducembryos (Keith et al 1994; Meinke et al 1994). These tion pathways during seedling growth (Castle and characteristics indicate that a functional LEC gene Meinke 1994; Misera et al 1994). Baumlein et al is necessary for the development of cotyledons (1994) have shown that when storage protein and from the primordia initiated early in embryogene- nonstorage protein promoters are tested in transsis and that genes active in desiccation and dor- genic Arabidopsis plants in a fus-3 genetic backmancy are independent of those that specify ground, transcription of only the storage protein cotyledon characters. promoters is inhibited; this has led to the view that In a unifying model, West et al (1994) have sug- the FUSS gene product might function as a trangested a specific role for the LEC-1 gene not only in scription regulator of storage protein genes. the regulation of late embryogenic development but also in the transition of embryos to a germina6. REGULATION OF EXPRESSION OF tion mode. The basis for this model is the observaDEFENSE-RELATED GENES tion that although lec-1 mutation affects desiccation The role of specialized proteins synthesized by tolerance and storage protein accumulation associated with late embryogenesis, some postgermina- embryos in preventing bacteria and fungi from tive traits, such as the expression of lipid transfer attacking seeds was not recognized until relatively protein gene at a level characteristic of seedlings, recently. Because the production of these proteins are initiated prematurely in the mutant embryos. exploits the genetic information possessed and Interestingly, some aspects of the maturation and expressed by cells of the embryo, some attention
Genetic and Molecular Analysis of Embryogenesis
has been devoted to cloning the genes involved and identifying their protein products. (i) Lectin Gene Expression in Embryos of Cereals and Grasses
One of the most extensively studied defenserelated proteins in embryos is lectin. Although a role for lectins in the defense against bacterial and fungal infection of plants has been mired in controversy, the bulk of the evidence is in favor of such a role (Stinissen, Peumans and Carlier 1983; Raikhel and Lemer 1991). The importance of other proteins, such as catalase and superoxide dismutase, in the defense of embryos against stress tolerance has barely been commented upon and, consequently, the regulation of expression of their genes deserves to be investigated (Acevedo and Scandalios 1990; M. Suzuki et al 1995). Lectin of wheat embryos, known as wheat germ agglutinin (WGA), is an embryo-specific protein whose synthesis commences about midway during grain development (Triplett and Quatrano 1982; Morris et al 1985). At this point in development, WGA is first detected immunocytochemically in the epidermal cells of the radicle, root cap, and coleorhiza. In the mature embryo, WGA is also found in the epiblast and in the surface layers of the coleoptile and scutellum. At the subcellular level, it is confined to the matrix of the protein bodies, the periphery of electron-translucent regions of the cytoplasm, and the cell wall-protoplast interface (Mishkind et al 1982; Raikhel and Quatrano 1986). Thus, lectin is localized in those cells and tissues of the embryo that establish direct contact with the soil during germination and its presence in the cell wall conforms to the expectation of it as a component of the defense arsenal of the embryo. Molecular studies of expression of a cDNA clone (WGA-B) encoding WGA isolectin B in many respects supports this view of a rather restricted temporal and spatial presence of lectin. Northern blot hybridization revealed that WGA-B transcripts increase significantly in the embryos between 10 and 40 days after anthesis and decrease markedly in mature embryos. The major sites of persistent accumulation of WGA-B transcripts are the epidermal layers of the radicle and coleorhiza of developing embryos (Figure 14.16). The expression patterns of WGA-B mRNA and WGA have much in common because both are present in the same cell types of the embryos (Raikhel and Wilkins 1987; Raikhel, Bednarek and Wilkins 1988). The expres-
435 sion pattern of barley lectin mRNA transcripts in the outermost layer of cells of the embryonic root tip and coleorhiza is similar to that for lectin accumulation (Mishkind et al 1983; Lerner and Raikhel 1989). Posttranslational modifications play a key role in the processing of rice lectin from a molecular mass of 36 kDa into small polypeptides of molecular masses 18,10, and 8 kDa. The large molecule is synthesized on the rough ER whence it is transported to the site of accumulation. It is believed that the processing to the mature form occurs outside the site of synthesis (Stinissen, Peumans and Chrispeels 1984). A surprising observation made with regard to rice lectin gene is that its expression is governed by two mRNA transcripts (0.9 kb and 1.1 kb). Both genes begin to function at high levels
Figure 14.16. Autoradiographic localization of transcripts of WCA-B in the embryo of dry grain of Triticum aestivum and immunolocalization of WGA in the embryo of a 40day-old caryopsis. (a) Localization of mRNA in the epidermal layer of the radicle; dark-field photograph. (b) Localization of WGA in the epidermal cells of the radicle and coleorhiza; Nomarski optics photograph. C, coleorhiza; R, radicle. Scale bar = 100 pm. (From Raikhel, Bednarek and Wilkins 1988. Planta 176:406-414. © Springer-Verlag, Berlin.)
436
early during embryogenesis in a tissue-specific manner and decline slightly at embryo maturation. A localized pattern of expression of the transcripts confined to the cells of the root cap, to the outer cell layers of the coleorhiza, radicle, and scutellum, and to all of the coleoptile is first observed in embryos as early as five days after anthesis. What fraction of the lectin expression can be attributed to each mRNA during embryogenesis? Data from gel-blot analysis of RNA isolated from embryos of different stages of development show that whereas both lectin mRNAs are present in high levels in developing embryos, the 1.1-kb species is the more prevalent one in mature embryos (Wilkins and Raikhel 1989). The selective accumulation of lectins in maturing embryos in a manner analogous to LEA proteins supports the notion that lectin gene expression is controlled by ABA in a developmental context. Additional support for this has come from the demonstration that exogenous ABA enhances the synthesis and tissue-specific accumulation of WGA and its mRNA in immature wheat embryos (Triplett and Quatrano 1982; Raikhel and Quatrano 1986; Mansfield and Raikhel 1990). As revealed by nuclear run-off transcription assay, the distinguishing feature of the action of ABA is the transcriptional induction of lectin steady-state mRNA levels (Mansfield and Raikhel 1990).
Zygotic Embryogenesis
(Riggs, Voelker and Chrispeels 1989; Riggs et al 1989). The latter approach has identified positive regulatory elements between -550 bp and -125 bp in the upstream region of the DLEC-2 gene that lead to enhanced levels of the gene product. Since gel retardation assays have indicated that this region contains nuclear protein binding sites, it is possible that the cellular machinery of the tobacco embryo recognizes the DLEC-2 gene and modulates its expression by protein-DNA interactions (Riggs, Voelker and Chrispeels 1989). Two other products of the lectin gene are an aamylase inhibitor purified from cotyledons of P. vulgaris (Moreno and Chrispeels 1989) and arcelin, found in certain wild relatives of P. vulgaris (Osborn et al 1988; Anthony, Vonder Haar and Hall 1991). Additional insight into PHA formation has been derived from the observation that in P. vulgaris and P. coccineus, PHA genes are expressed abundantly at the mRNA level in the cotyledons during seed maturation and at very low levels in the embryo axis (Hoffman, Ma and Barker 1982; Chrispeels, Vitale and Staswick 1984; Voss, Schumann and Nagl 1992). Biochemical and electron microscopic characterization has provided a nearly complete, but complex, picture of the synthesis, processing, and accumulation of PHA. The protein is synthesized exclusively on polysomes attached to the ER; biosynthesis is complete with the cotranslational glycosylation of the protein within the lumen of the (ii) Expression of Defense-related Genes in ER. The Golgi apparatus mediates in the transport Embryos of Other Plants of PHA from the ER to the vacuole for storage; this The initial diversification of organs in develop- transport involves dense vesicles with an electroning embryos of legumes is accompanied by the syn- dense matrix (Bollini and Chrispeels 1978; thesis of storage proteins and lectin in the protein Chrispeels 1983). Immunocytochemical localizabodies of cells of the cotyledons. In Phaseolus vul- tion has shown that PHA is associated with the garis, the storage protein phaseolin accounts for protein bodies of the cotyledonary cells, although about 50% of the total embryo protein, whereas the in the cells of the embryo axis the same protein is lectin PHA accounts for 5%-10% of the total pro- found floating in the cytoplasm. This is surprising teins. PHAs are tetrameric proteins composed of since the biosynthetic steps in the formation of the two different subunits known as PHA-E (erythro- mature protein body are the same in the cotyledons cyte-agglutinating) and PHA-L (lymphocyte agglu- and the embryo axis (Manen and Pusztai 1982; tinating), which are encoded respectively by two Chrispeels, Vitale and Staswick 1984). tandemly arrayed genes, DLEC-1 and DLEC-2. The fact that constructs of the DLEC-2 gene result These genes are intronless and their coding regions in the developmentally regulated expression of the exhibit 90% homology (Hoffman an Donaldson protein in embryos of transgenic tobacco seeds evi1985). The presence of regulatory factors in trans- dently shows that the gene product is correctly glygenic tobacco that recognize the promoter of DLEC- cosylated, assembled into tetramers, and targeted to 2 for the proper spatial and temporal expression of the vacuole-based protein bodies; this has been conthe gene has been established by using the reporter firmed by isolation and biochemical analysis of progene luciferase to assay for the activity of the pro- tein bodies from transgenic tobacco seeds (Sturm et moter and by deletion analysis with respect to al 1988). To test the possible role of glycan side expression in the seeds of transgenic tobacco chains in the targeting of proteins, one or both of the
Genetic and Molecular Analysis of Embryogenesis
glycosylation sites were disrupted by site-directed mutagenesis. When tobacco plants were transformed with the mutated gene, the protein was correctly targeted and localized in the protein body of the embryo as with the unmodified gene (Voelker, Herman and Chrispeels 1989). Thus, the immediate function of glycans does not appear to be one of targeting the signal of PHA to the vacuole, although other functions might be unearthed later. Like PHA, soybean lectin is also localized in protein bodies of the cotyledonary cells (Vodkin and Raikhel 1986). The nucleotide sequence of the gene encoding soybean seed lectin (JLE-1) shows that the gene is devoid of introns and directs the synthesis of a 1.1-kb mRNA. Gene transcripts are most prevalent at the midmaturation stage of the embryo, representing about 0.75% of its total mRNA mass, and are present only in insignificant amounts in late maturation stage embryo. This is similar to the regulation of other seed protein genes of soybean that are transcriptionally inactive or weakly transcribed in the maturation stage embryos (Goldberg, Hoschek, and Vodkin 1983; Vodkin, Rhodes and Goldberg 1983; Walling, Drews and Goldberg 1986). When the soybean lectin gene is introduced into the tobacco genome, lectin transcripts are found to show the same type of quantitative fluctuation in transformed seeds as in developing soybean embryos (Okamuro, Jofuku and Goldberg 1986; Lindstrom et al 1990). In any event, it should not seem peculiar that these genes no longer transcribe efficiently in the embryos of normal or transgenic plants after the cells are filled with all the gene products they can hold. Additional insight into the transcriptional apparatus that regulates lectin gene expression during soybean embryogenesis has been derived from experiments on lectin DNA-binding activity of proteins extracted from nuclei isolated from embryos of different stages of development (Figure 14.17). From the outset, a high degree of correlation was observed between lectin gene transcription and the presence of lectin DNA-binding activity. Thus, embryo stages showing a high degree of lectin gene transcription have high DNA-binding protein activity; in embryos with low levels of lectin gene expression, the protein activity is reduced accordingly. In fact, the lectin gene binds to a 60-kDa nuclear protein that exhibits a developmentally regulated program paralleling the lectin gene transcription rate. This protein is able to bind to a fragment of lectin gene sequences -77 bp to -217 bp relative to the transcription start site (Jofuku, Okamuro and Goldberg 1987). It follows that the
437
binding of the lectin gene to the nuclear protein may be among the parameters that determine its highly regulated expression during embryogenesis; a new basic understanding of the regulation of lectin gene expression during embryogenesis could come from the identification of this protein. The lectin gene in pea embryos is encoded by a small multigene family but, due to the presence of many stop codons, only one gene is considered functional (Gatehouse et al 1987; Kaminski, Buffard and Strosberg 1987). The lectin is localized in the protein bodies of the cotyledons and embryo axis (van Driessche et al 1981). De Pater et al (1993) showed that nuclear proteins from leaves and embryos of pea bind to a small fragment of the promoter of the functional lectin gene containing three overlapping TGAC-like motifs. Introduction of constructs containing this fragment into tobacco confers a high level of expression in transgenic embryos and, to a lesser degree, in the endosperm. These results show that embryo-specific expression of the lectin gene in pea can be identified with a group of specific nucleotides. Proteinase inhibitors are widely distributed in angiosperms, including legumes, in which they are localized in the embryos. These compounds defy attempts to define an accurate function during embryogenesis, but are apparently effective in roles as widely disparate as storage proteins, regulators of endogenous proteinases, and factors that confer disease resistance. KTI is a common proteinase inhibitor found in the protein bodies of cotyledonary cells of soybean embryos in association with storage proteins and lectins. It is encoded by a multigene family consisting of at least 10 members. Three KTI genes characterized (KTI-1, KT1-2, and KTI-3) and sequenced are intronless and encode distinct proteins, of which the one encoded by KTI-3 is identical to the major KTI found in embryos. General features of the expression programs of these three genes have been extensively investigated and the current picture can be summarized as follows. All three genes follow the same temporal profiles of expression in soybean embryos and in seeds of transformed tobacco plants, but their mRNAs accumulate to different levels during embryogenesis and seed formation. KTI-3 mRNA is the most prevalent and is followed by KTI-1 and KTI-2 in abundance. Other differences delineated by in situ localization, such as the greater abundance of KTI3 mRNA compared to KTI-1 and KTI-2 transcripts in the embryo axis, the appearance of KTI-3 transcripts in the globular and heart-shaped embryos
438
Zygotic Embryogenesis
0 + 0
+
o + o
+
0
+
o
+
o + o
Binding Activity -522 to +212 -217 to+212 - 77 to+212 +130 to +2)2 -522 to -334 -305 to -2)7 -2)4 to - 4 4
+ +
- 44 to + 64 Figure 14.17. Interaction of soybean lectin DNA sequences with nuclear proteins, (a) Gel retardation showing the interaction of embryo nuclear protein with the lectin 5'-deletions. Reactions 1-4 used probes 1-4 shown schematically in (c). (b) Gel retardation showing the interaction of embryo nuclear protein with 5' lectin gene fragments. Regions represented by probes 5-8 are shown in (c). In both (a) and (b): 0, without nuclear protein; +, with nuclear
protein, (c) Localization of the DNA-protein binding region. A schematic representation of the lectin gene is shown. The 5'- and 3'-probe termini are designated relative to the transcription start site. Summary of the binding activity from the gel retardation assays is represented by + or - . (From Jofuku, Okamuro and Goldberg 1987. Reprinted with permission from Nature 328:734-737. © Macmillan Magazines Ltd.)
and the absence of KTI-1 and KTI-2 transcripts in to midmaturation stage embryos. However, MA these embryos, and the generally distinct spatial transcripts are found mainly in the cells of the patterns of the accumulation of transcripts of all embryo axis, and those of MB are preferentially three genes in the cells of the cotyledons, seem to expressed in the cotyledons (Xue, Pihlgren and highlight the qualitative aspects of gene expres- Rask 1993). To place these observations in context, sion (Vodkin and Raikhel 1986; Jofuku and it may be noted that myrosinase is detected only in Goldberg 1989; Perez-Grau and Goldberg 1989). scattered cells of the cotyledons and embryo axis Another KTI is the 21-kDa albumin found in during later stages of embryogenesis in Brassica embryos of Theobroma cacao; this protein is local- napus (Thangstad et al 1990; Hoglund et al 1991; ized in membrane-enclosed organelles of the Falk, Ek and Rask 1995). cotyledonary cells (Spencer and Hodge 1991). Analysis of defense-related genes of embryos 7. GENERAL COMMENTS has not been extended very much beyond a few species of cereals and legumes. That such genes are The generalization emerging from the account active in the embryos of other plants is indicated by presented in this chapter is that virtually all phases the temporal, cell-specific, and tissue-preferential of embryogenesis are under genetic control. In the expression of two myrosinase genes (MA and MB) past, approaches like wide hybridization, radiation during embryogenesis in Sinapis alba. Myrosinases treatment of embryos, and the analysis of spontaare thioglucosides of potential importance in the neously occurring embryo mutants and patterns of interaction between plants and pathogens. nucleic acid and protein synthesis in developing Accumulation of transcripts of both genes is regu- embryos, have offered promising designs for studylated in a similar temporal pattern and is restricted ing gene activity during embryogenesis. The effec-
Genetic and Molecular Analysis of Embryogenesis
tive use of these methods has provided the necessary background information to apply gene cloning and mutant isolation techniques to the analysis of the molecular biology of embryogenesis. Despite their morphological simplicity, embryos are complex in molecular terms. The investigations described here clearly show that embryos express an enormously diverse array of genes at defined stages of development. It must be borne in mind that most of these genes code for the abundant proteins for housekeeping functions and are generally identified by screening cDNA libraries. In contrast, morphogenetic conversions of cellular phenotypes at different stages of embryogenesis are obviously under the control of regulatory genes whose detection is difficult by screening cDNA libraries. On this premise, the emergence of sophisticated mutational analysis has helped to identify a few genes that affect the basic body plan and the formation of specific organs of the embryo. Thus, when more information about the proteins encoded by the genes becomes available, the functions of structural and regulatory genes can be considered together in
439 a broader view of embryogenesis. The problems posed in the control of embryogenesis by structural and regulatory genes are old, but the insights they will open up are likely to be new. It remains to ask how transcriptional activation processes regulate developmental expression programs during embryogenesis. Current molecular models call for interactions between trans-acting proteins and as-acting regulatory sequences as the first step in the cascade of events. The information derived from the identification of promoter sequences of the genes and from the characterization of the nuclear proteins that interact with the promoters during embryogenesis, although limited, has broadened our concepts of the regulation of progressive embryogenesis and holds real promise of a breakthrough in molecular embryogenesis. It is too early to say whether common themes to explain transcriptional activation of the wide array of genes expressed during embryogenesis will emerge from future studies, or whether a diversity of mechanisms will be concealed behind seemingly simple promoter sequences.
Chapter 15
Storage Protein Synthesis in Developing Embryos Cytology of Reserve Deposition (i) Origin of Protein Bodies (ii) Origin of Lipids Regulation of Storage Protein Synthesis (i) Developmental Changes in Storage Protein Synthesis and Accumulation (ii) Regulation of Storage Protein mRNAs (iii) Modulation of Storage Protein Synthesis by Abscisic Acid Isolation and Characterization of Storage Protein Genes (i) Phaseolin Genes (ii) Glycinin and Conglycinin Genes (iii) Legumin, Vicilin, and Convicilin Genes (iv) Cruciferin and Napin Genes (v) Storage Protein Genes of Other Plants (vi) Genes for Enzymes of Lipid Bodies and for Oleosins 4. General Comments
As discussed in Chapter 14, the embryo responds to changing genetic and physiological pressures by initiating a series of developmental and molecular changes. These are reflected in the abrupt switch from a period of cell divisions to one of cell expansion, desiccation of cells, changes in growth hormone levels, and synthesis and accumulation of defense-related proteins and storage compounds. Storage materials are constituted primarily of an acervate complex of proteins known as storage proteins, in addition to starch and lipids. Like the endosperm storage products discussed in Chapter 12, embryo storage compounds play an important role in seedling survival by providing the source of carbon and nitrogen skeletons to the embryo during seed germination. Many aspects of the synthesis of storage pro440
teins during embryogenesis have been investigated using embryos of various agronomically important plants, such as bean, pea, soybean, and rapeseed. What these embryos have in common is that they represent cases in which cellular metabolism of the cotyledons is deflected from a programmed synthesis of housekeeping proteins to one concerned with the production of storage proteins. In recent years, considerable progress has been made toward understanding the regulation of the expression of storage protein genes in plant embryos. Progress was made possible not only by the desire to decipher the often complex biosynthetic mechanisms of the accumulation of embryo storage proteins that constitute a significant part of our diet, but also by the hope that the isolation and characterization of storage protein genes would provide clues to genetically engineer their synthesis in other plants. This chapter will explore the present state of knowledge on the cytology of storage protein accumulation and the biochemistry and molecular biology of storage protein synthesis in developing embryos. General accounts of the molecular control of storage protein synthesis during seed development are found in the reviews by Larkins (1981), Brown, Ersland and Hall (1982), Higgins (1984), Miintz (1987), and Shotwell and Larkins (1989), and in the volumes by Shannon and Chrispeels (1986) and Bewley and Black (1994); reviews by Casey, Domoney and Ellis (1986) and Gatehouse et al (1986) are addressed to the problems of gene expression during storage protein synthesis in seeds of legumes. A useful review covering the state of embryo storage protein research before the recent flurry of gene cloning work is by Millerd (1975).
Storage Protein Synthesis in Developing Embryos
441
supplies of reserve proteins are stored in the parenchyma cells of the cotyledons in the form of The major embryo storage proteins fall into the large protein bodies. The protein body consists of globulin and albumin groups. The two components an amorphous protein matrix bounded by two of globulins, which differ in molecular mass and close, nearly parallel electron-dense phospholipid sedimentation coefficients, are the 7S and IIS pro- layers of a limiting unit membrane. Lipid or oil teins. Both are holoproteins and, like endosperm bodies found in the cells of embryos storing fat storage proteins, are sometimes given names contain a matrix of triglycerides surrounded by a derived from the family, genus, or species from unit or by a half-unit membrane of one phosphowhich they were first described. Thus, names such lipid layer. Carbohydrates are generally stored as as vicilin, convicilin, and legumin (from Vicia faba, free starch grains in the cytoplasm or in the plasPisum sativum); conglycinin and glycinin (from tids. Considerable anatomical and ultrastructural Glycine max); phaseolin (from Phaseolus vulgaris); information has been published that supports these cruciferin (from Brassica napus); alfin and medica- general statements about the final form of the gin (from Medicago sativa); canavalin and the lectin reserve bodies in the cells of embryos of various con-A (from Canavalia sp., Fabaceae); and angiosperms. helianthinin (from Helianthus annuus) are in comHowever, there is some controversy as to how mon use. A comparison of the coding nucleotide the protein and lipid bodies are formed and how and amino acid sequences of several globulin stor- they are released from the site of their deposition. age proteins of angiosperm embryos has revealed Common to most plants examined is the initial that they have vestigial sequence homology to deposition of the storage protein body in the ceneither the 7S or IIS legume storage proteins; this tral vacuole of the cells of the cotyledons. The prohas led to the view that as a group, globulin storage tein body may increase in size to fill most of the proteins have evolved from two genes that existed vacuole. Alternatively, numerous protein bodies at the beginning of angiosperm evolution (Borroto appear in the cells later in embryogenesis, probably and Dure 1987). as the large vacuole becomes chopped up into Although albumins were long considered as smaller ones by cytoplasmic strands (Harris and metabolic proteins, they have now been upgraded Boulter 1976; Wenzel et al 1993). The enormity of to storage proteins because of their wide distribu- this change has been quantified in the developing tion, especially in embryos of oil seeds, and their cotyledons of Pisum sativum. About 8 days after amino acid composition with a high amide content. flowering, coincident with the appearance of proAlbumins generally migrate with a 2S sedimenta- teinaceous materials, cells of the cotyledon contain tion coefficient and are different from globulins in one or two vacuoles of an average diameter of 39 having a high cysteine content (Youle and Huang ^m. During the next 12 days, the average vacuole 1981). As will become evident later in this chapter, diameter falls to 1 fim, with a concomitant increase napin, an albumin from embryos of B. napus, has to 175,000 per cell in the number of vacuoles been studied extensively at the molecular level. enclosing protein bodies. The reality of a massive Ferns are another group of vascular plants that con- deposition of protein bodies is also underscored by tain albuminlike storage proteins. A 2S storage pro- the observation that during embryogenesis the survalue tein designated as matteuccin has been face area of 2vacuoles increases from an initial 2 characterized from haploid spores of the fern of 5,500 fim to a final value of 550,000 f/m (Craig, Matteuccia struthiopteris and shown to be closely Goodchild and Hardham 1979). The flow of events related to napin and a 2S albumin from sunflower connected with protein body formation described embryos (Templeman, DeMaggio and Stetler 1987; in P. sativum can be considered as representative of Templeman, Stein and DeMaggio 1988; Rodin and other plants. Rask 1990). The first indication of storage protein synthesis is the appearance of small clumps of dense staining material on the inner surface of the vacuolar mem(i) Origin of Protein Bodies brane. Later, this material becomes diffuse and Modern investigations into the cytology of the almost fills the entire vacuole; eventually it deposition of storage reserves in the cells of devel- assumes a dense form as the protein body. An oping embryos began with the advent of electron increase in protein body number might also occur microscopy. Initial results showed that in the by budding from the vacuole (Craig, Goodchild embryos of legumes and other plants, abundant and Hardham 1979; Craig, Millerd and Goodchild 1. CYTOLOGY OF RESERVE DEPOSITION
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Zygotic Embryogenesis
1980; Hinz et al 1995). A dual origin of protein bod- transport, or once the protein is in the ER lumen, ies has been described in developing cotyledons of the signal sequence is cleaved off. Cotranslational Glydne max. During the early stages of cotyledon glycosylation by the addition of mannose-rich development, protein bodies directly aggregate in sugar residues is another intrinsic modification the vacuoles from precursor proteins, whereas in that changes the structure of the protein molecules maturing cotyledons the lumen of rough ER is before their transport to the site of storage. The filled with proteinaceous material. Subsequently, occurrence of glycosylation has been supported by this is transformed into a large number of small experiments in which incubation of cotyledons protein bodies. It appears that irrespective of with the glycosylation inhibitor tunicamycin was whether they are fabricated in the vacuole or in the found to cause a decrease in glycosylated polypeplumen of the ER, no further transport of protein tides (Badenoch-Jones et al 1981; Bollini, Vitale and bodies is involved (Zheng et al 1990, 1992). Chrispeels 1983). The presence of unglycosylated Although in some plants both ER and Golgi bodies translation products in Xenopus laevis eggs injected have been implicated in the synthesis of storage with phaseolin mRNA and tunicamycin also proteins, the prevalent view is that they are synthe- argues for the involvement of a glycosylation step sized on polysomes bound to the ER and trans- in the final assembly of the storage protein ferred to the vacuole by the Golgi system. (Matthews, Brown and Hall 1981). Another line of Supporting a role for ER in reserve protein synthe- evidence that leads to the view of glycosylation as sis, close correlations have been observed between a general feature of embryo storage protein synthethe proliferation of rough ER and the onset of stor- sis is the presence of glycosylation enzymes such as age protein accumulation in the cotyledons of glycosyltransferases in particulate fractions of the Crambe abyssinica (Brassicaceae) (Smith 1974), Viciaembryos of pea (Nagahashi and Beevers 1978), soyfaba (Neumann and Weber 1978; Harris 1979; Adler bean (Bailey et al 1980), and bean (Davies and and Muntz 1983), Lupinus albus (Davey and van Delmer 1981; Chrispeels 1985), among others. Staden 1978), Phaseolus vulgaris (Briarty 1980), Glycosylation may be an important clue to the Sinapis alba (Rest and Vaughan 1972; Bergfeld, function of the protein or to its stabilization during Kiihnl and Schopfer 1980), Arabidopsis (Mansfield desiccation of the embryo in the seed, although at and Briarty 1992), and Picea glauca (Krasowski and present we do not know the full meaning of this Owens 1993). In legumes such as P. vulgaris step. (Baumgartner, Tokuyasu and Chrispeels 1980; The synthesis of storage proteins in the ER or Greenwood and Chrispeels 1985b), V. faba (Bailey, other organelles naturally raises the question as to Cobb and Boulter 1970; zur Nieden et al 1982), and how the proteins are transported to the vacuoles G. max (Nishizawa et al 1994), the structural analy- for storage. As in the case of lectins, discussed in sis has been combined with immunocytochemical Chapter 14, one idea that has good experimental and autoradiographic techniques to support the support is that the relocation of storage proteins origin of storage proteins in or close to the ER. from the ER to the protein bodies is mediated by More direct evidence linking rough ER with stor- the Golgi apparatus. The general experimental age protein synthesis has come from studies show- strategies upon which a role for the Golgi apparaing that the main components of storage proteins of tus is based are electron microscopy to demonstrate P. vulgaris (Bollini and Chrispeels 1979), Pisum the presence of electron-dense vesicles in close sativum (Chrispeels et al 1982), and Cucurbita sp. proximity to the Golgi complex and protein bodies, (Hara-Nishimura, Nishimura and Akazawa 1985) as in developing embryos of Capsella bursa-pastoris, are made exclusively on membrane-bound Arachis hypogaea (peanut; Fabaceae) (Dieckert and polysomes that can be recovered from rough ER by Dieckert 1976), and Fagus sylvatica (Fagaceae) sucrose-gradient centrifugation. There is evidence (Collada et al 1993); high-resolution electron microto show that in developing cotyledons of P. vul- scopic immunochemistry to demonstrate the presgaris, the newly synthesized phaseolin is tran- ence of storage proteins in the Golgi apparatus and siently stored in the lumen of the ER before it associated transport vesicles, as in Phaseolus vulcommences its journey to the protein bodies garis (Greenwood and Chrispeels 1985b), Pisum (Bollini, van der Wilden and Chrispeels 1982). sativum (Craig and Goodchild 1984a), Viciafaba (zur Storage proteins thus far characterized are asso- Nieden et al 1984), and Glydne max (Nishizawa et ciated with a signal sequence that is necessary for al 1994); and biochemical techniques to show the targeting proteins into the lumen of the ER as the presence in the Golgi complex of specific enzymes first step in intracellular transport. During protein for the glycosylation of storage proteins, as in P.
Storage Protein Synthesis in Developing Embryos
vulgaris (Chrispeels 1985). Craig and Goodchild (1984b) have shown that the ionophores monensin and nigericin interfere with the transport of newly synthesized vicilin in the cotyledons of P. sativum, causing its buildup in the Golgi apparatus and its diversion from the vacuole tonoplast to the plasmalemma. From these observations and by analogy to protein secretion in animal systems, a similar mechanism, involving the Golgi bodies, might be envisaged to account for protein transport in developing plant embryos. Although the transport of lectins and storage proteins using the Golgi system suggests common pathways, only further work will reveal whether the behaviors of the two kinds of molecules have distinctive features. The complete sequence of events by which the storage protein becomes visible in the cells of the mature embryo involves several changes that are not detected cytologically. Nonetheless, they have been analyzed at the molecular level, so that a model of the assembly of the protein has emerged. Soon after glycosylation, both IIS and 7S precursors are forged into trimers within the ER and transported to the protein bodies. A further change in the molecule occurs at this stage by the addition or deletion of sugar residues from the oligosaccharide side chains. In the vacuole, the trimer is proteolytically cleaved, which leads to the separation of the acidic and basic subunits. In general, the US trimers are fashioned into hexamers whereas the 7S protein is assembled as a trimer, and both appear as spherical structures enveloped by a single membrane (Sengupta et al 1981; Dickinson, Hussein and Nielsen 1989). (ii) Origin of Lipids
Based on the close association of nascent lipid or oil bodies in developing embryos with ER (FreyWyssling, Grieshaber and Miihlethaler 1963), plastids (Bergfeld et al 1978), or both (Mansfield and Briarty 1992), current views on the ontogeny of lipid bodies are that precursors for their synthesis are provided either by one or by both of these organelles. However, it is doubtful whether these organelles are the site of synthesis of lipids; it appears more likely that during storage lipid formation, oil bodies are secreted as small, electrondense, particulate masses into the cytoplasm, where they fuse to form larger oil bodies (Smith 1974; Herman 1987; Murphy, Cummins and Kang 1989). Because of the presence of the proteinaceous membrane (or oleosin), oil bodies that store triglycerides are different from the lipid or oil bodies referred to
443 previously (Herman 1987; Murphy and Cummins 1989a; Tzen et al 1990). Interestingly enough, like storage proteins, oleosins appear to be synthesized in the ER, and a conformational change presumably accounts for their integration as a membrane around the oil body (Loer and Herman 1993). The fairly constant association between oleosins and phospholipids of the unit membrane surrounding the triglyceride-containing oil bodies implies a role for oleosins in stabilizing the oil bodies as discrete organelles and in safeguarding them from coalescence (Tzen and Huang 1992). 2. REGULATION OF STORAGE PROTEIN SYNTHESIS
The regulation of storage protein synthesis can be viewed as part of an overall process controlled by gene action. At a gross level, the effects of gene action are seen in the timing and kinetics of storage protein accumulation in different genotypes of bean (Mutschler, Bliss and Hall 1980), and in the variations in protein characteristics, including the electrophoretic patterns of legumin and vicilin in different varieties of pea (Thomson and Schroeder 1978; Davies 1980; Schroeder 1982). Molecular approaches necessary to develop a conceptual understanding of the varietal and genotypic effects on storage protein synthesis and accumulation have not, however, been developed. This section of the chapter will discuss the temporal and spatial patterns of protein deposition and synthesis in developing embryos and the expression of their genes. The multisubunit combinations of storage proteins have offered a rich source of materials for such studies. (i) Developmental Changes in Storage Protein Synthesis and Accumulation
Deposition of storage proteins in developing embryos has been followed quantitatively by the incorporation of radioactive precursors, and qualitatively by SDS-PAGE analysis of accumulated polypeptides and by immunochemical methods. From various data it appears that developing cotyledons of Pisum sativum accumulate US legumin and 7S vicilin to a considerable extent, with the synthesis of vicilin generally preceding that of legumin. In terms of the timing, legumin accumulates throughout embryogenesis, whereas several periods of intensive accumulation of vicilin are spread over the whole time course. It is also found that, in contrast to vicilin subunits, which are pre-
444
Zygotic Embryogenesis
^-urease — 7S «' -7S B - I I S A-4 - I IS a c i d i c s
-24K protein
• §••• E
F
G
H
I
3
4
5
6
7
11
13
15
17
18
19
K 9 ao
- I I S basics
M O De - S t a g e n 12 14 - Seed length (mm) 22 >28 >so - Age (days)
Figure 15.1. Changes in the extant storage protein profiles of soybean embryos of different stages of development. Samples E-l and J-De were prepared from whole seeds and from isolated cotyledons, respectively. Stages of seed or
embryo development are indicated in the lower part of the figure. (From Meinke, Chen and Beachy 1981. Planta 153:130-139. © Springer-Verlag, Berlin. Photograph supplied by Dr. D. W. Meinke.)
sent in appreciable amounts at the early stages of embryogenesis, legumin and a 7S-type convicilin are prominent later in embryogenesis. Another apparent change in protein accumulation pattern includes a depression in the synthesis of all major storage polypeptides in the cotyledons with maturity of embryos (Millerd and Spencer 1974; Guldager 1978; Millerd, Thomson and Schroeder 1978; Gatehouse et al 1982; Chandler et al 1984). Several 2S albumins are also major components of the cotyledons and embryo axes of pea. Like globulins, they are broken down during germination, suggesting that they have a storage function (Higgins et al 1986). Vicilin is formed prior to legumin in developing embryos of Viciafaba; however, starting in a modest way, legumin synthesis overtakes vicilin production, thereby accounting for the ability of cotyledons to accumulate nearly four times more legumin than vicilin. Most legumin accumulation occurs during the period of cell elongation between embryo lengths of 10 and 25 mm, and embryos less than 4 mm do not contain any legumin at all. Fractionation of legumin and vicilin into component polypeptides showed that they consist of three and four subunits, respectively. The subunit composition of vicilin is found to change during embryogenesis, whereas that of legumin remains constant.
This suggests that the synthesis of individual subunits is under separate genetic control (Graham and Gunning 1970; Millerd, Simon and Stern 1971; Wright and Boulter 1972; Bassuner et al 1983). Both temporal and rate differences are apparent in the accumulation of three major fractions of seed proteins and their subunits during soybean embryogenesis. The bulk of the storage proteins, accounted for by 7S conglycinin and US glycinin, are synthesized late during embryo development; a minor 2S component predominates at very early stages, decreasing proportionately throughout embryogenesis. Two a-subunits of conglycinin and most of the acidic and basic subunits of glycinin begin to appear shortly after the termination of cell divisions in the developing cotyledons, whereas (3-subunits of conglycinin and a cluster of acidic subunits of glycinin begin to accumulate still later, coincident with the maturation of the cotyledons (Figure 15.1). An interesting inference about conglycinin accumulation is that the turnover of this protein is very rapid during the phase of active cell divisions in the cotyledons and that the protein becomes stable later in development. The embryo axis, unlike the cotyledons, is not committed to the accumulation of massive amounts of storage protein subunits and, due to rapid degradation, the P-subunit of conglycinin is virtually undetectable
Storage Protein Synthesis in Developing Embryos
445
in the embryo axis. The development of the embryo storage protein deposition; subsequently, it levels axis and cotyledons entails other changes, which off during the desiccation phase of embryos. The result in the former's containing different isomers synthesis of a 19-kDa oleosin associated with of conglycinin from those found in the cotyledons. triglycerides is initiated late in embryogenesis, In addition, the spatial pattern of accumulation of after the synthesis of cruciferin and napin is under storage proteins is somewhat selective, with higher way (Figure 15.2). The deposition of oleosin overlevels of glycinin and conglycinin being present in laps with the period of maximum storage protein the abaxial, as compared to the adaxial, part of the accumulation, but is independent of the synthesis cotyledons (Hill and Breidenbach 1974; Gayler and of triglycerides (Murphy and Cummins 1989a; Sykes 1981; Meinke, Chen and Beachy 1981; Murphy, Cummins and Kang 1989). The time Sengupta et al 1981; Shuttuck-Eidens and Beachy course of storage lipid synthesis in maize embryos 1985; Ladin et al 1987; Sugimoto et al 1987; Tang et closely parallels increases in embryo fresh weight, al 1993). Synthesis of storage proteins slightly although 90%-95% of the lipids and lipid bodies increases during heat-shock treatment of develop- are localized in the scutellum, the remainder being ing soybean embryos; no explanation is known for confined to the embryo axis. It has been claimed this phenomenon, other than that HSPs and stor- that embryos of maize, rapeseed, and certain other age proteins share common control mechanisms plants begin to accumulate triglycerides and oleosins concomitantly during the initial stages of (Altschuler and Mascarenhas 1982). Qualitative and quantitative developmental development; this observation is in agreement with changes in the synthesis and accumulation of stor- a model that implicates ER as the site of their synage proteins also occur during embryogenesis in Zea mays (Khavkin et al 1977, 1978; Cross and Adams 1983; Sanchez-Martinez et al 1987; Kriz and Schwartz 1986; Kriz 1989), Vigna unguiculata (Carasco et al 1978; Khan, Gatehouse and Boulter 1980), Phaseolus vulgaris (Hall, McLeester and Bliss 1972; Sun et al 1978; Murray and Kennard 1984), Gossypium hirsutum (Dure and Chlan 1981; Dure and Galau 1981), Brassica napus (Crouch and Sussex 1981; Murphy and Cummins 1989b; Murphy, Cummins and Ryan 1989; Hoglund et al 1992), Helianthus annuus (Allen, Nessler and Thomas 1985; This et al 1988), Triticum durum (Brinegar, Stevens and Fox 1985), Canavalia gladiata (Yamauchi and Minamikawa 1986, 1987), Sinapis alba (Fischer et al 1988), Medicago sativa (Xu et al 1991; Krochko, Pramanik and Bewley 1992), Amaranthus hypochondriacus (Amaranthaceae) (Raina and Datta 1992), and Cicer arietinum (Mandaokar et al 1993), and in the gymnosperms Pinus monticola (Gifford 1988), Picea abies (Hakman et al 1990), P. glauca (Misra and Green 1991), and P. glauca/P. engelmannii (Flinn et al 1991). Despite the inevitable variations encountered with species and with each storage protein, it can be stated in a general way that the synthesis and the accumulation of storage proteins go hand in hand beginning with the early development of embryos and tend to level off as embryos enter the maturation phase. U 6 8 10 12 Only limited information is available on the timing and pattern of accumulation of storage lipids in developing embryos. Typically, in embryos of Brassica napus, the most rapid phase of storage lipid formation occurs before the onset of
Weeks after anthesis Figure 15.2. Synthesis of storage proteins and oleosins in developing embryos of Brassica napus. D, cruciferin; A, napin; O, oleosin; • , other proteins. (From Murphy and Cummins 1989b.)
446
thesis (Qu et al 1986; Tzen et al 1993). In conifer embryos, triglycerides are found to accumulate continuously during the course of development and to account for nearly 50% of the fresh weight of the mature embryos (Feirer, Conkey and Verhagen 1989). Clearly, before a general picture of the gene expression program for storage lipids can emerge, more work is needed to document the temporal and spatial patterns of lipid accumulation in developing embryos of additional species. Lipoxygenase is an enzyme that catalyzes the oxidation of polyunsaturated fatty acids and other compounds and is abundant in seeds of leguminous plants. In pea (Domoney et al 1990) and soybean (Hildebrand, Versluys and Collins 1991), the enzyme is expressed in the embryo only during the cell expansion phase, coincident with the accumulation of storage proteins. Suggestive of gene duplication is the observation that three isozymes of lipoxygenase are coordinately expressed in the cotyledons and embryo axes of soybean. The ability of cultured cotyledons or cotyledons of cultured embryos to synthesize storage proteins is consistent with the notion that the switching on of genetic information for this activity is independent of any signal provided by the parent plant. Cotyledons excised from young embryos of pea, containing no trace of vicilin, begin to synthesize this protein in culture. However, legumin does not accumulate in excised and cultured cotyledons unless its synthesis is already under way before culture. Although a trigger provided by the pod was invoked to account for legumin synthesis in isolated cotyledons, a later work showed that cotyledons of cultured young embryos synthesize and accumulate legumin in higher quantities than cotyledons of embryos of similar fresh weight growing in vivo (Millerd et al 1975; Domoney, Davies and Casey 1980). Immature embryos of Theobroma cacao (A. J. Kononowicz and Janick 1984), Simmondsia chinensis (Buxaceae) (Wang and Janick 1986), and Brassica napus (Avjioglu and Knox 1989) cultured in vitro synthesize storage lipids in response to high concentrations of sucrose in the medium. Thus, for both storage proteins and lipids, the use of an embryo culture system is more amenable to control than embryos growing in vivo. Analysis of the DNA content of cells and storage protein accumulation in the cotyledons of cultured pea embryos has provided additional evidence showing that there is no selective amplification of the genes for storage proteins (Chapter 13). In the cotyledonary cells of cultured embryos, the maximum cellular DNA level increases, yet the mini-
Zygotic Embryogenesis
mum DNA level at which vicilin is detected remains at the 5C level, as in embryos growing in vivo. Evidence is also at hand showing that when embryos are treated with aphidicolin, an inhibitor of DNA replication, even cells of the cotyledons at the 4C DNA level synthesize vicilin (Corke, Hedley and Wang 1990a, b). From these observations, it seems likely that the cessation of mitosis, along with accompanying cell expansion, rather than elevated cellular DNA levels, accounts for the onset of vicilin synthesis in the cells of cotyledons. (ii) Regulation of Storage Protein mRNAs
Given the background of the pattern of accumulation of storage proteins in developing embryos, it is now possible to examine the relationship of the accumulation patterns to the levels of mRNA species and assess the extent of operation of transcriptional and translational controls. Changes in storage protein mRNAs of developing embryos have been assayed by cell-free translation of poly(A)-RNA, by competitive nucleic acid hybridization with labeled cDNA clones, by analysis of run-off transcripts from isolated nuclei, and by Northern blot, dot-blot, and in situ hybridization techniques. Polysome fractions or poly(A)-RNA isolated from developing embryos of Glycine max (Beachy, Thompson and Madison 1978; Beachy 1980; Beachy, Jarvis and Barton 1981; Sengupta et al 1981; Turner, Thanh and Nielsen 1981), Phaseolus vulgaris (Hall et al 1978; Sun et al 1978; Chappell and Chrispeels 1986), Pisum sativum (Evans et al 1979; Croy et al 1980a, b; Gatehouse et al 1981; Higgins and Spencer 1981; Higgins et al 1986; Domoney and Casey 1987; Yang et al 1990), Gossypium hirsutum (Dure and Galau 1981; Dure et al 1983), Viciafaba (Bassiiner et al 1983), Cucurbita sp. (Hara-Nishimura, Nishimura and Akazawa 1985), Lupinus angustifolius Qohnson, Knight and Gayler 1985), Triticum durum (Brinegar, Stevens and Fox 1985), Raphanus sativus (LarocheRaynal and Delseny 1986), Canavalia gladiata (Yamauchi and Minamikawa 1987), Brassica napus (Rodin et al 1990), and Picea glauca (Leal and Misra 1993a) have been shown to synthesize the appropriate storage proteins or their precursors when translated in a cell-free system (Figure 15.3). In some of these investigations, immunoprecipitation with antibodies combined with polypeptide mapping has led to the characterization of mRNA fractions coding for specific polypeptide subunits of storage proteins. A general conclusion from these studies is that mRNAs coding for storage proteins increase
447
Storage Protein Synthesis in Developing Embryos
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Figure 15.3. In vitro translation profiles of proteins of pea embryos of different stages of development. Arrowheads indicate the position of proteins of the different molecular
masses given on the left. Age of embryos is given at the bottom as days after flowering. (From Yang et al 1990. J. Expt. Bot. 41:283-288, by permission of Oxford University Press.)
from very low levels to become the dominant species in the cells of the cotyledons during the period of peak protein accumulation, and then they decline to low, near undetectable levels at later stages of maturity. Measurements of cDNA-RNA reassociation kinetics have shown that the decrease in storage protein synthesis during late embryogenesis in cotton cotyledons occurs simultaneously with the loss of a superabundant class of mRNAs. In this experiment, a cDNA probe made to storage protein mRNA was reassociated with mRNAs isolated from embryos of different stages of development. Results showed that mRNA sequences for the abundant storage protein decrease and are almost totally obliterated as the embryo progresses from the cell division phase to phases of cell expansion and maturation, indicating that the storage protein mRNA is active only during a transient phase of embryogenesis (Dure and Galau 1981; Dure et al 1983). In a similar way, using cloned cDNA probes made to storage protein mRNAs of soybean embryos, Goldberg et al (1981a, b) found that the appearance of complementary mRNA sequences in high frequency closely corresponds to the period of maximum storage protein accumulation and that the messages decay late in embryogenesis as dehydration sets in and embryos become mature. These superabundant messages, which number about 7-10, are probably transcribed exclusively in the
embryos, because they are undetectable in the leaf polysomes. The pattern of changing levels of storage protein mRNAs shows that the amount of protein synthesized is determined by the amount of its mRNA and suggests that storage protein synthesis in developing embryos is controlled at the transcriptional level. This idea has been pursued by extensive hybridization analyses using cloned probes made to different storage protein genes isolated from embryos of plants belonging to Fabaceae and Brassicaceae. Fabaceae. Changes in the abundance of transcripts for storage proteins during embryogenesis in pea have been well characterized by Northern and dot-blot hybridizations using cDNA clones for vicilin (Croy et al 1982), convicilin (Domoney and Casey 1983), and legumin (Chandler et al 1983; A. J. Thompson et al 1991). The different mRNAs that encode storage proteins are present at very low concentrations during early stages of embryo development. They reach their maximum at slightly different times during embryogenesis and subsequently decrease in a manner that correlates with the accumulation of their respective proteins; vicilin mRNAs, in general, accumulate more rapidly and decrease earlier than either legumin or convicilin mRNAs (Gatehouse et al 1982, 1988; Chandler et al 1984; Boulter et al 1987; Domoney
448
and Casey 1987; Bown, Ellis and Gatehouse 1988; Yang et al 1990; A. J. Thompson et al 1991). From assays for legumin and vicilin in runoff transcripts of nuclei isolated from cotyledons at different stages of embryogenesis, it is seen that the synthesis of vicilin transcripts precedes that of legumin during early embryogenesis, whereas legumin transcripts, in the virtual absence of vicilin, predominate during later stages. These results essentially confirm the changes in specific mRNA levels during embryogenesis in pea noted previously and suggest that storage protein synthesis in this system is dominated by transcription-driven controls (Evans et al 1984). This does not, however, rule out some posttranscriptional control of the level of expression of these proteins. The basis for this assertion is another work, which revealed no clear correlation between the steadystate levels of mRNAs for the different classes of legumin genes and the transcription rates of these genes in embryos of different ages. This suggested that the accumulation of vicilin and legumin mRNA in pea embryos is a two-step process in which temporal increases in transcription rates are followed by some posttranscriptional effects (Thompson et al 1989). At the cellular level, legumin and vicilin mRNAs are expressed in the storage parenchyma cells of the cotyledons and, depending upon the growing conditions of the plant, they may be further restricted to the nondividing cells. An association of legumin mRNA with the ER has been noted at the electron microscope level using a biotinylated probe complexed with avidin-ferritin (Harris and Croy 1986; Harris et al 1989; Hauxwell et al 1990).
Zygotic Embryogenesis
a
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i-Si 25 35 55 91 25 35 55 91 DAYS AFTER FLOWERING Figure 15.4. Regulation of conglycinin gene expression in developing soybean embryos, (a) A dot-blol hybridization showing differential accumulation of oc'/ct- (2.5 kb) and (3subunits (1.7 kb) of (3-conglycinin mRNAs. RNA blotted on nitrocellulose filter was hybridized with labeled sequences complementary to the two conglycinin subunits. Days after flowering are indicated below the dot-blots, (b) Transcription of a'/a- and P-conglycinin genes from nuclei isolated from soybean embryos of different ages. The relative transcription rates were quantified by hybridizing labeled nuclear RNAs with filter-bound linearized sequences complementary to the two glycinin subunits. Ranges of values are indicated by the vertical lines. (From Harada, Barker and Goldberg 1989. Plant Cell 1:415-425. © American Society of Plant Physiologists.)
embryogenesis within the temporal framework already noted (Nielsen et al 1989). In contrast, mRNAs encoding the a'/a- and (3-subunits of the conglycinin gene family accumulate and decay at In soybean, genes for glycinin and conglycinin different times during embryogenesis; compared are transcriptionally activated to peak levels dur- to the a' / a-subunit, the (3-subunit typically exhibits ing the midmaturation stage of embryogenesis and delayed accumulation and decay. When the tranare repressed in the mature embryos. Although scription rates of the a'/a- and (3-subunits of mRNAs for both genes are detected early in the conglycinin in embryos of different ages were development of the embryo, in the timing of determined, genes encoding the two subunits were mRNA appearance, conglycinin transcription pre- found to be transcribed at roughly the same relacedes that of glycinin by a few days (Goldberg et al tive rates, indicating that despite changes in the 1981a; Meinke, Chen and Beachy 1981; Scallon et al mRNA levels, the cells appear to use the templates 1985; Walling, Drews and Goldberg 1986). We do at the same rate (Figure 15.4). Thus, it has been not have a comparative picture of the cellular local- argued that embryos rely on posttranscriptional ization of glycinin and conglycinin in the develop- mechanisms in the accumulation of mRNAs for the ing embryos, but it has been shown that two conglycinin subunits (Naito, Dube and Beachy conglycinin mRNA first accumulates not only in 1988; Harada, Barker and Goldberg 1989). the cotyledons but also in the embryo axis (PerezIn developing cotyledons of Phaseolus vulgaris, Grau and Goldberg 1989). both transcriptional and posttranscriptional events Expression patterns of members of the glycinin such as mRNA stability appear to be important in and conglycinin gene families have provided some controlling gene expression for the 7S protein surprising findings. Five different glycinin genes, phaseolin. This is based on the observation that the designated GY-1, GY-2, GY-3, GY-4, and GY-5, accu-transcription rate of phaseolin is high early in mulate and decay coordinately during soybean embryo development and then declines, although
Storage Protein Synthesis in Developing Embryos
Figure 15.5. Dark-field images of autoradiographs of sections of Brassica napus embryos hybridized with 35SUTPlabeled napin antisense RNA probes, (a) Early heart-shaped embryo, (b) Late heart-shaped stage embryo. Label is seen in the cortex of the axis (ca). (c) Torpedo-shaped embryo. Label is in the axis, immediately below the incipient shoot apex (arrow) and in the outer face (arrowhead) of the cotyle-
449
dons, (d) Another torpedo-shaped embryo showing the absence of label in the provascular tissue and in a set of cells in the upper axis (above the arrow), a, axis; c, cotyledon; e, embryo; s, suspensor; v, provascular tissue. Scale bars = 100 urn. (From Fernandez, Turner and Crouch 1991. Development 111:299-313. © Company of Biologists Ltd. Photographs supplied by Dr. D. E. Fernandez.)
high mRNA levels are maintained (Chappell and embryogenesis and then declining to barely Chrispeels 1986). During embryo development in detectable levels in the mature embryos Canavalia gladiata, mRNA levels for canavalin and (Finkelstein et al 1985; DeLisle and Crouch 1989). In con-A are differently regulated, with canavalin situ localization profiles are in agreement with blot transcript expression preceding that of con-A. The hybridization data in showing temporal differences accumulation of transcripts of canavalin also in the first appearance of cruciferin and napin tranceases early in embryogenesis even as the expres- scripts in developing embryos and in showing their sion of con-A messages continues (Yamauchi et al simultaneous disappearance in mature embryos 1988). The abundance of mRNAs for the US pro- (Figure 15.5). Beginning with the detection of trantein medicagin, 7S protein alfin, and a 2S protein in scripts in the cortex of heart-shaped (napin) and developing embryos of Medicago sativa, as deter- torpedo-shaped embryos (cruciferin), transcript mined by Northern hybridization using probes for accumulation spreads to the outer and inner faces pea vicilin and legumin and a synthetic oligomer of the cotyledons until most cells and tissues of the for the 2S protein, appears to coincide with the embryo are enriched for both transcripts periods of maximum synthesis of the respective (Fernandez, Turner and Crouch 1991; Sjodahl et al proteins (Krochko, Pramanik and Bewley 1992). 1993). Comparison of the abundance of mRNAs for According to Bassiiner et al (1988), transcripts of a napin and cruciferin in developing embryos of gene that encode an unknown seed protein are Raphanus sativus has shown that napin mRNA is present during early development of embryos of synthesized a few days ahead of cruciferin message, Vicia faba with a time profile similar to that of but the latter is slow to decay in the mature embryos vicilin mRNA. However, the gene for narbonin, a (Laroche-Raynal and Delseny 1986). 2S protein from V. narbonensis, is expressed Investigations of the transcription rates of napin throughout most of the period of embryogenesis and cruciferin genes in B. napus embryos have (Nong et al 1995). highlighted the fact that cruciferin mRNA, which accumulates to a higher level, is more stable than Brassicaceae. Through the use of cDNA clones, napin mRNA during the peak periods of their accumRNA for 1.7S napin is detected by dot-blot mulation. Later in embryogenesis, stability of both hybridization in embryos of Brassica napus several mRNAs is eroded because comparable transcripdays before 12S cruciferin mRNA. Transcripts for tion rates result in reduced levels of both mRNAs both genes share similarities with glycinin and (DeLisle and Crouch 1989). Since the relative stabilconglycinin expression in soybean embryos in ity of mRNA sequences determines their abunaccumulating approximately halfway through dance in the cell, it appears that cruciferin and
450 napin genes are controlled differently at different stages of embryogenesis. There are three major subunits of cruciferin (CRU-1, CRU-2/3, and CRU-4), each encoded by a small gene family of about 5 members, whereas napin is encoded by a multigene family of at least 16 members. Regulatory trends in the study of individual members of the cruciferin family and members of the three different subunits have revealed similar developmental patterns of expression during embryogenesis of B. napus (Breen and Crouch 1992; Sjodahl et al 1993); on the other hand, the expression of members of one class of the napin gene peaks and declines earlier than expression of the other members of the family (Blundy, Blundy and Crouch 1991). The reasons for the different expression patterns for members of the napin gene family in embryos of B. napus are not presently known. It is possible that expression patterns of subunits of the same gene are genus specific, because two members of a napin gene family are coordinately expressed during embryogenesis in R. sativus (Raynal et al 1991).
Zygotic Embryogenesis
state level during the period of rapid cotyledon growth, and fall precipitously thereafter. Since no significant changes are detected in the pattern of accumulation of the three transcripts, the transcription of the genes appears to be regulated coordinately ( Dure et al 1983; Galau, Chlan and Dure 1983; Galau, Bijaisoradat and Hughes 1987). The maximum level of accumulation of mRNAs of a 12S protein helianthinin in the embryos of sunflower is observed several days before the protein level peaks (Allen, Nessler and Thomas 1985; This et al 1988). A gene that encodes a 35-kDa protein rich in essential amino acids isolated from seeds of Amaranthus hypochondriacus follows a course of
expression characterized by transcript synthesis during early embryogenesis, reaching a maximum at the midmaturation stage and decreasing during maturation (Raina and Datta 1992). During the prolonged period of development (extending to nearly six months) of embryos of Prunus amygdalus, expression of transcripts corresponding to the globulin, prunin, occurs in the cotyledons around 110 days after flowering (Garcia-Mas et al 1995, 1996). The time courses of expression of transcripts of Unlike storage proteins in the seeds of legumes and three 12S globulin storage protein genes and four other plants considered here, the storage proteins 2S albumin genes in embryos of Arabidopsis are of Bertholletia excelsa (Brazil nut; Lecythidaceae), basically invariant from one another, showing a constituted of 2S albumins, accumulate in the peak midway through embryogenesis and declin- embryonic hypocotyl, and expression of the genes ing during embryo maturation. Whereas tran- for these proteins is restricted to embryos that scripts of 12S proteins genes appear equally approach maturity (Gander et al 1991). Another abundant at different stages of embryogenesis, pattern of gene expression confined to most of the those of albumin genes AT2S-2 and AT2S-3 are embryo developmental period is displayed by expressed at significantly higher levels than are globulin genes GLB-1 and GLB-2 of maize embryos those of AT2S-1 and AT2S-4. The spatial pattern of (Belanger and Kriz 1989; Kriz and Wallace 1991). expression of AT2S-1 is unique and, unlike the Transcripts of vicilin, albumin, and a crystalloid other members of this gene family, which are storage protein are present at maximum levels in expressed both in the cotyledons and in the embryo the developing embryos of Picea glauca and P. axis, AT2S-1 is expressed mostly in the embryo axis glauca/P. engelmannii complex preceding the period (Pang, Pruitt and Meyerowitz 1988; Guerche et al of rapid accumulation of the proteins (Flinn et al 1990b). How the four closely related genes in a fam- 1993; Leal and Misra 1993a). In Pseudotsuga menily encoding the same protein are differentially reg- ziesii (Pinaceae), mRNAs corresponding to a leguulated is puzzling, although the use of different minlike storage protein begin to accumulate in the combinations of ris-acting elements to achieve this early to midstage embryos several days after the peak accumulation is observed in the megagametohas been suggested. phytic tissues and then decline during embryo maturation (Leal and Misra 1993b). Other Plants. The principal storage proteins of cotton embryos have apparent molecular masses of 48 kDa and 52 kDa and are encoded by three subStorage Lipid Transcripts. What is the relationfamilies of genes, one subfamily coding for 48-kDa ship between the rate of transcription and the proteins and two subfamilies coding for 52-kDa expression of genes of the enzymes involved in the proteins. Dot-blot hybridization with cDNA clones synthesis of oil body fatty acids and of the oleosins representing the three mRNA subsets showed that that enclose the oil bodies? In Brassica napus, a patmRNAs for the storage proteins increase rapidly in tern of a relatively early peak in transcript expresyoung embryo cotyledons, maintain a high steady- sion and the gradual loss of transcripts as embryos
451
Storage Protein Synthesis in Developing Embryos
become desiccated has been observed for genes for ACP (de Silva et al 1990), enoyl-ACP-reductase (Kater et al 1991), stearoyl-ACP-desaturase (Slocombe et al 1992), and acyl-ACP thioesterase (Loader et al 1993). Transcripts for these genes are detected before the onset of, or coincident with, the peak of storage lipid synthesis in the embryos, and their disappearance corresponds with the attainment of steady-state levels of lipids. Late transcription, apparently independent of the transcription of genes for oil body enzymes, is seen for the oleosin gene. Early stage embryos lack oleosin transcripts, which are found at only a low level until the embryo reaches its maximum fresh weight and begins to desiccate (Keddie et al 1992b). At the organ level, transcripts of stearoyl-ACP-desaturase and oleosin genes are restricted to the embryonic cotyledons and radicle. Consistent with their temporal mode of expression, transcripts of stearoylACP-desaturase come into the limelight in the embryonic organs earlier than those of oleosins and they disappear when the oleosin message level is still high (Slocombe et al 1992; Cummins et al 1993). Changes in mRNA levels of the 16-kDa and 18-kDa oleosin isoforms of maize embryo fall into a clearly recognizable category of developmental regulation as they increase during embryo development and, following a peak at midpoint of embryogenesis, decrease to low values. The close parallel noted between the rate of transcription of the 16-kDa oleosin isoform gene and the steady-state level of the corresponding mRNA in developing embryos (Figure 15.6) suggests that the transcript level of this gene is controlled at least in part by the rate of transcription (Vance and Huang 1988; Qu, Vance and Huang 1990). A similar temporal pattern of expression is seen for oleosin mRNAs from cotton embryos (Hughes and Galau 1989). The pattern of accumulation of oleosin mRNAs in developing embryos of Prunus amygdalus follows that for storage proteins, although oleosin transcripts are still detectable at late embryo developmental stages (Garcia-Mas et al 1995).
(iii) Modulation of Storage Protein Synthesis by Abscisic Acid There are several indications that the deposition of storage reserves in maturing embryos is linked to changes in their ABA content. Although the bulk of evidence for this has come from studies on cultured immature embryos, this discussion will begin by referring to a work done in vivo (Schroeder 1984). The possible intervention of exogenous hor-
20 25 30 35 40 Days after pollination Figure 15.6. Time course of transcript accumulation of the 16-kDa oleosin gene (L-3 gene) and the rate of gene transcription in developing maize embryos, (a) Densitometric scanning of a Northern blot using labeled probe for the 16kDa oleosin gene, (b) Relative levels of gene transcription determined by run-off transcription assays using nuclei isolated from embryos at the stages shown. (From Vance and Huang 1988.)
mones in storage protein synthesis in the embryos appeared to be an attractive idea that could be tested by injecting the hormones into developing pea pods. This experiment showed that the total protein content and, in particular, the vicilin content of embryos increase in response to ABA injections whereas other hormones, such as NAA and benzylaminopurine, induce increases in the legumin content. The most extensive series of experiments to establish a connection between ABA and storage protein synthesis has been done with cultured embryos of Brassica napus. The results are basically straightforward in showing that when cultured in media containing ABA or a high osmoticum secured by the addition of sucrose or mannitol, embryos at the early cotyledon stage proceed through a normal developmental program and continue to synthesize cruciferin and napin; in contrast, in the absence of ABA or high osmotic environment in the medium, cultured embryos germinate precociously and synthesize storage proteins at barely detectable levels. The effects of these
452
same treatments on the developmental regulation of storage lipid synthesis were less clear-cut (Crouch and Sussex 1981; Finkelstein and Crouch 1984; Finkelstein and Somerville 1989). Consistent with a role for ABA in stimulating cruciferin and napin synthesis, the mRNA levels for these proteins and the message transcription rates are appreciably higher in embryos cultured in ABA than in those grown in the basal medium (DeLisle and Crouch 1989). Seeds of transgenic tobacco harboring the promoter of a napin gene linked to the GUS gene also respond to ABA by showing enhanced reporter gene activity (Jiang, Abrams and Kermode 1996). However, in assigning a role to ABA in mediating gene expression for cruciferin and napin synthesis, we are confronted with two other observations. One is that the hormone could not effectively sustain the accumulation of cruciferin or its mRNA in cultured older embryos that are on the verge of desiccation. Secondly, osmotic effects on cruciferin synthesis are not associated with elevated levels of ABA in the embryos. Results like this have led to the conclusion that ABA plays an indirect role in storage protein accumulation, perhaps by inhibiting water uptake (Finkelstein et al 1985; Finkelstein and Crouch 1986). How this is translated into an effect on gene expression is difficult to comprehend at present.
Zygotic Embryogenesis
of fluridone, which reverses the effect of the hormone and causes a decrease in the P-subunit level (Bray and Beachy 1985). The promotive effects of ABA on the synthesis of storage proteins and their mRNAs have been carefully documented in embryos of germinating grains of Zea mays (Vance and Huang 1988; Qu, Vance and Huang 1990); cultured embryos of Sinapis alba (Fischer, Bergfeld and Schopfer 1987), Triticum aestivum (Williamson and Quatrano 1988), and Z. mays (Paiva and Kriz 1994); cultured cotyledons of Vicia faba (Barratt 1986a, b), Helianthus annuus (Goffner, This and Delseny 1990), and Medicago sativa (Xu and Bewley 1995a, b, c); and cultured embryo axes of Phaseolus vulgaris (Long, Dale and Sussex 1981). The results with V. faba cotyledons are especially interesting in showing that not only does a high osmotic medium mimic the effect of ABA in promoting the accumulation of legumin and vicilin, but the osmotic effect is reversed by fluridone (Barratt 1986a).
The effect of ABA on the fate of oleosins of germinating embryos has been the focus of some investigations. That the expression of a maize 18kDa oleosin gene is positively regulated by ABA is seen from the observation that a decrease in the transcript level of this gene noted in germinating embryos is reversed by the addition of ABA (Vance Some of the best evidence for the role of ABA in and Huang 1988; Qu, Vance and Huang 1990). the synthesis of storage proteins comes from the According to van Rooijen et al (1992), the expreswork on cultured embryos of soybean. As in the sion of oleosin genes in embryos of B. napus is sencase of B. napus, the effects of ABA on the synthesis sitive to both ABA and osmotic stress, although the of storage proteins and their mRNAs in cultured level of mRNA accumulation under osmotic stress soybean embryos are resoundingly correlated with is higher than with ABA. Dramatic increases in oil their stage of development. The presence of ABA in body proteins and oleosin mRNA, comparable to the medium potentiates the expression of genes for those obtained with ABA, are also observed in 7S and IIS storage proteins in midmaturation stage embryos of B. napus grown in the presence of jasembryos, but the hormone has the opposite effect monic acid (Wilen et al 1991; van Rooijen et al in cultured early cotyledon stage embryos. The 1992). Culture of embryos of transgenic B. napus decline in IIS protein mRNA level seen in cultured plants harboring the oleosin gene promoter from late maturation stage embryos is not, however, Arabidopsis in media containing ABA, jasmonic completely reversed by ABA (Eisenberg and acid, and osmoticum has given some interesting Mascarenhas 1985). Although the ABA effect must results. Basically, this work showed that transgenic involve much more than mere differences in the embryos respond to both ABA and osmoticum by age of embryos, it would appear from these results increased GUS gene expression, with an additive that ABA is necessary to maintain the high levels of effect when both are applied together, but no storage protein mRNAs. Another work showed appreciable response is seen with jasmonic acid that synthesis of the different subunits of 7S protein (Plant et al 1994). Although these results affirm that is not coordinately regulated by ABA in cultured ABA may not be the sole mediator of osmotically cotyledon explants, because the hormone specifi- induced gene expression, they also show that any cally promotes accumulation of the P-subunit and effect of jasmonic acid may be indirect or posttranits mRNA without any effect on the a- and a'-sub- scriptional. One of the few studies on the role of units. The involvement of ABA in the synthesis of ABA on lipid synthesis in embryos has shown that the fi-subunit is readily confirmed by the addition in a medium containing sucrose, the addition of
453
Storage Protein Synthesis in Developing Embryos
ABA or osmoticum restores the ability of cultured wheat embryos to conserve and accumulate triacylglycerol (Rodriguez-Sotres and Black 1994). The hypothesis favoring a role for ABA in modulating gene activity for storage protein synthesis did not remain uncontested and, on the balance, the available evidence is conflicting as to whether or not endogenous ABA is involved in storage protein synthesis in the embryos. In retrospect, it seems that in some cases the ability of ABA to prevent precocious germination and induce a maturation program in cultured immature embryos misled investigators to link the hormonal effect to storage protein synthesis. A few observations in favor of the negative view of ABA's effect on storage protein synthesis will be considered here. Reference was made earlier to a study that showed that a high osmoticum reinstates storage protein accumulation in cultured embryos of B. napus without increasing the level of endogenous ABA. Somewhat similar results were obtained with cotyledon explants of H. annuus incubated in a high osmoticum (Goffner, This and Delseny 1990). These observations are inconsistent with any possible consequence of ABA on storage protein synthesis and gene expression. Contrary to the effects of ABA on storage protein synthesis in cultured soybean embryos, in pea embryos grown in a high osmoticum sufficient to forestall precocious germination, addition of ABA does not enhance storage protein accumulation (Davies and Bedford 1982). Similarly, although ABA prevents precocious germination of cotton embryos, the hormone does not reinstate the levels of 48-kDa and 52-kDa storage proteins. This is not surprising, since synthesis of these proteins is seen to begin, and to reach a maximum, when ABA levels are low in the embryo and to decline during the period of ABA increase (Dure and Galau 1981; Galau, Bijaisoradat and Hughes 1987). Finally, analysis of several Arabidopsis mutants deficient in ABA or insensitive to ABA has shown that these mutations are not sufficient in themselves to alter storage protein gene expression, which appears comparable to that in the wild type (Pang, Pruitt and Meyerowitz 1988). It will be recalled that Chapter 14 discussed at some length the problems in assigning a role to ABA as a developmental regulator of those aspects of embryogenesis related to the accumulation of LEA mRNAs and proteins. Clearly, based on the information presented here, we are not in a position to generalize on the relationship between ABA and storage protein synthesis and gene expression in the embryos. The reasons for this uncertainty
seem to reside in the multiple effects of ABA, its relation to water stress, and the variety of proteins whose synthesis is affected by the hormone. 3. ISOLATION AND CHARACTERIZATION OF STORAGE PROTEIN GENES A number of embryo storage protein genes have been cloned and sequenced, and important information about the regulatory constraints for their expression has been gathered. The cloning of storage protein genes, like the cloning of other developmentally regulated genes that has been considered in this book, has traditionally begun with the isolation of their corresponding cDNAs. For characterization of the genes, cDNA clones are used for the isolation of genomic clones and for determination of the amino acid sequence of proteins and the number of gene copies encoding the proteins. As discussed in the previous section, the cDNA clones have been used to monitor the spatial and temporal expression of their transcripts in the developing embryos. To date, storage protein genes from embryos of approximately 10 species have been cloned, and their genetic organization has been defined to varying degrees. The following account represents, on a case-by-case basis, a summary of the more important characteristics of these genes, their products, and their regulatory mechanisms. (i) Phaseolin Genes
Phaseolin is one of the first storage proteins well characterized at the DNA and protein levels. An initial study reported the isolation and partial nucleotide sequence of a phaseolin genomic clone and a cloned cDNA that contained about 40% of the mRNA transcript (Sun, Slightom and Hall 1981). The complete nucleotide sequence of the genomic clone subsequently showed that it includes 1,990 bp of DNA, consisting of 80 bp of 5'-untranslated sequence, 1,263 bp of protein-encoding region interrupted by five introns, 135 bp of 3'-untranslated sequence, a promoter sequence, and two ATrich regions. The protein encoded by the gene has a very hydrophobic amino-terminal segment typical of signal peptides (Slightom, Sun and Hall 1983). On the basis of solution hybridization kinetics and Southern blot analysis, it was estimated that phaseolin is encoded as a small multigene family of about seven members and, considering the enormous amount of phaseolin synthesized during growth of the cotyledons, these genes appear to be
454 overworked (Talbot et al 1984). However, organization of the gene varies among members of the phaseolin multigene family. Complete and partial nucleotide sequences of several phaseolin cDNA clones have shown that the protein is encoded by two distinct subfamilies that differ in their coding regions by the presence or absence of direct repeats of two different sizes (Slightom et al 1985). This probably provides a reason for the heterogenous nature of the proteins encoded by the phaseolin gene. The phaseolin gene has been widely used to study the temporal and spatial regulation of expression of a cloned storage protein gene in transgenic plants. A native phaseolin genomic fragment consisting of the 782-bp upstream, 1,990-bp coding, and 1,100-bp downstream regions fused to
Zygotic Embryogenesis
fers the correct spatial and temporal regulation of GUS gene activity in developing embryos of transgenic tobacco, similar to the pattern characteristic of phaseolin accumulation in bean embryos. Subsequently it was found by 5'-deletion analysis that the upstream region of the phaseolin gene has at least three positive and two negative regulatory elements that interact to confer embryo specificity in transient assays using bean protoplasts and in stable assays of seeds of transformed tobacco. This work also showed that sequences between -295 bp to +20 bp from the upstream DNA region of the gene suffice to provide the genomic regulatory elements required for specific developmental activation and detectably high expression in both assays (Bustos et al 1989,1991). How the developmental control of the phasethe T-DNA gene of Agrobacterium tumefaciens was olin gene follows from the interplay of positive and shown to induce expression in tumor tissues negative elements has turned out to be a complex regenerated on infected sunflower stems (Murai et story in itself. Analysis of the level of expression of al 1983). When the same gene was integrated into phaseolin mRNA and protein in the seeds of the tobacco genome, levels of phaseolin in the tobacco plants transformed with additional 5'-deleembryonic tissues of seeds of transformed plants tion mutants has identified a major positive regulaare found to be enormously higher than those in tory element (-295 bp to -228 bp), a minimal the endosperm and seed coat, thereby attesting to promoter (-64 bp to -14 bp), a minor negative elethe tissue-specific and developmentally regulated ment (-422 bp to -296 bp), and a minor positive expression of the gene (Sengupta-Gopalan et al element (-782 bp to -423 bp) in the upstream 1985). Immunocytochemical techniques revealed sequence. A negative role has also been assigned to that, as in the case of bean embryos, the protein is a stretch of nucleotides between -295 bp and -107 bp targeted primarily to the protein bodies of the in repressing the expression of phaseolin mRNA embryos of transgenic tobacco seeds (Greenwood and protein in the stems and roots of transgenic and Chrispeels 1985a). Other experiments have tobacco. These results have raised the possibility demonstrated expression of the normal phaseolin that a boost in the accumulation of phaseolin gene in the cells of an insect, Spodoptera frugiperda mRNA and protein in the embryo is the net result (Bustos et al 1988), and expression of modified of a combination of transcriptional activation and phaseolin genes in callus tissues and seeds of trans- repression, such as promotion by a strong positive genic tobacco (Cramer, Lea and Slightom 1985; element (-295 bp to -228 bp) and a minimal proChee, Klassy and Slightom 1986; Hoffman, moter (-64 bp to -14 bp) in the targeted embryo Donaldson and Herman 1988; Chee, Jones and cells, and inhibition of expression in the nontarSlightom 1991; van der Geest et al 1995) and alfalfa geted vegetative parts by a negative element (-295 (Bagga et al 1992), in caryopses of rice (Zheng et al bp to -107 bp) (Burow et al 1992). 1995), and in seeds of tobacco plants regenerated As a diagnostic characteristic of cis-acting DNA from callus cultures of transgenic plants (Frisch et regulatory elements, four distinct nuclear proteins al 1995). In all cases, the native and modified genes that recognize in vitro the -295 bp to +45 bp were fully integrated into the genomes of the sequences of the phaseolin promoter have been respective heterologous hosts and they invariably identified (Figure 15.7). One DNA-binding protein, specified the phaseolin transcripts or their protein designated as CAN, binds motifs present in three products. locations, CACGTG (-248 bp to -243 bp), CACCTG Considerable work has been done to analyze (-163 bp to -158 bp), and CATATG (-100 bp to -95 the regulatory elements that control embryo-spe- bp), in the 5'-proximal region. A second protein, cific expression of phaseolin in transgenic systems. AG-1, binds to the two AT-rich sequences at -376 An initial investigation showed that an 802-bp bp to -367 bp and -356 bp to -347 bp and to a fragment of phaseolin genomic clone 5'-flanking nearly complementary sequence at -191 bp to -182 sequences that extend from -782 bp to +20 bp con- bp. Of these, the CACGTG motif or the G-box core
Storage Protein Synthesis in Developing Embryos
Temporal and Spatial Regulatory Elements (Lite) SEED
SEED
-422
Target sequence:
A AQ-1
AAAAAOACAA (-1JIJ147)
STEM ROOT
(Early?)
-£U
-227
-295
DNA binding protein:
STEM ROOT
-106
-64
O D CAN
CA-1
CACQTO
CA-rlch
(•175'-110) CACCTQ (-37t/-M7) AAAAAOOCAA (-1I1/-1 St) CATATQ {"i • i ; -1 • i)
-13
TATA-QOR
binding proton
TATAAA (-111-17)
Figure 15.7. Regulatory elements in the phaseolin gene. A major positive element (++) (-295/-228J and a minimal promoter (-64/-14) are shown; others not shown are a minor positive element (-783/-423J and a negative element (-422/-296). The sequence upstream of -107 contains a negative element repressing phaseolin expression in the stem and root. A positive element (+) for stem and root expression lies within the -106/-65 region. Also indicated is a temporal element for late expression and a possible early expression element (-64/-14). DNA-binding proteins, AG-1, CAN, and CA-1, and TATA-box-binding protein and their target sequences are shown at the bottom. (From Kawagoe, Campbell and Murai 1994.)
455
by this segment of the gene make an important contribution toward gene expression has come from the observation that a 55-bp, AT-rich sequence motif in the 5'-region of the gene (-682 bp to -628 bp) interacts with nuclear proteins from immature bean cotyledons and enhances the expression of a chimeric GUS gene in transgenic tobacco embryos (Bustos et al 1989). A later work showed that this enhancer is located in a 1,047-bp fragment in the 5'-region of the phaseolin transcription initiation site and that sequences 3'- of the polyadenylation site of the gene also contain motifs that bind to matrix preparations from tobacco nuclei. The presence of the matrix attachment regions easily accounts for the high expression in transformed tobacco seeds of phaseolin promoter-G US gene constructs containing these elements (van der Geest et al 1994). Very little attention has been paid to the endogenous factors in developing embryos that enhance or repress storage protein gene expression. A gene (PVALF) (for Phaseolus vulgaris ABAI-3-like
factor) whose transcripts are expressed in abundance in bean embryos during the few days preceding the accumulation of phaseolin and later sequence has been proposed as a major positive cis- during seed desiccation has been shown to up-regacting element in the phaseolin gene (Kawagoe and ulate phaseolin promoter in transient assays of Murai 1992). Introduction of plasmid constructs embryos. The explanation of this effect doubtless with substitution mutations in the G-box into pro- lies in the transcriptional activation domain of the toplasts derived from bean cotyledons substan- protein product of the gene in promoting phaseolin tially reduces the G-box activity in GUS gene expression during embryogenesis. The functional assay. Substitution mutations in the AG-1 binding homology of the protein to ABI-3 and VP-1 proteins sequences act both as positive and as negative reg- seems to assign it a role as a regulatory factor durulators, depending upon the position to which ing embryo maturation processes, including storthey bind. Other results have revealed that the age protein synthesis (Bobb, Eiben and Bustos G-box and CACCTG motif affect each other's 1995). A regulator that is isolated from embryonic expression synergistically. The implication of the nuclei and that in part constitutes a basic/leucinesubstitution mutation experiments is that both zipper protein by binding to a hybrid G-box/C-box CAN and AG-1 are transcription factors, although (GCCACGTCAG or CACACGTCAA) on the the mechanism of their interaction is at present phaseolin gene antagonizes the transactivation of unknown (Kawagoe, Campbell and Murai 1994). A the phaseolin promoter by PVALF (Chern, Eiben cDNA clone for one of the G-box-binding proteins and Bustos, 1996). This observation helps explain has been shown to encode a basic region /helix- the silencing of storage protein gene transcription loop-helix protein (bHLH), a member of a family of during late embryogenesis. ubiquitous transcription factors found in many The preceding account of the regulation of eukaryotes (Kawagoe and Murai 1996). The identi- genes that encode phaseolin may provide a prefication of this new transcription factor is consis- view of the complexity of the transcriptional appatent with current models of gene regulation in ratus of other storage protein genes. Both G-box eukaryotes involving the binding of a multiplicity and matrix attachment regions are common to the of proteins to different regulatory sequences in the genes isolated from many plant and animal sysgene. tems and, although their functions are not comThe molecular evidence relating to the role of pletely understood, available evidence indicates the upstream regions of the phaseolin gene will that they make the entire genetic information easily now be considered. That the functions mediated accessible for replication and transcription. The
456 presence of these motifs in the phaseolin gene suggests that when the mechanism of action of the gene is fully known, it may not be very different from that which regulates other genes.
Zygotic Embryogenesis
subunits has been attributed to a recombinational event, such as a large insertion in the first exon of the a'-gene (Schuler, Schmitt and Beachy 1982; Doyle et al 1986; Schuler et al 1982). Amino acid composition also illustrates considerable similarity between the a- and a'-subunits in their being rich (ii) Glycinin and Conglycinin Genes in aspartate, glutamate, leucine, and arginine, and Five glycinin genes (GY-1, GY-2, GY-3, GY-4, and low in methionine and cysteine. In comparison to GY-5) from soybean embryos, including one (GY-2) the a- and a'-subunits, the P-subunit is devoid of from embryos of wild soybean (Glycine soja), have methionine and cysteine, and is more hydrophobic been cloned, sequenced, and compared in attempts in nature (Thanh and Shibasaki 1977). to understand their structure and the mechanisms The degree to which soybean protein genes are that regulate their expression. Each member of this expressed in transgenic systems in terms of tissue family is encoded by two to four genes constituting specificity and developmental regulation has been a small multigene family. The genes are separated studied extensively following the report that traninto two subfamilies, designated as Group I (GY-1, scripts of the cc'-subunit of conglycinin accumulate GY-2, GY-3) and Group II (GY-4, GY-5) based on temporally in developing embryos of Petunia percent sequence homology. All five genes share a hybrida (Beachy et al 1985). Other investigations common structure characterized by the presence of that closely followed this work demonstrated four exons interrupted by three introns. Potential expression of the P-subunit of conglycinin in developmental control sequences identified in the embryos of tobacco and P. hybrida (Bray et al 1987; 5'-regions of the genes include a TATA-box and Barker, Harada and Goldberg 1988), and expresCAAT-like boxes located 25-30 bp and about 100 sion of glycinin genes in embryos of tobacco bp, respectively, upstream from the transcription (Lelievre, Oliveira and Nielsen 1992; Itoh et al 1993) start sites and multiple polyadenylation sequences and soybean (Iida, Nagasawa and Oeda 1995), and (Fischer and Goldberg 1982; Scallon et al 1985; in transgenic potato tubers (Utsumi et al 1994). Nielsen et al 1989; Xue et al 1992; Weng et al 1995). When the a'- and p-subunit genes of conglycinin Before it. is cotranslationally processed, each were introduced into P. hybrida in a single conglycinin gene encodes a precursor polypeptide that struct, time-course studies showed that the order of contains an amino-terminal leader sequence fol- expression of their transcripts in developing lowed by an acidic polypeptide, a short peptide embryos of the transgenic plant mimics the expreslinker, and a basic polypeptide (Ereken-Tumer, sion pattern during soybean embryogenesis (Naito, Richter and Nielsen 1982; Marco et al 1984; Dube and Beachy 1988). The predictable outcome Staswick, Hermodson and Nielson 1984). The dif- of experimental operations using the oc'-subunit ference in the amino acid sequences of glycinin promoter fused to the GUS gene in the antisense encoded by the different glycinin genes that is most orientation was the suppression of reporter gene impressive is that the amino-terminal sequences activity in the seeds of transgenic tobacco during show only about 50% homology (Moreira et al the period when expression of the oc'-promoter was 1979,1981). high (Fujiwara, Lessard and Beachy 1992). According to Fujiwara et al (1992), accumulation of Genes encoding a-, a'-, and p-subunits of soy- the P-subunit of conglycinin in embryos of transbean conglycinin have been cloned and analyzed genic P. hybrida is repressed by application of (Schuler, Schmitt and Beachy 1982; Schuler et al exogenous methionine to developing pods and 1982; Tierney et al 1987). Soybean conglycinin enhanced by subjecting plants to sulfur deficiency belongs to a multigene family that contains about in a manner similar to that in which embryos of 15-20 genes encoding the different subunits (Ladin, soybean respond to the same nutritional stimuli. Doyle and Beachy 1984; Harada, Barker and Exogenously applied methionine down-regulates Goldberg 1989). The overall organization of the expression of the P-subunit gene in the seeds of subunits has revealed close homologies between wild-type transgenic Arabidopsis and in seeds of members of the gene families throughout the 3'-ter- mutants of transgenic plants that overaccumulate minal half of the coding sequences and the 3'-non- methionine; accumulation of both P-subunit coding sequences. From these homologies it has mRNAs and their protein products is enhanced, been suggested that the a- and oc'-subunits of con- however, in the seeds of transgenic plants grown glycinin diverged from a common ancestral gene. under sulfur deficiency (Hirai et al 1994; Naito et al The major difference in size between the a- and oc'-
457
Storage Protein Synthesis in Developing Embryos Lsgumio box LaB4
|GAA
lagl
PAAGCCA?HA
G1
CCCAGttECTt}-
EAAGCCAK
1 ATCAa7?ArAj--HGTGTA>«gC>rTA|HGCTTCCATAGCCATGCA:a£cTCicACAATGTa-CAA
ECTCfrcacCATgAacftc
jCACccuSGGCTTCCATAGCCATGCATaCTGAAGAATGTCfrCAApCtcagca
r 1.64 last Gl
HTTCTGTfcACAcqTOTt-- ACTcTCt |TTCH-caACA
CACtgTTC
ITCCTCTfr C--HCTAT AAAlfcACCdCGCClaC
|TCCC[ACTt TCaatgacgtgt cCACc-TnCAC&cECTCrCTTqtlC t ATAAAlftUCCAqt tCTC»t t jAGGTTCTCCGCUTCACl-
ccctacftTCTGTfaACgt-TdTCCCJ
tCA-- •Irf'cAC^tltCCTCTCTTc'ifcTATAAATUKCCACGCClTC
signaI LaB4 Lag I Gl
ACTtcj»CCA?cTTC«ctcAC»«|rCCtlti:ATTAG'l}teTTt-*Cr«TCA-p>GTC<| AIACCA! ACTcAAaCA-TTC
A*CAtTCtc
ITTAG1V
elCTC-TCt-TC
|AGGTTCTCCGCTTCAda
p e p i i da
CaAT6
pCClKCAfTfaGTccTTaaACaCTCAIt:AGTC«ltcaCifrfGCCCAAGCT|
AGTtTlT TTtjctTTfottfrTCTGiff TTTa-AaT-
Figure 15.8. Major conserved elements, including the legumin-box, in the 5'-flanking regions of three legumin genes. Matches of four or more bases between any two genes are boxed; all matches are capitalized. Substitutions are shown in lower case; deletions are shown as dashes. Nucleotides represented in all three genes are indicated by dots below the Gl (glycinin 1) sequence. Brackets identify the signal
peptide coding region, the TATA element, and the leguminbox, as indicated. Heavy overlining indicates the experimentally determined 5'-ends of the LEG1 and Gl genes. The proposed mRNA start site of LEB4 (nucleotide +1) corresponds to the major 5'-end of the Gl gene. (From Baumlein et al 1986. Nucl. Acids Res. 14:2707-2720, by permission of Oxford University Press.)
1994a, b). These observations suggest that the inductive or repressive specificity of exogenous chemicals in the regulation of storage protein synthesis can function even in transgenic systems whose storage proteins are not regulated by these chemicals. A deletion series of conglycinin a'-subunit promoter analyzed in transgenic P. hybrida has revealed that the first 257-bp segment contiguous to the transcription start site contains sufficient regulatory information to program gene expression to a level almost equal to that of the complete promoter (Chen, Schuler and Beachy 1986). In addition, a fragment at -78 bp to -257 bp in the upstream region of the a'-promoter was identified as a cis-acting element enhancing embryo-specific activity in transgenic tobacco when inserted into the constitutive CaMV 35S promoter (Chen, Pan and Beachy 1988; Chen et al 1989). In vitro studies using crude nuclear extracts from immature soybean embryos have shown the presence of several proteins that interact with upstream sequences of the a'- and (3subunits of conglycinin. Two of these proteins, designated as soybean embryo factor-3 (SEF-3) and SEF-4, bind to the promoter fragment of the a'-subunit that has been shown to confer full gene expression in transgenic tobacco. During soybean embryo development, activities of SEF-3 and SEF-4 generally parallel the accumulation of a'-subunit mRNA
(Allen et al 1989; Lessard et al 1991). Specific mutations introduced into the sequences that bind SEF-3 and SEF-4 generally abolish the in vitro binding of these embryo factors (Fujiwara and Beachy 1994). Seen in the light of these results, it appears that certain SEFs play a specific role in the expression of the conglycinin gene. A novel aspect of the regulation of transcription of globulin genes in embryos of leguminous plants is the possible enhancer role of a 28-bp DNA element known as the legumin-box (Figure 15.8) about -45 bp to -65 bp upstream of the TATA-box identified in the promoter region of all globulin genes so far sequenced (Lycett et al 1985; Baumlein et al 1986). The key sequences in the legumin-box responsible for regulating the expression of glycinin genes in transformed tobacco have been found to be 5'-CATGCAT-3', 5'-CATGCAC-3', and 5'-CATGCATG-3' motifs known as RY-repeats (Dickinson, Evans and Nielsen 1988; Lelievre, Oliveira and Nielsen 1992; Itoh et al 1993). For conglycinin, the presence of the legumin-box and a core of the 3'-part of the promoter of the a'-subunit of the gene appears to be hardly required to direct a low level of embryo-specific gene expression in transgenic tobacco. Disruption of the RY-repeat elements alone also reduces expression of the gene (Chamberland, Daigle and Bernier 1992; Fujiwara and Beachy 1994).
458
Zygotic Embryogenesis
translational processing sites. Similarity in the basic structure of the genes also extends to the presence of five introns. Unlike legumin, vicilin has a predicted Genes for legumin and vicilin have been iso- amino acid sequence that allows for a signal peptide lated mostly from embryos of pea and broad bean, (Gatehouse et al 1983; Lycett et al 1983; Domoney in which they occur in families of 8-12 per haploid and Casey 1985; Higgins et al 1988). Comparable to genome. Four members of the legumin A-type gene the legumin-box, a highly conserved C-rich region at family sequenced thus far from pea (LEG-A, LEG- approximately 120 bp from the transcription start A1-LEG-A2, LEG-B, and LEG-C) have nearly identi- site in the vicilin genes of Pisum sativum and cal promoter regions characterized by TATA- and Phaseolus vulgaris, designated as the vicilin-box, is CAAT-boxes and sequences homologous to an ade- another example of a conserved upstream sequence novirus enhancer. The sequences of the coding in the storage protein gene determining tissue speciregions of the genes that are interrupted by three ficity of expression (Gatehouse et al 1986; Bown, introns, however, show slight differences due to Ellis and Gatehouse 1988). insertions or deletions (Croy et al 1982; Lycett et al The coding sequence of a convicilin gene {CVC1984, 1985; Rerie, Whitecross and Higgins 1991). A A) cloned from pea is similar to that of vicilin and is fifth legumin A-type gene (LEG-D) is relegated to interrupted by five introns. The major difference the status of a pseudogene because of the presence between the two sequences is the insertion of a very of two stop codons that could impede the synthesis hydrophobic 121-amino acid sequence near the Nof any functional polypeptides (Bown et al 1985). In terminus of convicilin. The predicted amino acid addition, genes for two minor B-type subunits of sequence of convicilin also confirms the presence of legumin, LEG-] and LEG-K, have also been fairly a leader sequence (Bown, Ellis and Gatehouse 1988; well characterized (Gatehouse et al 1988; A. J. Newbigin et al 1990). Thompson et al 1991). The predicted amino acid In attempts to determine the controlling sequences of LEG-A, LEG-B, and LEG-C genes gen- sequences of the legumin gene, the promoter of erally matched the sequences obtained by direct LEG-A gene from pea was found to function only protein sequencing, and no clear evidence was marginally in an in vitro transcription system obtained for the presence of signal peptides in the (Evans et al 1985). However, LEG-A, vicilin, and proteins (Croy et al 1982; Lycett et al 1984, 1985). convicilin genes from pea, and LEB-4 gene and the The structure of two B-type legumin genes (LEB-4 gene encoding an unknown seed protein in broad and LELB-3) cloned from embryos of Viciafaba dif- bean that contains most of the 5'-flanking fers from that of A-type legumin genes cloned from sequences are able to direct the synthesis of seedthe same species and from pea in having only two specific proteins in transgenic Nicotiana tabacum introns in the coding region; moreover, the pre- and N. plumbaginifolia (Baumlein et al 1987, 1988, dicted polypeptide of LEB-4, as well as that of the 1991a; Croy et al 1988; Ellis et al 1988; Higgins et al gene that codes for an unknown seed protein in V. 1988; Rerie, Whitecross and Higgins 1991; faba embryos, comes equipped with signal Manteuffel and Panitz 1993). Perhaps an exciting sequences (Baumlein et al 1986; Wobus et al 1986; result with potential application in increasing the Bassiiner et al 1988; Heim, Baumlein and Wobus level of sulfur-containing amino acids of embryo 1994). The amino acid sequences of leguminlike storage proteins is the demonstration of stable storage proteins deduced from cDNA clones iso- expression in transgenic tobacco seeds of a vicilin lated from embryos of Magnolia salicifolia (Fischer et gene from Viciafaba modified with eight additional al 1995) and seeds of Pseudotsuga menziesii (Leal methionine residues (Saalbach et al 1995). In tranand Misra 1993b), Ginkgo biloba (Ginkgoales) sient assay, vicilin gene driven by barley hordein (Arahira and Fukazawa 1994; Hager et al 1995), B-l gene promoter is also expressed in the and Calocedrus decurrens (Cupressaceae) (Hager endosperm of barley (Heim et al 1995). A fulland Dank 1996) show reasonable homology with length LEG-A cDNA from pea engineered to pea legumin and with 12S globulins from other encode legumin in yeast cells has been shown to plants. direct the protein to Golgi-associated vesicles of the Vicilin subunits of pea embryos are encoded by a heterologous host (Croy et al 1988). small multigene family. Sequence comparison of Whereas the 5'-flanking sequence of pea LEG-A three vicilin genes has revealed more than 80% gene containing the complete legumin-box homology among them, with most of the differences sequence failed to initiate expression in transgenic being confined to the sequences coding for the post- systems, correct spatial and temporal expression (iii) Legumin, Vicilin, and Convicilin Genes
Storage Protein Synthesis in Developing Embryos
459
was detected in a promoter construct that contained made from the CRU-1 gene has implicated different 549 bp of 5'-flanking sequence. The evidence for the elements of the promoter in regulating transcrippresence of controlling elements in the 5'-flanking tional activity in transgenic embryos during early sequence demonstrated by transgenic plant expres- stages and late stages of development (Sjodahl et al sion has been strengthened by the identification of 1995). The three major groups of cruciferin genes sequence-specific interactions between pea embryo are also represented in radish seeds and, based on nuclear proteins and the specific promoter frag- sequence comparison of the members of the two ment (Shirsat et al 1989; Shirsat, Meakin and gene families with one another and with rapeseed Gatehouse 1990; Meakin and Gatehouse 1991). This genes, it appears that there is more homology supports the idea that the legumin-box is not the between cruciferin genes from the two different sole promoter determinant in LEG-A gene expres- species than between the two genes from radish sion. An extensive analysis of the expression of (Depigny-This et al 1992). Analysis of partial or functionally important DNA sequences in the 5'- complete nucleotide sequences of four genomic flanking region of legumin gene LEB-4 from V. faba clones isolated from Arabidopsis supports the view in transgenic N. tabacum has yielded results similar that the clones are members of three cruciferin subto those obtained with LEG-A gene, implying that families; the presence of putative introns in the the legumin-box core element CATGCATG func- Arabidopsis sequence in the same positions as in the tions properly in cooperation with additional pea legumin sequence, and their absence in the upstream promoter elements (Baumlein et al 1991b, homologous B. napus sequence, seem to indicate 1992). Finally, is promotion of growth of the the close relationship of Arabidopsis cruciferins to embryo linked with the expression of storage pro- the 12S globulins of other angiosperms (Pang, tein genes? The extreme case of an experimental Pruitt and Meyerowitz 1988). Expression of a cruapproach to answer this question was the introduc- ciferin-GLZS chimeric gene in embryos of transtion of a pea vicilin gene promoter fused to DT-A genic tobacco appears to follow a pattern similar to gene into transgenic Arabidopsis and tobacco; the that of the native gene in B. napus embryos. results showed that the final form of the embryos Analysis of the expression of the truncated gene was only marginally affected by the toxic gene, promoter showed that positive regulatory elements which drastically affected storage protein accumu- containing sequences sufficient for seed-specific lation (Czak6 et al 1992). activity and a region for enhanced activity in the root meristem of transgenic embryos are dispersed over the 5'-flanking sequence (Bilodeau, Lafontaine and Bellemare 1994). (iv) Cruciferin and Napin Genes Although enormous quantities of cruciferin are synthesized during embryogenesis in members of Brassicaceae, there are no more than three gene families active in the process. As expected, each major group of cruciferin subunits is encoded by a distinct gene family (CRU-1, CRU-2/3, and CRU-4). Our knowledge of the molecular structure of the cruciferin gene is based on nucleotide sequence analyses of several cDNA clones of the three subunits and two genomic clones isolated from Brassica napus. Comparative data from cDNA clones have shown a high degree of similarity in the 5'-flanking and 3'-untranslated regions between gene members of a family and between members of different families (Simon et al 1985; Rodin et al 1990; Sjodahl, Rodin and Rask 1991; Breen and Crouch 1992). Features typical of a eukaryotic genome, such as the TATA-box, transcription start site, and polyadenylation signals, are present in the genomic clones of CRU-1, which, however, lack the legumin-box (Ryan et al 1989; Rodin et al 1992). Transformation of tobacco with deletion constructs
Napin is a well-characterized albumin-type of seed storage protein and cDNA and/or genomic clones have been isolated and characterized from seeds of Brassica napus (Crouch et al 1983; Ericson et al 1986, 1991; Josefsson et al 1987; Scofield and Crouch 1987; Baszczynski and Fallis 1990), Raphanus sativus (Raynal et al 1991), and Arabidopsis
(Krebbers et al 1988; van der Klei et al 1993). It is a member of a small to large gene family, with estimates of the number of genes in the family ranging from 5 in Arabidopsis to more than 10 in B. napus and R. sativus. An important conclusion from nucleotide sequence data is that, as in other 2S storage proteins, there are no introns. Repeated CATGCA sequences, sometimes reiterated as overlapping tandem repeats, and stretches of TACACAT consensus motifs have been identified in many napin genes. The presence of these sequences may have gross molecular consequences; in line with this possibility, the TACACAT and other sequences of a napin gene were found to interact with a nuclear protein from developing seeds of B. napus (Ericson
460
et al 1991; Gustavsson et al 1991). Although napin sequence data available for the three genera have afforded an opportunity to trace the evolution of this gene family in Brassicaceae, analysis of the coding and 3'-noncoding sequences has led to contradictory models (Raynal et al 1991). Tagged napin genes are expressed in the embryos of transformed B. napus but are silent in the leaves; the amount of napin is reduced in embryos of transgenic plants transformed with an antisense napin gene. These results are in agreement with the essential role of the promoter in the differential regulation of gene activity (Radke et al 1988; Kohno-Murase 1994). Based on 5'-deletion analysis, two portions of the 5'-upstream region of the promoter of a napin gene have been identified as important for the transcriptional regulation of the gene. These include an AT-rich region between -309 bp and -211 bp that confers strong expression of GUS gene in transgenic tobacco and conserved sequences in the region between -148 bp and -120 bp that are necessary for temporal and spatial, as well as quantitative, regulation of the gene. Whereas globular and heart-shaped embryos of transgenic plants exhibited no GUS gene activity, it was expressed in the cotyledons and axes of late stage embryos. Through the use of gene constructs containing a 196-bp fragment between -152 bp and +44 bp, napin-gene-promoter-directed GUS gene was also detected in the cells of the endosperm of transgenic seeds at all stages of development (Stalberg et al 1993). It appears that the same motifs that enhance endosperm-specific expression of the zein gene in maize endosperm are responsible for napin gene expression in the endosperm of transgenic tobacco seeds. (v) Storage Protein Genes of Other Plants
Consistent with the specialized role of storage proteins in the nutrition of the embryo during its development and germination, the structure of storage protein genes isolated from embryos of other plants is not radically different from those described already. Based on analyses of cDNA and genomic clones and deduced protein sequences, three sets of genes representing two gene families are believed to encode storage protein genes of cotton embryos. These families, known as the cc-globulin family and the (3-globulin family, are further subdivided into two subfamilies each. Mapping of genomic DNA has revealed that members of the two subfamilies of the cc-globulin family are arranged in tandem, with genes of each subfamily
Zygotic Embryogenesis
showing considerable divergence (Chlan et al 1986, 1987; Galau, Wang and Hughes 1991, 1992a). The existence of genes from both subfamilies adjacent to each other in the genome is traced to the amphidiploid nature of cotton. Although there is considerable homology between nucleotide sequences of cotton a-globulins and vicilins from legumes, cotton genes contain but four introns, compared to the presence of five introns in vicilin genes. The primary difference between the structure of genes encoding canavalin and that of genes encoding con-A in embryos of Canavalia ensiformis and C. gladiata lies in the presence of five introns in canavalin and the absence of introns in con-A (Takei, Yamauchi and Minamikawa 1989; Yamauchi and Minamikawa 1990; Ng, Ko and McPherson 1993). The DNA sequences of canavalin and con-A genes contain a number of A/T-rich regions and the core motif of the legumin-box (Figure 15.9); assay of transient GUS gene expression by deletion analysis of 5'-upstream regions of the genes has suggested that the A/T-rich regions and leguminbox contain positive regulatory elements for transcriptional activation (Yamamoto et al 1995). Genes encoding helianthinin (Vonder Haar et al 1988; Bogue et al 1990) and a 2S albumin (Allen et al 1987; Kortt et al 1991) from embryos of Helianthus annuus, 7S globulins GLB-1 and GLB-2 from maize embryos (Belanger and Kriz 1989; Wallace and Kriz 1991), a 7S protein from embryos of Theobroma cacao (Spencer and Hodge 1992), and narbonin from Vicia narbonensis (Nong et al 1995) belong to small families; indeed, there is no evidence for the presence of more than one 7S protein gene in T. cacao. Cloning and sequencing of ubiquitin from alfalfa embryos has shown that, despite its well-known role as a carrier protein, ubiquitin has a high degree of homology to 2S albumin from Brazil nut embryos (Pramanik and Bewley 1993). Characterization of two allelic variants (GLB-1L, GLB-1S) and a null allele (GLB-10) of GLB-1 gene from maize embryos has provided new insight into the molecular basis for allelic polymorphism of this gene; to the extent that the nature of this polymorphism is reflected in identifiable defects in the gene, GLB-1L and GLB-10 differ from the functional GLB-1S allele by the presence of small nucleotide insertions. These changes result in the production of a deranged protein in GLB-1L and in a translational frameshift that introduces a premature termination codon in GLB-10. However, the low steady-state level of GLB-10 transcripts in embryos homozygous for the GLB-10 allele does not reconcile with the observation that
Storage Protein Synthesis in Developing Embryos
461
50
o M
e>
raoo
ata a
a •*
Distance to transcription start site (bp) 60 r
o
Distance to transcription start site (bp)
Figure 15.9. A diagrammatic representation of the correlation between 5'-deletion of the con-A gene (a) and canavalin gene (b) and transient GL/Sgene expression levels in embryos of Canavalia gladiata. Stippled boxes and black boxes represent regions containing more than 15 bp
and more than 30 bp of AT-sequences, respectively. The sequence motifs are CATCCAT/A (black diamond); CANNTC (triangle); vicilin-box (crossed box); TATA-box (black dot). (From Yamamoto et al 1995.)
both GLB-10 and GLB-1L genes maintain comparable levels of transcription as determined by nuclear run-off transcription assays. It might appear that mRNA instability impairs the proper functioning of the GLB-10 transcripts (Belanger and Kriz 1991). A 404-bp DNA fragment located 322 bp upstream of a helianthinin gene (HAG-3D) has been shown to contain nuclear protein binding sites and to enhance GUS gene expression in embryos of transgenic tobacco (Jordano, Almoguera and Thomas 1989). Bogue et al (1990) found that transfer of 5'flanking sequences of another helianthinin gene (HAG-3A) is sufficient to confer developmentally regulated helianthinin expression in embryos of transgenic tobacco and to correctly target the protein to protein bodies of the transformed embryo. In both HAG-3A and HAG-3D genes, a region designated as the proximal promoter region contains identical sequences that confer significant reporter gene expression in the embryos of transgenic tobacco; identification of sequence motifs in the proximal region that bind to seed nuclear proteins has taken us a step closer to determining the basis of expression of sunflower storage protein genes (Nunberg et al 1994). The observations described here emphasize again that the ease of manipulation
of chimeric genes in transgenic plants offers a fruitful approach to studying the control of gene expression during embryogenesis. Molecular approaches combined with transgene technology have been employed to identify ris-regulatory domains involved in differential expression of Arabidopsis 2S storage protein genes and to offer a shortcut to alter the amino acid composition of transformed embryos. De Clercq et al (1990a) showed that the promoter of the ATZS-1 gene, which encodes a 2S albumin isoform of Arabidopsis, drives expression of a reporter gene in both transformed Brassica napus and Nicotiana tabacum. When the entire AT2S-1 gene was transferred into tobacco, not only was gene expression found to be tissue specific and developmentally regulated, but the proteins were correctly processed and targeted to the protein bodies. In a previous section, it was pointed out that in contrast to AT2S-2, which is expressed throughout the embryo, AT2S-1 is expressed only in the embryo axis of Arabidopsis. That a specific sequence in the genome is responsible for this differential expression pattern was indicated by the demonstration that replacement of a 67-bp region within a conserved 300 bp 5' of the initiation codon of AT2S-1
462
with a corresponding fragment from ATZS-2 was sufficient for expression of the chimeric AT2S-1 gene throughout the embryo. A significant feature of the 67-bp cotyledon regulatory region of AT2S-2 is the CATGCA motif found in the legumin-box of seed protein genes. Abolition of cotyledon expression by the AT2S-1 gene has been traced to a mutation in the CATGCA motif to CAAGTA (Conceicao and Krebbers 1994; Conceicao, van Vliet and Krebbers 1994). Cloning of genes encoding a protein rich in methionine from embryos of Brazil nut has afforded exceptional opportunities to manipulate the amino acid composition of embryo storage proteins of other plants (Altenbach et al 1987; de Castro et al 1987; Gander et al 1991). A beginning in the development of a system for the genetic manipulation of the 2S albumin gene of Arabidopsis has been made with the demonstration that the level of methioninecontaining proteins of transgenic embryos can be raised by chimeric genes consisting of parts of AT2S1 fused with Brazil nut 2S albumin gene (de Clercq et al 1990b; Conceicao, van Vliet and Krebbers 1994). Chimeric genes driven by regulatory regions of phaseolin gene, soybean lectin gene, or CaMV 35S promoter attached to the cDNA clone encoding Brazil nut protein have been similarly used to change the quality of the proteins expressed in transformed embryos of legumes and rapeseed (Altenbach et al 1989, 1992; Guerche et al 1990a; Saalbach et al 1994). The potential benefits of reshuffling genes in combinations not possible in nature to limit or to increase the levels of specific amino acids of storage proteins in embryos is enormous.
Zygotic Embryogenesis
(vi) Genes for Enzymes of Lipid Body Biosynthesis and for Oleosins
Understanding the genetic basis of the regulation of storage lipid synthesis in developing embryos has been possible by cloning the genes of enzymes involved in the fatty acid biosynthesis machinery and by manipulating the genes in transgenic systems. The structure of the genes characterized and the ease of their manipulation support the general conclusions drawn previously from analyses of storage protein genes. cDNA clones for ACP genes have been isolated from embryos of Brassica campestris (Rose et al 1987; Scherer et al 1992) and B. napus (Safford et al 1988). ACP in embryos of B. napus is encoded by a heterogeneous, multigene family of some 35 genes per haploid genome. Immunocytochemical studies have shown that the enzyme is localized exclusively in the plastids of the embryo cells of B. napus; to accommodate this observation, it is believed that the protein is probably encoded by the nuclear DNA and directed into plastids (Safford et al 1988; de Silva et al 1990). Nucleotide sequences of three genomic clones encoding ACP in B. napus and B. campestris appear to show a high degree of homology in the coding and 5'-noncoding regions and in the location of three introns in identical positions. The promoter region of the B. campestris gene is characterized by the presence of three different types of repeats. The coding sequences of ACP genes of B. napus and B. campestris have much in common, implying that the genes may have evolved from a common ancestral gene (de Silva et al 1990; Scherer et al 1992). Four other enzymes of the fatty acid synthetase family Reference was made earlier to changes in tran- from embryos for which some nucleotide data are script abundance of a gene that encodes an available are lipoxygenases from Pisum sativum unknown protein in embryos of Viciafaba. Although (Casey, Domoney and Nielsen 1985) and soybean it is doubtful whether the protein has storage func- (Shibata et al 1988; Yenofsky, Fine and Liu 1988); tions, characterization of the gene has shown that it enoyl-ACP reductase and acyl-ACP thioesterase has two introns and belongs to a family of 10-20 from B. napus (Kater et al 1991; Loader et al 1993); members. The gene is expressed in the embryos of and stearoyl-ACP desaturase from B. napus tobacco and Arabidopsis transformed with a frag- (Slocombe et al 1992) and Carthamus tinctorius (safment containing 637 bp of the 5'-flanking region flower; Asteraceae) (G. A. Thompson et al 1991). together with the complete 51 bp of the 5'-untrans- The most striking feature of the genes isolated from lated sequence (Baumlein et al 1991b). Based on the embryos is their considerable identity to sequences expression of smaller deletion constructs made of the same genes characterized from nonembryfrom the promoter region in seeds of transgenic onic organs of other plants. When soybean embryo tobacco, it appears that the promoter function is lipoxygenase gene was transferred into tobacco, not mediated in a combinatorial way by the interaction only was the gene expressed in the seeds, but it also of several positive and negative elements. affected fatty acid oxidative metabolism in the Surprisingly, a CATGCATG motif of the legumin- leaves of the transformed plants (Deng et al 1992). box found in the promoter acts as a negative regu- The time course of expression of the promoter of latory element controlling gene expression (Fiedler stearoyl-ACP desaturase gene in embryos of transet al 1993).
Storage Protein Synthesis in Developing Embryos
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-949 -900 -810 -720 -630 -540 -450 -360 -270 -180 -90 1
CTTTCGGGATAAAGCAATCACCTGGCGATTCAACGTGGTCGGATCATGA CGCITCCAGAAAACATCGAGaVAGCTCrCGAAGCAACaVAAGCTGACCTCTTTCGGATCGTACAGAACCCGAACAATCTCGTTATGTCCC GTCGTCTCCGAACAGACATCCTCGTAGCTCGGATTATCGACGAATCCATGGCTATACCCAACCTCCGTCTTCGTCACGCCTGGAACCCTC TGGTACGCCAATTCCGCTCCCCAGAAGaUkCOKKXXrCGAATTGCGCGAATTGCTGACCTGGAGACGGAACATaSTaSTCGGGTCCTTGC GCGATTGCGGCGGAAGCCGGGTCGGGTTGGGGACGAGACCCGAATCCGAGCCTGGTGAAGAGGTTGTTCATCGGAGATrrATAGACGGAG ATGGATCGAGCGGTTTTGGGGAAAGGGGAAGTGGGTTTGGCTCTTTTTCATAGAGAGAGTGCAGCTTTGGAGAGAGACTGGAGAGGTTTA GAGAGAGACGCGGCGGAGATTACCGGAGAGAGGCGACGAGAGATAGCATTATCGAAGGGAAGGGAGAAAGAGTGACGTGGAGAAATAGAA AACCGTTAAGAGTCGGATATTTATCATATTAAAAGCCCAATGGGCCTGAACCCATTTAAACAAGACAGATAAATGGGCCGTGTGTTAAGT TAACAGAGTGTTAACGTTCGGTTTCAAATGCCAACGCCATAGGAACAAAACAAA£GiGl£aCAAGTAAACCCCTGCCGTTTAC»CCT£A ATGCGTGCATGTGAAGCCATTAACACGTGGCGTAGGATGCATGACGACGCCATTGACACCTGACTCTCTTCCCTTTCTTCATATATCTP TAATCAATTOACTACT(aTTGTCATAGCTATTCGGAAAATACATACACATCCTTTTCTCTTCGATCTCTCTCAATTCACAAGAAGCAAA ATGACGGATACAGCTAGAACCCATCACGATATCACAAGTCGAGATCAGTATCCCCGAGACCGAGACCAGTATTCTATGATCGGTCGAGAC M T D T A R T H H D I T S R D Q Y P R D R D Q Y S M I G R D 9 1 CGAGACAAGTATOCCATGATTGGCCGAGGCCGAGACCAGTACAACATGTATGGTCGAGACTACTCCAAGTCTAGACAGATTGCTAAGGCT R D K Y S M I G R D R D Q . Y N M Y G R D Y S K S R Q I A K A
V T A V T A G C g L L V L S S I . T L V O T V I A I . T V A T P 2 7 1 CrGCTTGTTATCTTTAGTCCAATCCTTGTCCCTGCTCTCATCACCGTTGCATTC L L V I F S P I L V P A L I T V A L L I T G F L S S G G F G 361 ATTGCAGCTATAACCGTCTTCTCTTGGATCTACAAgtaagtggacatttaaacatatatttcaagttgtacaatatgttttaagaagcgg I A A I T V F S W I Y K 451 taatttattttttttttttttttgaattttaagaattcagggtttccccaaaggctttctaggcccaaaggactggtccccctcctggcg 541 ctgacgagctccatgtaataatgcccoagtggcccgagagaattgtttgcagcgtgaggcttcgaacccgggcgtattgggaagcggtaa 631 tttattaaataaaacataatggttgaatatagcgacatgccttgtgaggggaaaaaaagtacaaaccataaaattatacataaccgacaa 721 gtggattttagatattacattaaaatgccgttttacatcatcattttggctagctatacacaagacttgacttagctagcttgatacgta 811 cgtgtagtatatgtagcatgcacgtgtgtgtgaattgtgatgaatagGTATGCAACGGGAGAGCACCCACAAGGGTCAGATAAACTGGAC Y A T G E H P Q G S D K L D 901 AGTGCAAGGATGAAGCTGGGAGGCAAAGTTCAGGATATGAAGGACAGAGCTCAGTACTATGGACAACAGCAAACAGGTGGGGAAACGAC S A R K K L G G K V Q D M K D R A Q Y Y G Q Q Q T G G E H D 991 CGTGACCGTACCCGTGGAACCCAG<»CACTACCTAAATTACGCCATGACTATTTTCATAGTCCAATAAGGCTGATGTCGGGAGTCCAGTT R D R T R G T Q H T T * 1 0 8 1 TATGAGCAATAAGGTGTTTAGAATTTGATCAATGTTTATAATAAAAGGGGGAAGATGATAT<»<»GT^^ 1 1 7 1 TAAATTTGTGTGTTTCTATTTGTAAACCTTCTGTATATGTTGTACTTCTTTCCCTTTTTAAGTGGTATCGTCTATATGGTAAAACGTTAT 1 2 6 1 GATTGGTCTTTCCTTTTCTCTGTTTAGGAIAAAAAGACTG<»TGTTTTATCTTTAGTTATATTATGTTGAGTAA^ 1 3 5 1 CTGGTTCCGTAGAGTAGACTAGCAGCCGAGTTGAGCTGAACTGAACTCCTGGCAATGTGAACACTGGATGCAAGATCAGATGTGAAGATC 1 4 4 1 TCTAATATGGTGGTGGGATTGAACATATCGTGTCTATATTTTTG1TGGCATTAAGCTCTTAACATAGATATAACTGATGCAGTCATTGGT 1 5 3 1 TCATACACATATATAGTAAGGAATTACAATGGCAACCCAAACTTCAAAAACAGTAGGCCACCTGAATTGCCTTATCGAATAAG AGTTTGT 1 6 2 1 TTCCCCCCACTTCATGGGATGTAATACATGGGA'CTTGGGAGTTTGAATGAACGTTGAGACATGGCAGAACC Figure 15.10. A 2,640-bp oleosin gene from Brassica napus. The transcription initiation codon, ATG, is numbered as the first three nucleotides. The putative regulatory sequences such as TATA-box, CAAT-boxes, RY-repeats, CCboxes, octamer, putative ABA-binding sites (ACGTGGCGT and ACGTCTCCT), and polyadenylation signal are under-
lined. The intron sequence is given in lower case. The predieted amino acid sequence, including the sequence determined by peptide microsequencing, is underlined, (From Lee and Huang 1991. Plant Physiol. 96:1395-1397. © American Society of Plant Physiologists.)
genic tobacco shows that activity of the gene increases during the early period of development in a way similar to the accumulation of transcripts in embryos of B. napus (Slocombe et al 1994). Oleosin cDNA or genomic clones have been isolated and characterized from embryos of maize (Vance and Huang 1987; Qu and Huang 1990; Qu, Vance and Huang 1990; Lee and Huang 1994), Glycine max (Kalinski et al 1991), B. napus (Lee and Huang 1991; Keddie et al 1992a, b, 1994), Helianthus annuus (Cummins and Murphy 1992; Thoyts et al 1995), Arabidopsis (van Rooijen, Terning and Moloney 1992), Gossypium hirsutum (Hughes, Wang and Galau 1993), and Hordeum vulgare (Aalen 1995). The conclusion drawn from a comparison of the sequences of maize, B. napus, and Arabidopsis is that versions of the gene both with and without introns are known; whereas the maize genes are intronless, there is just one intron in the genes of the other two genera. A striking property of oleosin genes from maize and Arabidopsis is the presence of RY-repeat
sequences, usually a characteristic displayed by legume storage protein genes (Qu and Huang 1990; van Rooijen, Terning and Moloney 1992; Lee and Huang 1994). It is also worth noting that promoters of maize (Qu, Vance and Huang 1990), B. napus (Lee and Huang 1991; Keddie et al 1992a), and Arabidopsis (van Rooijen, Terning and Moloney 1992) oleosin genes have sequences that match the ABA-responsive element from the 5'-regulatory region of wheat EM gene (Chapter 14); the interspecific ubiquity of this element suggests that it might play a fundamental role in the expression of ABA-regulated genes (Figure 15.10). Chimeric constructs of 5'-flanking sequences of the Arabidopsis oleosin gene introduced into B. napus plants after transcriptional fusions to GUS gene have exposed both positive and negative regulatory elements involved in modulating quantitative levels of expression. Assay of embryos of transgenic plants for reporter gene activity has identified sequences between -1,100 bp to -600 bp
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and -400 bp to -200 bp as positive regulatory components and those between -600 bp to -400 bp as down-regulators of gene expression. A clear confirmation that the oleosin gene is expressed during early embryogenesis is provided by the demonstration that its promoter directs intense gene expression in both cotyledons and the embryo axis beginning at the heart-shaped stage (Plant et al 1994). A similar analysis with the promoter of a B. campestris ACP gene showed that the direct repeats are not essential for its embryo-specific expression in transgenic plants (Scherer et al 1992). Illustrative of the developmentally modulated expression of the oleosin gene of B. napus is the observation that its promoter fragment fused with GUS gene is expressed specifically in the embryo and endosperm of transgenic tobacco throughout seed ontogeny (Keddie et al 1994). Other experiments with chimeric oleosin genes and genes involved in fatty acid biosynthesis have been directed toward engineering the fatty acid composition of embryos by antisense expression (Knutzon et al 1992), targeting of heterologous oleosin proteins (Lee et al 1991; Batchelder, Ross and Murphy 1994), production of recombinant protein bodies (van Rooijen and Moloney 1994), and overexpression of bacterial fatty acid synthetase genes (Verwoert et al 1995) in transgenic plants. 4. GENERAL COMMENTS
Of the variety of metabolic changes that occur in developing angiosperm embryos, storage protein synthesis is very much a central one. The voluminous data reviewed in this chapter show that embryos vary enormously in the type of proteins they stockpile in the cells; yet, all storage proteins are probably synthesized in the rough ER and stored in protein bodies. Although there is some evidence justifying a role for Golgi vesicles in the transport of proteins from the site of their synthesis to the site of storage, the precise way this occurs in the large majority of embryos remains to be elucidated. Once the synthesis of storage macromole-
Zygotic Embryogenesis
cules is set in motion, measurements of mRNA synthesis, polysome formation, and a variety of other parameters indicate that cells of the embryo committed to the accumulation of storage proteins are metabolically different from the rest of the cells of the embryo. Embryos of most of our economically important pulses, cereals, and oilseeds have provided the experimental material for a series of interactive investigations into storage gene expression patterns. Foremost among these has been the isolation and characterization of genes encoding storage proteins. The transcriptional activation of these genes, whether mediated by hormones such as ABA or some other mechanisms, essentially accounts for the massive accumulation of storage proteins in the embryos. In this sense the genes can be considered to function as the ultimate controlling agent in the synthesis of proteins in the conventional way, by the interaction of as-acting sequences with DNA-binding factors. However, transcriptional control alone does not fully explain the mechanism controlling storage protein synthesis. Although not emphasized in this chapter, a cascade of posttranscriptional and translational control mechanisms also regulate the expression of storage protein genes studied. Expression of storage protein genes in transgenic plants has been a direct result of research on the cloning and sequencing of these genes. The formation of protein bodies in the embryo cells of heterologous transformants exhibits all the basic phenomena associated with their development in embryos of homologous mother plants: the synthesis of proteins in the rough ER, their targeting to vacuoles, and the embryo specificity of their expression. Because storage proteins constitute the major nitrogen reserve of the embryos of seeds that form an integral part of the diet of humans and animals, in one important way the expression of storage protein genes in transgenic plants promises immense potential of a practical nature. This relates to the means to produce genetically engineered seeds with desirable storage protein contents.
SECTION IV
Adventive Embryogenesis
Chapter 16
Somatic Embryogenesis
1. Historical Background 2. Dynamics of Somatic Embryogenesis (i) Embryogenesis from Callus (ii) Direct Somatic Embryogenesis (iii) Recurrent Embryogenesis (iv) Embryogenesis from Isolated Protoplasts 3. Cell Biology and Physiology of Somatic Embryogenesis (i) Cytology of Somatic Embryogenesis (ii) Attainment of Polarity (iii) Nutritional Factors (iv) Hormonal Factors (v) Role of Secreted Molecules 4. Biochemistry of Somatic Embryogenesis (i) Changes in Macromolecule Synthesis (ii) Role of Embryonic Proteins (iii) Polyamine Metabolism during Embryogenesis 5. Regulation of Gene Expression (i) Genes of Somatic Embryogenesis and Their Proteins (ii) Storage Protein and Other Genes (iii) Genetic Analysis of Somatic Embryogenesis 6. General Comments
This chapter will examine the cell and molecular biology of the embryogenic development of somatic cells of angiosperms and gymnosperms, or the phenomenon of somatic embryogenesis. Embryogenic development of somatic cells can be contrasted to the rigorously programmed development of the embryo from the zygote (zygotic embryogenesis) insofar as virtually any somatic cell of the plant body can, under certain experimental conditions, behave like a zygote and faithfully replay a developmental program leading to the production of embryolike structures while remaining innocent of sex. Thus, somatic
embryogenesis provides the most clear-cut demonstration of the dictum that all plant cells except those that have undergone irreversible differentiation are totipotent and retain the developmental potential to proliferate into an adult plant. Compared to the limited number of embryos arising from gametic fusion and the difficulty of extracting them from the confines of the ovule, the enormous number of somatic cells potentially capable of embryogenic development ensures the availability of an equally staggering number of embryolike structures by simple experimental manipulations. Despite the fact that zygotic embryos and embryolike structures formed from somatic cells are identical in appearance and possess the same morphogenetic potential, to emphasize the divergent pathways through which they have evolved, the term "embryoid" is generally used to refer to the latter. Although embryogenic development of somatic cells appears simple, unhindered by the sorting of genes that occurs in the zygote and limited by the genetic information parceled out to the cells from generation to generation, there are a number of cellular and physiological factors that influence unfolding of the developmental program. Some of these factors will be discussed in the early part of the consideration of somatic embryogenesis in this chapter. Later, the biochemical and molecular changes of somatic cells that enable them to organize into stable tissues and organs of embryos will be examined. The reader is referred to the reviews by Merkle, Parrott and Flinn (1995) and Nomura and Komamine (1995) for a comprehensive discussion of the physiological and tissue-culture aspects of this topic, and to those by Dudits, Bogre and Gyorgyey (1991), de Jong, Schmidt and de Vries (1993), and Dudits et al (1995) for a discussion of the 467
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cellular and molecular investigations. However, first the discussion will navigate swiftly through the story of the breakthrough that led to the discovery of somatic embryogenesis. 1. HISTORICAL BACKGROUND The discovery of somatic embryogenesis in plants is ineluctably tied to the demonstration of totipotency of plant cells. Totipotency has been viewed as the quintessential problem in plant cell biology ever since Haberlandt prophesied at the turn of this century that it would be possible to grow embryos from the vegetative cells of plants. The pioneering experiments that led to a demonstration of the totipotency of plant cells were initiated by Steward and co-workers (Steward, Mapes and Smith 1958; Steward, Mapes and Mears 1958) using the secondary phloem of domestic carrot. These investigators found that culturing slabs of the carrot tissue in a solidified tissue culture medium that supplied the requirements of essential salts, vitamins, and organic nutrients supplemented with coconut milk typically produces a proliferating mass of callus constituted of simple parenchymatous cells. Whereas previous investigators using the same system were preoccupied with the formulation of culture conditions for the indefinite growth of the callus, Steward's group found that it was possible to produce a suspension culture of individual cells and small cell clusters by growing the callus with gentle agitation in a liquid medium of the same composition as the solid medium. Although the origin of the cell clusters appeared to be a vexing problem at first, careful examination of a range of cells and cell aggregates established that individual cells dissociating from the callus divide and the daughter cells formed remain attached to one another as cell clusters. Subsequent growth of this totally disorganized population of cells and cell clumps in the liquid medium without subculture leads to lignification of the inner cells of the clump, the formation of cambiumlike derivatives, and, eventually, the appearance of lateral root primordia. A normal carrot plant is assembled in the culture flask when the rooted aggregate is subsequently grown on a solid medium lacking coconut milk. In these studies, the arduous task of monitoring a single cell in isolation during its transformation into a plantlet was not attempted; it remained for Vasil and Hildebrandt (1965) to show that a single cell of a hybrid tobacco nurtured in isolation from other cells in a defined medium forms a completely organized plant. This cell, grown in a drop of the liquid
Adventive Embryogenesis
medium in a microculture chamber, divides repeatedly to form a callus that is subsequently induced to form roots and shoots on a solid medium. The sweeping transformation of a single cell into a whole new plant is a remarkable developmental feat. In contrast to the preceding, Reinert (1959) found that a callus originating from a strain of carrot root, following a long period of culture in a medium containing coconut milk and IAA, differentiates bipolar embryolike structures upon transfer to a synthetic medium enriched with an elaborate mixture of amino acids, amides, vitamins, hypoxanthine, and IAA. Although it was hard to specify from this work what caused some callus cells to behave differently from the rest of the proliferated mass and whether the embryoids originated from single somatic cells, the observation nonetheless dispelled the notion that the formation of embryolike structures is the monopoly of the zygote. It also took us a step closer to fulfilling Haberlandt's prophesy. A natural consequence of these observations was an intensification of the efforts to induce an embryogenic type of development in identifiable, single somatic cells by manipulations of culture techniques and medium composition. The breakthrough occurred when, almost simultaneously, Steward (1963) and Wetherell and Halperin (1963) reported that single cells and cell clusters in a cell suspension culture of carrot regenerate an enormous number of embryoids, which are faithful replicas of zygotic embryos. In Steward's original method, embryos isolated from seeds of domestic or wild carrot are allowed to germinate in a medium containing coconut milk. The free cells that slough off from the seedling hypocotyl are a lavish source of totipotent cells that exhibit a typical embryogenic type of development. The effect is even more dramatic when a cell suspension originating from an immature embryo of wild carrot is plated on a nutrient agar plate; here, virtually every cell of the suspension yields an embryoid (Steward et al 1964). Wetherell and Halperin (1963) extended these observations to calluses originating from other parts of wild carrot, such as the root, petiole, and peduncle. A follow-up work also explicitly showed that coconut milk is not a factor in inducing embryogenic development of the somatic cells of carrot and that cells of the callus reared in a medium containing a moderate dose of 2,4-D form embryoids upon transfer to the same medium containing a reduced concentration of auxin. This new idea introduced interpretations that led for a period of time to lively disagreements
Somatic
Embryogenesis
in the literature on somatic embryogenesis. At about the same time that these discoveries came to light, in a note that has often been ignored, Kato and Takeuchi (1963) described a balanced embryogenic sequence of development in single cells of carrot root callus sloughed off into the growth medium. Finally, the transformation of a single cell of the callus, nurtured in isolation from other cells, into an embryoid was also demonstrated, reinforcing the conclusion that embryoids formed in cell suspension cultures indeed have their origin in single cells (Backs-Hiisemann and Reinert 1970). One of the important questions considered in the wake of the discovery of somatic embryogenesis was the criteria to define an embryoid. In the work on carrot just described, structures appearing in the cell suspension were designated as embryoids on the basis of their bipolarity and their close resemblance to the organogenetic part of typical zygotic embryos of dicotyledons. Since superficial resemblance alone may be deceptive, it was suggested that besides acquiring early bipolarity, an embryoid should have had its origin in a single cell and should not be connected to a preexisting vascular strand in the mother tissue (Street and Withers 1974; Haccius 1978). However, these criteria have generally been ignored in the majority of contemporary publications that describe somatic embryogenesis in cultured plant cells, tissues, and organs; even to the present day, it is the acquisition of bipolarity that is used by investigators to describe as embryoids structures that originate in tissue culture. The division sequences during the transformation of somatic cells into embryoids appear to be different from the pattern of divisions of the zygote to form an embryo. As seen in carrot, these differences place limitations on the reliance on external morphology alone in comparative studies of zygotic and somatic embryogenesis in a given species. Development of the embryo of carrot follows a pattern typified by the Solanad type and begins with an unequal transverse division of the zygote. Both resulting cells divide by transverse walls to form a file of four cells. A third round of transverse divisions in each of the four cells gives rise to eight cells held together in a filament. The first longitudinal divisions occur in the three cells farthest from the micropyle, whose descendants eventually generate the organogenetic part of the embryo except the root tip. A limited number of divisions in the other five cells of the filament produce the root tip and a massive suspensor (Borthwick 1931; Lackie and Yeung 1996).
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Unraveling the division sequences of somatic cells of carrot during their transformation into embryoids appeared to be difficult because it had not been possible to identify by any unique cytochemical or ultrastructural features those cells in a suspension culture that were destined to divide in the embryogenic pathway. Moreover, the occurrence of potential embryogenic cells as part of a cell cluster and as single cells seemed to demand the presence of more than one division sequence to effect the transformation into embryoids. Halperin (1966) found that in carrot cell suspension culture, the presumptive embryoid appears as a globular structure from a disorganized cellular mass without displaying any definite sequence of cell lineage associated with early segmentation. Although a suspensor comparable in stature to that subtending the zygotic embryo is absent, cells of the original aggregate often remain attached to the globular embryoid as a suspensorlike appendage. In the only instance in which a single somatic cell of carrot was followed in isolation during embryogenesis, it was found that in contrast to the zygote, which gives rise to a polarized bicellular structure by an asymmetric division, the somatic cell yields a conglomerate of cells by random divisions; the embryoid emerges from this cellular mass by an additional series of random divisions (BacksHiisemann and Reinert 1970). The main problem with these investigations was that it was not possible to establish the normal fates of the division products of the cell as the cell was transformed into an embryoid. This problem was overcome in a detailed study of the early development of somatic embryos of carrot by McWilliam, Smith and Street (1974). This work showed that although the first division of the presumed embryogenic initial in a cell aggregate is in the transverse plane, the daughter cells that are formed diverge along different pathways. The terminal cell divides longitudinally to form the embryo proper, and the cell closest to the cellular aggregate divides transversely to form an incipient suspensorlike structure. This is enough discrimination for our purposes to say that early division sequences of the somatic embryo of carrot appear to have more in common with the Crucifer type than with the Solanad type of embryo formation. Although these results point to a fundamental difference in the segmentation patterns of zygotic and somatic embryos of carrot, it is important to recognize that both sequences create developmentally programmed cell lineages that result in identical structures. The fact that embryolike structures are induced in free cells of carrot by simple manip-
470
ulative techniques seems to suggest that somatic embryogenesis in this plant is not an aberrant form of development displayed by cultured cells but is comparable in many respects to zygotic embryogenesis. Subsequent demonstration of somatic embryogenesis in a variety of plants under different cultural conditions has tended to reinforce this proclivity. 2. DYNAMICS OF SOMATIC EMBRYOGENESIS
As a means of generating embryolike structures from unspecialized somatic cells, manipulation of various plant organs in culture has been a favorite subject of research. Embryogenesis has been induced in a variety of explants, such as the stem, hypocotyl, root, leaf, inflorescence axis, pedicel, floral bud, individual floral parts including anther filament, excised seed embryo and its parts, endosperm, and nucellus. Relatively few studies have been concerned with the cellular origin of embryoids, and only a limited number of comparative studies have been made to assess the embryogenie potential of the different parts of the same plant. Moreover, meristematic cells, such as the cambium and the sites of bud and root initiation, are not generally excluded from the cultured explants; it is therefore not established whether embryoids arise from differentiated cells that acquire totipotency or from meristematic cells that have no prior commitment toward differentiation.
Adventive Embryogenesis
from immature embryos (Hakman et al 1985). In later years, the methodology has been extended to obtain embryoids from several additional species of Picea and from Zamia, Ceratozamia, Cycas (Cycadales), Larix, Pinus, Pseudotsuga (Pinaceae), and Sequoia (Taxodiaceae). The potential use of the technique of somatic embryogenesis for clonal propagation in forestry and as a means of guaranteeing the survival of species on the verge of extinction (for example, Cycadales) has generated a great deal of interest. For a review of the work on somatic embryogenesis in gymnosperms, reference is made to the book edited by Jain, Gupta and Newton (1995).
Earlier it was mentioned that in the successful induction of embryoids from wild carrot, aseptic culture of the tissue explant to produce a callus is the essential first step and that transfer of the callus to a medium of a different composition elicits initiation of embryoids. Although this classical protocol became practicable with many species, in others it is possible to bypass the callus stage and to induce embryogenesis directly on the cultured explant. With certain plants, recurrent embryogenesis constitutes the basis for mass production of embryoids; this occurs when the first generation of somatic embryos produced on an explant gives rise to successive cycles of embryoids, most commonly from the superficial cells of the cotyledons or hypocotyl. It is as if when cells of an explant are determined as embryoids, the embryoid nuclei experience an irreversible embryonic epigenetic No useful purpose will be served by listing the state that continues to activate the same block of number of species and genera in which somatic genetic information, generation after generation. embryogenesis has been reported, because new In still other cases, somatic cells enzymatically cases are described in almost every issue of jour- stripped of their cell walls regenerate a new cell nals devoted to plant tissue culture and related wall and pursue an embryogenic type of developfields. From published lists and reports, it appears ment. These modes of somatic embryogenesis that somatic embryogenesis is well spread among a have no species barriers, since certain organs in a respectable number of diverse angiosperm fami- plant may respond to culture by direct somatic lies, although plants belonging to Apiaceae, embryogenesis whereas others produce may Ranunculaceae, Solanaceae, and Fabaceae among embryoids from a regenerated callus, or the same dicotyledons, and to Poaceae among monocotyle- organ may produce embryoids through both pathdons, exhibit a greater propensity for somatic ways. Similarly, some embryoids produced on an embryogenesis than do members of other families. explant might germinate into seedlings, whereas It is not herbaceous angiosperms alone that display others might enter a recurrent cycle of embryogesomatic embryogenesis by appropriate tissue cul- nesis. In many cases in which embryoids are proture stimuli; in the last few years, regeneration by duced by way of naked protoplasts of somatic somatic embryogenesis has been reported in an cells, the embryoids themselves regenerate addiincreasing number of woody angiosperms (Brown, tional embryoids easily. What are the critical Finstad and Watson 1995; Dunstan, Tautorus and developmental aspects of somatic embryogenesis Thorpe 1995; Krishnaraj and Vasil 1995). Among by these pathways? A few widely investigated gymnosperms, somatic embryogenesis was first examples will be used to examine this question demonstrated in Picea abies from a culture initiated now.
Somatic Embryogenesis
Figure 16.1. Somatic embryogenesis in carrot, (a) Cell clusters representing early stage globular embryoids formed upon transfer of the cell suspension to a medium lacking 2,4-D (induction medium), (b) Formation of late
(i) Embryogenesis from Callus
The carrot system has been the most comprehensively studied with respect to the developmental aspects of somatic embryogenesis from a callus. For experimental purposes, the callus is initiated from hypocotyl or root segments excised from seedlings of domestic carrot germinated under aseptic conditions and cultured on the surface of a high nitrogen-containing solidified mineral-salt medium such as Murashige-Skoog medium supplemented with sucrose, an organic addendum including mj/o-inositol, a source of iron such as ferric ethylenediaminetetraacetic acid (FeEDTA), a cytokinin [kinetin, zeatin, or N6(A2-isopentenyl) adenine (2iP)] and 2,4-D. A suspension culture is initiated by transferring a piece of the callus to a liquid medium of the same composition, but with a reduced level of 2,4-D, and is gently agitated on a horizontal shaker. This results in the production of a suspension culture consisting of single cells and proliferating cell clusters known as proembryogenic masses. Embryogenesis is induced when an aliquot of the suspension is transferred to a basal medium from which 2,4-D is omitted (induction medium). Under these conditions, the first signs of embryogenesis are noticed in four to five days, when the cell aggregates are transformed into globular embryoids. Subsequent processes of regeneration proceed rapidly and involve the appearance of cotyledons (which initiate bipolar differentiation), formation of the shoot apex, initiation of the root apex, and maturation of embryoids (Figure 16.1). Some investigators have established conditions for obtaining high yields of synchronously developing carrot embryoids starting with a cell suspension (Fujimura and Komamine 1979b; Giuliano, Rosellini and Terzi 1983). It is also possible to frac-
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globular and heart-shaped embryoids in the induction medium, (c) Torpedo-shaped and later stages of embryoids formed in the induction medium. (Photographs supplied by Dr. S. C. Minocha.)
tionate the cell suspension by sieving on nylon screens, followed by density gradient centrifugation in Percoll solution to obtain a population of embryogenic single cells and to identify certain spherical cells as potentially embryogenic. These cells proceed with the normal developmental tempo to form proembryogenic masses and to differentiate into embryoids in high frequency (Nomura and Komamine 1985). Another line of investigation using video cell-tracking has identified five different types of embryogenic single cells in a carrot cell suspension that differentiate into somatic embryos with varying frequencies (Toonen et al 1994). These experiments represent the nearest thing to identifying the somatic cell equivalent of the zygote in a population of cells. In hypocotyl explants of carrot, the embryogenic cells are believed to arise from certain cells of the central cylinder that divide asymmetrically upon exposure to 2,4-D (Guzzo et al 1994,1995). Given the fact that carrot cells newly liberated from an explanted tissue or a callus do not have the capacity to produce embryoids, but acquire this capacity only after a certain period of growth in the 2,-4-D-containing medium, it seems reasonable to conclude that auxin induces the development of embryogenic potential in the cells. This would presumably be accomplished by a specific gene expression program under the influence of 2,4-D. The work on somatic embryogenesis in carrot has taken on yet another dimension with the demonstration that tissues of the mericarp (oneseeded half of the ovary) of carrot, as well as mechanically wounded mature zygotic embryos, can provide the starting material for the production of embryoids in a completely hormone-free medium. Small cell clusters generated from these explants are maintained as preglobular stage
472 proembryos by continuous subculture in a buffered medium (pH 4.0) containing 1-5 mM NH4+ as the sole source of nitrogen. The determining factor that allows continued embryogenic development of the cell clusters appears to be an increase in the pH of the medium to 4.5 or above (Smith and Krikorian 1988,1989,1990). Other options available to induce somatic embryogenesis in carrot in a hormone-free medium include culture of seedling meristems in media of high osmolarity or in those containing heavy metal ions, although under these conditions, embryoids are formed without a visible callus phase (Kamada et al 1989; Kiyosue et al 1990b). A possible mechanism based on the release of cells from positional or chemical restraints, which allows the cells to express their innate embryogenic potential, is suggested to account for these results. Later in this chapter it will be seen that the role of auxin in the establishment and maintenance of embryogenic cell cultures of carrot has occasioned intense debate. The embryogenic cell culture developed in the hormone-free medium offers a new level of opportunity for a renewed investigation of the seemingly intractable problem of auxin action in somatic embryogenesis.
Adventive Embryogenesis
Hernandez-Fernandez and Christie 1989). Apparently, when a favorable genotype is selected, the nature of the explant is of secondary importance in determining the potential for somatic embryogenesis, and a wide range of explants including cotyledons, hypocotyl, shoot tip, petiole, leaf, root, and stem - are responsive in culture (Brown, Finstad and Watson 1995). There have been several experimental studies on the choice of the culture medium and the specific components of the medium necessary to induce embryogenic development in cultured explants of M. sativa. There is evidence to show that the growth hormone component and nitrogen supply in the medium are the critical factors that affect somatic embryogenesis. The explants become most susceptible to the inducing effects of an auxin such as 2,4-D, 2,4,5-trichlorophenoxyacetic acid, or (2,4dichlorophenoxy) propanoic acid, and a cytokinin like kinetin, which in combination can increase appreciably the number of cells becoming embryogenic (Meijer and Brown 1987b; Brown, Finstad and Watson 1995). Turning to nitrogen supply, both induction of embryoids and their subsequent development appear to be controlled by the quality and quantity of nitrogen supplied in the medium. The Medicago System. The development of The nitrogen requirement can be met by optimizing with an amino acid in combinaprotocols for regeneration of somatic embryos in the concentration + Medicago sativa illustrates the need for protracted tion with NH+4 or by maintaining an appropriate studies using various genotypes, explants, and ratio of NH4 :NO3~ in the medium (Stuart and medium formulations to obtain suspension cul- Strickland 1984a, b; Meijer and Brown 1987a). tures that yield embryoids reproducibly and in Based on the data, a widely used protocol to large numbers. The first report of somatic embryo- obtain embryogenic cell suspensions and embrygenesis in M. sativa showed that a callus originat- oids of M. sativa for biochemical and molecular ing from cultured embryos regenerates embryoids investigations (Figure 16.2) involves the induction upon transfer to a medium containing an optimum of a callus in a medium containing a weak auxin level of 12.5 mM NH4+ (Walker and Sato 1981). (such as NAA) and kinetin, maintenance of the tisComing in the wake of a previous report of plant sue as a suspension of microcalluses in the same regeneration by organogenesis from calluses initi- medium, initiation of embryogenesis by a short ated from immature ovaries and anthers cultured pulse treatment with 2,4-D, and the subsequent in a medium containing kinetin and/or 2,4-D and growth of embryoids in a hormone-free medium NAA and transferred to a medium lacking auxin (Dudits, Bogre and Gyorgyey 1991). In another proand cytokinin, this suggested that shoots observed tocol, callus initiated on a solid medium containing in the earlier study were actually embryoids 2,4-D and kinetin is transferred to a suspension cularrested in development (Saunders and Bingham ture in which kinetin is replaced by NAA. 1972). Later investigations with different genotypes Embryogenesis is initiated in cell suspensions colof M. sativa showed that the genetic background of lected on a 200-ji m mesh by transfer to a hormonethe donor plants clearly betrays the regeneration free medium (Pramanik, Krochko and Bewley response of the explants (Mitten, Sato and Skokut 1992). 1984; Brown and Atanassov 1985). This principle was recognized in breeding experiments in which Cereals and Grasses. Because of their economic it was possible to transfer the trait for somatic importance, members of Poaceae have been special embryogenesis within populations of plants targets of attempts to induce somatic embryogene(Brown 1988; Wan, Sorensen and Liang 1988; sis. In the early period of this work, success or its
Somatic Embryogenesis
473
explant types, medium composition, and hormonal additives that promote callus growth and subsequent embryogenesis completely understood; indeed, a feature that gripped the imagination of workers in the field was that the first notable success was achieved by judicious choice of explants, use of selected hormonal additives, and unorthodox manipulation of the medium at critical stages of culture. The example that vividly illustrates this is Pennisetum americanum. Culture of immature embryos excised from 10-15-day-old caryopses or culture of segments of unopened inflorescence on a solid medium supplemented with 2,4-D is found to regenerate a callus. Transfer of the callus to a liquid medium supplemented additionally with coconut milk produces a suspension culture consisting predominantly of richly cytoplasmic embryogenic cells that spontaneously develop into globular or heart-shaped structures. Even at this stage, the multicellular units are not fully committed to the embryogenic pathway; embryoids with the characteristic morphology of cereal embryos are obtained when the suspension culture is plated on an agar medium containing ABA but lacking 2,4-D (Vasil and Vasil 1981,1982b). In an alternative pathway of somatic embryogenesis displayed by cultured immature embryos, some of the superficial cells of the scutellum divide repeatedly to form embryoids or they regenerate an embryogenic callus from which embryoids subsequently arise (Vasil and Vasil 1982a). With some minor adjustments in protocol, surgically separated parts of mature embryos of P. americanum consisting of the shoot apex and subjacent leaf primordia are also found to yield an embryogenic callus and embryoids in culture (Botti and Vasil 1983).
Figure 16.2. Induction of somatic embryos in alfalfa, (a) A microcallus suspension grown in the presence of NAA and kinetin. (b) A purified fraction of early globular stage embryoids formed in a hormone-free medium following pulse treatment with 2,4-D. (c) A purified fraction of late torpedo-shaped embryoids. (Photographs supplied by Dr. D. Dudits.)
lack thereof with cereals and grasses was critically noted, but less so today since it is now a routine practice to obtain continuously growing embryogenie cell suspensions from a wide range of cereals and grasses and to induce somatic embryogenesis in them. In no case are the specific requirements of
There are indications in other members of Poaceae that the maturation of embryoids is a critical stage in embryogenesis that is controlled by a set of nutrients different from those that induce embryogenesis. This may be illustrated by somatic embryogenesis in wheat. Embryoids are formed with relative ease on calluses raised from young inflorescence axes and immature embryos grown in a medium containing 2,4-D, but precocious germination resulting in a leafy scutellum and multiple shoots is rampant in these embryoids. However, an embryogenic callus regenerated from explants cultured in a double-strength mineral-salt medium supplemented with myo-inositol, glutamine, casein hydrolyzate, and 2,4-D readily forms normal, full-term embryoids (Ozias-Akins and Vasil 1982,1983). During the years of, or closely following, these investigations, embryogenesis was
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induced in calluses derived from a variety of explants of a host of other members of Poaceae by subtle manipulations of the medium composition. These reports have become far too numerous to catalogue here, but they have been tabulated elsewhere recently (Krishnaraj and Vasil 1995). The successful reports of somatic embryogenesis in Poaceae have led to the view that cultured explants produce different types of calluses, which differ in their regenerative potential, and that selection of the appropriate callus (embryogenic callus) is an important step in inducing embryogenesis. Two aspects of somatic embryogenesis in this group of plants that we do not understand at all and that have limited the use of the system in biochemical and molecular investigations are the slow growth of the callus and the rapid loss of its embryogenic competence. (ii) Direct Somatic Embryogenesis
The induction of a callus on the cultured explant and the transfer of the loosely dissociated cells to a medium of a different composition to induce embryogenesis present a complicated situation, particularly in light of the bewildering hormonal requirements of the different species thus far investigated and the need in some cases to select embryogenic calluses. The justification for the continued use of carrot and a few other plants is partly that they provide elegant embryoid induction systems but also that they offer favorable material for molecular studies. However, the feasibility of somatic embryogenesis directly on the cultured explant has simplified the system and offers the prospect of gaining a better understanding of the cell-level controls that operate during embryogenic induction. This has been demonstrated in a most dramatic way by the culture of young flower buds of Ranunculus sceleratus in a medium supplemented with coconut milk and IAA; the dense callus that forms on the cut end of the explant subsequently differentiates numerous embryoids. Some of the embryoids germinate precociously into seedlings and bear a second generation of embryoids freely exposed along the entire length of the hypocotyl and stem (Konar and Nataraja 1965). Much of what we know about medium requirements for direct somatic embryogenesis has been gleaned from other studies, which have used explants of leaves, ovules, nucellar tissues, embryos, somatic embryos, and haploid pollen embryoids of various plants. The summary of evidence as it stands now suggests that the culture of
Adventive Embryogenesis
explants in a medium containing coconut milk or in a medium supplemented with 2,4-D alone or in combination with a cytokinin, elicits the formation of embryoids directly from the superficial differentiated cells of the explant or from an incipient callus regenerated on the cut surface of the explant. In leaf explants of Dactylis glomerata, anatomical studies have unequivocally demonstrated that the embryoid arises by dedifferentiation of a differentiated cell of the explant (Conger et al 1983). In most other cases, direct evidence of this kind is lacking. There is a further aspect to the analysis of D. glomerata that provides an underlying basis for direct somatic embryogenesis and embryogenesis through the intervention of a callus. This aspect is in evidence when the medium is confronted with explants taken from different parts of the same leaf or with segments from the same region of different leaves. Segments taken from the more basal parts of the leaf are so dominated by a callusforming habit that they never display direct somatic embryogenesis; by contrast, those excised from the distal portions form embryoids directly. Comparison between the responses of leaves of two different ages shows that the potential for direct embryogenesis shifts slightly toward the base of the older of the two leaves. These observations carry important implications for the role of the meristematic tissues of the leaf in promoting callus growth versus embryogenesis. A juvenile grass leaf has an increasing basipetal gradient of meristematic activity, with the meristematic area in a mature leaf very much restricted to its base. Apparently, direct somatic embryogenesis is perpetuated by a recrudescence of growth in the fully differentiated cells in the more apical part of the leaf, whereas cells that are actively dividing tend to lapse into a callus-forming mode (Conger et al 1983). It is a prediction from this work that the cytological state of the leaf cells at the time of culture is more important than any special additives to the medium in choosing the pathways toward morphogenesis. This said, the process of direct somatic embryogenesis still remains obscure because we do not know the genetic, epigenetic, and physiological changes in the cultured leaf segments that affect the outcome. (iii) Recurrent Embryogenesis
Current views concerning the formation of repeated cycles of embryoids from a single explant are derived from the observations on Ranunculus sceleratus referred to previously. Although this
Somatic Embryogenesis
work did not explicitly demonstrate the formation of embryoids for more than one cycle from the cultured explant, subsequent work has shown that in some cases, the primary somatic embryos fail to mature but give rise to successive cycles of embryoids directly from the superficial cells. This phenomenon of recurrent or secondary embryogenesis is well studied in Medicago sativa, in which the initiation of recurrent embryogenesis requires transferring a callus formed in a hormone-enriched medium to a hormone-free medium. When the regenerated somatic embryos are maintained in the same medium or transferred to fresh hormone-free medium, senescence of the cotyledons and formation of a second generation of embryoids from the hypocotyl are observed. The newly formed embryoids can be used to start the cyclic regeneration of fresh crops of embryoids almost indefinitely (Lupotto 1983, 1986). There are aspects of this process that do not provide easy explanations, notably the relationship between the senescence of the cotyledons and the activation of cells of the hypocotyl. This might involve the loss of intercellular coordination due to cotyledonary senescence or a block in the maturation pathway of the embryoid that leads to embryogenic divisions in the hypocotyl cells. (iv) Embryogenesis from Isolated Protoplasts
Embryoid development from protoplasts isolated directly from somatic cells of explants and from embryogenic cells maintained in suspension cultures has been studied under standard tissue culture conditions. In the first reports of embryogenesis from isolated protoplasts, pieces of carrot root and aliquots of suspension cultures from root and petiole of carrot seedling were used as starting materials (Grambow et al 1972; Kameya and Uchimiya 1972). The protoplasts were cleared by centrifugation and suspended in an isotonic culture medium. They were subsequently cultured in the same medium, the osmolarity of which was reduced in a step-wise fashion with the progress of culture. The naked protoplasts formed embryoids only after they re-formed a cell wall. Generally, the period up to the formation of a new cell wall was marked by changes in the size and shape of the protoplasts and, irrespective of the protoplasts' origin, the first one or two divisions in the isolated protoplasts were observed in about six days after culture. Various fates can befall a re-formed cell, one of which is repeated divisions to form a cell
475
cluster and the transformation of daughter cells into embryolike structures. Characteristically, the requirements for embryogenic development of cells originating from either cell type by way of isolated protoplasts reflect those of the respective parent cells. Cell clusters originating from protoplasts isolated from suspension cultures regenerate embryoids upon transfer to a medium lacking 2,4D, whereas those formed from protoplasts of root cells require a medium enriched with coconut milk. Overall, these results show that despite the completely different pressures to which isolated protoplasts are exposed, once they re-form a wall their subsequent morphogenesis is as highly precise and predictable as that of a totipotent cell. The amazing progress in protoplast technology has led to the development of protocols to generate calluses and plantlets from protoplasts isolated from a number of plants. In many cases, protoplasts isolated from leaf mesophyll cells have been found to undergo embryogenic development with relative ease. However, it has not been clearly established that, in the majority of species thus far studied, isolated protoplasts regenerate plantlets by embryogenesis. 3. CELL BIOLOGY AND PHYSIOLOGY OF SOMATIC EMBRYOGENESIS
Two essential facts about somatic embryogenesis that have withstood the test of time well are that (a) dedifferentiation and division of the cell induced by a strong auxin, such as 2,4-D, is the invariant first step in the process, and (b) embryogenic development is initiated in the absence of auxin or in the presence of a reduced concentration of auxin. Although auxin carries the overtone of a producer of proembryogenic masses in the sense of assembling the newly formed cells into a cluster, it has become clear that the role of auxin is somewhat limited. A simple illustration of this is provided by the observation that if carrot explants are exposed to 2,4-D for a short period of time, potentially embryogenic single cells can be subsequently induced to differentiate into proembryogenic masses in a medium lacking auxin; treatment of the explant for a prolonged period with auxin also results in the formation of proembryogenic masses. Thus, a knowledge of the source of the embryo mother cell, particularly with regard to its encounter with auxin, becomes an important consideration in the study of the cell biology of somatic embryogenesis. Most of the work in this area has been done with cell suspensions propagated by
476 repeated subculture in an auxin-containing medium. A wide array of observations made on carrot cell cultures over the years has helped to define precisely the types of single cells and inchoate cell aggregates found in a suspension culture (Emons 1994). In the aggregate state, proembryogenic masses consisting of small, peripherally disposed, cytoplasmic meristematic cells surrounding a mass of large vacuolate, nondividing cells are of central importance in embryogenesis. Here, the behavior of the meristematic cells can be clearly distinguished from that of the nonmeristematic cells. Embryoids are formed from the peripheral cells, whereas the large nonmeristematic cells break away from the aggregate and become elongate, free-floating cells. The significance of the peripheral cells becoming embryo mother cells is clear; these cells are freely exposed and are not restricted, on topological ground, by surrounding cells of a cluster. The suspension culture also contains embryogenic single cells, which may divide to form proembryogenic masses, and elongate cells, which are the progenitors of the small embryogenic cells. It is from the behavior of the small peripheral cells of the cluster and the embryogenic single cells that the most obvious features of embryoid formation become clear. (i) Cytology of Somatic Embryogenesis Most published accounts of the ultrastructural cytology of somatic embryogenesis in carrot are dominated by comparative studies of identifiable potentially embryogenic and nonembryogenic cells bathed in an auxin-containing medium and challenged in a medium devoid of auxin. The importance of the cell surface for embryogenesis was revealed by differences in the wall architecture, plasma membrane rosettes (putative microfibril synthesizing complexes), plasmodesmata number, and exocytosis configuration (related to secretion and membrane cycling events) seen in freeze-fracrure studies of the peripheral and central cells of a proembryogenic mass. An ordered wall texture, the presence of many plasmodesmata, and exocytosis configurations on the plasma membrane are characteristic of the embryogenic cells of carrot, whereas nonembryogenic cells have unordered wall texture, few plasmodesmata, and limited involvement in exocytosis. No differences are seen between the number of plasma membrane rosettes in the two cell types (Emons, Vos and Kieft 1992). Similar observations on cell wall architecture and plasmodesmata have been made in maize cell sus-
Adventive Embryogenesis
pension culture (Emons and Kieft 1990). This circumstantial evidence suggests that intercellular communication and limited elongation-prone growth are characteristic of embryogenic cells in the suspension. Our minimal ultrastructural image of an embryogenic cell has a munificent supply of starch-filled plastids; round, oval, or irregular mitochondria; a small number of lipid bodies; free ribosomes; abundant ER studded with polysomes; and multivesicular bodies of presumed Golgi origin (Halperin and Jensen 1967; Wochok 1973a; Street and Withers 1974; Wilson, Israel and Steward 1974). Although not seen in carrot cells, a striking organization of rough ER, stacked in extensive parallel sheets associated with numerous ribosomes, has been found in the potentially embryogenic cells of Corylus avellana (Vujicic, Radojevic, and Neskovic 1976). Presumably, these subcellular features are indicative of an enhanced metabolic activity of the embryogenic cells as they embark on a new round of divisions. The ultrastructural consequence of embryogenic induction in carrot cells transferred to a medium lacking auxin is implicit in the increase in free ribosomes, the decrease in ER, a loss of polysomes, and an increase in the number of Golgi vesicles and lipid bodies (Halperin and Jensen 1967; Wochok 1973a; Street and Withers 1974; Wilson, Israel and Steward 1974). There are both similarities and differences between the ultrastructural profiles of embryogenesis in cells in suspension culture and embryogenesis in callus cells immobilized on a solid medium or callus arising directly from the superficial cells of cultured explants. In the habituated nucellar callus of Citrus sinensis, virtually all cells are embryogenic, thus making it relatively easy to characterize the progenitor cells of embryoids. Embryogenesis is usually initiated in single cells located on the periphery of, or within, an existing cluster of cells referred to as a proembryoid. Although the embryoid mother cells are ultrastructurally undistinguished, the cells are characterized by the presence of a thick wall lacking plasmodesmata. A globular embryoid is formed by divisions of the cell within the confines of this wall, which disintegrates to release the embryoid (Button, Kochba and Bornman 1974). A special callose wall has been shown to regenerate around the epidermal and cortical cells of root explants and mesophyll cells of leaf explants of Cichorium intybus, which directly give rise to embryoids (Dubois et al 1990, 1991). The disruption of plasmodesmatal connections and the insulation by callose indicate that the cell
Somatic
Embryogenesis
embarking on embryogenesis is physically isolated from its neighbors; we do not have any basis to explain how such activity can induce embryogenic divisions. The involvement of regular epidermal cells in embryogenesis has been demonstrated in the hypocotyl of embryoid-derived seedlings of Ranunculus sceleratus. The embryoid mother cells scattered singly or in small groups in the epidermis are each typically blessed with a large central nucleus surrounded by a dense cytoplasm rich in ribosomes or polysomes, amyloplastids, and spherosomes. By contrast, nonembryogenic epidermal cells have a peripheral cytoplasm with sparse organelles. The ultrastrucrural profile of embryogenic epidermal cells corresponds closely with that of the superficial cells of an embryogenic callus obtained from cultured flower buds of R. sceleratus; the latter cells seem to reflect and accommodate the subcellular changes seen in embryoids (Konar, Thomas and Street 1972; Thomas, Konar and Street 1972). These observations carry the implication that at some stage in seedling ontogeny, a few epidermal cells of the hypocotyl are reprogrammed as embryoid mother cells, whereas other continue to bide their time as regular epidermal cells. Thus the same genome is able to display two different phenotypic expressions. In plantlets of Medicago sativa regenerated from a callus, single epidermal cells of the cotyledons and hypocotyl usually function as embryoid mother cells (Dos Santos, Cutter and Davey 1983). Rice scutellum provides an example in which the epidermal cells of the cultured explant directly differentiate into embryoids. These columnar cells, known as epithelium, lie at the interface between the scutellum and the endosperm. At the time of culture, the epithelial cells contain a centrally placed nucleus surrounded by numerous lipid and protein bodies and underdeveloped plastids and mitochondria. The further development of plastids and mitochondria, the elaboration of the ER and Golgi bodies, and an increase in the ribosome density of the epidermal cells follow in quick succession in response to culture preparatory to embryogenic divisions. Whereas depletion of lipid bodies is characteristic of germinating embryos, in cultured scutellum most of the lipid bodies are retained in the epithelium (Jones and Rost 1989a). These observations show that cytoplasmic reorganization and changes in metabolic activity accompany the reprogramming of cells in the embryogenic pathway. Embryogenesis of the hypocotyl epidermis of cultured embryos of
477
Trifolium repens differs in one significant respect from that of rice; the first sign of embryogenic induction is a shift from regular anticlinal divisions to irregular periclinal and oblique quantal divisions (Maheswaran and Williams 1985). Any changes in the ultrastrucrural profile of the cell preparatory to division remain unclear; in T. repens, such changes, if they occur, may be viewed as characteristic of a cell induced to divide in a plane different from that of its neighbors and may not be related to embryogenesis. Of interest in the morphogenetic context are the patterns of microtubule organization in the cells. Studies that have best documented them at the ultrastructural level show that random arrays of microtubules predominate in the cytoskeletal framework of carrot cells grown in the presence of auxin, whereas they appear to be arranged in multiple layers in the cells of the early stage embryoids (Halperin and Jensen 1967; Wochok 1973a; Street and Withers 1974; Wilson, Israel and Steward 1974). Indirect immunofluorescence has shown that in the embryogenic cells of Picea marina, microtubules form an extensively branched structural meshwork that reflects a provision for limited cell expansion (Tautorus et al 1992). Reorientation of microtubules is also characteristically noted during embryogenic development of isolated protoplasts of Medicago sativa (Dijak and Simmonds 1988), P. glauca (Fowke et al 1990), and Larix X eurolepis (Staxe'n, Klimaszewska and Bornman 1994). It might be thought that microtubule orientation would provide clues to cell polarity and expansion; unfortunately, the different patterns of microtubule arrangement indicate that embryogenic cells are not committed, at least initially, to any specific direction or type of growth. Another aspect of the cell biology of somatic embryogenesis that has been studied is the cell wall carbohydrate complexes that may be involved in the embryogenic determination of cells. By screening with a monoclonal antibody, Pennell et al (1992) identified a carbohydrate epitope in the somatic cells of carrot and showed a direct correlation between expression of the epitope and embryogenic potential of the cell. As a marker for an early determinative event in embryogenesis, this carbohydrate is believed to act transiently before the somatic cell becomes embryogenic. Attempts to correlate embryogenic induction in carrot and maize cells to the distribution of other biopolymers and to calmodulin have not been successful. Although embryogenic and nonembryogenic cells react positively to the presence of carbohydrates
478
and calmodulin, the distribution patterns of these markers do not reveal a critical role in embryogenic induction (Timmers, de Vries and Schel 1989; Emons, Mulder and Kieft 1993; Timmers, Kieft and Schel 1995). Analysis of the nuclear cytology of certain carrot cell lines has indicated their preference for alternatives to simple mitotic divisions in culture. These abnormal divisions, referred to as reductional grouping or prophase chromosome reduction, mimic meiosis and generate a limited number of cells with a reduced number of chromosomes, mostly at the haploid level (Nuti Ronchi et al 1992a, b). If these haploid cells are in fact the potential embryogenic cells, as segregation at the DNA level indicates, we can assume that reduction in chromosome number must be an essential first step in the process whereby somatic cells assume an embryogenic role (Giorgetti et al 1995). Other studies have noted correlations between the tetraploid state attained by carrot cells during prolonged serial subculture and their inability to produce embryoids (Smith and Street 1974; Coutos-Thevenot et al 1990). Although the possible effect of the ploidy level of cells cannot be dismissed lightly, it would appear that the cells in a callus or suspension culture are very dissimilar from one another and therefore need not follow identical nuclear changes during embryogenesis. (ii) Attainment of Polarity It was seen in Chapter 13 that during zygotic embryogenesis, polarity is already established in the zygote by the concentration of cellular components toward one end of the cell. The first division of the cell separates an organelle-enriched terminal cell that forms the embryo proper from a basal cell that forms the suspensor. It is certain that polarity is already inherent in embryogenic cells attached to a proembryogenic mass or in superficial cells of an explant that directly differentiate into embryoids; here, daughter cells formed away from the point of attachment of the mother cells invariably differentiate into embryoids. Since no signs of polarity are evident in the free-floating embryogenic cells, polarity is believed to arise during subsequent differentiation of the cell into an embryoid. Attempts to demonstrate polarity in embryogenic single cells have not been entirely successful because no consistent changes in the generation of electric current or in the intracellular redistribution of specific ions that are important in the fixation of polar axis have been documented. The symmetric,
Adventive Embryogenesis
globular embryoid is the earliest recognizable stage of somatic embryogenesis in carrot in which electrical polarity is first detected. Currents are found to enter the apical pole of the embryoid and to leave the region near the presumptive radicle pole; this pattern remains constant up to the torpedoshaped stage. The traffic of electric current through the cells apparently results in ionic gradients, the inward current at the apical pole of the embryoid being carried by K+ influx and the outward current at the radicle end resulting from active H+ efflux (Brawley, Wetherell and Robinson 1984; Rathore, Hodges and Robinson 1988). If one considers that ionic currents are important in the accumulation of specific metabolites or in the facilitation of the polar transport of auxin, their role in the fixation of polarity in the embryoids becomes clear. In related studies, the application of a low-voltage electrical field and an increase in Ca2+ in the medium have been shown to greatly increase embryoid formation from alfalfa mesophyll protoplasts and carrot cell suspension cultures, respectively (Dijak et al 1986; Jansen et al 1990). The simplest interpretation of these observations is that the electrical pulse might impose polarity in the cells, which then leads to embryogenic development, and the redistribution of a specific ion that accompanies the fixation of polarity is attained by perturbing the Ca2+ concentration in the cells. The attractiveness of the role of Ca2+ in fixing polarity prompted a further study, which showed that globular embryoids arising from proembryogenic masses of carrot cells had increased levels of this ion (Timmers et al 1996). Electrical polarity similar to that detected in embryogenic cell clusters and embryoids also prevails in cell clusters of carrot proliferating in a medium containing 2,4-D. In molecular terms, this may mean that the cell clusters are embryogenically determined but are restrained from further development by the presence of 2,4-D in the medium (Gorst, Overall and Wernicke 1987). The distribution of activated calmodulin in cell clusters growing in a medium containing 2,4-D and the localized presence of calmodulin in a polar fashion in embryoids also suggest that embryogenic determination is already attained in the cell clusters (Timmers, de Vries and Schel 1989). The distribution of plasma membrane-associated arabinogalactan protein is a reliable indicator of polarity in embryogenic cells and embryoids of carrot, although the function of this protein is not well understood. The protein epitope defined by a monoclonal antibody is first expressed in the surface cells of proembryogenic masses and thus iden-
Somatic Embryogenesis
tities cells in terms of their position. The incipient polarity in the globular stage embryoid is reflected in the presence of the protein in several surface layers at the potential shoot end (Stacey, Roberts and Knox 1990). These and the other data considered earlier may constitute the beginnings of a mechanistic explanation of how certain cells of the proembryogenic mass begin to function as embryoid mother cells. Examination of some other parameters that characterize somatic embryogenesis gives few helpful insights into the basis of polarity. Serial observations have shown that in single embryogenic cells of carrot, the first division is unequal, giving rise to a small cytoplasmic cell and a large vacuolate cell. Further divisions are confined to the cytoplasmic cell and lead to the formation of a proembryogenic mass. Polarity of DNA synthesis and mRNA accumulation is observed in the proembryogenic masses when they are transferred to the embryoid-inducing medium (Nomura and Komamine 1986a, b). In the globular embryoid of carrot, polarity is first evident at the shoot pole by the appearance of esterase activity (Caligo, Nuti Ronchi and Nozzolini 1985), whereas in transformed carrot cells harboring GUS gene under the control of Agrobacterium tumefaciens
T-DNA gene-5 promoter, polarity is first noted in the root pole of the globular embryoid by the expression of GUS activity (Mattsson, Borkird and Engstrom 1992). Enzyme systems sensitive to regulatory functions concerned with establishment of the polarity of the shoot and of the root poles thus appear to be different. (iii) Nutritional Factors
The requirements for the most rapid growth of an embryogenic callus may not necessarily be the same as those that promote somatic embryogenesis in the tissue. Nevertheless, some components of the medium are important in causing enhanced growth of the callus and, secondarily, in increasing the number of embryoids formed. Nearly all the work on somatic embryogenesis has involved continued subculture of the callus in a medium that has mineral salts and sucrose as its base. The importance of the type of nitrogen present in the mineral-salt mix used in the medium in inducing somatic embryogenesis has generated much discussion. Even from the early days of plant tissue culture, the secondary phloem of carrot was famous for its ability to proliferate uncontrollably in a medium containing 2,4-D and nitrate as the sole source of nitrogen. Halperin and Wetherell (1965) showed that this callus dis-
479
plays little inclination to form embryoids when challenged in an induction medium unless it was originally grown in a medium containing a small amount of NH4+ as well. Using petiole-derived callus of carrot, it was found that cells from the callus grown in a medium containing NO3" fail to form embryoids in the induction medium even if the medium contains NH4+, whereas callus cells grown in an ammonium-containing medium form embryoids in large numbers in the induction medium with or without NH4+. The conclusion from these results was that NH4+ is essential for the expression of embryogenic competence in carrot cells, representing a contrast with NO3~, which probably facilitates the subsequent development of embryoids. But the role of NH4+ in somatic embryogenesis came under suspicion when it was claimed that the actual concentration of nitrogen in the medium, and not its form, is important in the somatic embryogenesis of carrot (Reinert, Tazawa and Semenoff 1967). A subsequent work clearly demonstrated the importance of reduced nitrogen by showing the contrast between very low embryogenesis in the presence of 5-95 mM KNO3 and high embryogenesis in a medium containing 12-40 mM KNO3 and 10 mM NH4C1. Ammonium ion alone is also effective when the pH of the medium is kept constant (Wetherell and Dougall 1976; Dougall and Verma 1978). Additional evidence for a specific requirement for NH4+ in the medium for somatic embryogenesis has come from work on Medicago sativa and other legumes (Walker and Sato 1981; Meijer and Brown 1987a; Greinwald and Czygan 1991). Several exogenously supplied amino acids regulate somatic embryogenesis in carrot to varying degrees, the most effective one being a-alanine. Glutamine, asparagine, glutamic acid, arginine, proline, and serine also promote initiation and development of embryoids to some extent, whereas histidine, leucine, valine, and methionine are considered inhibitory (Kamada and Harada 1979b; Nuti Ronchi et al 1984). Based on the lack of accumulation of a-alanine and the presence of an active alanine amino transferase system in carrot cells supplied with a-alanine, the effectiveness of this amino acid as a nitrogen source for somatic embryogenesis is attributed to its rapid incorporation and its utilization for protein synthesis during the period of cell divisions (Kamada and Harada 1984a, b). As compared to glycine and glutamic acid, a-alanine and proline induced a 5-10-fold increase in the number of embryoids in embryogenic cell cultures of Medicago sativa. NMR determination of the utilization of nitrogen by the cells has led to the inter-
480
esting finding that all four amino acids are utilized equally for protein synthesis by cells of both embryogenic and nonembryogenic genotypes. The only consistent difference is that the nonembryogenic cells have an increased level of free amino acids (Skokut, Manchester and Schaefer 1985). Although the use of NMR has eliminated the possibility of preferential uptake or metabolism of amino acids by the embryogenic cells, the real reasons for the effectiveness of oc-alanine and proline remain elusive. (iv) Hormonal Factors
A consideration of the critical issues relating to the role of hormones in somatic embryogenesis will begin here by reviewing the well-established facts. A key question is whether a cell is programmed for embryogenesis before it encounters auxin in the medium. Although it has been difficult to dissociate the role of auxin in promoting callus growth from its role in conferring embryogenic competence in the cell, the demonstration of direct embryogenesis in cultured epidermal strips of carrot has placed the issue in a clear light. The fact that the epidermal cells are not transformed into embryoids unless 2,4D is present in the medium, or unless they are exposed to the auxin for at least 12-24 hours, suggests that auxin treatment is necessary to bestow embryogenic competence on the cells (Kato 1968; Masuda et al 1995). In the more widely investigated cell suspension culture of carrot, somatic embryogenesis is induced readily by transferring cells from an auxin-containing medium to an auxin-free medium. The main focus of many of the physiological investigations on the role of hormones in somatic embryogenesis has been on the dilemma posed by the requirement for auxin to make cells embryogenically competent and, yet, the subsequent inhibition of embryogenesis by auxin. It has repeatedly been observed that 2,4-D is more potent than other auxins in making carrot cells embryogenically competent. Does 2,4-D modify the endogenous auxin levels of the cells to bestow embryogenic competence and later allow them to complete embryogenic development? This question is at present unanswerable; however, there is sufficient evidence of extensive changes in the endogenous auxin levels of embryogenic cells to implicate auxin in the somatic embryogenesis of carrot. Fujimura and Komamine (1979a) demonstrated the presence of IAA in carrot cells by the classical Avena coleoptile curvature method, but there was little change in auxin activity during
Adventive Embryogenesis
embryogenesis. Another part of this work showed that treatment of carrot cell suspension in the embryogenic mode with 2,4-D or the antiauxin 2,4,6-trichlorophenoxyacetic acid (2,4,6-T) during the first four days inhibits embryogenesis when the cell suspension is examined seven days later, whereas these same compounds do not affect embryogenesis when supplied during the last seven days of culture. In a similar way, proembryogenic masses of carrot cells proceed up to the globular stage and then cease to grow further in the presence of l,2-benz-isoxazol-3-acetic acid (a weak antiauxin) added at the beginning; the inhibitory effect is much diminished by later additions (Baldan et al 1995). Like 2,4-D, IAA also inhibits organized growth in embryogenically determined cells and causes them to revert to a callus mode of growth (Schiavone and Cooke 1987). The sensitivity of embryogenically determined cells to antiauxins suggests that a certain critical level of endogenous auxin may be important in the subsequent development of embryoids. Another work showed that carrot callus cells initiated in a medium containing 2,4-D had high levels of native IAA. When such embryogenically determined cells are transferred to an auxin-free medium, high steady-state levels of IAA are maintained during the first seven days, after which there is a decline at subsequent stages of embryogenesis; the cells also register a rapid decline in both free and conjugated 2,4-D during the first seven days. A nonembryogenic cell line of carrot analyzed simultaneously had similar 2,4-D levels but higher IAA levels than the embryogenic cells throughout the experimental period. At least two roles for auxin in somatic embryogenesis can be envisaged as a modest conclusion from this work. The transition of embryogenic cells to globular embryoids requires metabolic events mediated by high levels of endogenous IAA, whereas completion of embryogenesis has a lower threshold auxin requirement (Michalczuk, Cooke and Cohen 1992). Analysis of a variant cell line of carrot also confirms the hypothesis that high levels of IAA suppress the development of embryoids. This cell line, selected for resistance to 5-methyltryptophan, accumulates an excessive amount of IAA and continues to proliferate as a callus in a medium lacking 2,4-D; like wild-type cells grown in the presence of 2,4-D, cells resistant to 5-methyltryptophan do not produce embryoids (Sung 1979). How 2,4-D induces embryogenic competence in cells by increasing the level of native IAA is a difficult question to answer, but pointers have come from a variety of observations. For example, in cells
Somatic Embryogenesis
proliferating in the presence of 2,4-D, a pathway of synthesis of IAA from tryptophan predominates, whereas a nontryptophan pathway is the primary mode of IAA synthesis in embryoids developing in the absence of 2,4-D in the medium (Michalczuk et al 1992). Another view is that in the presence of 2,4D, part of the IAA is metabolized into indoleacetylaspartic acid. This would accord with a model in which IAA must suffer one of two fates: It is utilized for inducing embryogenic competence or it is converted into an inactive compound (Sasaki et al 1994). Other changes suggested by the presence of 2,4-D in the medium are decreases in the level of auxin-binding proteins (Lo Schiavo et al 1991) and embryogenic protein markers (Borkird, Choi and Sung 1986) and modifications of DNA methylation of key loci (Lo Schiavo et al 1989; Vergara et al 1990; Munksgaard, Mattsson and Okkels, 1995). We do not have any overall picture of the extent to which these mechanisms of 2,4-D action are utilized by carrot cells on their way to becoming embryoids. Role of Cytokinins. Cytokinins have generally found their way into culture media used to induce somatic embryogenesis in explanted tissues and organs, and a cytokinin has been an invariant component of the standard carrot cell suspension medium. Among the beneficial effects attributed to cytokinins is a partial alleviation, by the addition of low concentrations of kinetin, of the decline in embryogenic potential of carrot cells during prolonged growth in an auxin-containing medium (Wochok and Wetherell 1972). Addition of 2iP causes an increased frequency of embryogenesis in carrot cells grown in a low concentration of 2,4-D, which is suboptimal for embryogenesis, and in 5methyltryptophan-resistant cells, in which the process is impaired by high levels of endogenous IAA. Embryoids formed by this treatment comprise mostly globular and heart-shaped stages in the wild-type cells, but embryoids in the variant cell line do not develop beyond the globular stage (Sung, Smith and Horowitz 1979). It seems reasonable to postulate that cytokinin treatment promotes embryogenesis in carrot cells by stimulating divisions in the proembryogenic masses. This view finds support in studies showing that the capacity of cell clusters to undergo embryogenesis is enhanced when zeatin is present continuously in an auxin-free induction medium. If the hormone is added at different times after transfer of cells to the induction medium, high-frequency embryogenesis occurs in cells exposed to zeatin during the first three to four days after trans-
481 fer. It is from the behavior of cells in culture that the most obvious role of zeatin becomes clear; by enhancing cell divisions that occur in the proembryogenic masses during the first three to four days after their transfer, zeatin causes a change in the direction of differentiation of the cell clusters (Fujimura and Komamine 1980a, b). The use of cytokinins alone or in combination with auxin to induce somatic embryogenesis from various explants is now well established in several other species, including gymnosperms (Maheswaran and Williams 1984; Mo and von Arnold 1991; Norgaard and Krogstrup 1991). There is also a second role for cytokinins in somatic embryogenesis, one that causes a decrease in embryogenic response. Some investigators have pointed out that the failure of embryogenic development in explanted tissues of certain members of Poaceae is due to high endogenous levels of cytokinins (Rajasekaran et al 1987; Wenck et al 1988). The effect of cytokinins used in conjunction with auxin in callus and cell clusters of pea (Kysely and Jacobsen 1990), soybean (Lippmann and Lippmann 1984), and species of Lupinus (NadolskaOrczyk 1992) is to discourage them from differentiating into embryoids. Such effects of cytokinins are not all-or-none phenomena, because some embryoids are formed even in media containing optimum inhibitory levels of the hormone; in lupine, cytokinins inhibit embryogenesis when added along with NAA as the auxin but not when 2,4-D is used. The one question concerning cytokinin action in these systems that may be important in future research is whether the hormone interrupts embryogenic determination or inactivates embryogenic processes at later stages. Role of Other Hormones. A critical assessment of the role of GA in somatic embryogenesis is hampered by the conflicting reports of its effects on embryogenesis and by the difficulty of relating changes in the levels of endogenous GA to embryogenic events. In carrot cell cultures (Fujimura and Komamine 1975; Kamada and Harada 1979a) and in nucellar tissues of Citrus sinensis (Kochba et al 1978), GA inhibits embryogenesis by reducing the number of embryoids formed, whereas in a callus strain originating from cotyledons of cacao, the hormone promotes embryogenesis (H. Kononowicz and Janick 1984). Although characteristic changes in the concentrations of several endogenous gibberellins are observed during embryogenic transformation of carrot cells, the most striking one is the presence of severalfold higher levels of less-polar
482
Adventive Embryogenesis
GA in the embryoids than in the nonembryogenic phology, although a few are normal or near normal. cells proliferating in the auxin-containing medium. Abnormalities have probably arisen by imbalances In contrast, the nonembryogenic cells had high lev- in the hormonal constituents of the medium or in els of polar GA (Noma et al 1982). On a per embry- the culture environment. A considerable restoraoid basis, an increase in the concentration of free tion of the form of the embryoids to that of normal and water-soluble GA-like substances is found seed embryos is achieved if cells are subcultured in during embryogenesis in a hybrid grape cell sus- the dark in a medium containing ABA; in the prespension culture (Takeno et al 1983). From these ence of ABA in the light, restoration is less comobservations one is left with the feeling that changes plete (Ammirato 1974). This interesting result led to induced during somatic embryogenesis of carrot and studies that demonstrated an interaction between grape may be targeted at some aspect of GA metabo- ABA, zeatin, and GA during the normal maturalism, converting the hormone to a form that permits tion of somatic embryos from caraway cells embryogenesis. (Ammirato 1977). Supplementation of the medium We are also confronted with a set of conflicting with ABA also causes an increase in the number of data on the effects of ethylene on somatic embryoge- morphologically normal embryoids formed nesis because both promotory and inhibitory effects in somatic cell cultures of carrot (Kamada and of the hormone have been noted in the same system. Harada 1981), Picea abies (von Arnold and Hakman The main point of contention is that in carrot cell sus- 1988), wheat (Carman 1988), and Digitalis lanata pension, some ethylene-releasing agents stimulate (Scrophulariaceae) (Reinbothe, Diettrich and embryogenesis whereas others inhibit it; similarly, Luckner 1990). There have been attempts to relate some presumed inhibitors of ethylene biosynthesis globally the changes in ABA content of embryopromote embryogenesis whereas others are genic cells and embryoids to different phases of inhibitory (Roustan, Latch<§ and Fallot 1989a, b, 1990; somatic embryogenesis, but the data do not supNissen 1994). Nissen (1994) has pointed out the pit- port a connection (Kamada and Harada 1981; falls of using inhibitors of ethylene biosynthesis to Rajasekaran, Vine and Mullins 1982; Kiyosue et al monitor somatic embryogenesis because their effects 1992a; Etienne et al 1993a, b). may not be related to the inhibition of ethylene production. If ethylene stimulates somatic embryogene(v) Role of Secreted Molecules sis as claimed, it is clear that inhibition of embryogenesis by 2,4-D and other auxins cannot be The centerpiece of somatic embryogenesis is the due to auxin-induced ethylene production. Ethylene- utilization by cultured cells of metabolites supplied induced inhibition and promotion of somatic in the medium to evoke totipotent development. embryogenesis have been reported in cell cultures of Yet, it is ironic that as single cells and cell clusters Hevea brasiliensis (Auboiron, Carron and Michaux- metabolize nutrient substances, they release into Ferriere 1990) and Medicago sativa (Ke.pczyriski, the medium components that favor cell proliferaMcKersie and Brown 1992), respectively. tion and somatic embryogenesis. This was first The deepening understanding of the role of borne out by the observation that there is a quantiABA during maturation and dormancy of seed tative reduction in embryogenesis when carrot cell embryos has underlined the importance of this hor- suspension is diluted to low cell density and that a mone in somatic embryogenesis. As anticipated partial alleviation of this is achieved by the addifrom the work on seed embryos, exogenous addi- tion of conditioned cell suspension medium (Hari tion of ABA inhibits somatic embryogenesis in car- 1980). Later years have witnessed intense activity rot cell clusters (Fujimura and Komamine 1975; to characterize the secreted molecules, and a role Kamada and Harada 1979a, 1981) and prevents for extracellular proteins in somatic embryogenesis precocious germination of Brassica napus embry- in carrot and other plants is now firmly entrenched. oids (Maquoi, Hanke and Deltour 1993). Glycoproteins such as peroxidase and phosSuggestive of the involvement of protein kinase or phatase have been identified among the comprotein phosphatase in the ABA signal transduc- pounds released by embryogenic carrot cells into tion pathway, the levels of several acidic phospho- the medium, although it is doubtful whether they proteins are found to decline when embryogenic have any regulatory function in embryogenesis carrot cell suspension cultures are treated with (Ciarrocchi, Cella and Nielsen 1981). An interesting ABA (Koontz and Choi 1993). The majority of relationship exists between the release of a 65-kDa embryoids formed in a cell suspension of Carum glycoprotein and somatic embryogenesis in a carcarvi (caraway; Apiaceae) have an abnormal mor- rot cell suspension, whereas nonembryogenic cells
483
Somatic Embryogenesis
10 10
80
Figure 16.3. Rescue of embryogenic potential of a carrot cell suspension by conditioned medium. Open symbols represent the number of suspension cells per ml of culture medium containing 2,4-D. Closed symbols represent the embryogenic potential expressed as the number of embryoids of all stages produced by 10" suspension cells with additions as indicated: • , no addition; • , addition at day 19 of 50% (v/v) medium conditioned by a 7-day-old embryogenic suspension culture; A, addition at day 19 of a 50% (v/v) equivalent high-molecular-mass fraction (over 5 kDa) obtained from the conditioned medium of 7-day-old
embryogenic suspension culture; • , addition at day 19 of a 50% (v/v) equivalent boiled high-molecular-mass fraction obtained from the conditioned medium of 7-day-old embryogenic suspension culture; K, addition at day 19 of 50% (v/v) conditioned medium devoid of material over 5 kDa, obtained from a 7-day-old embryogenic suspension culture. The arrow at day 19 indicates removal of the hypocotyl explant in all cultures; at day 30 (arrow), all cultures were transferred to fresh medium with 2,4-D. (From de Vries et al 1988b. Planta 176:196-204. © SpringerVerlag, Berlin. Print supplied by Dr. S. C. de Vries.)
excrete a 57-kDa protein into the medium (Satoh et al 1986). The most promising attempt to implicate extracellular proteins in somatic embryogenesis in carrot cell suspension was the demonstration that addition of a crude, high-molecular-mass, heatlabile protein from a conditioned medium accelerates the process (Figure 16.3). Cell lines impaired in somatic embryogenesis fail to excrete one or more of these proteins, identified as glycoproteins, but their inability to regenerate embryoids is not inevitable, because it is partially reversed by the addition of a protein preparation from embryogenic cell lines (de Vries et al 1988a, b). The use of the glycosylation inhibitor tunicamycin illustrates a fruitful approach to identifying the excreted protein. This fungal antibiotic inhibits somatic embryogenesis, but not unorganized proliferation of carrot cells cultured in an induction medium (Lo Schiavo, Quesada-Allue and Sung 1986). Embryogenic competence of the cells is restored when the inhibitor is supplied along with
an adequate dose of the concentrated conditioned medium. If a protein present in the conditioned medium were to alleviate the effect of the inhibitor, the protein may have the same composition as the wild-type metabolite. The component in the conditioned medium that restored embryogenic competence in tunicamycin-inhibited cells was identified as a 38-kDa cationic peroxidase whose catalytic properties, rather than the carbohydrate moiety, were found to be decisive in the complementation assay. An explanation sought at the cellular level for the tunicamycin effect and its reversal by peroxidase is along the lines of the promotion of cell expansion by the inhibitor and its prevention by the secreted peroxidase (Cordewener et al 1991). Additional evidence linking extracellular proteins to somatic embryogenesis is provided by the analysis of a temperature-sensitive carrot variant cell line (ts-llc) isolated by chemical mutagenesis. Cells of the variant line usually grow and regenerate embryoids when propagated at 24 °C but are
484
unable to do so at 32 °C, in which environment they do not progress beyond the globular stage. Moreover, embryoids formed at the nonpermissive temperature display pronounced abnormalities in their morphology. Defects in the variant cells are overcome simply by growing them at the nonpermissive temperature in a medium conditioned by wild-type somatic embryos (Lo Schiavo et al 1990). Purification of the conditioned medium has led to the demonstration that rescue of embryoids in the ts-llc cell lines is accomplished by a 32-kDa glycosylated acidic endochitinase, which is apparently secreted at a reduced level at the nonpermissive temperature (de Jong et al 1992,1995).
Adventive Embryogenesis
Mention should also be made of secreted proteins that are not considered to be directly involved in promoting somatic embryogenesis in carrot cell cultures because their addition does not alleviate the inhibitory effects of tunicamycin. One such protein, designated as EP-1, belongs to a family of heterogeneous proteins and is found localized specifically in the pectin-containing wall material of the vacuolate, nonembryogenic cells. At the nucleotide level, EP-1 shows limited homology to the SLG and SLR genes of the self-incompatibility series and to genes encoding S-locus putative receptor kinases (van Engelen et al 1991, 1993). Another extracellular protein (EP-2) shows homology to plant lipid transfer proteins. The The specific role of endochitinase in overcoming involvement of EP-2 protein in embryogenesis can the effect of the mutation has not yet been defined. be seen by the expression of its transcripts in seed Experiments in which compounds that contain embryos and in the proembryogenic mass of cells oligomers of N-acetyl glucosamine were tested for when they decide whether to become embryoids or their ability to promote embryoid formation in ts- to continue to proliferate. Localized expression of llc cells at the nonpermissive temperature showed transcripts of the gene encoding EP-2 protein in the that the metabolic products of Rhizobium legumi- peripheral protoderm cells of globular and heartnosarum nodulation (NOD) genes, identified as shaped embryoids has suggested a role for lipid lipooligosaccharides, are effective in mimicking the transfer proteins in the transport of cutin monomers. effect of endochitinase (de Jong et al 1993). But it is The requirement of embryoids to maintain a proper far from clear how chitinase and the nodulation fac- osmotic balance in the liquid medium in which they tors lead to organized growth in somatic cells, a are bathed explains the presence of cutinized walls problem that is difficult to investigate because of the (Sterk et al 1991). Nonembryogenic cells of carrot possible involvement of unidentified signal mole- cell suspension culture also secrete a 47-kDa glycocules. A set of arabinogalactan proteins secreted protein (EP-4). This protein is localized specifically into the medium by carrot embryoids have been in the walls of the clustered cells, with the highest shown to restore embryogenic potential in a non- level in the cross walls. The gene that encodes this embryogenic cell line and to accelerate the produc- protein has a sequence similarity with the early tion of embryoids in embryogenic lines. The activity nodulin gene from alfalfa (van Engelen et al 1995). of this crude mixture depends upon specific frac- Although one can argue that these proteins do not tions that inhibit or promote embryogenesis have a direct role in embryogenesis, an alternative is (Kreuger and van Hoist 1993, 1995). A role for to suppose that they specify the type of cells prosecreted xyloglucan endotransglycosylase in duced in a suspension culture (Figure 16.4). somatic embryogenesis has been inferred from its Extracellular proteins have been identified durability to contribute to cell elongation through wall ing somatic embryogenesis in barley (Nielsen and loosening. A corollary to the idea that a carrot cell Hansen 1992), grape (Coutos-Thevenot et al 1992), suspension enriched for nonexpanding cells, such Citrus aurantium (Gavish, Vardi and Fluhr 1991, as those undergoing embryogenesis, secretes less of 1992), Digitalis lanata (C. Reinbothe et al 1992), this enzyme into the medium is that expanding Pinus caribaea (Domon et al 1994), Picea glauca nonembryogenic cells should secrete more of the (Dong and Dunstan 1994), and P. abies (Egertsdotter enzyme, as shown by Hetherington and Fry (1993). and von Arnold 1995). Growth of cell suspension This observation is consistent with a model that cultures of grapevine is associated with an ordered substantiates the requirement for rapid cell divi- process of protein secretion that depends more sions during the initial phase of embryogenesis. In upon the presence of auxin in the medium than summary, somatic embryogenesis in carrot depends upon its absence (Figure 16.5). In formal terms, the on the activity of more than one secreted glycopro- grapevine cell suspension may be described as one tein that probably helps to maintain the proper bal- bearing the closest resemblance to the carrot cell ance between cell division and cell enlargement in culture system; as in the carrot system, the two single cells and cell clusters in a suspension culture. extracellular proteins secreted by embryogenic
Somatic
Embryogenesis
Figure 16.4. Immunofluorescence localization of EP-4 protein in the embryogenic cells of carrot, (a, b) Serum produced
8 days
against EP-4 protein, (c) Preimmune serum. (From van Engelen etal 1995; photographs supplied by Dr S. C. de Vries.)
11 days
15 days
kDa
Figure 16.5. A Western blot showing the accumulation of proteins secreted into the medium by embryogenic (-aux) and nonembryogenic (+aux) cells of grape. Proteins from the culture media were separated by SDS-PAGE and were revealed by a peroxidase-labeled con-A staining protocol. Arrows on the right indicate positions of prominent bands, stds, molecular mass standards. (Reprinted from P. Coutos-
Thevenot et al 1992. Extracellular protein patterns of grapevine cell suspensions in embryogenic and nonembryogenic situations. Plant Sci. 86:137-145, with kind permission of Elsevier Scientific-NL, Sara Burgerhartstraat 25, 1055 KV Amsterdam, The Netherlands. Photograph supplied by Dr. P. Coutos-Thevenot.)
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Adventive Embryogenesis
cells nurtured in an auxin-free medium are 32-kDa Undoubtedly, these proteins are the products of cationic peroxidase and 10-kDa lipid transfer pro- gene activity of the cells, but only in a few cases tein (Coutos-Thevenot et al 1992, 1993). By secret- have the genes been isolated and characterized. ing endochitinases into the medium, embryogenic cells of barley also bear some similarities to carrot cells (Kragh et al 1991). Compared to nonembryo- 4. BIOCHEMISTRY OF SOMATIC EMBRYOGENESIS genic cells, embryogenic cells of P. caribaea have a highly specific protein secretion profile in which A suspension of embryogenic cells is an excelthe synthesis of at least five proteins identified in lent model of a differentiating system in which all two-dimensional gels is modulated at different biochemical activities associated with somatic stages of early embryogenesis. One of these pro- embryogenesis can be studied in a clonal populateins is analogous to germin, a glycosylated protein tion, but the goal of understanding the biochemsynthesized during germination of wheat embryos istry of somatic embryogenesis has hardly been (Domon et al 1994, 1995). Germinlike proteins are achieved. Much emphasis in the biochemical analyprobably the first markers of somatic embryogene- sis of somatic embryogenesis has been placed on sis in P. caribaea, and their synthesis during the comparing the synthesis of nucleic acids and properiod of the most active growth of somatic teins during embryogenic and during nonembryoembryos is significant in the general context of cell genic development of the cells. The results have division and cell elongation. By taking advantage made it possible to delve into the causal mechaof an observation that in P. abies, concentrated nisms of the process and to provide a baseline level extracellular proteins from embryogenic cell lines of information for investigations of somatic influence the morphology of less-developed embryogenesis at the molecular level. somatic embryos, researchers found an arabinogalactan protein and a protein showing antifungal activity to be the active components stimulating the (i) Changes in Macromolecule development of embryoids of different morphologSynthesis ical stages (Egertsdotter, Mo and von Arnold Some early experiments give us a look at the 1993; Egertsdotter and von Arnold 1995; Mo, metabolism of DNA, RNA, and proteins during the Egertsdotter and von Arnold 1996). Embryogenic growth and embryogenic development of carrot cells of maize and carrot secrete mucilaginous cells in suspension culture. Because the most funpolysaccharides into the medium, but it is not damental event in the cell suspension culture is an established whether they have a role in embryoincrease in cell number, DNA synthesis is essential genic induction (Everett, Wach and Ashworth 1985; for the process. This conclusion is based on data on Kikuchi et al 1995). changes in the DNA content of cells and the incorAlthough the terms in which somatic embryo- poration of labeled precursors of DNA synthesis. In genesis is viewed here attribute a promotory role cells growing in a medium lacking auxin, a rapid for secreted proteins, not all secreted proteins pro- replication of chromatin and other nuclear materimote embryogenesis. Nucellar cell cultures of C. als is a biochemical transition point in anticipation aurantium accumulate several glycoproteins in the of embryogenic induction (Wochok 1973b; Verma medium in a stage-dependent manner; of these, and Dougall 1978; Fujimura, Komamine and certain 53-57-kDa proteins secreted by proembryo- Matsumoto 1980; Kononowicz and Janick 1988). genic masses inhibit embryogenesis in newly initiOne interesting question is the extent to which ated proembryogenic masses and prevent the changes in cytoplasmic DNA are associated with subsequent transition of embryoid initials to glob- the acquisition of embryogenic potency. Consistent ular embryoids (Gavish, Vardi and Fluhr 1991, with a body of work showing that novel mitochon1992). This is unlikely to cause practical difficulties drial genome reorganization is seen in long-term in the continued growth of somatic embryos that is plant tissue cultures, there is some evidence suggenerally accomplished by transfer of the proem- gesting that similar changes may be correlated with bryogenic masses to another medium, in which embryogenic development of somatic cells. secretion of inhibitory proteins ceases. Changes are generally reflected in the abundance Summarizing, the potential of the extracellular of certain forms of mitochondrial genome, a proteins is that it is possible to purify the crude decreased abundance of certain hybridizing fragextracts and test the proteins for their ability to pro- ments and an amplification of others, or the mote or inhibit embryogenesis in vitro. appearance or disappearance of some hybridizing
Somatic
487
Embryogenesis
bands (Rode et al 1988; DeVerno, Charest and Bonen 1994). The specialization of carrot cells in the embryogenie pathway involves not only increased DNA synthesis but also increased RNA and protein synthesis. The augmentation of RNA and protein synthesis is noted beginning 2-4 hours after the transfer of cells to an auxin-free medium, although the amounts synthesized are too small to have an impact on their accumulation in the cells. Because no new cells are formed in the suspension culture during the first 12 hours of growth in the auxin-free medium, it is clear that the increases noted are due to cellular RNA and protein synthesis. Another small peak in the RNA synthetic activity of embryogenic cells is seen at 96 hours, coincident with the formation of the first embryoids (Fujimura, Komamine and Matsumoto 1980; Sengupta and Raghavan 1980a). Accompanying active RNA synthesis is also an accelerated synthesis of enzymes of the pyrimidine nucleotide pathway (Ashihara, Fujimura and Komamine 1981). On the whole, these data are merely suggestive and do not prove that RNA and protein synthesis is essential for the transition of carrot cells to the embryogenic mode. A determination of the RNA species synthesized by carrot cells during the early hours of embryogenic induction is important in establishing the role of mRNA in the process. It was found that as early as 6 hours after the transfer of cells to a hormone-free medium, there is a decrease in the synthesis of rRNA and a concomitant increase in the synthesis of minor species of RNA in the 12S and 18S regions. Prompted by the discontinuation of hormone treatment, embryogenic cells synthesize more mRNA than do nonembryogenic cells, and the periods of observed increase correlate well with embryogenic induction, which is when RNA synthesis appears to pick up (Sengupta and Raghavan 1980b). That no significant replication of rRNA genes occurs during somatic embryogenesis in carrot is supported by the results of experiments in which the amounts of rDNA in embryogenic and nonembryogenic cells were determined by DNA-rRNA hybridization (Masuda, Kikuta and Okazawa 1984). The experimental evidence favors the view that removal of auxin from the medium controls the biochemical events of embryogenesis by eliciting the synthesis of mRNA. Yet, it has not become unequivocally clear that gene activity for embryogenesis in carrot cells is initiated upon transfer of the cells to a medium lacking auxin. The issue is
whether proteins synthesized in cells nurtured in the auxin-free medium are coded on newly formed mRNA or on mRNA transcribed when the cells are in contact with auxin. Based on the striking similarities noted between the in vitro translation products of mRNA populations of proembryogenic masses and torpedo-shaped embryoids of carrot, Wilde et al (1988) have suggested that the gene expression program for embryogenesis occurs when cells are growing in the auxin-containing medium. As will be discussed later, many proteins necessary for embryogenic induction are already synthesized in the proembryogenic masses, probably using mRNA transcribed under the influence of auxin (Figure 16.6). (ii) Role of Embryonic Proteins
Analysis of protein metabolism during somatic embryogenesis has concentrated on isozyme changes and on the characteristic appearance and disappearance of a number of proteins and enzymes during embryogenesis. Results of these investigations have an important bearing on understanding the molecular events that trigger embryogenic development in somatic cells and in the formation of embryoids. An early electrophoretic analysis showed different patterns of expression of isozymes of peroxidase, esterase, and glutamic dehydrogenase in embryogenic and nonembryogenic cells of carrot. Especially significant was the presence of only slow-migrating isozymes of glutamic dehydrogenase in the embryogenic cells as compared to the presence of additional, fast-moving ones in the nonembryogenic counterparts (Lee and Dougall 1973). In this context there is now evidence showing that activities of protease (Carlberg et al 1984) and nitrate reductase (Rambaud and Rambour 1989) and isozymes of esterase (Chibbar et al 1988) and isoperoxidase (Joersbo et al 1989) can be used as potential markers of somatic embryogenesis in carrot cells. The phenomenon of isozyme changes is perhaps best characterized in an embryogenic line of Citrus sinensis that shows an upsurge of peroxidase activity concomitant with the appearance of embryoids. At the time when the rise in enzyme activity is under way, a new band typifying embryogenic activity appears in the isozyme profile (Kochba, Lavee and Spiegel-Roy 1977). Selective expression or repression of proteins or of isoforms of various enzymes has been noted during somatic embryogenesis in the following: explants of Brassica alboglabra (Chinese celery) petiole (Zee, Wu and
488
Adventive Embryogenesis
a Figure 16.6. Two-dimensional SDS-PACE of in vitro translated proteins from (a) proembryogenic masses and (b) torpedo-shaped embryoids of carrot. Closed arrows show translation products present in both; arrowheads show pro-
teins specific to torpedo-shaped embryoids. (From Wilde et al 1988. Planta 176:205-211. © Springer-Verlag, Berlin. Photographs supplied by Dr. T. L. Thomas.)
Yue 1979); embryos of maize (Everett, Wach and Ashworth 1985; Fransz, de Ruijter and Schel 1989), rice (Abe and Futsuhara 1989), barley (Coppens and Dewitte 1990), soybean (W. Liu et al 1991; Stejskal and Griga 1995), and wheat (Bapat, Rawal and Mascarenhas 1992); immature maize glumes (Rao, Suprasanna and Reddy 1990); lettuce cotyledons (Zhou et al 1992); ovules of Cercis canadensis (Fabaceae) (Buckley and Trigiano 1994); and apical meristem of date palm (Baaziz et al 1994). The relevance of biotin enzymes in somatic embryogenesis in carrot has been highlighted by the observation that the activity of acetyl-coA carboxylase increases substantially during embryogenesis. This could well play an important role in the accumulation of biotincontaining polypeptides associated with the transformation of embryogenic cell clusters into embryoids (Wurtele and Nikolau 1992).
cells as early as 4 hours of growth in fresh media but, in the presence of 2,4-D, they gradually diminished and finally disappeared (Sung and Okimoto 1981). From this it appears that the synthesis of embryonic proteins is an early event of somatic embryogenesis triggered by auxin; however, by its very presence in the medium, auxin also bars the continued synthesis of these proteins. Among other biochemical functions that change their expression during callus and embryogenic modes of growth in carrot cells is the synthesis of two callus-specific proteins (C-l and C-2) by cells grown in high-density cultures in a medium containing 2,4-D. Particular insight into the functions of embryonic proteins and callus-specific proteins has been provided by the protein synthetic profile of a mutant cell line of carrot that is resistant to cycloheximide in both callus and embryogenic modes of growth. Despite the inhibition of protein synthesis, the mutant cells growing as callus in the presence of 2,4D in the medium synthesize the embryonic proteins, but not the callus-specific proteins (Sung and Okimoto 1981, 1983; Borkird, Choi and Sung 1986). The corollary to these findings is that callus growth can proceed even in the absence of synthesis of callus-specific proteins, whereas the synthesis of embryonic proteins does not necessarily induce embryogenesis. The evidence based on these studies supports the concept that the callus trait and the embryogenic trait are coordinately expressed by a common mechanism, the activation of one function being accompanied by the elimination of the other. If
The concept of embryonic proteins as early developmental markers of somatic embryogenesis arose from a comparative two-dimensional SDSPAGE analysis of newly synthesized proteins of carrot cells growing in the presence or absence of 2,4-D in the medium. Comparison of the spectrum of proteins synthesized in cells growing for 12 days showed no pronounced differences in the nearly two hundred or so polypeptides spotted on the gels, except that the embryogenic cells had two additional proteins, E-l and E-2, designated as embryonic proteins. The surprising finding was that regardless of the presence or absence of auxin in the medium, these two proteins were synthesized in the
Somatic
Embryogenesis
489
this interpretation is valid, it might appear that in Reinbothe, Tewes and Reinbothe 1992b; C. somatic cells of carrot, the action of a single gene Reinbothe et al 1992), protein related to the onset provides the signal either for continued proliferative of cell divisions (Smith et al 1988), membrane proteins (Slay, Grimes and Hodges 1989), LEA progrowth or for embryogenesis. Specific proteins have been identified as molec- teins (Reinbothe, Tewes and Reinbothe 1992b), ular markers during somatic embryogenesis in pea oleosins (Ross and Murphy 1993), lipids (Turnham (Stirn and Jacobsen 1987), rice (Chen and Luthe and Northcote 1982; Feirer, Conkey and Verhagen 1987), Dactylis glomerata (Hahne, Mayer and Lorz 1989; Dutta et al 1991), and storage proteins 1988), Trifolium rubens and T. pratense (McGee et al(Crouch 1982; Shoemaker, Christofferson and 1989), soybean (Dahmer, Hildebrandt and Collins Galbraith 1987; Hakman et al 1990; Roberts et al 1992), Nicotiana plumbaginifolia (Reinbothe, Tewes 1990; Dahmer, Hildebrand and Collins 1992; and Reinbothe 1992b), Digitalis lanata (C. Krochko, Pramanik and Bewley 1992; Lai, Reinbothe et al 1992), Cichorium intybus (Hilbert, Senaratna and McKersie 1992; Misra et al 1993; Dubois and Vasseur 1992; Boyer, Hilbert and Burns and Wetzstein 1994; Krochkov et al 1994) are Vasseur 1993), and Coffea arabica (coffee; major chores of progressive somatic embryogeneRubiaceae) (Yuffa, de Garcia and Nieto 1994). In sis in cell and callus cultures of different species. callus lines of rice, several polypeptides in the The transient appearance and disappearance of 40-44-kDa range are detected more abundantly in subsets of proteins in globular and late stage the embryogenic callus than in the nonembryo- embryoids formed in cell suspensions of Nicotiana genic callus; these polypeptides also have counter- plumbaginifolia and Digitalis lanata provide an parts in the extant proteins of grain embryos of rice unusually clear picture of the transcriptional and (Chen and Luthe 1987). In leaves of chicory, in posttranscriptional controls of protein synthesis which embryoids are formed directly by the trans- during somatic embryogenesis. Changes in only formation of mesophyll cells, synthesis of the first less than 10% out of the approximately 250 embryonic proteins occurs within two days of cul- polypeptides identified in the in vivo protein synture of the explant, before any overt morphological thetic profile of both species were associated with or cytological signs of change are detected (Figure an altered in vitro translation profile (Figure 16.8). 16.7). During a five-day induction period when the Although these proteins are under developmental mesophyll cells acquire embryogenic competence, regulation at the transcriptional level, a posttranat least 15 new proteins that appear transiently or scriptional control operates for the majority of proaccumulate steadily have been identified (Boyer, teins of embryogenesis whose synthesis does not Hilbert and Vasseur 1993). A rather complex pat- correlate with corresponding changes in in vitro tern of protein changes is seen during somatic translated mRNAs (Reinbothe, Tewes and embryogenesis in callus cultures of Dactylis glom- Reinbothe 1992b; C. Reinbothe et al 1992). When erata. On the basis of an analysis of extant, in vivo the apical and basal halves of torpedo-shaped synthesized and in vitro translated proteins, undif- embryoids of carrot are analyzed separately, the ferentiated single cells and embryoids are found to stage-specific proteins are found to be asymmetripossess their own set of characteristic proteins, cally distributed in the two segments. This observawhich do not change during proliferative and tion might be reconciled with the existence of embryogenic modes of growth (Hahne, Mayer and biochemical differences between the two cell types or Lorz 1988). Thus, if embryonic marker proteins are spatially polarized gene expression patterns in the synthesized, they are made very early, at a stage developing embryoid (Racusen and Schiavone 1988). Compared to seed embryos, somatic embryos of that could not be identified precisely. A major reason for this is that, unlike in carrot, somatic different species show varying degrees of defects in embryos develop spontaneously in D. glomerata storage protein accumulation that generally result and cannot be reliably induced or suppressed by in lower amounts of protein in the embryoids (Crouch 1982; Shoemaker, Christofferson and changes in the medium composition. Galbraith 1987; Roberts et al 1990; Flinn, Roberts The production and accumulation of histone and Taylor 1991; Dahmer, Hildebrand and Collins and nonhistone chromosomal proteins (Gregor, 1992; Misra et al 1993; Burns and Wetzstein 1994; Reinert and Matsumoto 1974; Fujimura, Krochko et al 1994). Treatment of somatic embryos Komamine and Matsumoto 1981; Sanchez-Pina, with glutamine (Lai, Senaratna and McKersie Kieft and Schel 1989), cytoskeletal proteins (Cyr et 1992), maltose and sucrose (Burns and Wetzstein al 1987), embryoid-specific proteins (Dong, Satoh 1994), and more reproducibly with ABA (Dutta and Fujii 1988; Racusen and Schiavone 1988;
490
Adventive Embryogenesis
Figure 16.7. SDS-PAGE showing the changes in extant proteins associated with somatic embryogenesis in leaf segments of Cichorium intybus. (a) Noninduced leaves, (b) Leaves induced for 3 days, (c) Leaves induced for 5 days, (d) 5-day induced leaves after transfer to an expression medium (same as induction medium, but without glycerol) for 3 days. A, polypeptides showing increased accumulation; T , polypeptides showing decreased accumulation or not detected; -», polypeptides that remained constant during embryogenesis.
Embryogenesis-related polypeptides are indicated by large arrows and their absence is noted by circles. Polypeptides of general interest are numbered 1 to 51. (Reprinted from C. Boyer, J.-L. Hilbert and J. Vasseur 1993. Embryogenesisrelated protein synthesis and accumulation during early acquisition of somatic embryogenesis competence in Cichorium. Plant Sci. 93:41-53, with kind permission of Elsevier Scientific-NL, Sara Burgerhartstraat 25, 1055 KV Amsterdam, The Netherlands.)
and Appelqvist 1989; Roberts et al 1990; Kim and Janick 1991; Maquoi, Hanke and Deltour 1993; Misra et al 1993) leads to accumulation of storage products at levels closely approaching those of mature seed embryos.
with those in nucleic acids, have made them prime candidates for gene regulation in a variety of physiological processes in plants. The importance of polyamine metabolism in somatic embryogenesis was highlighted by the observation that induction of embryogenesis in carrot cells is accompanied by increased levels of the three polyamines within one to two days of their transfer to the hormone-free medium. On the other hand, a mutant cell line of carrot that fails to form embryoids but exhibits a growth rate similar to that of the wild-type cells does not show increased polyamine levels (Montague, Koppenbrink and Jaworski 1978; Fienberg et al 1984; Mengoli et al 1989). In line with
(iii) Polyamine Metabolism during Embryogenesis Polyamines, of which putrescine, spermidine, and spermine are the best known, are characteristically basic in nature and are distinguished from amino acids by the absence of a carboxyl group. Their ability to bind with acidic groups, especially
Somatic Embryogenesis
Figure 16.8. Composite diagram of polypeptides translated in a cell-free system by RNA isolated from somatic embryos of Digitalis lanata. Filled spots indicate polypeptides for which an exact counterpart was detected in the extant protein profile of embryoids; numbering is arbitrary. All other polypeptides synthesized in vitro had no counterpart in the extant protein profile. In vitro translated polypeptides whose mRNA levels changed during somatic embryogenesis are indicated by N and a number. Polypeptides indicated by arrowheads represent mRNAs that were developmental^ regulated and reached maximum level in the embryoids. WK indicates polypeptides translated in the absence of exogenous RNA. (From C. Reinbothe et al 1992; print supplied by Dr. S. Reinbothe.)
these observations, activities of arginine decarboxylase (an enzyme involved in the synthesis of putrescine from arginine) and of S-adenosylmethionine decarboxylase (which catalyzes the decarboxylation of S-adenosylmethionine in the putrescine-to-spermidine and the spermidine-tospermine pathways) are found to be higher in the embryogenic cells than in their nonembryogenic counterparts (Montague, Armstrong and Jaworski 1979; Fienberg et al 1984). A role for arginine decarboxylase and S-adenosylmethionine decarboxylase in somatic embryogenesis, suggested by these results, has also been substantiated by experiments that showed a significant reduction in embryogenic potential, with a concomitant decrease in the levels of polyamines, in carrot cells treated with oc-difluoromethylarginine (an inhibitor of arginine decarboxylase) or methylglyoxal bis(guanylhydrazone) (an inhibitor of S-adenosylmethionine decarboxylase). The inhibition of embryogenesis cannot be considered an indirect effect of inhibitors because it occurs without any decrease in the growth rate of cells and is reversible by the simultaneous addition of putrescine with a-difluoromethylarginine and
491
of spermidine with methylglyoxal bis(guanylhydrazone) (Feirer, Mignon and Litvay 1984; Minocha et al 1991). Another investigation showed that difluoromethylornithine, an inhibitor of ornithine decarboxylase, has the opposite effect on somatic embryogenesis in carrot because it accelerates the development of embryoids in a medium containing inhibitory levels of 2,4-D. This is accompanied by an increased accumulation of polyamines, especially putrescine, and by high arginine decarboxylase activity in the cells (Robie and Minocha 1989). An approach involving the expression of a mammalian ornithine decarboxylase gene in transgenic carrot cells has shown that the increased biosynthesis of putrescine leads to the improved ability of cells to regenerate somatic embryos (Figure 16.9). The presumed mechanism here is the provision of an effective alternative, provided by the mammalian enzyme, to the usual arginine decarboxylase pathway for the biosynthesis of putrescine (Bastola and Minocha 1995). Other examples of embryogenic systems in which polyamine levels and/or levels of corresponding enzymes correlate highly with somatic embryogenesis are Medicago sativa (Meijer and Simmonds 1988), Hevea brasiliensis (El Hadrami et al 1989), Vitis vinifera (Faure et al 1991), Picea abies (Santanen and Simola 1992), Cichorium intybus (Helleboid et al 1995), and P. glauca X P. engelmannii (Amarasinghe, Dhami and Carlson 1996). 120
3
5 Time (days)
Figure 16.9. Number of globular and torpedo-shaped embryoids in a control (N) and in a transgenic line of carrot (N14) transformed with a mouse ornithine decarboxylase cDNA. The cells were grown in a liquid medium without 2,4-D and embryoids were counted at different times after transfer. (From Bastola and Minocha 1995. Plant Physiol. 109:63-71. © American Society of Plant Physiologists.)
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Since the effects of polyamines on somatic embryogenesis in carrot cells appear to be antagonistic to those of ethylene, a close physiological link between these two compounds and somatic embryogenesis has been suspected. In a carrot cell line sensitive to ethylene, ethylene-induced inhibition of somatic embryogenesis is accompanied by a reduction in spermidine and spermine synthesis and in the activities of polyamine biosynthetic enzymes (Roustan, Latche and Fallot 1992). Consequently, ethylene might be considered to inhibit somatic embryogenesis in carrot indirectly by interfering with polyamine metabolism. 5. REGULATION OF GENE EXPRESSION
The logistical problems of somatic embryogenesis in the widely investigated carrot system suggest that one of the traits immediately affected during embryogenic induction of cells is the differential utilization of genetic information, which leads to the synthesis of new mRNAs and proteins. The complex morphology attained by embryoids also demands the operation of a program of continued gene expression with long-term implications. The carrot somatic embryo system, used extensively to help analyze questions concerning gene regulation, has proved to be unusual in one fundamental respect: Given the extensive overlap between in vivo synthesized proteins of nonembryogenic and embryogenic cells, there is the possibility that only minor changes in gene expression accompany embryogenic induction. Another option is to suppose that genes involved in embryogenesis are already expressed in cells during their proliferative growth. Both these possibilities are borne out by the identification and characterization of genes from embryogenic cells and somatic embryos of carrot. Despite the uncertainty about the relevance to the induction process of genes isolated from embryogenic cells, much of the thinking about gene activation during somatic embryogenesis has been focused on these genes. A group of three cDNA clones that are homologous to mRNAs preferentially expressed in embryoids of carrot were first isolated by screening an expression library with embryoid-enriched antibodies. The transcript levels of these clones are severalfold higher in the embryoids than in nonembryogenic cell clusters. A further embryo-related aspect of these recombinants concerns the similarity in the expression patterns of their transcripts in somatic and seed embryos of carrot (Choi et al 1987). The increasing
Adventive Embryogenesis
accumulation of transcripts of two of the clones (DC-8 and DC-59) in the heart-shaped and older embryoids has emphasized the developmental regulation of these genes. Although the genes are first expressed in the globular embryoids, they are not required for the formation of globular proembryogenic masses in the auxin-containing medium (Borkird et al 1988). Subsequent molecular and genetic analyses have shown that DC-8 and DC-59 genes both encode proteins showing characteristics of LEA proteins. Thus, the DC-8 gene encodes a hydrophilic protein that does not possess an obvious signal sequence at the N-terminal end (Franz et al 1989). As with other LEA genes, the embryoid-specific expression of both DC-8 and DC-59 genes is enhanced by the presence of ABA in the medium (Hatzopoulos, Fong and Sung 1990; Hatzopoulos et al 1990; Goupil et al 1992). Additional LEA genes isolated from embryogenic cells of carrot include DC-3 (Wilde et al 1988; Vivekananda, Drew and Thomas 1992), ECP-31, ECP-40 (Kiyosue et al 1992c, 1993), and EMB-1 (Wurtele et al 1993). The isolation and characterization of other genes - represented by clones DC-5 and DC-U (Wilde et al 1988), DC-2.15, DC-7.1, and DC-9.1 (Aleith and Richter 1990), AX110 (Nagata et al 1993), CEM-1 (Kawahara et al 1994), and CEM-6 (Sato et al 1995) - storage protein genes (Newton, Flinn and Sutton 1992), and genes for protein kinase (Lindzen and Choi 1995) and for GTP-binding proteins (Kiyosue and Shinozaki 1995) allow comparisons to be made between the expression patterns of different mRNA populations and their protein products during somatic embryogenesis. (i) Genes of Somatic Embryogenesis and Their Proteins
As indicated, the early investigations on gene expression that seemed pertinent to somatic embryogenesis concerned the two LEA genes, DC8 and DC-59. The latter encodes a basic polypeptide homologous to the lipid body membrane protein L-3 (16-kDa oleosin isomer) from maize embryos. Molecular analysis has shown that DNA-binding factors from embryoids interact with sequences within the 5'-flanking regions of the L-3 gene. These segments show similarity to the ABA-responsive sequences of the DC-8 gene; their relationship with LEA genes is probably similar, if not identical, to that seen in seed embryos (Hatzopoulos, Fong and Sung 1990; Hatzopoulos et al 1990). Clear evidence that initiation and main-
493
Somatic Embryogenesis
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tenance of the embryoid gene expression program occurs under conditions that allow cell proliferation in the auxin-containing medium was provided by Northern blot hybridization of RNA with probes made to cDNA clones DC-3, DC-5, and DC13. The pattern of expression of these recombinants is similar in the proembryogenic masses and embryoids of different ages. It is a prediction of the embryoid-specific nature of the transcripts that they would be expressed only in the embryogenic cells and embryoids and not in other parts of the plant; transcripts of the DC-3 gene meet this requirement (de Vries et al 1988b; Wilde et al 1988). DC-3 is a member of group 3 LEA genes belonging to a small gene family. Suggestive of the physiological role of DC-3 proteins in conferring desiccation tolerance in a way analogous to LEA proteins, expression of GUS gene linked to the promoter of DC-3 gene is found to increase in transgenic tobacco seedlings in response to water deficit and ABA application (Seffens et al 1990; Vivekananda, Drew and Thomas 1992).
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flit Figure 16.10. Northern blot showing the accumulation of EMB-1 mRNA during somatic embryogenesis in carrot cell suspension. Poly(A)-RNA was obtained from cell cultures and parts of the carrot plant as indicated. Cells lacking embryogenic capacity were grown to the log phase in media with or without 2,4-D. A single EMB-1 mRNA is present in somatic and zygotic embryos. (From Wurtele et al 1993. Plant Physiol. 102:303-312. © American Society of Plant Physiologists. Photograph supplied by Dr. E. S. Wurtele.)
The LEA genes ECP-31 and ECP-40, which belong to small gene families, have much in common between them. Nucleotide and amino acid sequences showed that ECP-32-encoded protein is homologous to a cotton LEA protein, whereas the ECP-40 gene has regions of homology in the ABAinducible RAB-16 gene from rice. Transcripts of lus coincides with the accumulation of transcripts both genes are found at high levels in the embryo- homologous to the wheat EM gene. genic cells and at low levels in developing somatic By hybridizing RNA gel blots with oligonuembryos; treatment of somatic embryos with ABA cleotide probes, relatively few differences are found leads to an increase in the accumulation of tran- in the temporal expression pattern of LEA transcripts of ECP-31 and ECP-40. The peripheral cells scripts during somatic embryogenesis in Nicotiana of carrot proembryogenic masses are the sites of plumbaginifolia and Digitalis lanata. In both species, accumulation of transcripts or protein products of transcript levels peak in globular embryoids and these genes (Kiyosue et al 1990a, 1991, 1992b, c, decline during subsequent embryoid develop1993). Another LEA gene that is highly expressed ment; this implies the possible existence of comin the somatic embryos of carrot, but is virtually mon regulatory pathways in the expression of undetectable in the proembryogenic masses, is LEA genes even in taxonomically unrelated species EMB-1. Transcripts of this gene are absent in non- (Reinbothe, Tewes and Reinbothe 1992b). A gene embryogenic cells invariably carried along with isolated from embryogenic cells of Betula pendula embryogenic cells into the induction medium, was found to be homologous to carrot DC-8 gene indicating that they are not induced by the transfer at the protein level; despite this, no positive signals of cells to an auxin-free medium but are directly of the gene were seen in B. pendula cells representrelated to embryogenesis (Figure 16.10). The in situ ing different stages of somatic embryogenesis pattern of accumulation of the transcripts of this (Puupponen-Pimia et al 1993). According to gene is very similar in both seed embryos and Reinbothe et al (1994), LEA transcripts are induced embryoids. The protein encoded by the EMB-1 upon treatment of leaf segments of N. plumbaginigene displays homologies to both EM protein of folia with jasmonic acid; this has led to the suggeswheat and a LEA protein of cotton (Ulrich, Wurtele tion that this compound alone, or along with ABA, and Nikolau 1990; Wurtele et al 1993). Corre et al may have a specific role in regulating the expres(1996) have shown that the acquisition of potency sion of LEA genes during somatic embryogenesis. for somatic embryogenesis by wheat embryo calThe transfer of proembryogenic cells of carrot to
Adventive Embryogenesis
494
* -*
Figure 16.11. Autoradiographs of globular embryoids of carrot showing localization of 35S-labeled antisense (a) and sense
(b) CEM-1 RNAby in situ hybridization. (From Kawaharaetal 1992; photographs supplied by Dr. R. Kawahara.)
a hormone-free medium also elicits transient expression of genes that probably have a role in triggering embryogenesis. The number of these genes is quite small and they even bear similarity to those expressed in the proembryogenic mass. Transcripts of several clones isolated from a cDNA library made to poly(A)-RNA of carrot cells grown for 8 days in the hormone-free medium show a transient accumulation from 3 days up to 16 days after transfer, coinciding with the globular or, rarely, with the heart-shaped embryoids. Thereafter, the prevalence of transcripts reaches such low values as to render them nonfunctional. The deduced proteins of three clones from this group (DC-2.15, DC-7.1, and DC-9.1) show unusual amino acid sequences, such as the presence of the core of a proline-rich domain in DC-2.15 and a large number of glycine residues in DC.7.1 and DC.9.1. Although these proteins bear some resemblance to cell wall proteins, the lack of clearly demonstrable function raises the question of their relevance to the onset of embryogenesis (Aleith and Richter 1990).
ribosomes during the synthesis of proteins for housekeeping chores in the cell, transcription of the gene is developmentally regulated and is confined to the cells of embryoids that are in an active state of division (Kawahara et al 1992,1994). This makes a good case that large amounts of the elongation factor are necessary to support cell divisions during embryogenesis although they may not be involved in triggering the signals for embryogenesis (Figure 16.11). Genes encoding a variety of proteins, including LEA, EM, and HSPs, have been isolated from somatic embryos of Picea glauca; preliminary evidence shows that these proteins function in the same way in somatic embryos as they do in zygotic embryos (Dong and Dunstan 1996). (ii) Storage Protein and Other Genes
The expression of storage protein genes in developing seed embryos of various angiosperms and gymnosperms was discussed in Chapter 15. The pattern of expression of storage protein genes in developing somatic embryos differs in some Screening a cDNA library of 3-day-old proem- respects from that described in the corresponding bryogenic masses of carrot has also identified genes seed embryos. Analysis of mRNA accumulation encoding a glycine-rich protein (CEM-6) as well as a and synthesis of storage proteins in somatic protein homologous to the eukaryotic translation embryos of Medicago saliva showed that although elongation factor la (CEM-1) (Kawahara et al 1992; the periods of accumulation of 7S and US storage Sato et al 1995). Although the latter protein func- proteins generally coincide with the availability of tions in the interaction of the amino acyl tRNA with their mRNAs, the accumulation of transcripts of 2S
Somatic Embryogenesis
storage protein precedes the first appearance of the protein by at least seven days (Krochko, Pramanik and Bewley 1992). The perfectly normal translation of 2S storage protein mRNA in a cell-free system confirmed that there is no internal damage in the messengers; from this, the repression of 2S storage protein synthesis during the early stages of somatic embryogenesis in M. sativa has been visualized as a translational control effect (Pramanik, Krochko and Bewley 1992). In contrast to the earliest expression of napin gene transcripts in the torpedo-shaped seed embryos of Brassica napus, transcripts of this gene tend to be present in somatic embryos of all stages. However, the up-regulation of napin gene expression in somatic embryos by ABA is similar to that seen in seed embryos (Fleming and Hanke 1993). The effects of ABA in stimulating and maintaining steady-state levels of various storage protein mRNAs are retained in somatic embryos of certain species of the gymnosperms (Flinn et al 1993; Leal et al 1995). These results were anticipated to some extent from reports of physiological studies of somatic embryos. The work reviewed has some theoretical interest since it shows that somatic embryos simulate zygotic embryos even at the molecular level. Although carrot embryoids of certain stages respond to a heat shock by synthesizing specific proteins, an observation that heat shock to globular embryoids can permanently arrest their development whereas embryoids of other developmental stages subjected to the same stress recover fully, led to a detailed analysis of the regulation of heatshock genes during somatic embryogenesis in this system. Using a cloned heat-shock gene encoding a small HSP, it was found that globular embryoids synthesize and accumulate significantly less heatshock mRNAs than do either embryoids of other developmental stages or undifferentiated callus cells. Despite this, globular embryoids synthesize the same spectrum of HSPs as those produced by cell suspensions or embryoids of other stages of development. The failure of globular embryoids to synthesize heat-shock mRNA has been attributed to the absence of transcriptional induction of the heat-shock gene. Globular embryoids appear to respond to heat shock by translating the existing heat-shock mRNAs at an elevated rate; in all probability, the ability of these embryoids to regulate mRNA sequestration or availability compensates for the low level of available mRNAs (Pitto et al 1983; Zimmerman et al 1989; Apuya and Zimmerman 1992). In the general context of a sen-
495
sitive developmental window, it is the dominance of translational control without the production of essential stage-specific transcripts that seems most significant in protecting globular embryoids from heat shock while curtailing their development. Transcripts of two HSP cDNAs isolated from alfalfa callus share the property of being induced in developing embryoids at noninductive temperatures although their accumulation is also temperature dependent (Gyorgyey et al 1991). Transcripts of both actin and tubulin genes are present in nonembryogenic cells, and the requirement for the cytoskeletal proteins associated with embryogenic induction is generally accompanied by an increased transcript accumulation (Cyr et al 1987; Reinbothe et al 1994). Studies on histone gene expression during somatic embryogenesis in alfalfa have revealed significant differences in the regulation of the transcript levels of two clones of histone H3 gene. In callus suspension cultures, histone H31 gene transcripts showed a sharp increase one day after a short pulse in a high concentration of 2,4-D and then decreased to very low levels, although differentiated embryoids contained significant levels of transcripts. Despite the presence of consistently high levels of H3-11 gene transcripts in the noninduced cells, 2,4-D treatment activated higher levels of expression of this gene in induced cells and in embryoids (Wu et al 1988; Kapros et al 1992). An underlying element of similarity in the expression of the two variant genes is that it is broadly confined to the cell cycle, but distinct mechanisms of regulation of expression probably prevail. A key regulator of the eukaryotic cell cycle is a 34-kDa phosphoprotein; during somatic embryogenesis in alfalfa, the time course of expression of a cDNA encoding an alfalfa protein kinase homologous to the yeast cell division cycle phosphoprotein (p34cdc2) coincides with the expression of H3-1 gene (Hirt et al 1991). The protein product of a homeotic gene has been implicated in soybean somatic embryogenesis. Homeotic genes of the type described in floral organ initiation (Chapter 2) contain the consensus homeobox domain and are also important in early embryo development in insects, amphibians, and mammals. A cDNA was isolated by screening a soybean somatic embryo cDNA library with a maize KN-1 cDNA clone. Although the deduced protein of the isolated clone shares a high amino acid identity with the KN-1 protein, it has also some unique features of its own, such as the presence of a long leader sequence prior to the translation start site of the protein (Figure 16.12). The
496
Adventive Embryogenesis
AATAAGAGAATTGTGTGTCGTGTTTGTTTTTGTTTGGTTTGTTGTAAGGTTAGCTAGTG AGTATTCTACACAAGGGTGGTGGTAGGGCAAAAAGGATAAGACAGTGAATGTGTGTGTGT GTGTGTGTGTGTGTGTGTTGTTGACAAGCAAAAGCTATGGAGGGTGGTAGTAGTAGCTCT M E G G S S S S AATGGCACTTCTTATCTGTTGGCTTTTGGAGAAAACAACAGTGGTGGGCTATGCCCAATG N G T S Y L L A F G E N N S G G L C P M ACGATGATGCCTTTGGTGACTTCCCATCACGCTGGTCATCATCCAATAAATCCTAGTAAT T M M P L V T S H H A G H H P I N P S N AATAATAATGTAAACACAAACTGTCTCTTCATTCCCAACTGCAGTAACAGTACTGGAACT N N N V N T N C L F I P N C S N S T G T CCTTCTATCATGCTCCACAATAATCACAACAACAACAAAACTGATGATGATGATAACAAC P S I M L H N N H N N N K T D D D D N N AACAACACTGGGTTAGGGTACTATTTCATGGAGAGTGACCACCACCACCATCACCACGGC N N T G L G Y Y F M E S D H H H H H H G AACAACAACAACAATGGAAGCTCCTCCTCCTCCTCCTCTTCTGCTGTCAAGGCCAAGATC N N N N N G S S S S S S S S A V K A K I ATGGCTCATCCTCACTATCACCGTCTCTTGGCAGCTTACGTCAATTGTCAGAAGGTTGGG M A H P H Y H R L L A A Y V N C Q K V G GCCCCGCCTGAAGTGGTGGCAAGGTTAGAAGAAGCATGTGCTTCTGCAGCGACAATGGCT A P P E V V A R L E E A C A S A A T M A GGTGGTGATGCAGCAGCTGGATCAAGCTGCATAGGTGAAGATCCAGCTTTGGATCAGTTC G G D A A A G S S C I G E D P A L D Q F ATGGAGGCTTACTGTGAGATGCTCACAAAGTATGAGCAAGAACTCTCCAAACCCTTAAAG M E A Y C E M L T K Y E Q E L S K P L K GMGCCATGCTCTTCCTTCAAAGGATCGAGTGCCAGTTCAAAAATCTTACAATTTCTTCC E A M L F L Q R I E C Q F K N L T I S S TCCGACTTTGCTAGCAATGAGGGTGGTGATAGGAATGGATCGTCTGAAGAGGATGTTGAT S D F A S N E G G D R N G S S E E D V D CTACACAACATGATAGATCCCCAGGCAGAGGACAGGGATTTAAAGGGTCAGCTTTTGCGC L H N M I D P Q A E D R D L K G Q L L R AAGTATAGCGGATACTTGGGCAGTCTGAAGCAAGAATTCATGAAGAAGAGGAAGAAAGGA K Y S G L G S L K Q E F M K K R K K G AAGCTACCTAAAGAAGCAAGGCAACAATTACTTGAATGGTGGAACAGACATTACAAATGG K L P K E A R Q Q L L E W W N R H Y K W CCTTACCCATCCGAATCCCAGAAGCTGGCTCTTGCAGAGTCGACAGGTCTGGATCAGAAG P Y P S E S Q K L A L A E S T G L D Q K CAAATCAACAACTGGTTTATTAATCAAAGGAAACGGCACTGGAAGCCTTCflSAGGACATG Q I N N W F I N Q R K R H W K P S 2 D M CAGTTTGTGGTGATGGATCCAAGCCATCCACACTATTACATGGATAATGTTCTAGGCAAT Q F V V M D P S H P H Y Y M D N V L G N CCATTTCCCATGGATCTTTCCCATCCCATGCTCTAGAAAATTATCCCTCGTTTGTGGGCT P F P M D L S H P M L * GCTGATAATAGATTCATAAACTCGTGCTGTCACTTATTAAAACCTTACAATTATTAATAT TAATTAATATGCATTCTAAGAAATCCTAGATTGCTATACTATAATATAGTACGCAGGTGT ATCCCTTGCTAGCTTTTTAGACGGTCCTTTGTGTGGATCATCTAGTTGAAGGAGTTATGA ATAAATAAAATTCCAT 1515
59 119 179 239 299 359 419 479 539 599 659 719 779 839 899 959 1019 1079 1139 1199 1259 1319 1379 1439 1499
Figure 16.12. Nucleotide sequence and the deduced amino acid sequence of a homeobox-containing gene {SBH-1) from somatic embryos of soybean. The homeobox nucleic acid sequence and the homeodomain protein are boxed. Putative polyadenylation signals are underlined.
The suggested acidic region is noted by double underline, The nucleic acid sequence that possibly encodes the short leader peptide is in bold type. (From Ma, McMullen and Finer 1994; print supplied by Dr. J. J. Finer.)
expression of transcripts of this gene is relatively specific for embryoids, in which it is developmentally regulated. It has been suggested that the protein encoded by the gene might play an important part as a transcription factor during embryoid development, especially at the critical globular to heart-shaped transition stage (Ma, McMullen and Finer 1994). A potentially interesting aspect of cell suspension cultures is that they can be used to follow for-
eign gene expression during the proliferative growth of cells and during embryogenic development. With carrot cell suspension cultures, an obvious hypothesis that can be tested is that certain genes are expressed in transformed embryoids but not in the nonembryogenic cells. This was found to be true in a transgenic carrot cell suspension harboring the chimeric gene construct containing the ROL-C promoter of the Ri plasmid of Agrobacterium rhizogenes and the GUS gene. Transformed embry-
Somatic Embryogenesis
oids showing high levels of reporter gene expression were produced when cells were transferred to a hormone-free medium, whereas proliferative growth of cells in a medium containing 2,4-D led to decrease in gene activity with increasing time in culture (Fujii and Uchimiya 1991). Studies on GUS gene expression using different promoter constructs demonstrated that a combination of several regulatory regions in the ROL-C promoter is required for the activation of GUS gene during somatic embryogenesis (Fujii, Yokoyama and Uchimiya 1994). These results show that activation of the ROL-C promoter occurs as a consequence of the developmental switch that leads to embryogenie divisions of the somatic cells. A highly active expression pattern is also directed by the promoter of A. tumefaciens T-DNA gene-5 during somatic embryogenesis in transformed carrot cells (Mattsson, Borkird and Engstrom 1992). (iii) Genetic Analysis of Somatic Embryogenesis
The foundations of our present concepts on the utilization of genetic information by developing systems rest not only on the isolation and characterization of genes and the protein products involved in the specification of organs, but also on the analyses of sophisticated mutant phenotypes. As seen in previous chapters, some of the best insights into the gene expression programs during floral organogenesis, pattern formation in embryos, progressive embryogenesis, and endosperm development have been gleaned from the study of mutants. Despite the obvious advantages of a mutational approach to studying gene action during somatic embryogenesis, no true genetic mutants have been isolated from embryogenic cell suspension cultures. In a broader sense, this is due to the spectrum of genetic and epigenetic variations that one encounters in the currently used somatic embryogenesis systems that obscure interesting mutant phenotypes (Zimmerman 1993). In an attempt to understand the genetic basis of somatic embryogenesis, some attention has been paid to variant cell lines and conditional mutants generated in the carrot system. In a previous section, reference was made to 5-methyltryptophanresistant cells of carrot, which present a striking contrast to the wild-type cells in their inability to regenerate embryoids in the induction medium. How can this effect on embryogenesis be explained by biochemical alterations modulated by a genetic mutation? This happens simply because in the vari-
497
ant cell line, the enzyme anthranilate synthetase, which catalyzes the synthesis of tryptophan, is resistant to feedback inhibition by the end product, tryptophan. Consequently, there is an accumulation of tryptophan, in turn leading to an increase in the endogenous level of IAA in the cells to the point at which embryogenesis is impaired (Sung 1979). It seems quite likely that similar mechanisms may exist in other auxin-resistant, variant cell lines that are impaired in embryogenesis; however, direct evidence is lacking (Filippini et al 1992). The extent of alterations in cellular properties during somatic embryogenesis as a function of gene regulation is best revealed by the responses to cycloheximide of wild-type cells and cycloheximide-resistant cells of carrot. In the former cell line, callus proliferation but not somatic embryogenesis is sensitive to the drug, whereas in the variant cell line both functions are insensitive to the drug. A central question is whether there are fundamental differences in the cycloheximide insensitivity of the two cell lines or whether a common mechanism operates differently in the two lines. It has been shown that cycloheximide resistance in the embryoids of both cell lines and in the callus of the variant line is caused by internal cellular mechanisms that inactivate the drug (Sung, Lazar and Dudits 1981). This observation falls in line with the notion that the general form of organization of the gene encoding for cycloheximide resistance is the same, but its expression is regulated differently in the two cell types. In oc-amanitin-resistant carrot cell lines that inactivate the drug during both the proliferative and the embryogenic phases of growth, the lesion appears to be due to changes in the properties of RNA polymerase II (Vergara et al 1982; Pitto, Lo Schiavo and Terzi 1985). Another mutational approach for the genetic dissection of somatic embryogenesis is the isolation of conditional mutants that are unable to produce embryoids at high temperature but that remain wild type at low temperature (Breton and Sung 1982; Giuliano, Lo Schiavo and Terzi 1984; Schnall, Cooke and Cress 1988). The growth of carrot cell suspension culture at 32 °C has allowed the retrieval of three primary classes of cells, which are impaired at different stages of embryogenesis (Figure 16.13). They are distinguished when nurtured in the embryogenic medium at the restrictive temperature by their phenotypes, such as continued growth as callus without producing embryoids and growth blocked at the globular stage of embryogenesis (Breton and Sung 1982). A systematic analysis of the variant cell line blocked at the
498
Adventive Embryogenesis
HA
ts-59
ts-85
24°C
32°C
Figure 16.13. Temperature-sensitive mutants of carrot impaired at different stages of embryogenesis during growth in media lacking 2,4-D. Parental line HA produces embryoids at both temperatures when transferred to a medium lacking 2,4-D, although later stages of embryoids are slightly affected at the higher temperature. Phenotypes of ts-59 and ts-85 are similar to the wild type at 24 CC in
media without 2,4-D. At 34 °C, ts-59 embryoids are arrested at the globular stage whereas ts-85 cells continue to grow as callus in media without 2,4-D. ->, embryoids at globular and heart-shaped stages; > , embryoids at torpedo-shaped stage; ->, undifferentiated callus; —», enlarged globular embryoids. (From Breton and Sung 1982.)
globular stage showed that the developmental block can be imposed by exposure to the high temperature either at the globular stage or at the globular to heart-shaped transition stage. The impact of this mutation is seen in the HSP profiles of the wild-type and mutant cells because the latter apparently fail to phosphorylate some HSPs in the 36^45-kDa range. Whether the differential modification of HSPs is timed to coincide with the arrest in development of the embryoids is not known (Lo Schiavo, Giuliano and Sung 1988). It is, however, doubtful whether genetic changes in the temperature-sensitive mutants can be discerned by protein analysis. Schnall et al (1991) showed that differences in the protein profiles of two conditional mutants generally reflect cell line differences, age differences, or random variations, rather than variations due to genetic lesions.
6. GENERAL COMMENTS An important concept that emerged in the wake of the discovery of somatic embryogenesis is the totipotency of plant cells. Totipotency invokes the ingrained genetic information in the cell as an essential element in the cell's ability to recapitulate the stages of embryogenesis in a different environment. If one compares the origin and position of a seed embryo and an embryoid, there is no commonality between them. Moreover, as the progenitor of the zygote and embryo, the egg contains specialized proteins that determine its functional competence after fertilization, whereas the somatic cell is almost at the end of its life as a differentiated cell. Yet, the final forms of embryos and embryoids are identical, suggesting that attainment of form in terms of the hierarchy of decisions is similar in both
Somatic
Embryogenesis
systems. Thus, the implications of the genomic information in the nucleus and its mediation through the environment of the cell in somatic embryogenesis are immense, but have not been properly appreciated. Carrot cell suspension culture continues to retain preeminence as the workhorse in much of the research into the physiology and molecular biology of somatic embryogenesis. That gene expression for embryogenesis is initiated even when the cells are undergoing proliferative growth instead of embryogenic growth is a particularly valuable insight into the mode of function of the genes in the carrot system. The expression of additional genes, which specify the function of the embryogenic cells, appears to proceed when the cells are induced in the embryogenic pathway. Also expressed are genes for the array of secreted proteins that have become recognized as a common feature of several somatic embryogenesis systems.
499 On the applied front, the virtually unlimited production of embryoids in some of our Economically important crop species has stimulated interest in plant propagation using automated systems to generate somatic embryos by cloning. Considerable progress has also been made in the production of synthetic seeds and desiccation-tolerant somatic embryos that can undergo dehydration without loss of viability. In recent years the technology of somatic embryogenesis from protoplasts is finding an increased application for the production of transgenic plants by the introduction of foreign DNA. Although numerous refinements await to be made in the manipulation of cell and protoplast culture systems to suit particular crops and in the encapsulation systems to produce mature, quiescent somatic embryos that functionally mimic seeds, new information is also needed to understand the basis of somatic embryogenesis in a large number of plants.
Chapter 17
Embryogenic Development of Pollen Grains 1. Historical Background (i) A Survey of Pollen Sporophytic Development (ii) Ploidy Level of Plants 2. Factors Controlling Pollen Sporophytic Growth (i) Genotypic Effects (ii) Stage of Pollen Development (iii) Medium Composition (iv) Thermal Stress (v) Role of the Anther Wall and Tapetum (vi) Embryogenic Competence of Pollen Grains 3. Cytology of Pollen Embryogenesis (i) Pollen Division Pathways (ii) Ultrastructural Cytology 4. Molecular Biology of Pollen Dedifferentiation (i) Dynamics of DNA, RNA, and Protein Metabolism (ii) Genes Expressed during Pollen Embryogenesis 5. General Comments
In the reproductive biology of angiosperms, the microspore and pollen grain represent paradigms for studies on cell differentiation. As described in Chapter 4, an important reason for this is that by a program of gene expression regulated in space and time, the unicellular microspore matures into the pollen grain and embarks on a pathway leading to terminal differentiation by the production of two cells with divergent developmental potential. A second reason is that the induction of embryogenic divisions in microspores or pollen grains of certain plants by tissue culture methods makes it possible to gain insight into mechanisms involved in the deflection of a typical gametophytic program into an atypical sporophytic pathway. This chapter will focus on this alternative developmental pathway of microspores and pollen grains, which gives rise to 500
embryoids and plantlets with the haploid or gametic number of chromosomes. This phenomenon is known as androgenesis, haploid embryogenesis, or pollen embryogenesis, but the last-mentioned term is preferred for use in this book. Irrespective of how the morphological and cytological changes converge to cause gametophytic or sporophytic types of growth in pollen grains, it seems certain that the same genetic blueprint is utilized to meet the informational demands of these transformation episodes. However, as will be shown in the following account, the biochemical and molecular basis for the embryogenic growth of pollen grains is much less understood than that for gametophytic development. The topic as developed here is based on the work done on representative species, and no attempt is made to cover the numerous reports of the induction of sporophytic divisions in pollen grains of diverse species or the manipulations of tissue culture systems required to induce such divisions or improve the efficiency of pollen embryogenesis. This chapter will evaluate the subcellular, biochemical, and molecular changes that are triggered as pollen grains enter the sporophytic program and also the intracellular and extracellular control mechanisms that modulate embryogenic induction. For comprehensive reviews of the tissue culture aspects of pollen embryogenesis, the reader is referred to Bajaj (1983, 1990); although these reviews give access to the names of genera and species in which pollen embryogenesis has been demonstrated, it should be noted that the list has grown substantially since these reviews were written. Condensed treatments of some segments of the topic covered here can be found in several other surveys (Raghavan 1986a; Sangwan and SangwanNorreel 1987b; Vicente, Benito Moreno and
Embryogenic Development of Pollen Grains
501
inducing the embryogenic development of pollen grains is that they are encased within the anther locule during the crucial period when the initial divisions in the embryogenic pathway occur. This makes it difficult to dissociate any influence of the anther wall and tapetum on embryogenic divisions from the more obvious effects of the components of the medium. For an analysis of the sporophytic development of pollen grains in the absence of any influence from the somatic tissues of the anther, the introduction of a method for inducing embryogenesis in isolated pollen grains proved to be a signifi1. HISTORICAL BACKGROUND cant improvement of the anther culture system The history of research on the transformation of (Nitsch and Norreel 1973). In this method, pollen pollen grains into embryoids began with a seminal grains from the anthers of flower buds of D. innoxia discovery by Guha and Maheshwari (1964). These pretreated at 3 °C for 48 hours are released into an investigators found that when excised anthers of isotonic medium and cleaned by filtration and cenDatura innoxia at the pollen grain stage were cul- trifugation. Aliquots of the pollen suspension are tured in a mineral-salt medium supplemented cultured in a liquid medium conditioned by the either with casein hydrolyzate, IAA, and kinetin or addition of an extract of cultured anthers of D. with coconut milk, grape juice, or plum juice, innoxia. Under these conditions, embryogenic embryolike structures appeared from the sides of pollen grains in the aliquot divide and form structhe anther in about six or seven weeks. Although it tures resembling zygotic embryos. was initially suspected that the embryoids might Viewed from a developmental angle, these have had their origin in the somatic tissues of the observations offer tangible evidence showing that anther, persuasive ontogenetic evidence of their differentiation of the angiosperm pollen grain is origin from the pollen grains, and hence of their not accompanied by an irreversible change in the haploid nature, was soon forthcoming (Guha and genome; rather, the same genome is reprogrammed Maheshwari 1966). It was found that in cultured to evoke totipotent development. The acquisition anthers, a variable but substantial number of of stable dedifferentiative changes by animal nuclei pollen grains acquired a selective advantage over irreversibly blocked from inducing morphogenesis the rest of the population and began to divide was demonstrated by the nuclear transplantation repeatedly to form multicellular masses. Later, the experiments of King and Briggs (1953). In these exine gave way, releasing the cellular masses that studies, it was shown that nuclei removed from a organized into typical bipolar embryoids. lethal frog hybrid (Rana pipiens X R. catesbeiana) and Moreover, had the embryoids originated from the transplanted into enucleate eggs of R. pipiens replisomatic tissues of the anther, they would be cate in the alien cytoplasm and promote hybrid diploid, rather than haploid, as was the case. development. Like the lethally blocked frog nuclei, A variation of the mode of pollen sporophytic the block to morphogenesis in the terminally difdevelopment described in D. innoxia was reported ferentiating pollen grain is removed by the simple in cultured anthers of Oryza sativa. Niizeki and expedient of culturing anthers in a mineral-salt Oono (1968) found that when anthers of rice were medium. The ability to produce haploid plants cultured at the mature pollen grain stage in a from pollen grains has made it possible to obtain medium supplemented with IAA, 2,4-D, and stable inbred lines in large numbers in a single step kinetin, multicellular bodies appeared from within for breeding purposes. Homozygous diploid lines the anther in about four to eight weeks. These sub- are generated by treating the haploid plants with sequently yielded dense calluses. Cytological colchicine. examination of the callus confirmed its haploid nature and, hence, its origin from the pollen grain. (i) A Survey of Pollen Sporophytic Transfer of the callus to a medium containing IAA Development and kinetin elicited the regeneration of shoot and root systems and the formation of plantlets by During the last three decades, research in pollen organogenesis. embryogenesis has proceeded along the three One problem with anther culture as a means of major lines just described and has generated a long Heberle-Bors 1991; Ferrie, Palmer and Keller 1995). Since this chapter follows immediately the one on somatic embryogenesis, it is reasonable to hope that embryogenic development of somatic cells may help to illuminate the development of pollen grains along a similar pathway. Compared to somatic embryogenesis, pollen grains employ a different strategy to become embryogenic, but the final products generated by dedifferentiated somatic cells and pollen grains are tantalizingly similar.
Adventive Embryogenesis
Figure 17.1. Pollen embryogenesis in cultured anthers of Nicotians tabacum. (a) A cultured anther split open to show the developing embryoids. (b) Representative embry-
oids from a cultured anther, (c) Origin of plantlets from a cultured anther. (Photographs supplied by Dr. R. S. Sangwan.)
list of plants in which pollen sporophytic growth is initiated in the embryogenic or callus pathways in cultured anthers and in isolated pollen cultures. Plants belonging to Solanaceae have consistently proved a good source of anthers for induction of the embryogenic type of development in pollen grains (Figure 17.1). Great success has been achieved with anthers of several species of Capsicum, Datura, Hyoscyamus, Lycopersicon, Nicotiana, and Petunia. Although the pioneering study on the anthers of D. innoxia included hormones and undefined natural plant extracts in the medium, later work showed that perfectly normal embryoids of pollen grain origin appear in anthers of this species cultured in a mineral-salt-sucrose medium (Nitsch 1972). It is also clear that hormonal additives are not essential ingredients of the medium for inducing pollen embryogenesis in cultured anthers of several species of Nicotiana (Sunderland 1971; Collins and Sunderland 1974). Since the first report of pollen embryogenesis in cultured anthers of N. tabacum, this species has been intensively studied with respect to the factors inducing embryogenic development of pollen grains (Nitsch and Nitsch 1969). Investigations on Hyoscyamus niger have testified to the great potential of the anther culture approach in this species for a molecular-cytological analysis of the dedifferentiation of pollen grains in the embryogenic pathway (Raghavan 1990c).
type of growth with a high degree of fidelity. Typical bipolar embryoids appear in cultured anthers of Brassica campestris, B. napus, B. nigra, B. pekinensis, B. juncea, Raphanus sativus, and Sinapis alba. A systematic study of the requirements for pollen embryogenesis in cultured anthers of B. campestris and B. napus showed that an elevated temperature of 30-35 °C during the early period of anther culture appreciably increases the yield of embryogenic pollen grains, embryoids, and plantlets for use in molecular and mutagenic investigations and for breeding programs (Keller and Armstrong 1978, 1979; Chame and Beversdorf 1988; Polsoni, Kott and Beversdorf 1988). Only isolated genera and species from other dicotyledonous families are reported to initiate the sporophytic type of growth from pollen grains of cultured anthers. Selected families, with the names of genera in which pollen grains of cultured anthers undergo a partial or complete embryogenic type of growth to yield embryoids given in parentheses, are Asteraceae (Helianthus), Bignoniaceae (Jacaranda), Caprifoliaceae (Sambucus), Caricaceae (Carica), Cucurbitaceae (Luffa), Euphorbiaceae (Hevea), Fabaceae (Arachis, Cajanus, Phaseolus, Pisum, Psophocarpus, Vigna), Hippocastanaceae (Aesculus), Liliaceae (Asparagus), Malvaceae (Gossypium), Arecaceae (Cocos), Primulaceae (Primula), Ranunculaceae (Anemone), Rutaceae (Citrus, Poncirus), Sapindaceae (Litchi), Scrophulariaceae (Digitalis), and Vitaceae (Vitis). For a period of time, it was believed that in mono-
Pollen grains of cultured anthers of plants belonging to Brassicaceae display the embryogenic
Embryogenic Development of Pollen Grains
503
cotyledons the usual mode of pollen dedifferentia- strated by spikelet culture in barley and rice, the tion involved an intermediate callus phase, and the enclosed pollen grains begin to divide in the embryogenic development of pollen grains from embryogenic pathway and appear outside the flocultured anthers of this group was known only in a rets as undifferentiated callus at a speed comparagenotype of Oryza saliva var. indica (Guha- ble to that attained by anther culture (Wilson 1977; Mukherjee 1973). In later years, with the help of Datta 1987; Kim and Raghavan 1988). specially concocted media and the careful selection Experimental approaches in the embryogenesis of genotypes grown under controlled temperature of isolated pollen grains spawned by the work on and photoperiodic conditions, pollen embryogene- Datura innoxia described earlier were designed to sis has been reported in cultured anthers of several develop a medium of known chemical composition members of Poaceae, such as maize, barley, wheat, to replace the anther extract and to induce embryoand rye. genic divisions in pollen grains cultured in a singlePollen grains of cultured anthers of many dicots step method. Although varying degrees of success and monocots have attracted attention for the ease have been reported in inducing embryoid or callus with which they form calluses; upon transfer of the formation in isolated pollen grains of several callus to a medium of a different composition, species investigated, it is in Nicotiana tabacum and plantlet regeneration occurs by organogenesis. For Brassica napus that the procedures have been certain genera included in Apiaceae (Coriandrum), streamlined to ensure a high frequency of embryoAsteraceae (Gerbera, Helianthus), Chenopodiaceae genesis (Figure 17.2). The method developed for N. (Beta), Convolvulaceae (Pharbitis, Ipomoea), tabacum begins with Percoll density gradient cenFabaceae (Albizzia), Gesneriaceae (Pelargonium), trifugation to obtain a homogeneous population of Iridaceae (Freesia, Gladiolus), Myrtaceae (Psidium),embryogenic pollen grains. If these pollen grains Onagraceae (Oenothera), Papaveraceae (Papaver), are first subjected to a starvation diet by being culRosaceae (Fragaria, Malus), Rutaceae (Citrus, tured in a medium lacking sucrose and glutamine Coffea), Salicaceae (Populus), Scrophulariaceae and are then transferred to a regular enriched (Digitalis), and Ulmaceae (Ulmus), this is perhaps medium, they divide in the embryogenic pathway the only route available to produce plants of pollen in high numbers (Kyo and Harada 1986). With a grain origin. Generally, the medium requirements slight modification involving culture of the Percoll for pollen embryogenesis or for the induction of fraction in a starvation medium of just water for pollen callus growth in cultured anthers of plants three days at 30 °C followed by transfer to a regulisted here do not differ materially from those con- lar medium, a high number of embryogenic divisidered earlier; for extremely recalcitrant species, sions are easily induced in pollen grains of N. they include a combination of plant hormones with rustica (Kyo and Harada 1985). The critical step in or without additives like coconut milk, yeast inducing embryogenic divisions in homogeneous extract, or potato extract. However, in the absence suspensions of pollen cultures of B. napus is a highof hard evidence, the reasoning advanced in sup- temperature shock by culture in the dark at 33 °C port of the role of specific components of the for three days followed either by a more comfortmedium in triggering embryogenic development able temperature of 25 °C or by incubation in colchicine for 18-42 hours and subsequent culture of pollen grains can be misleading. in the regular medium, both at 25 °C (Huang 1992; It is evident from this survey that pollen grains Zhao, Simmonds and Newcomb 1996). Detailed of the cultured anthers of plants belonging to a protocols for the pollen culture of both species have wide spectrum of families in both dicots and mono- been recently described (Raghavan 1995). With the cots respond to reprogramming cues by the pro- advent of techniques for the introduction of foreign duction of embryoids or calluses, but we still have DNA into plant cells, isolated pollen culture offers little inkling of how the dedifferentiation program singular advantages over anther or inflorescence is triggered. A major obstacle to the production of a culture for genetic transformation and in vitro large number of pollen embryoids and plantlets by selection of transformants. anther culture lies in excising the required number of anthers for culture, especially from small flower Another procedure for generating high-effibuds. A method of generating pollen-derived ciency cultures of embryogenic pollen grains relies embryoids and calluses in a labor-intensive way on the fact that in some plants, like tobacco, wheat, involves the culture in a liquid medium of pieces of barley, rice, and D. innoxia, anthers dehisce soon the inflorescence with attached flower buds of the after culture, allowing all or some of the enclosed appropriate stage of development. As demon- pollen grains to escape. If anthers are cultured in a
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Adventive Embryogenesis
Figure 17.2. Embryogenic development of isolated pollen grains of Nicotiana tabacum. (a) Nonvacuolate pollen grains with no starch grains at the time of culture, (b) 2 days after culture in the starvation medium, (c) 10 days after cul-
ture in the enriched medium showing multicellular pollen grains. Scale bars = 50 urn. (From Kyo and Harada 1986. Planta 168:427-432. © Springer-Verlag, Berlin. Photographs supplied by Dr. M. Kyo.)
liquid medium, the shed pollen can be handled for all intents and purposes as a typical pollen culture. By serially transferring cultured anthers at intervals to fresh media, embryogenic pollen grains and embryoids of different ages are obtained (Sunderland and Roberts 1977). The use of shed pollen culture to study the molecular biology of pollen embryogenesis leaves much to be desired because, in many cases, pollen grains are shed only after the initial events of embryogenic induction are completed inside the anther. In terms of the specific nutritional requirements for the dedifferentiation of pollen grains into embryoids, the role of unknown substances leaching out from the anther into the medium cannot be discounted. Pollen grains of gymnosperms have remained recalcitrant to tissue culture manipulations designed to induce embryogenic development. A recent work has shown that isolated microspores of Ginkgo biloba are able to form embryoids directly when nurtured in a medium with or without auxin supplementation (Laurain, Tremouillaux-Guiller and Ch6nieux 1993). This must surely require that under otherwise propitious conditions, microspores can regulate their dedifferentiation program using endogenous factors.
genetic variation observed among the progeny of plants originating from gametic cells cultured in vitro is known as gametoclonal variation (Morrison and Evans 1987, 1988). The frequency of nonhaploids in anther cultures varies in the different species. For example, in Brassica campestris and rye, no haploids have been reported, whereas other genera generally yield some haploids with a preponderance of nonhaploids (Sunderland and Dunwell 1977; Bajaj 1983). Because the origin of embryoids and calluses from the somatic tissues of the anther has been rigorously excluded in these studies, the reasons for the increased chromosomal complement of the cells must lie in the pollen grains or in the products of their division. It has been noted that in plants such as Datura innoxia, the proportion of nonhaploids regenerated from pollen grains varies with the age of anthers at the time of culture, haploids being predominant in anthers enclosing early stage pollen grains, but decreasing with later stages of pollen along with the concomitant appearance of nonhaploids of higher levels of ploidy (Engvild, Linde-Laursen and Lundqvist 1972; Sunderland, Collins and Dunwell 1974). There is little or no evidence that these pollen grains at the time of culture are anything but haploid, thereby strengthening the conclusion from cytological studies that nonhaploid embryoids might have arisen by nuclear fusion between division products of the pollen cells during embryogenic development (Sunderland, Collins and Dunwell 1974).
(ii) Ploidy Level of Plants
Given the interest in obtaining haploids, it is commonly observed in anther culture investigations that embryoids and plants formed directly from pollen grains, or formed through the intermediary of a callus, are not always haploids, but constitute a mixture of different levels of ploidy. The
Endopolyploidy seems to be the primary mechanism by which nonhaploid calluses are formed in anther cultures. Even though prolonged growth of
Embryogenic Development of Pollen Grains
the callus seems to increase the ploidy level of cells, this does not rule out the possibility that the initial growth of the callus is accompanied by chromosomal changes of a less extreme form. Not infrequently, calluses and plants regenerated from cultured anthers appear to be mixoploids constituted of cells of different ploidy levels. In Lycopersicon peruvianum (Sree Ramulu et al 1976)
and Hordeum vulgare (Mix, Wilson and ForoughiWehr 1978), in which the origin of mixoploids has been analyzed in some detail, evidence seems to indicate that more than one pollen cell is involved in mixoploid formation. Apart from changes in the chromosome number of regenerated plants, the varied cytological syndromes of the pollen cells and their progenies, such as chromosomal rearrangements, differential DNA amplification, deletions, and inversions, have been little investigated in pollen-derived plants. 2. FACTORS CONTROLLING POLLEN SPOROPHYTIC GROWTH A wide range of extracellular and intracellular factors appears to affect the induction of pollen sporophytic growth in cultured anthers. In the most widely investigated tobacco anther culture system, the following parameters have been considered important in optimizing anther and pollen efficiency: physiological state and conditions of growth of the donor plants, plant age, stage of pollen development, pretreatment of flower buds, temperature and photoperiodic regime of cultures, nature of the nutrient medium, composition of the culture vessel atmosphere, and the presence of nutrient and nonnutrient substances in the culture medium. Pollen sporophytic growth in some plants, especially those belonging to Poaceae, has shown a certain degree of genotypic specificity, although in others a discrete effect of the somatic tissues of the anther has been detected. A consideration of the effects of all of the factors that impinge on pollen embryogenesis is beyond the scope of this section; the intent here is to concentrate on a few selected factors that have contributed in important ways to the transformation of the incipient gametophyte to an independent sporophyte. As will be seen later, many questions remain to be answered before we have a complete understanding of the role of the various physiological factors in pollen dedifferentiation in anther cultures. For example, the relationship between the selection of anthers for culture and the stage of the mitotic cycle of the enclosed pollen grain is essentially
505
unknown. What are the critical ingredients of the nutrient medium that trigger sporophytic divisions? Is the embryogenic competence of pollen grains restricted by their development in the anther? (i) Genotypic Effects
From a historical perspective, the idea that hereditary differences between cultivars of a species play a role in the regenerative ability of pollen grains of cultured anthers arose from the work of Gresshoff and Doy (1972a, b). These investigators found that out of 43 cultivars of Lycopersicon esculentum and 18 lines of Arabidopsis
tested, only 3 within each species gave rise to haploid tissues of pollen grain origin. Subsequent investigations have shown that most plants, including cereals, show genotypic differences in pollen sporophytic growth from cultured anthers. A case of extreme genotypic specificity is illustrated in the protracted experimentation that ensued before suitable maize germ plasms were identified for anther culture. In the first reports of pollen sporophytic development in cultured anthers of maize published in China, only 9 out of 159 genotypes tested were found to yield embryoids or calluses from pollen grains (Miao et al 1978). Subsequent reports of the induction of pollen embryogenic or callus types of growth from maize anthers from outside China have invariably used Chinese or U.S.-Chinese hybrid germ plasms. In one investigation, using 12 U.S. genotypes, anthers of 4 responded positively to culture (Genovesi and Collins 1982). In another work, anther culture-responsive germ plasms that are of interest to maize breeders were identified, although only slightly over 50% of the genotypes tested responded positively (Petolino and Jones 1986). From an analysis of the response frequencies of crosses between high-yielding and low-yielding genotypes of maize, it seems that breeding for improved anther culture responsiveness appears possible through selection (Petolino and Thompson 1987). Based on these investigations, selection of high-yielding genotypes has gone hand in hand with improvements in the culture medium and pretreatment of spikes to obtain embryoids and calluses from pollen grains of cultured anthers of maize routinely and reproducibly. Since each cultivar is formed from the clonal descendants of a single fertilized egg, whatever may be the mechanism that accounts for the regenerative ability of pollen grains, it must be precise
506 enough to single out individual cultivars. A trend toward species specificity is observed with regard to the percentage of embryogenic pollen grains in anthers of different species of the same genus. This observation indicates that although genotypic differences may not account for all known cases of failure to obtain pollen dedifferentiation in cultured anthers, this is a factor to be reckoned with in highly heterogeneous populations of plants. (ii) Stage of Pollen Development
Adventive Embryogenesis
(Kott, Polsoni and Beversdorf 1988; Pechan and Keller 1988; Binarova et al 1993). Ultrastructural studies combined with various cytochemical, immunochemical, and in situ hybridization approaches have shown that at the most favorable stage for embryogenic induction, the microspore nucleus in Capsicum annuum has features associated with active transcription (Gonzalez-Melendi et al 1995). A challenging problem is to establish these relationships in a satisfactory way for pollen embryogenesis in a wide range of plants.
It is now accepted that the cytological stage of (iii) Medium Composition the pollen grain in the anther at the time of culture is critical for the induction of sporophytic diviAccording to our current understanding, sions. Although this stage varies between species, anthers of a few species respond to embryogenic the developmental window lies between the early induction when they are cultured in a simple unicellular and the bicellular stage of the pollen medium containing only macronutrient salts, grain. In Nicotiana tabacum, anthers are generally micronutrients, vitamins, myo-inositol, and responsive when they are cultured at periods sucrose. Despite its widespread use as a carbon beginning with the liberation of microspores from energy source in supporting plant regeneration in the tetrad and ending with bicellular pollen grains, tissue cultures, sucrose has not been favored in but embryoids are readily formed in high numbers anther cultures of barley, potato, and wheat, in when anthers are cultured at the unicellular which it has been replaced by maltose. The differmicrospore stage or as the microspore begins to ences in the metabolism of sucrose and of maltose divide (Sunderland and Wicks 1971). However, in microspore cultures of barley have been used to with shed pollen cultures and isolated pollen cul- weigh the reasons for the superiority of maltose tures of tobacco, bicellular pollen grains yield over sucrose. It appears that compared to the slow embryoids with high frequency (Sunderland and metabolism of maltose, sucrose is metabolized Roberts 1977; Heberle-Bors and Reinert 1979). The rapidly, leading to the accumulation of toxic cytologically responsive stage in Beta vulgaris amounts of ethanol within the microspores (Scott, extends from close to the end of meiosis to the early Lyne and ap Rees 1995). unicellular microspore (van Geyt, D'Halluin and It is now known that anthers of the vast majorJacobs 1985), whereas the mid-unicellular stage is ity of plants studied require either an auxin or a optimal for barley (Wheatley, Marsolais and Kasha cytokinin or a combination of both in the medium. 1986). When embryoid formation was quantified as There is strong evidence indicating the existence of a function of pollen developmental stage, the best wide variations in the kind and concentrations of results are obtained at the unicellular stage of both these groups of hormones that are necessary pollen development in cultured anthers of for inducing and sustaining the pollen sporophytic Hyoscyamus niger (Raghavan 1978), Peltophorum pte-type of growth in cultured anthers of various rocarpum (Fabaceae) (Rao and De 1987), Brassica species. It is of interest to note that when pollen napus (Kott, Polsoni and Beversdorf 1988), grains of cultured anthers form calluses instead of Simocalamus latiflora (bamboo; Poaceae) (Tsay, Yeh embryoids, they do so in a medium enriched with and Hsu 1990), and Arachis hypogaea (Willcox et al auxins and cytokinins; subsequent regeneration of 1991), and rarely have anthers containing pollen plants by organogenesis occurs when the callus is grains at earlier or later stages of development been transferred to the basal medium, to a medium conas bountiful. This raises the question of whether taining cytokinin with a low level of auxin, or to the embryogenic fate of pollen grains is deter- one containing a weak auxin. The hormones thus mined by the stage of the mitotic cycle of the first perform a morphogenetic function, since their haploid microspore mitosis. It has been claimed absence from the medium does not provoke pollen that in B. napus, embryoids are induced in isolated dedifferentiation. pollen grains cultured from Gl through G2 phases, A new level of understanding of the role of auxbut the development of the dividing cells was not followed far enough to relate this to embryogenesis ins and cytokinins in inducing the embryogenic development of pollen grains in cultured anthers
Embryogenic Development of Pollen Grains
has been achieved in some cases in which these hormones have been found to modify the process. For example, although auxins such as IAA, NAA, and indolebutyric acid (IBA), and cytokinins such as zeatin, kinetin, and benzylaminopurine, are effective in enhancing the production of pollen embryoids in cultured anthers of Datura innoxia, the best response is obtained with zeatin and kinetin. This has led to the suggestion that embryogenic divisions are related to the maintenance of a certain level of endogenous hormones in the pollen grain (Sopory and Maheshwari 1976). Pollen grains in cultured anthers of Hyoscyamus niger respond to moderately high concentrations of 2,4-D in the medium by regenerating calluses rather than embryoids. In cultured anthers, compared with the basal medium, auxin does not affect the number of pollen grains dividing in the sporophytic pathway or the patterns of early division sequences involving the vegetative and generative cells. However, after release from the exine, undifferentiated growth prevails in the cellular mass and results in a small nodule of callus (Raghavan 1978). These results augment the general impression that 2,4-D does not interfere with the embryogenic induction of the pollen grain but affects the subsequent growth of embryoids. In contrast, the role of cytokinins such as kinetin, benzylaminopurine, zeatin, zeatin riboside, and 2iP is one of reducing pollen efficiency in cultured anthers of H. niger. During prolonged growth of anthers in media containing cytokinins, there is a tendency for the transformation of all or parts of the surviving embryoids into calluses (Raghavan and Nagmani 1989). The deleterious effects of cytokinins on pollen efficiency in cultured anthers of H. niger raise questions about the wisdom of their widespread and arbitrary use in anther culture media.
507
lated ethylene production in cultured anthers (Reynolds 1987). Unlike IAA, 2,4-D does not stimulate ethylene biosynthesis by cultured anthers, nor does ethylene cause callus growth in the absence of 2,4-D in the medium (Reynolds 1989). Since artificially increasing the concentration of intracellular free Ca2+ in cultured anthers by the ionophore A23187 enhances IAA action in inducing embryogenic divisions in pollen grains and in partially substituting for 2,4-D in promoting pollen callus growth, the involvement of Ca2+ in signal transduction during pollen sporophytic growth has been suggested (Reynolds 1990). Among other additives to the culture medium, activated charcoal has beneficial effects on pollen embryogenesis that seem to be well established. The most convincing explanation of these effects stems from the assumption that charcoal might adsorb inhibitors that are released from the anther, or present in the agar, or produced by the degradation of other metabolites present in the medium. Phenolic substances appear to be prime candidates for the role of anther-derived inhibitors. In the anther culture of Anemone canadensis (Ranunculaceae), the concentration of phenolic substances is found to be high in the culture medium lacking activated charcoal. The addition of activated charcoal, which reduces the concentration of phenolics to less than 20% of the original value, also enhances embryogenesis in cultured anthers (Johansson 1983). It is likely that an increase in the frequency of pollen embryogenesis reported to be produced by polyvinylpolypyrrolidone in Datura innoxia anthers is attributable to its well-known action in adsorbing phenolic compounds (Tyagi, Rashid and Maheshwari 1981). Whether this compound, like activated carbon, can be used on a wider level to enhance anther productivity remains to be seen.
Several workers have suggested a role for ethylene in pollen embryogenesis in cultured anthers (iv) Thermal Stress (Dunwell 1979; Babbar and Gupta 1986; Reynolds 1987; Biddington, Sutherland and Robinson 1988; In several plants, temperature shocks are effecCho and Kasha 1989). In Datura metel, the ethylene tive in inducing a parthenogenetic activation of the precursor, methionine, and the ethylene-releasing egg that leads to the production of haploid plants. compound, ethrel, were found to stimulate embry- Similarly, a cold treatment given to plants has been oid formation, whereas ethylene antagonists like known to alter the plane of division of the pollen Co++ and Ag+ decreased anther productivity in cul- grains and to result in symmetrical cells. In the ture (Babbar and Gupta 1986). According to wake of the discovery of pollen embryogenesis, the Reynolds (1984a, 1986), the presence of a moderate administration of temperature shock has become a concentration of IAA in the medium leads to routine procedure to enhance the embryogenic embryogenic development in pollen grains of cul- response of pollen grains in cultured anthers. The tured anthers of Solanum carolinense, whereas the first reports of favorable effects of low temperature addition of 2,4-D causes pollen callus formation. on pollen embryogenesis were concerned with the The effect of IAA was attributed to auxin-stimu- induction of embryogenic divisions in the isolated
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Adventive Embryogenesis
pollen culture of Datura innoxia described in a pre- cold treatment of anthers of D. metel and tobacco vious section (Nitsch and Norreel 1973). By follow- leads to a rapid accumulation of proline, y-aminobuing a rigid program of cold stressing the flower tyric acid, and glutamic acid, possibly for the nutribuds of tobacco grown under short-day conditions, tion of embryogenic pollen grains (Sangwan and researchers have formulated procedures to ensure Camefort 1978). That the subcellular morphology high-frequency ab initio pollen embryogenesis. This of the pollen grains and tapetum may exist in an is accomplished by selecting anthers from plants altered state in cold-treated anthers of D. metel is induced to flower in short days at 15 °C, subjecting suggested by observations that pollen vacuoles them to an additional cold treatment at 10 °C for 10 become filled with bands of material of alternating days in the dark, and isolating embryogenic pollen high and low electron density and that the tapetum grains by sucrose-Percoll gradient centrifugation is disorganized (Camefort and Sangwan 1979; (Rashid and Reinert 1981, 1983). Sporophytic divi- Cadic and Sangwan-Norreel 1983; Sangwan and sions are initiated only rarely in pollen grains of Camefort 1984). Although these observations do barley anthers cultured without a temperature not establish a causal relationship between the stress, whereas culture of anthers excised from ultrastructural and biochemical changes and the spikes or tillers pretreated at 4 °C induces pollen increase in anther response, the ideas must be sporophytic divisions in high numbers (Huang and included in any model that attempts to explain the Sunderland 1982). Two striking effects of cold effect of thermal shock on pollen embryogenesis. stress are seen in anthers of Petunia hybrida. First, there is an increase in anther productivity when cold-stressed anthers are cultured; secondly, sim(v) Role of the Anther Wall and Tapetum ply incubating cold-stressed buds at a slightly Circumstantial evidence for a role for the higher temperature for 24 hours without any culsomatic tissues of the anther in pollen embryogenture medium is sufficient to induce embryogenic esis was provided by the observation that embryodivisions in pollen grains (Malhotra and genic development is more readily induced in Maheshwari 1977; Babbar and Gupta 1980). Among pollen grains enclosed within the wall layers and other plants, a cold shock to flower buds increases tapetum of an intact anther than in isolated pollen the yield of embryoids and plantlets in cultured cultures. A variety of related experiments subseanthers of Hyoscyamus niger (Sunderland and quently indicated that chemical factors emanating Wildon 1979), D. metel (Gupta and Babbar 1980), from the wall layers and tapetum of the anther, Lycium barbarum, L. chinense (Solanaceae) (Gu, Gui rather than a physical constraint provided by these and Xu 1984), and Papaver somniferum (Dieu and cells, trigger sporophytic divisions of pollen grains. Dunwell 1988). Especially in several species of An early transplantation experiment showed that Brassica, a short period of culture at a higher temembryogenic development of free pollen grains of perature before incubation at 25 °C generally stimNicotiana tabacum is assured if they are incubated in ulates embryoid formation in anther cultures the locules of anthers of the same species, or of N. (Dunwell, Cornish and de Courcel 1985; Arnison et glutinosa or Petunia hybrida (Pelletier and Ilami al 1990). 1972). It will be recalled that in the first successful induction of embryogenesis in isolated pollen culThe underlying mechanism by which thermal tures of Datura innoxia, the regular mineral-salt shock increases the frequency of pollen sporophytic growth in cultured anthers is unknown, but medium was supplemented with an extract of culsome information is available to narrow down the tured anthers of the same species. In line with the possibilities. One view is that a cold shock causes a postulated role for substances released from the transient increase in the formation of pollen grains anther is another observation, namely, that a with two symmetrical cells and that these pollen medium conditioned by the growth of barley grains are probably more favorably disposed to anthers improved the yield of pollen calluses in enter the embryogenic pathway than pollen grains cultured barley anthers (Xu, Huang and with asymmetrical cells (Nitsch and Norreel 1973). Sunderland 1981). Fractionation of the medium led Another interpretation is that a low temperature to the isolation of two low-molecular-mass commight impede the senescence of somatic tissues of pounds possessing a conditioning effect on culthe anther and thus ensure the survival of a great tured pollen grains of barley (Kohler and Wenzel number of pollen grains that embark on embryo- 1985). Vergne et al (1993) have found that a 32-kDa genic divisions (Sunderland and Roberts 1979). protein accumulates in the somatic tissues of Data from amino acid analysis have shown that anthers of maize during cold pretreatment of the
Embryogenic Development of Pollen Grains
tassel; the conclusion that this protein is synthesized in the anther wall or tapetum relys on the assumption that it is not secreted by the microspores. At the histological level, in cultured anthers of Hyoscyamus niger and of barley, pollen grains lying in the vicinity of the tapetum are found to dedifferentiate into embryoids and calluses, respectively (Raghavan 1978; Idzikowska, Ponitka and Mtodzianowski 1982). This leads to a consideration of the chemical nature of the substances produced in the somatic tissues of the anther. Based on amino acid analyses, it has been claimed that the culture of anthers of N. tabacum is associated with a rapid increase in the levels of serine and glutamine (Nitsch 1974b; Homer and Pratt 1979); interestingly, supplementation of the medium with these compounds led to the first successful demonstration of embryogenesis in isolated pollen grains of tobacco cultured in a defined medium (Nitsch 1974b). The formation of inhibitory substances in the somatic tissues of the anther can also explain some of the aforementioned results. Inhibitors are apparently the byproducts of the senescence of anther tissues and they cause degeneration of the pollen grains or retardation of the growth of embryoids. Compared to the continuous culture of D. innoxia anthers in one medium, the serial transfer of the anthers to fresh liquid medium results in an increase in the number of embryoids harvested (Tyagi, Rashid and Maheshwari 1979). One can explain these results by assuming that the anthers produce an inhibitor whose concentration is diluted by the serial transfer. Older pollen grains appear to be the source of inhibitory substances in isolated pollen culture of Brassica napus. Toxic effects are generally observed when potentially embryogenic unicellular pollen grains are cultured along with bicellular pollen; the negative effects are mimicked by a medium conditioned by the growth of older microspores (Kott et al 1988). On the whole, the role in pollen embryogenesis of substances emanating from the pollen or from the somatic tissues of the anther has been something of an enigma because of the apparently competing effects of promotory and inhibitory substances. (vi) Embryogenic Competence of Pollen Grains
A stumbling block in the analysis of the factors controlling pollen embryogenesis is our inability to determine when pollen grains become competent to produce embryoids and what causes them to
509
attain competency. Because most of the cases of pollen sporophytic growth reported have been those induced in cultured anthers, a considerable literature may be said to testify to the attainment of competence as a result of the trauma of the excision and culture of anthers. Overt signs of dedifferentiation in pollen grains of cultured anthers are the regression of the gametophytic cytoplasm and the loss of RNA and protein in tobacco (Bhojwani, Dunwell and Sunderland 1973; Dunwell and Sunderland 1974b), the increased stainability for cytoplasmic RNA and histones in Datura innoxia (Sangwan-Norreel 1978), and the increased RNA and protein synthesis and increased mRNA accumulation in Hyoscyamus niger (Raghavan 1979a, b, 1981b, 1984). In tobacco and H. niger, it is even possible to identify changes conducive to embryogenesis in a developmental bias, such as differences in staining intensity between gametophytic pollen grains and potentially embryogenic pollen within a few hours to a few days after culture of anthers (Sunderland and Wicks 1971; Raghavan 1979a). Thus, viewed from the perspective of the pollen grains and what goes on within them, diversity rather than similarity marks their development in cultured anthers; some become embryogenically competent, whereas others mark their time as gametophytic pollen or disintegrate. Another current view is that pollen grains attain competence for sporophytic growth naturally during anther development and that subsequent culture of the anther provides the environment for expression of this predetermined potential. The production of variant types of pollen grains that differ from the main population with respect to their size and staining properties is known as dimorphism and is a way of life for anthers of many plants. The most compelling evidence in support of the origin of embryoids from variant pollen grains in barley and tobacco is the correlation observed between the number of such pollen grains in the anther and the number of pollen sporophytic units formed in representative samples of cultured anthers (Dale 1975; Horner and Street 1978; Horner and Mott 1979). A case for the origin of embryoids from small variant pollen grains in wheat is supported by the similarities between the development of these pollen grains and the early stages of embryoid formation in cultured anthers (Zhou 1980). The conditions for the formation of dimorphic pollen grains in plants have been reviewed by Heberle-Bors (1989). As shown in tobacco, the photoperiodic and temperature regimes of the growth of the donor
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Adventive Embryogenesis
plants play a key role in the evolution of embryo- phytic pathway - is the unicellular stage. The first genic competence in pollen grains. Generally, division of the pollen grain in culture is cytologiplants raised under short days (8-hour photope- cally and morphologically very similar to the diviriod) and low temperature (18 °C) produce small sion that occurs in vivo. However, from this point anthers that have a high percentage of embryo- onwards, there are striking differences in the mangenically competent pollen grains, which are ner in which the daughter cells participate in referred to as premitotic pollen (P-grains); high embryoid or callus formation. As one might expect, yields of embryoids are obtained when these several ultrastructural changes suggestive of the anthers, or pollen grains isolated from them, are replacement of the gametophytic program with a cultured (Heberle-Bors and Reinert 1980, 1981). It sporophytic imprint should also be occurring in the turns out that P-grains in tobacco are functionally microspores during this period. Several key studies sterile pollen grains and that the frequency of their that are largely responsible for our current underformation is in some way linked to the develop- standing of the cellular and subcellular cytology of ment of sexuality in flowers. It has been proposed pollen embryogenesis are reviewed in this section. that P-grains are produced under conditions that induce high pollen sterility and that when the sex (i) Pollen Division Pathways balance of the flower is tipped toward femaleness there is also a tendency for large-scale pollen sterilBased on the assumption that a vegetative cell ity (Heberle-Bors 1982a, b, 1983). Although not and a generative cell are born out of the first hapwidely investigated, genetic factors are also per- loid mitosis, current theory considers three general haps responsible for P-grain formation in anthers. routes to the multicellularity of bicellular pollen The primary reason for the presence of high fre- grains to be important: (a) repeated divisions of the quencies of P-grains in anthers of different geno- vegetative cell; (b) repeated divisions of the genertypes of wheat has been traced to certain cms lines ative cell; and (c) divisions of both vegetative and with different cytoplasmic and nuclear back- generative cells. grounds (Heberle-Bors and Odenbach 1985). All this is impressive in support of the concept Division of the Vegetative Cell. The essential of embryogenic predetermination of pollen grains cytological details of the origin of embryoids from in anthers. In the light of this hypothesis, reports of the vegetative cell were first traced in a cultivar of the spontaneous formation of multicellular pollen Nicotiana tabacum. By about the sixth day after culgrains in anthers of hybrids of Solarium (Ramanna ture of the anther, the vegetative cell loses its mor1974; Ramanna and Hermsen 1974), Narcissus phogenetic individuality and is partitioned by a biflorus (Koul and Karihaloo 1977), and Paeonia (Li series of internal walls until a mass of cells typical 1982) and the induction of multiple pollen grains of somatic cell size is produced. Unable to contain by treating plants with chemicals, such as ethrel in the burgeoning growth within, the exine breaks wheat (Bennett and Hughes 1972), can be consid- open, liberating the cellular mass into the anther ered in a new light. These represent cases in which locule where it passes through the typical stages of embryogenically competent pollen grains formed globular, heart-shaped, torpedo-shaped, cotyledon in the anther have apparently escaped from the initiation, and mature embryoids. The culmination intracellular control system and express incipient of this developmental feat is the germination of the sporophytic features. On the whole, evidence for embryoid and its appearance outside the anther pollen predetermination is scanty in the large wall as a plantlet. A restrictive feature of this pathmajority of plants that yield embryoids or calluses way is that the generative cell either remains undifrom cultured anthers with relative ease, and so it vided or undergoes but a few divisions and, cannot be said that the concept of embryogenic pre- without contributing to the formation of an embrydetermination of pollen grains has been proved in oid, these cells simply wilt away (Sunderland and Wicks 1971). With minor differences in detail, a rigorous sense. embryogenesis by the division of the vegetative cell has been described in other cultivars of tobacco 3. CYTOLOGY OF POLLEN (Bernard 1971; Mii 1973), Datura metel (Iyer and EMBRYOCENESIS Raina 1972), D. innoxia (Sunderland, Collins and Dunwell 1974), Capsicum annuum (George and In most species studied, the vulnerable stage in Narayanaswamy 1973), Solarium surattense, Luffa pollen development - the one at which the gamecylindrica (Sinha, Jha and Roy 1978), Brassica napus tophytic program is deflected toward the sporo-
Embryogenic Development of Pollen Grains
(Fan, Armstrong and Keller 1988), and wheat (Reynolds 1993). The reality of the vegetative cell giving rise to a callus was first described in anther cultures of a cultivar of barley, in which the initial dispositions of the vegetative and generative cells are the same as in tobacco. The cellular mass released by the breakage of the exine continues unorganized growth as a callus rather than differentiates into an embryoid (Clapham 1971). Investigations that followed this work furnished additional examples of pollen callus formation from the vegetative cell in other cultivars of barley (Sun 1978; Wilson, Mix and Foroughi-Wehr 1978; Zhou and Yang 1980), as well as in cultured anthers of wheat (Wang et al 1973; Pan, Gao and Ban 1983; Szakacs and Barnabas 1988), rice (Chen 1977; Yang and Zhou 1979), maize (Miao et al 1978), and rye (Sun 1978). Extremes of behavior of the vegetative cell during its transformation into a callus have been noted in some cases. One is the appearance of micronuclei in the division products of this cell in pollen grains of wheat (Zhu, Sun and Wang 1978). In a cultivar of barley, nuclear divisions in the vegetative cell are not accompanied by cytokinesis and lead to the formation of multinucleate pollen grains (Zhou and Yang 1980). In another cultivar, fusion between two or three nuclei occurs during the intermediate free nuclear stage of the vegetative cell, or sometimes the nucleus of the generative cell is incorporated into a derivative of the vegetative cell by fusion (Sunderland and Evans 1980). These examples draw attention to the extent of cytological abnormalities in the division of the vegetative cell that may contribute to the formation of a mixoploid callus from a single pollen grain. Pollen sporophytic growth in cultured anthers by the activity of a functional vegetative cell is designated as the A pathway. From a detailed analysis of pollen embryogenesis in cultured anthers of tobacco, a further subdivision of this pathway into A-l, A-2, and A-3 has been proposed. The primary considerations for this are the different fates of the generative cell in the final products formed and the occurrence of an intermediate free nuclear or cellular phase during the division of the vegetative cell (Misoo, Yoshida and Mastubayashi 1979). Unfortunately, these variations have been little investigated in other plants and thus we do not know for sure whether they are widely applicable. Divisions of the Generative Cell or of Both Generative and Vegetative Cells. The role of the generative cell in pollen sporophytic growth was
511 slow to be recognized, but it appears that this cell is capable of independent divisions as an embryoid or a callus mother cell. The first hint about the developmental significance of the generative cell in pollen embryogenesis came from observations on the division patterns of pollen grains in cultured anthers of Nicotiana tabacum (Bernard 1971; Sunderland and Wicks 1971; Nitsch 1972; Rashid and Street 1974), N. sylvestris (Rashid and Street 1974), and D. metel (Iyer and Raina 1972) showing that as the embryoid is formed from the vegetative cell, the generative cell divides at a slow pace to produce a small population of free nuclei or cells. Conclusive evidence for the formation of embryoids by the division of the generative cell came from the work on cultured anthers and anther segments of Hyoscyamus niger. Periodic cytological examination revealed the occurrence of embryogenic divisions delimiting the generative and vegetative cells as early as 12 hours after culture. Further divisions are confined to the generative cell, which initially forms a group of cells within the exine. Following its release from the exine, the multicellular mass faithfully recapitulates the stages of zygotic embryogenesis to form embryoids and plantlets. Various fates can befall the vegetative cell, one of which is to form a two- to many-celled suspensorlike structure subtending the organogenetic part of the embryoid formed by the division of the generative cell (Raghavan 1976a, 1978). The concept of an active role for the generative cell in the presence of a passive or less active vegetative cell in pollen sporophytic growth has received support from anther culture investigations in maize (Guoetal 1978; Miao et al 1978), barley (Sunderland et al 1979), wheat (Pan, Gao and Ban 1983), and rice (Qu and Chen 1984). In cultured anthers of H. niger (Raghavan 1978), N. tabacum (Anand, Arekal and Swamy 1980), cereals (Bouharmont 1977; Miao et al 1978; Sun 1978; Sunderland et al 1979), and Cocos nucifera (Monfort 1985), it is also not unusual to find embryogenesis by the repeated divisions of both generative and vegetative cells. The resulting structures, being amalgams of unlike parts, are chimeras at the cellular level. Although nuclear divisions are invariably followed by cytokinesis, the vegetative cell or generative cell may occasionally produce several free nuclei, which are incorporated into the embryoid or callus (Sun 1978; Sunderland et al 1979). Pollen sporophytic growth by the exclusive division of the generative cell or by the division of both generative and vegetative cells is included in the E pathway. A unique type of pollen embryogenesis involv-
512
Adventive Embryogenesis
ing both the generative and vegetative cells or their nuclei occurs in cultured anthers of Datura innoxia. What we find here are not independent divisions of these cells in a strict sense, but their fusion to form an abnormal cell; both nuclei then divide simultaneously on a common spindle. The fusion might occur between one or two haploid vegetative cell nuclei and a haploid or endopolyploid generative cell nucleus. Fusions of this kind allow the relationship between parental genomes to change, so that nuclei with nonhaploid chromosome numbers are formed (Sunderland, Collins and Dunwell 1974). This pathway of embryogenesis, known as the C pathway, may have widespread significance in accounting for the frequent occurrence of embryoids with nonhaploid chromosome numbers.
sion of microspores may underlie their potential to develop into embryoids, whereas asymmetrically dividing microspores almost always regenerate calluses from the vegetative cell (Szakacs and Barnabas 1988,1995). Since colchicine treatment has been shown to lead to an increase in the number of symmetrically dividing embryogenic pollen grains in anther and pollen cultures of Brassica napus, it has been suggested that cytokinetic symmetry is an important factor in deflecting the gametophytic program of the pollen grain toward the embryogenic pathway (Zaki and Dickinson 1991,1995). Another scenario, which is not yet fully documented, appears to be a variation of the B pathway and has been described in cultured anthers of wheat. Here, the two identical nuclei divide repeatedly to generate a cluster of free nuclei, but it is not Symmetrical Division of Pollen Grains. In certain whether cells are forged out of these nuclei some culture systems, a symmetrical division of and whether the multicellular mass so produced the pollen grain to produce two identical cells or proceeds to form an embryoid or a callus (Zhu, Sun nuclei is the cue that sets the division products off and Wang 1978; Pan, Gao and Ban 1983). This is into the differentiative pathway, known as the B known as the D pathway. pathway. As described in cultured anthers of The disposition of cells and nuclei in the differAtropa belladonna (Solanaceae) (Narayanaswamy ent pathways of pollen embryogenesis described and George 1972; Rashid and Street 1973), wheat here is illustrated in Figure 17.3. In cultured (Ouyang et al 1973; Pan, Gao and Ban 1983; anthers, pollen sporophytic units originate by mulReynolds 1993), rice (Guha-Mukherjee 1973), tiple pathways. The relative abundance of the units Datura innoxia (Sunderland, Collins and Dunwell formed by a specific division sequence varies not 1974), Nicotiana sylvestris, N. tabacum (Rashid and only in anthers of different flower buds of the same Street 1974), Hordeum distichum, H. vulgare plant but also in anthers of the same flower bud. (Matsubayashi and Kuranuki 1975; Zhou and Yang What prompts pollen grains to choose a particular 1980), Hyoscyamus niger (Raghavan 1978), Solarium division sequence is not understood. surattense, Luffa cylindrica, and L. echinata (Sinha,
Jha and Roy 1978), the two cells divide repeatedly to form the embryoid; the contributions of neither cell can be considered more important than those of the other. It should be added here that it has not been determined whether there are any cytochemical or histochemical differences between the two similar-looking cells or nuclei in the examples cited. Just because two cells look alike does not mean that they are not different at another level of organization. In cultured anthers of Triticale (Sun, Wang and Chu 1974) and H. vulgare (Wilson, Mix and Foroughi-Wehr 1978), despite the close similarity of the two nuclei formed, cytochemical and physiological observations seem to indicate that multicellular pollen grain is generated by the division of the nucleus, which appears to be similar to the nucleus of the vegetative cell of an asymmetrically partitioned pollen grain. This makes pollen grains with symmetrical cells prime suspects for housing disguised generative and vegetative cells. In cultured anthers of wheat, the symmetrical divi-
(ii) Ultrastructural Cytology
The ultrastructure of pollen embryogenesis has been studied in detail in only four species (Nicotiana tabacum, Datura innoxia, Hyoscyamus niger, and Brassica napus); in consequence, most of what can be said is based on these investigations. Despite this limitation, the starting premise is that there are no cellular changes more fundamental to pollen dedifferentiation than those that can be discerned in the electron microscope. An important line of enquiry is the fate of the microspore cytoplasm programmed for gametophytic differentiation. The distinctive ultrastructural features of the vegetative and generative cells as they are incorporated into an embryoid or a callus are also of interest. Nicotiana tabacum. Few plants have rivaled N. tabacum as a target for ultrastructural investigations of pollen embryogenesis. However, for all of the virtues of tobacco as a good experimental system,
Embryogenic Development of Pollen Grains
513
ance of zones of multivesiculate bodies analogous to lysozomes (Vazart 1971; Dunwell and Sunderland 1974a, b). Cytoplasmic regression stimulates a series of alterations in the structure, function, and developmental program of the vegetative cell. The earliest change seen in this cell is its division in the embryogenic pathway. As the newly formed cells accumulate their own cytoplasm, it becomes populated by a fresh array of organelles, including ribosomes arranged in polysomal profiles, lipid centers, and starch-containing plastids (Dunwell and Sunderland 1975). More than any other single event, the sequential disappearance and appearance of the cytoplasmic organelles has led to the view that embryogenic pollen is incarnated from the gametophytic pollen by a programmed destruction of the cytoplasm of the vegetative cell, which thereby depletes this cell of a specific phenotype, followed by the repopulation of the cell by a new set of organelles. Because the generative cell eventually degenerates, its ultrastructure has not been followed in much detail.
Figure 17.3. Diagrammatic representation of the different pathways of pollen embryogenesis. In A, C, and E, solid circles represent nuclei of the generative cell or its division products; hollow circles represent nuclei of the vegetative cell or its division products. In C, solid circles enclosed in hollow circles indicate fusions between the two nuclei. In B and D, circles represent symmetrical nuclei born out of the first haploid mitosis and their division products. (From Raghavan 1986c.)
contradictory observations exist, depending upon whether ultrastructural changes are followed in pollen grains of cultured anthers or in ab initio pollen cultures. In the sequence of changes described, no obvious features associated with embryogenesis are seen in pollen grains during the first five days of culture of anthers; rather, during this period, the gametophytic imprint appears more and more prominent in the vegetative and generative cells. This period in culture may well be critical in establishing that gametophytic differentiation proceeds to the maximum permissible extent before the sporophytic program takes over. A major structural upheaval associated with embryogenesis is seen seven to eight days after culture as a regression of the cytoplasmic organization of the vegetative cell resulting in the elimination of much of the gametophytic program. During this period, the organelles appear in complete disarray and in various states of disintegration; this is coupled with the disappearance of ribosomes and the appear-
Other studies have shown that subcellular changes during the embryogenic development of tobacco pollen grains must be viewed in the context of the cold stress accorded to flower buds to increase pollen efficiency. In ab initio pollen cultures originating from anthers of cold-stressed flower buds, embryogenic divisions begin with the laying down of a convoluted fibrillar wall between the plasma membrane and the inner layer of the intine, thus insulating the entire pollen cytoplasm. Embryogenic pollen grains do not display ultrastructural features that could be construed as organelle regression; rather, they appear to possess the hallmarks of metabolically repressed cells, such as an attenuated cytoplasm, condensed mitochondria, and sparsity of ribosomes. As the vegetative cell divides in the embryogenic pathway, the newly formed cells regain the ultrastructural profile of metabolically active cells; this is evidenced by an increase in the ribosome population in the cytoplasm sufficient to crowd out other organelles and by a change in the mitochondrial morphology (Rashid, Siddiqui and Reinert 1981, 1982). The presence of an increased number of ribosomes and complex mitochondria is also characteristically seen during pollen sporophytic development in Aesculus hippocastanum (Radojevic, Zylberberg and Kovoor 1980) and Hordeum vulgare (Idzikowska 1981). These observations support the notion that the metabolic requirements of the pollen grains during the initiation of embryogenic divisions are very demanding.
514 One plausible explanation for the conflicting reports on the ultrastructural changes during pollen embryogenesis in tobacco is that the reports do not take into account the effect of low-temperature stress on the donor plants in preventing organelle regression or in causing other ultrastructural anomalies in the cytoplasm of cultured pollen grains. A previous section referred to the changes in the vacuole in pollen grains of cold-stressed anthers of Datura metel (Camefort and Sangwan 1979; Sangwan and Camefort 1984). Similar changes that might obscure the regression of organelles due to embryogenic induction cannot be ruled out in tobacco pollen grains from coldstressed plants. In this context, attention may be drawn to the fact that signs of regression, such as chromatin condensation, a decrease in nucleolar size, and the loss of nuclear pores, are frequent in the vegetative cell of isolated pollen grains of tobacco subjected to a starvation diet as a prelude to embryogenesis (Garrido et al 1995). A comparative analysis of the ultrastructural changes associated with embryogenesis in ab initio cultures of tobacco pollen grains collected from plants grown under standard greenhouse conditions and from plants subjected to temperature stress will go a long way toward resolving this controversy. Datura innoxia. In contrast to the results from tobacco, there is no evidence for a breakdown of the gametophytic cytoplasm during the ultrastructural displacement that inevitably accompanies pollen embryogenesis in cultured anthers of D. innoxia. Prior to, or at the time of, culture of anthers, the enclosed pollen grains display an undistinguished ultrastructural profile characterized by the presence of structurally simple mitochondria and proplastids distributed in the sparse cytoplasm surrounding a large central vacuole. Although this profile remains essentially unchanged in the unicellular embryogenic pollen grains of cultured anthers, both vegetative and generative cells are enriched with ribosomes, plastids, and mitochondria following the first haploid mitosis. Changes in proplastids, such as their transformation into amyloplasts, and the appearance of numerous cristae in the mitochondria have been identified as being specially indicative of embryogenic induction of pollen grains. It is this basic subcellular organization of the vegetative cell that is partitioned repeatedly as embryogenic divisions are initiated (Dunwell and Sunderland 1976; Sangwan-Norreel 1978; Sangwan and SangwanNorreel 1987a, b). In D. metel, curious organelles
Adventive Embryogenesis
resembling groups of polysomes and rough ER surface as early as the 2-4-celled stage of the embryoid, but they disappear at the globular or heart-shaped stage. Although its relevance is not determined, the tonoplast of embryogenic pollen grains of D. metel possesses a tanninlike deposit that is absent in normal pollen grains and persists up to the stage of the globular embryoid (Sangwan and Camefort 1982, 1983). This change in the embryogenic pollen grain is of interest as a possible marker for the induction process. Hyoscyamus niger. By combining ultrastructural observations with stereological analysis, Reynolds (1984b) has shown that the early features that distinguish the embryogenic pollen grain from the normal pollen are an increased volume of the granular zone in the nucleolus and an increased amount of dispersed chromatin in the nucleus. These changes, visible in the unicellular pollen grains as early as six hours after culture of the anther, are indicative of enhanced nuclear and nucleolar activities. Following the first haploid mitosis, the nuclear and nucleolar changes are perpetuated in the generative cell, which functions as the embryo mother cell. Some differences are also noted between the volume-area fractions of the cytoplasm occupied by mitochondria, plastids, rough ER, and Golgi cisternae in the generative cells of embryogenic pollen grains and those in the nonembryogenic pollen grains. Since these changes occur in the absence of any noticeable regression of the cytoplasm in the generative cell, it has been emphasized that metabolic quiescence is not associated with embryogenic induction; rather, it appears that ultrastructurally there is a gradual assimilation of the gametophytic program into one that ensures a sporophytic mode of development. Another study by Reynolds (1985) has identified some pollen grains that divide symmetrically to yield two apparently identical nuclei, both of which contribute to the formation of the embryoid; in these pollen grains there is an abrupt cessation of gametophytic features before embryogenic division is initiated. Comparing the subcellular cytology of pollen grains that divide symmetrically and asymmetrically, a case can be made that the specific pathways of pollen embryogenesis in cultured anthers of H. niger are correlated with the timing and degree of repression of gene activity for gametophytic development. Other Plants. As one considers the ultrastructure of pollen embryogenesis in other plants, no
Embryogenic Development of Pollen Grains
common thread of events or sequence of changes appears to emerge. In cultured anthers of Brassica napus, the major changes that augur the division of the embryogenic pollen grain to form two identical cells are the movement of the nucleus from the peripheral to the central position, starch assembly by plastids, the formation of a thick fibrillar wall beneath the intine, an increase in ribosome number, and the appearance of large cytoplasmic aggregates of an unidentified globular material. Speculatively, the cytoplasmic granules have been considered as heat-shock granules that probably shield mRNAs from damage (Zaki and Dickinson 1990; Telmer, Newcomb and Simmonds 1993,1995; Testillano et al 1995). The cytoskeletal elements of the pollen grain do not seem to have a primary function in embryogenic induction, because the microtubules and microfilaments show the expected pattern of distribution that coincides with the position of the nuclei during embryogenic divisions (Hause et al 1993). In a continuation of the trends established in the daughter cells born out of the first haploid mitosis, organelles are distributed uniformly in all cells of the globular embryoid enclosed within the exine. Following the rupture of the exine, there is an asymmetric distribution of starch granules in the cells of the embryoid that is the first sign of polarity. Granules generally disappear from cells at the site of the exine break but they persist at the opposite end, which eventually becomes the root apex (Hause et al 1994). The main finding of light and electron microscopic studies of cultured anthers of barley is that embryogenic pollen grains display a proliferation of cytoplasmic organelles, followed by division to form the vegetative and generative cells or two symmetrical cells (Chen, Kasha and Marsolais 1984; Chen et al 1984). In barley anthers subjected to a cold stress, prior to the first haploid pollen mitosis, the cytoplasm of embryogenic pollen grains is depleted of storage materials and organelles, which for the most part display little differentiation (Huang 1986). Comparison of the subcellular cytology of pollen grains from normal and cold-stressed anthers shows that the pollen cytoplasm is not stable enough to withstand the stress. A highly differentiated cytoplasm in the vegetative cell sets the stage for embryogenic divisions in pollen grains of cultured anthers of Zea mays (Barnabas, Fransz and Schel 1987). The formation of new cells in a rapid burst of mitotic activity, and the subsequent burgeoning growth, begin to stretch the exine and, eventually, only remnants of sporopollenin are seen on the
515
pollen wall. The frequent occurrence of binucleate and multinucleate pollen grains in cultured anthers suggests that nuclear divisions during pollen embryogenesis are independent of cytokinesis. In such pollen grains, cell walls are formed subsequently by centripetal ingrowth from the intine or from the plasma membrane (Reynolds 1985; Huang 1986; Barnabas, Fransz and Schel 1987). Phragmoplasts with attached vesicles and attached spindle microtubules and with plasmodesmatarich walls have been noted in electron microscopic examination of pollen embryoids, indicating that the newly formed cells are partitioned by walls formed by conventional cell plates (Huang 1986; Barnabas, Fransz and Schel 1987; Zaki and Dickinson 1990). It is clear from the account presented in this section that ultrastructural analysis has not identified a subcellular marker of the sporophytic transformation of the pollen grain. Perhaps it is time now to take a close look at the first haploid mitosis that sets the stage for embryogenic division of the pollen grain. For example, are there changes in the positioning of the cytoskeletal elements of the pollen grain during normal gametophytic development and during induced embryogenesis? Is there a differential distribution of cytoskeletal proteins in the pollen grain during its developmental transformations? These questions are worth exploring if only to highlight what we need to know about the early cytological signals of embryogenic induction in the pollen grains. 4. MOLECULAR BIOLOGY OF POLLEN DEDIFFERENTIATION
The fundamental premise of the molecular biology of pollen embryogenesis is that pollen grains programmed for terminal differentiation into gametes are diverted to a pathway of continued divisions and increasingly complex morphology of an embryolike structure. From what we know about the basis for cell differentiation, it is an ingrained idea that these changes depend on the input of newly synthesized mRNAs and proteins. Unfortunately, little information is available on the nucleic acid and protein metabolism of the vegetative and generative cells of embryogenic pollen grains, and investigations that take into account the biochemical basis for the altered developmental expression seem to be a major lacuna in this topic. This is attributable in part to the small number of pollen grains in a given anther that become embryogenic and in part to the different pathways
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of pollen embryogenesis observed in one and the same anther in culture. It was therefore natural in the early stages to rely on cytochemical and autoradiographic data to evaluate the macromolecular changes associated with pollen sporophytic development. In recent years, a beginning has been made in the use of ab initio pollen cultures to follow the pattern of gene expression during the embryogenie transformation of microspores. For now, attention will be directed to the pattern of nucleic acid and protein synthesis in the pollen grains that heralds embryogenic development.
Adventive Embryogenesis
formed by the exclusive division of the generative cell. When the embryoid is derived from both generative and vegetative cells, silver grain density due to label incorporation is relatively low in the derivatives of the vegetative cell in comparison to that found in the generative cell derivatives (Raghavan 1977a). Overall, the patterns of DNA replication in the vegetative and generative cells provide useful models to establish the respective roles of these cells as embryoid mother cells.
RNA Metabolism. An early step in the embryogenic development of pollen grains is the transcription of DNA to form RNA. This might be (i) Dynamics of DNA, RNA, and Protein linked to the initiation of the sporophytic program Metabolism or might be directed to the elimination of the existAt the heart of the division of the vegetative cell ing gametophytic program. In cultured anthers of or the generative cell in the embryogenic pathway tobacco, depending upon the temperature of incuis the renewed DNA replication in these cells. bation, there is a lag period of 6-12 days before the Microspectrophotometric measurements of the vegetative cell divides in the embryogenic pathDNA content of embryogenic pollen grains in cul- way. The stainable RNA content of the pollen tured anthers of tobacco have shown that DNA grains stabilizes during this period to serve as a doubling occurs in the vegetative cell before any good marker for distinguishing between gametoovert signs of dedifferentiation; in contrast, the phytic and embryogenic pollen grains. Generally, vegetative cell of the normal gametophytic pollen the former display a four- to sixfold increase in grain retains the 1C DNA amount. The culture of RNA content, whereas the latter have a low level isolated tobacco pollen grains in a starvation of RNA. It appears that suppression of the gamemedium before transfer to an enriched medium for tophytic program, as reflected in the low RNA embryogenesis affects neither renewed DNA syn- level, is necessary to ensure that the genes for thesis in the vegetative cell nor ongoing DNA syn- embryogenic induction are fully expressed withthesis in the generative cell (Chu, Liu and Du 1982; out being masked by the simultaneous expression Zarsky et al 1992). In similar studies with cultured of genes for pollen maturation and germination anthers of Datura innoxia, variations are noted in (Bhojwani, Dunwell and Sunderland 1973). the DNA content of the nuclei of embryogenic Cytochemical study of pollen embryogenesis in pollen grains. Generally, unicellular embryogenic Datura innoxia has shown that compared to nonpollen grains have 1C and 2C DNA amounts, embryogenic pollen grains, embryogenic pollen whereas each of the two identical nuclei in a bicel- grains exhibit an increased stainability for cytolular pollen grain has 1C DNA. A modal 1C DNA is plasmic RNA prior to the first haploid mitosis also measured in the nuclei of vegetative and gen- (Sangwan-Norreel 1978). erative cells. The generation from cultured anthers Results from autoradiography of the incorporaof D. innoxia of plants with differing ploidy levels tion of 3H-uridine show that embryogenically has led to the suggestion that fusion between determined pollen grains of Hyoscyamus niger nuclei might account for this phenomenon begin to synthesize RNA as early as the first hour (Sangwan-Norreel 1979, 1981, 1983). Somewhat of culture of anthers. Although the generative cell similar conclusions were reached from observa- of normal bicellular pollen grains does not incortions of the changes in the DNA contents of porate any appreciable amount of the label into its embryogenic pollen grains of wheat and Petunia sp. RNA, there is an accelerated RNA synthesis in the (Raquin et al 1982). generative cell of the embryogenic bicellular pollen The cytological view of embryogenesis by the grains. During progressive embryogenesis, RNA division of the generative cell in cultured anthers of synthetic activity is finely balanced between the Hyoscyamus niger has been confirmed by autoradi- fates of the generative and vegetative cells (Figure ography of 3H-thymidine incorporation. Failure of 17.4). This ensures that pollen grains in which RNA DNA synthesis or cytokinesis in the vegetative cell synthesis occurs either in the generative cell or in and its division products is typical of embryoids both the generative and vegetative cells divide fur-
Embryogenic Development of Pollen Grains
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Figure 17.4. Autoradiographic incorporation of 3H-uridine into pollen grains of cultured anthers of Hyoscyamus niger. (a) A multicellular pollen grain with label in the division products of the generative cell nucleus, 72 hours after culture, (b) Another multicellular pollen grain, 96 hours after culture, (c) A globular pollen embryoid, 144 hours after culture. In all cases, the vegetative cell nucleus is indicated by the arrow, (d) A 3-celled pollen grain with label in the
division products of the generative cell and in the nucleus of the vegetative cell, 96 hours after culture, (e) A multicellular embryoid, 120 hours after culture, (f) Another multicellular embryoid with two nuclei formed by the division of the vegetative cell, 144 hours after culture. In d-f, division products of both vegetative (v) and generative (g) cells are labeled. Scale bars = 10 urn. (From Raghavan 1979b.)
ther along the embryogenic pathway, whereas those in which RNA synthesis occurs exclusively in the vegetative cell become starch filled and nonembryogenic (Raghavan 1979a, b; Reynolds and Raghavan 1982). Thus, transcription in the generative cell is a more important prerequisite for embryogenic divisions of pollen grains of H. niger than transcription in the vegetative nucleus. In spite of their small representation in the genome, mRNAs accumulate in a characteristic way in the embryogenic pollen grains of H. niger. By in situ hybridization using 3H-poly(U) as a probe, it was found that although unicellular pollen grains do not bind the probe during their normal ontogeny, the embryogenically determined pollen grains are labeled within a few hours of culture of anthers. This labeling is sensitive to actinomycin-D, which supports the idea that mRNA is newly synthesized by the pollen grains as the
anther establishes contact with the nutrient medium. Compared to the generative and vegetative cells of normal pollen grains, which are only transiently active in binding 3H-poly-(U), embryogenic bicellular pollen grains exhibit the striking feature of continued transcriptional activity in the generative cell. The vegetative cell of embryogenic bicellular pollen grains is generally starved for mRNAs and pollen grains that accumulate mRNAs exclusively in this cell pass their time in culture as starch-filled, nonembryogenic pollen (Raghavan 1981b). It would appear from these results that in cultured anthers, a small number of the enclosed pollen grains synthesize new mRNAs that apparently code for the proteins necessary to induce the first embryogenic mitosis. The trigger that sets in motion the synthesis of new mRNAs is possibly the trauma of the excision and culture of anthers. In vitro translation profiles of mRNAs isolated
578
Adventive Embryogenesis
66453629242014Figure 17.5. In vitro translation profiles of proteins from (a) bicellular and (b) embryogenic pollen grains of tobacco. Autoradiograms of translation products separated by twodimensional SDS-PACE are shown. Arrows in (B) point to
two major proteins not seen in (A). Molecular mass markers are shown on the left. (From Carrido et al 1993. Sex. Plant Reprod. 6:40-45. © Springer-Verlag, Berlin. Slide supplied by Dr. O. Vicente.)
from embryogenic pollen grains and embryoids generally support the results from cytological studies. Late unicellular to early bicellular pollen grains of Brassica napus undergo embryogenic divisions after a thermal shock at 32 °C for 8-12 hours. Pechan et al (1991) have shown that during the first 8 hours of high-temperature stress, a number of mRNAs that are absent, or present at low levels, in freshly isolated microspores appear in the embryogenic counterparts. Since some of these messages probably code for proteins homologous to HSPs of the 70-kDa class, their relevance to the embryogenic induction of pollen grains remains to be clarified (Cordewener et al 1995). That the synthesis of new mRNA is associated with embryogenic induction is suggested by the appearance of two abundant mRNAs when isolated pollen grains of tobacco are induced to form embryoids in an enriched medium after a period of starvation (Figure 17.5) (Garrido et al 1993). Some differences confined to the relative abundance of certain messages have also been noted between pollen embryoids of two different stages of development isolated from cultured anthers of barley (Higgins and Bowles 1990).
accumulate a large amount of protein (Bhojwani, Dunwell and Sunderland 1973). In the embryogenic bicellular pollen grains of D. innoxia, the nucleus of the generative cell is more enriched for histones than that of the vegetative cell; a possible role for histones in preventing the division of the generative cell in the embryogenic pathway is implied by this observation (Sangwan-Norreel 1978). A change in the spatial accumulation of 70-kDa HSPs may be achieved in the microspores of B. napus during the thermal shock preparatory to embryogenic induction (Figure 17.6). Compared to normal unicellular and bicellular pollen grains, the embryogenic counterparts display intense and differential labeling of nuclei with anti-HSP-70 antibodies, showing that embryogenic induction depends critically upon stage-specific changes in the intracellular distribution of HSP-70 (Cordewener et al 1995). The role demanded of the embryogenic proteins suggests that they should be present at the critical stage of induction. However, the hope of identifying specific proteins that are synthesized sufficiently early to have a role in the embryogenic determination and induction of microspore embryogenesis appears unrealistic. A two-dimensional electrophoretic analysis did not reveal any qualitative differences between the in vivo synthesized proteins of embryogenic and nonembryogenic pollen grains of B. napus; by a careful quantitative study, it was, however, found that some polypeptides are synthesized at a slightly higher rate in the embryogenic than in the nonembryogenic pollen grains (Cordewener et al 1994; Custers et al 1994). The most optimistic outlook is that the new proteins are synthesized transiently or in such small quantities
Protein Metabolism. Information available on the protein metabolism of embryogenic pollen grains lags behind that on RNA metabolism. The previously noted decrease in RNA accumulation in the embryogenic pollen grains of tobacco, suggestive of the elimination of the gametophytic program, is associated with an apparent lack of protein accumulation, whereas pollen grains that mature and complete the gametophytic program in culture
Embryogenic Development of Pollen Grains
519
that they cannot be detected by the techniques used. In H. niger, the embryogenic development of pollen grains is linked to the synthesis of certain general and basic proteins. Whereas unicellular pollen grains of intact anthers do not incorporate any 3H-tryptophan or 3H-leucine into proteins, culture of anthers is associated with the incorporation of these amino acids, as well as with the accelerated incorporation of 3H-arginine and 3H-lysine, into unicellular, potentially embryogenic pollen grains. Following the first haploid mitosis, these pollen grains incorporate 3H-leucine, 3H-arginine, and 3Hlysine into the generative cell or into both the generative and vegetative cells. The course of development pursued by the bicellular pollen grains is markedly affected by the protein synthetic activity in the constituent cells. Pollen grains that incorporate labeled arginine, leucine, and lysine into the vegetative cell fail to become embryogenic but continue the gametophytic program. When the generative cell or when both the generative and vegetative cells incorporate the labeled precursors, pollen grains continue embryogenic divisions. In the bicellular pollen grains, the incorporation of 3 H-tryptophan is sparse and is found only in the vegetative cell (Raghavan 1984). The pattern of protein synthesis in the unicellular and bicellular pollen grains of H. niger is remarkably similar to the pattern of RNA synthesis.
Figure 17.6. Immunofluorescence localization of 70-kDa HSP in pollen grains of Brassica napus immediately after isolation (a-e) and after culture for 8 hours at 32°C under embryogenic conditions (f-j). (a) Vacuolate, late unicellular pollen grain with label in the nucleoplasm and weak label in the cytoplasm, (b) Late unicellular pollen grain with label in the cytoplasm, less in the nucleoplasm, and none in the nucleolus. (c) Mitotic microspore with label throughout the cytoplasm and negative staining on the chromosomes, (d) Early bicellular pollen grain with the cytoplasm of both the vegetative and generative cells and the wall between both cells (arrow) labeled, (e) Late bicellular pollen grain with slightly more intense label in the cytoplasm of the generative cell than in the vegetative cell, (f) Stage corresponding to (a), with intense label in the nucleoplasm and weak label in the cytoplasm. Arrow points to an intensely labeled area, (g) Stage corresponding to (b), with intense label in the nucleoplasm and weak label in the cytoplasm, (h) Stage cor-
It is to be expected that increased protein synthesis detected autoradiographically in the embryogenic pollen grains will be matched at the biochemical level. In embryogenic pollen grains of Nicotiana tabacum, new sets of membrane-bound phosphoproteins and soluble and insoluble glycoproteins are synthesized, whereas different sets of these classes of proteins appear in the nonembryogenic pollen (Kyo and Harada 1990; Kyo and Ohkawa 1991; Rihova, Capkovd and Tupy 1996). In a restricted analysis, attempts to identify changes in mitosis-specific phosphoproteins during embryogenic induction of isolated microspores of B. napus were not, however, successful (Hause et al 1995). Increased levels of arginine and polyamines responding to (c), with label in the cytoplasm, (i) Stage corresponding to (d), with strong label in both the generative cell and vegetative cell nuclei, (j) Stage corresponding to (e), with diffuse label in the cytoplasm and nucleoplasm of both the generative and vegetative cells. Intensely labeled spots are also seen in the cytoplasm of the latter. Scale bar = 10 um. (From Cordewener et al 1995. Planta 196:747-755. © SpringerVerlag, Berlin. Photographs supplied by Dr. M. M. van Lookeren Campagne.)
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Adventive Embryogenesis
characteristically appear during embryogenic Embryoids accumulate little or no napin and crudevelopment of pollen grains of N. tabacum and D. ciferin mRNAs until they reach the globular or innoxia (Villanueva, Mathivet and Sangwan 1985; heart-shaped stages. From this period onward, Garrido, Chibi and Matilla 1995). The transient transcript levels also begin to increase through the nature of the phosphoproteins and the large inclusion of ABA in the medium. The accumulation increases in the arginine and polyamine levels in of cruciferin transcripts is particularly tardy, and the embryogenic pollen grains give these bio- even in torpedo-shaped embryoids the transcripts chemical parameters some legitimacy as part of attain detectable levels only in the presence of ABA the induction process in an as yet undetermined (Wilen et al 1990). Jasmonic acid is another hormonal substance that functions like ABA in inducway. Clark et al (1991) found that a 17-kDa embryo- ing napin and cruciferin mRNA accumulation. specific polypeptide reliably predicts the transi- Moreover, the natural occurrence of jasmonic acid tion of pollen embryoids in cultured anthers of in seed embryos and pollen embryoids of B. napus barley to the stage of embryonic organ produc- suggests that subtleties in the action of this hortion. The appearance of this protein in itself is not mone are important in the regulation of storage concerned with the regulation of pollen embryo- protein synthesis during pollen embryogenesis genesis, because near-identical proteins are seen (Wilen et al 1991). Oleosin mRNAs respond in a in seed embryos of barley and other cereals. The similar way to the treatment of embryoids with temporal distribution of storage proteins (Crouch ABA, sorbitol, or jasmonic acid (van Rooijen et al 1982; Taylor et al 1990; Holbrook et al 1991), stor- 1992; Zou et al 1995). Embryogenically induced microspores of B. napus also express a gene for aceage lipids (Pomeroy et al 1991; D. C. Taylor et al tohydroxyacid synthase, which codes for an essen1991; Wiberg et al 1991), and activities of enzymes tial enzyme in branched-chain amino acid involved in the glycolytic pathway (Sangwan et al biosynthesis (Ouellet, Rutledge and Miki 1992). On 1992) and in fatty acid biosynthesis (Weselake et al the other hand, transcripts of cloned nitrate reduc1991, 1993; Taylor et al 1992, 1993) in pollen tase genes are activated in the embryoids no earlier embryoids of Brassica napus and B. campestris gen- than the heart- or torpedo-shaped stages. Since erally resemble their patterns seen in zygotic gene expression is independent of the presence of embryos. nitrate in the medium, some nitrate-sensitive mechCertain stages of the dedifferentiation of pollen anism appears to be turned on in the embryoids in grains in cultured anthers of barley are associated a stage-specific manner (Fukuoka et al 1996). with the appearance of specific isozymes of acid phosphatase, esterase, NADH dehydrogenase, and There is good evidence linking the expression of peroxidase. The most distinct isozyme changes are a cloned napin gene with the embryogenic inducobserved concomitant with the transition of the tion of isolated microspores of B. napus. This evidisorganized cells into heart-shaped embryoids dence is based on the observation that transcripts and later with the formation of incipient embryonic of the gene that are present in small amounts in organs (Pedersen and Andersen 1993). Potentially, microspores at the time of culture are expressed at isozymes could act as markers of the embryogenic elevated levels after an inductive heat shock at transformation of microspores, but this possibility 32 °C for four days. Particularly interesting is the has not been considered as widely in pollen fact that napin transcripts decrease in abundance as the microspores divide in the embryogenic pathembryogenesis as in somatic embryogenesis. way and they do not reappear until the embryoids are phased into the cotyledonary stage of develop(ii) Genes Expressed during Pollen ment. High expression of a GiiS-napin gene conEmbryogenesis struct in embryogenically induced microspores of A global view of gene expression during pollen transgenic plants is considered a rigorous proof of embryogenesis has been derived from studies of the role of the napin gene as an early marker of the patterns of the appearance and disappearance pollen dedifferentiation (Boutilier et al 1994). In a of transcripts of genes for storage proteins in devel- similar way, following an inductive in vitro starvaoping embryoids of Brassica napus. As with seed tion treatment, there is a dramatic increase in the embryos, there are indications that the develop- level of a low-molecular-mass heat-shock protein ment of pollen embryoids is associated with the gene transcript in embryogenic tobacco pollen accumulation of storage protein gene transcripts grains (Zarsky et al 1995). Whether or not the napin and heat-shock protein genes have a functional role and that it is strongly influenced by ABA.
Embryogenic Development of Pollen Grains
521
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VMl pEMB 56 | § i j pEMB 98 pEMB 115
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o
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Figure 17.7. Expression of embryoid-abundant mRNAs during pollen embryogenesis in cultured anthers of Triticum aestivum. Data are based on densitometry of autoradiograms of Northern blot of RNAs from embryoids of different stages hybridized with cDNA clones. Stages represented: (1), unicellular pollen grain at the time of culture; (2), multicellular pollen grain, 2 days after culture; (3),
proembryoid, 7 days after culture; (4), globular embryoid, 10 days after culture; (5), bipolar embryoid, 14 days after culture; (6), young embryoid, 21 days after culture. Clones pEMB-4 and pEMB-94 are considered embryoid specific. (From Reynolds and Kitto 1992. Plant Physiol. 100:1744-1750. © American Society of Plant Physiologists. Print supplied by Dr. T. L. Reynolds.)
in embryogenesis in the respective systems is not clear at present. Analysis of cDNA clones from a library made to poly(A)-RNA from young pollen embryoids of Triticum aestivum promises to provide new insight into the genes that are activated during the early stages of pollen embryogenesis (Figure 17.7). Two embryoid-specific recombinants isolated by differential screening are expressed early during embryogenesis and are not detected in the nonembryogenic pollen grains or in the vegetative tissues of the plant (Reynolds and Kitto 1992). The same library also yielded an embryoid-abundant gene that encodes a metallothionein-like protein. Expression of the gene is restricted to embryogenic microspores, embryoids, and zygotic embryos and is modulated by ABA (Reynolds and Crawford 1996). This is attractive preliminary evidence indicating that some genes, which are not part of the gametophytic program, are associated with the dedifferentiation phase of the pollen grains. It is possible that in this species, in which pollen grains are not encumbered by thermal shock or starvation stress for embryogenic induction, an overlap
between the expression of low-abundance embryoid-specific genes and that of stress-related genes can be avoided. 5. GENERAL COMMENTS The concept that the vegetative and generative cells of the angiosperm pollen grain possess the innate potential to develop into whole new plants by a process of simulated embryogenesis has arisen largely from anther and pollen culture investigations reviewed in this chapter. To initiate embryogenic divisions, pollen grains enclosed within the anther, or isolated from it, must be cultured in a medium and sufficient time must elapse under specified conditions to allow the gametophytic program gradually to wane. We cannot identify with certainty any hormonal or chemical substances in the medium that stimulate embryogenic divisions of pollen grains. In the two most widely exploited systems, subjecting pollen grains to temperature or nutritional stress appears to be associated with the acquisition of totipotency by the vegetative cell. There is preliminary evidence that connects stress-
522 imposed embryogenic divisions to the activation of genes for storage protein and heat-shock protein synthesis but their relevance to the pollen embryogenic induction process remains to be established. Moreover, the changeover from the gametophytic program to the sporophytic program in pollen grains is so profound that it is not unreasonable to look for genes that might qualify as embryonic genes, as in somatic embryogenesis. A highly interesting finding is that in several species investigated, the stage of development of the pollen grains at the time of culture is critical in determining whether the pollen grains will continue as gametophytes or evolve into sporophytic units. Hence, the ultimate control of embryogenic induction might lie in the genes that are expressed during a narrow developmental window centering around
Adventive Embryogenesis
the time of the first haploid mitosis. Determination of the parameters of this stage in pollen cytology as it relates to the cell cycle is necessary to achieve synchrony in pollen development which can lead to increased yields of embryoids and plantlets. Before concluding this chapter, it is important to emphasize that the ease of production of isogenic diploids by anther culture has had a significant impact on genetic and breeding experiments in many plants. Although this area of work has not been stressed in the preceding pages, it should be kept in mind that haploid plants and isogenic diploids have been generated in most of our crops by anther culture technology and improved strains of plants using doubled haploids in breeding programs are already under cultivation in many parts of the world.
SECTION V
Applications
Chapter 18
Genetic Transformation of Embryos 1. Transformation through Pollen Grains and Ovules 2. Transformation of Developing Embryos (i) DNA Delivery by Microprojectiles (ii) Other Methods of Embryo Transformation 3. Transformation of Embryogenic Cells and Somatic Embryos (i) Monocots and Dicots (ii) Gymnosperms 4. General Comments
One of the most promising approaches currently available to improve the quality, diversity, and yield of our crop plants is genetic engineering. In a broad sense, the genetic engineering of plants connotes a manipulation of plant cells and organs at the molecular level leading to the introduction, integration, and expression of specific and useful segments of foreign genetic material in a host plant. The application of genetic manipulative techniques has resulted in the production of plants with altered metabolic pathways and useful agronomic traits such as insect, viral, or herbicide resistance; it has also generated custom-made male-sterile plants useful in hybrid seed production. Other genetic engineering approaches, such as improving the post-harvest qualities of fruits and vegetables and creating crops that synthesize useful vaccines, Pharmaceuticals, or chemicals, are already at an advanced stage on the drawing board or are well into the developmental phase. Historically, engineering of the first transgenic plants relied on the ability of the natural vector Agrobacterium tumefaciens to introduce recombinant
DNA molecules into plant cells; in later years, this has become the most widely used procedure for the genetic transformation of various dicotyledonous
plants. Cocultivation of isolated protoplasts with bacteria was the method of choice in some of the early attempts at Agrobacterium-mediated transformation. Currently, the less cumbersome and more reliable method of leaf disc transformation is routinely used to obtain transgenic dicots under standard tissue culture conditions. The limited susceptibility of monocotyledons to A. tumefaciens infection has severely restricted the use of this vector to transform monocots, particularly the cereals. Successful gene transfer methods developed for monocots involve direct DNA uptake by isolated protoplasts through osmotic shock, DNA transfer by electroporation of protoplasts and cells, and microprojectile-mediated DNA transfer into cells, tissues, and organs capable of regeneration by organogenesis or somatic embryogenesis. In the context of the discussion on various aspects of male and female gametogenesis and embryogenesis in flowering plants given in the previous chapters, the objective here is to review the work done on plant transformation using male and female sexual cells, zygotes, developing embryos, embryogenic cells, and embryoids instead of leaf discs as the starting material. It is now a relatively simple exercise in genetic engineering to produce transgenic tobacco plants by transplanting the appropriate cloned gene by Agrobacterium-mediated leaf disc transformation. The sexual cells, embryogenic cells, and embryos thus far investigated are not comparable to the tobacco leaf in its simplicity and efficiency in the uptake and integration of foreign genes. However, the original protocols referred to previously and their modifications to suit particular situations, used alone or in combination, have resulted in the introduction of alien genes into many plants through the sexual route. 525
526 As will be seen later, not all genes that have been successfully introduced are agronomically important; moreover, in many cases, only the expression of reporter genes has been followed by transient assays of regenerants. A further caveat regarding these investigations is that field trials to determine the stability of gene integration and expression through several sexual crosses have been conducted in only a few cases. The elegance of the use of the sexual route for plant transformation resides in the freedom to choose a single-celled sperm, egg, zygote, or microspore for the delivery and integration of foreign DNA. Embryogenic somatic cells and microspores offer potential targets for transformation comparable to the zygote in their ability to recapitulate embryogenic development with the introduced genes. Reviews by Christou (1995), Jahne, Becker and Lorz (1995), Ritala et al (1995), Sagi et al (1995b), and Sautter et al (1995) published in a symposium volume are considered especially relevant to the topics discussed here.
Applications
remains unproven in these experiments. Some investigators have reported a total lack of success in attempted pollen-mediated transformation in maize (Sanford, Skubik and Reisch 1985; Booy, Krens and Huizing 1989). Two protocols using electrotransformed pollen to produce transgenic plants have been described (Matthews, Abdul-Baki and Saunders 1990; Smith et al 1994). Plasmid constructs carrying the GUS gene were incorporated by electroporation into germinating pollen grains of Nicotiana gossei. Positive evidence of the production of transformed plants by pollination with the electroporated pollen has come mainly from Southern blot of genomic DNA of the putative transformants. Another method tested involved targeting male sporogenous cells, instead of pollen grains, as the recipients for gene transfer. This was done by the particle bombardment of tassels of maize with GUS gene or anthocyanin marker genes at the time of the initiation of stamen primordia and the culturing of the tassels to produce functional pollen grains in vitro. The positive GUS activity that developed in 1. TRANSFORMATION THROUGH POLLEN the tassels of the mature anthers showed that DNA GRAINS AND OVULES integration could be facilitated during anther There is considerable latitude for the investiga- ontogeny (Dupuis and Pace 1993). Assuming that in tor using pollen grains as vectors for transformation vitro matured and transformed pollen grains will purposes because DNA can be transferred to recip- be as effective as normal pollen grains in the fertilient plants in a variety of ways. These include mix- ization process, this technique offers some advaning foreign DNA with pollen grains, mixing tages when the collection of pollen grains from Agrobacterium containing the desired gene with small flowers presents a drawback. pollen grains, the microinjection of DNA into pollen An indirect pollen delivery system used with cells, the electroporation of germinating pollen some success followed from the demonstration of grains in the presence of DNA, the introduction of the transfer of the tumor-inducing gene from A. DNA into pollen grains by particle bombardment, tumefaciens into progenies of Petunia hybrida (Hess and allowing the uptake of DNA by pollen tubes. and Dressier 1989) and the A. tumefaciens-mediated The pollen grains and pollen tubes are subsequently transfer of the GUS gene into pollen grains used for pollination and fertilization in the normal (Siissmuth, Dressier and Hess 1991). In the protocol reproductive cycle. The possibility of introducing based on the Agrobacterium infection of pollen foreign genes into plants through pollen grains was grains to transfer the kanamycin-resistance gene to suggested by experiments on the transfer of antho- wheat, the pollen-bacterial suspension was pipetcyanin genes within and between species of Petunia ted into the spikelets, which were allowed to com(Hess 1980). Later investigators obtained some plete the fertilization cycle and form mature putative transformants using the male gametophyte caryopses. The kanamycin-resistant seedlings as a carrier of exogenous DNA into the genome of obtained were fully fertile and showed evidence of the egg or zygote of maize using the normal fertil- the integration of parts of the donor DNA in the ization cycle (de Wet et al 1985; Ohta 1986; Roeckel genome of the F2 generation (Hess, Dressier and et al 1988). Either the total genomic DNA of plants Nimmrichter 1990). However, problems concernpossessing dominant alleles of a few marker genes ing the stability of the foreign DNA were not comor plasmid DNA harboring a gene expressing pletely eliminated in this work and have continued kanamycin resistance was used in the transforma- to plague the approach. Another group of investition experiments. Since no molecular controls of gators have now claimed that the apparent transtransformation were included in the protocol, or formation of maize and other cereals by T-DNA controls turned out to be negative, the case for the from A. tumefaciens through the male sexual route unambiguous incorporation of foreign DNA is an artifact of the procedure and does not rigor-
Genetic Transformation of Embryos
ously prove transformation of the plant nuclear genome (Langridge et al 1992). The possibility of directly introducing genes into the egg, the zygote, or the endosperm nucleus through the pollen tubes is inferred from studies in which exogenous DNA was applied to the styles of pollinated flowers of cotton, rice, and wheat. It is believed that the injected DNA is taken up by the pollen rubes and transferred to the embryo sac before or after fertilization (Gong et al 1988; G. Y. Zhou et al 1988; F.-Y. Wang et al 1995). Changes observed in the protein profiles of endosperms of lines of wheat transformed with sorghum DNA suggest that the donor genes are probably expressed (F.-Y. Wang et al 1995). That DNA penetrates the germ cells of rye was shown by injecting the inflorescence, before male and female meiosis, with plasmids harboring a gene coding for resistance to kanamycin. Although molecular signals established the transgenic nature of the small number of kanamycin-resistant seedlings recovered, Mendelian inheritance of the gene was not demonstrated (de la Pefta, Lorz and Schell 1987).
527
medium were isolated and allowed to undergo organogenesis in a suitable medium, and the plantlets formed were reared to maturity. The expression of reporter genes in the transformants, presence of integrated copies of the inserted DNA in their genome, ability of explants of transformants to form calluses and roots on the selective medium, and transmission of the resistance gene according to Mendelian principles are considered to provide evidence of transformation (Sangwan, Bourgeois and Sangwan-Norreel 1991). Through variations of this protocol, transgenic plants have been regenerated from seed embryos of Vigna unguiculata (Penza, Lurquin and Filippone 1991), Cicer arietinum (Fontana et al 1993), Pisum sativum (Schroeder et al 1993), Datura innoxia (Ducrocq, Sangwan and Sangwan-Norreel 1994), and Panax ginseng (ginseng; Araliaceae) (Lee et al 1995). Investigations on D. innoxia and ginseng have taken advantage of the ability of the cells of the infected cultured embryos to regenerate somatic embryos directly on the explant or through a callus phase and thus produce a large crop of transformed plants.
With the demonstration of the Agrobacteriummediated transfer of viral sequence to the stem of maize seedlings (agroinfection), the competence of maize embryos for agroinfection has been established. Although the susceptibility of embryos varies with the genotype, the stage of embryogenesis endowed with a functional shoot apical meristem is thought to be the major factor conferring readiness for successful T-DNA transfer (Schlappi and Hohn 1992). Transgenic rice tissues were recovered from mature rice embryos inoculated with Agrobacterium, but the regeneration of plants was not reported from these cultures (Raineri et al 1990). A breakthrough in the Agrobacterium-mediated transformation of rice was made in two independent investigations using embryo explants to transfer marker genes. Chan et al (1993) obtained a few transgenic plants by infecting immature embryos with bacteria carrying chimeric GUS and NPTII genes under the control of promoters of the 2. TRANSFORMATION OF DEVELOPING rice oc-amylase gene and the Agrobacterium nopaEMBRYOS line synthase gene, respectively. Calluses regenerThere are indications that seed embryos of certain ated from embryos gave rise to plants by plants can be rapidly transformed by coculturing organogenesis. A higher rate of recovery of transwith A. tumefadens. In the protocol developed for the genic plants was obtained when calluses initiated transfer of two reporter genes into Arabidopsis, the from excised scutella of mature embryos were critical step for efficient transformation was a three- cocultured with the bacteria (Hiei et al 1994). Both day preculture of immature or mature embryos in a works clearly demonstrated the stable integration of medium supplemented with an auxin and a foreign DNA into the rice genome, as well as the sexcytokinin, followed by incubation with the bacterial ual inheritance of the genes in a Mendelian fashion. vector for two days. The kanamycin- or hygromycin- This technology has now been extended to maize resistant calluses that appeared on a regeneration embryos to produce high frequencies of transferIn another approach, followed by Luo and Wu (1988), plasmid DNA coding for neomycin phosphotransferase (NPTII) under the control of the CaMV 35S promoter was applied to decapitated stylar ends of rice florets after normal pollination, and both phenotypic and molecular proofs to support claims of transformation were observed. It has been proposed that the exogenous DNA is taken up by the pollen tubes and integrated into the egg during fertilization. A critique of this work, however, has pointed out that instead of integrating into the rice genome as a unique, high-molecular mass, the foreign DNA is found in Southern blots at different locations in different transformants (Potrykus 1990). So, although we have some glimpse of the fate of the donor DNA, it remains to be established whether it becomes part of the host genome without undergoing denaturation or degradation.
528
mants hitherto not possible with biolistic methods (Ishida et al 1996). Although the reasons for the success that had eluded previous investigators are not clear, the long controversy about the ability of Agrobacterium to transform monocotyledons may now be put to rest, at least for two major crops. Transformation using pollen embryoids has rarely been attempted, but its potential is great. The transformation of developing embryoids in cultured anthers will eliminate the need for the cumbersome isolation procedures usually necessary for seed embryos. Since the embryoids grow directly into plantlets, it is possible to bypass the problems of the regeneration of plantlets from transgenic calluses. The first, and the most direct, attempt to transform pollen embryoids was by Neuhaus et al (1987), who delivered DNA by microinjection into globular pollen embryoids of Brassica napus. The very nature of the technique, targeting only one cell for injection, resulted in numerous chimeric events in the regenerated embryoids and plantlets. When secondary embryoids, presumably formed from transformed single cells, were analyzed, it was possible to demonstrate the predicted segregation of the marker gene. Microinjection of chimeric DNA into unicellular microspores of B. napus has shown that the relatively small amount of DNA transferred is insufficient to attain stable transformation (Jones-Villeneuve et al 1995). Direct DNA uptake mediated either by polyethylene glycol or by electroporation was found to cause transient expression of reporter genes in microspores of barley (Kuhlmann et al 1991) and maize (Fennell and Hauptmann 1992), but the regeneration of transgenic plants was not achieved. Present indications are that Agrobacterium-mediated transformation is probably a more versatile way to transform pollen embryoids than either microinjection or direct DNA uptake. This became evident from the successful infection of proembryoids of B. napus by bacteria containing selectable genes for antibiotic resistance, although no transgenic plants showing integration of T-DNA into the nuclear genome were recovered (Pechan 1989). The production of transgenic haploid plants of Datura innoxia and Nicotiana tabacum
seems to provide good examples of the ease of transforming pollen embryoids by the cocultivation of anthers or embryogenic pollen grains with the bacterial vector. The data from this work imply that with both species, competence for transformation is correlated with the onset of the growth of cotyledons on the embryoids, because no transformants are recovered from embryogenic pollen grains or proembryoids cocultured with the bacteria (Sangwan, Ducrocq and Sangwan-Norreel 1993).
Applications
(i) DNA Delivery by Microprojectiles By direct microprojectile delivery of DNA into embryogenic pollen or embryoids, transgenic plants of barley, wheat, and tobacco have been produced. When freshly isolated pollen grains of barley were used as target cells, the transferred GUS genes and BAR genes (encoding phosphinothricin acetyltransferase [PAT], an enzyme that inactivates the herbicidal compound L-phosphinothricin [LPPT]) were inherited in all progeny plants of pollen grain origin, indicating the homozygous nature of the primary regenerants (Jahne et al 1994). In experiments targeting pollen embryoids of barley, the possibility of producing herbicide-resistant plants was clearly in view, although only resistant calluses were obtained in the limited trials conducted (Wan and Lemaux 1994). In work with tobacco, pollen grains induced to form embryoids by starvation were bombarded immediately before or after the first division with plasmid DNA bearing a GLZS-CaMV 35S promoter construct. The two transgenic plants obtained were analyzed by Southern blot to establish stable integration of the foreign DNA, and selfing and backcrosses of one diploidized progeny clearly demonstrated that the transgene was transmitted to the offspring in accordance with Mendelian predictions (Stoger et al 1995). Compared to CaMV 35S promoter, GUS gene under the control of maize ubiquitin promoter, when delivered to early stage pollen embryoids of wheat, produced a high level of expression in regenerated plants (Loeb and Reynolds 1994). The delivery of foreign DNA into developing seed embryos by microprojectile bombardment has been attempted mainly in cereals. In some dicots that are easily infected by Agrobacterium tumefaciens, regeneration by leaf discs is notoriously difficult and slow. Here also, as in the cereals, developing embryos have been used as the starting material for direct introduction of foreign genes. In some of the early works, using immature embryos of barley, pearl millet, and wheat, success was limited to demonstrating the transient expression of reporter genes under the control of appropriate promoters (Kartha et al 1989; Chibbar et al 1991; Taylor and Vasil 1991; Taylor, Vasil and Vasil 1993). Based on these results, transformation systems for cereals of major agricultural importance have been developed that involve the microprojectile bombardment of embryos, the subsequent culture of bombarded embryos to promote the formation of callus, the selection and maintenance of the embryogenic callus, the regeneration of plants, and segregation analysis of the progeny containing the
Genetic Transformation of Embryos
introduced gene. Included in the list of plants used for transformation by this route are sorghum (Casas et al 1993), several varieties of wheat (Lonsdale, Onde and Cuming 1990; Vasil et al 1992, 1993; Weeks, Anderson and Blechl 1993; Becker, Brettschneider and Lorz 1994; Iglesias et al 1994; Nehra et al 1994; Blechl and Anderson 1996), maize (Klein et al 1988; Koziel et al 1993), rice (Christou, Ford and Kofron 1991; L. Li et al 1993), and barley (Wan and Lemaux 1994; Ritala et al 1994). Most of the manipulations were carried out with embryos excised from grains midway through development. Embryos were bombarded for a brief period immediately after excision, or after culture, but before the initiation of callus. Microprojectiles were delivered from the scutellar side or, rarely, from the side of the embryo axis; in some cases, excised pieces of the scutellum were bombarded and cultured. Some of the results obtained bear out the prediction that it is possible to engineer insecticide or herbicide tolerance in plants by transferring into immature embryos the appropriate genes. For example, European corn borer, which attacks maize, is susceptible to insecticidal proteins produced by Bacillus thuringiensis. Koziel et al (1993) introduced into maize embryos a chimeric CKY-1A gene encoding a truncated version of the protein derived from B. thuringiensis. Transformed plants containing the chimeric gene were found to express high levels of the insecticide protein and to offer resistance to repeated heavy infestations by the European corn borer. In wheat transformation experiments, the marker gene BAR was introduced; stably transformed plants selected using L-PPT, an ingredient of the commercial herbicide basta, showed resistance to topical applications of the herbicide (Vasil et al 1993; Nehra et al 1994). Rice (Christou, Ford and Kofron 1991), wheat (Weeks, Anderson and Blechl 1993), sorghum (Casas et al 1993), and barley (Wan and Lemaux 1994) plants similarly transformed with the BAR gene were selected in the presence of another L-PPT-containing herbicide, bialaphos, and were found to be resistant to further applications of this compound. The feasibility of manipulating the levels and types of storage proteins in wheat endosperm appears close to reality by the introduction of a high-molecular-mass glutenin gene into embryos and by the subsequent regeneration of transgenic lines (Blechl and Anderson 1996). These findings have considerably broadened our understanding of the usefulness of organized tissues such as the embryo to act as transformation targets by particle acceleration. The protocol may also prove highly efficient in introducing other useful traits into additional varieties of these crops. Since regeneration of many crops in tis-
529
sue culture has a strong genotype-dependent component, the development of a genotype-independent transformation system for each crop would be a significant achievement. Among dicots, transgenic soybeans were the first to be regenerated by the particle bombardment of excised embryos. Plasmid DNA containing an NPTII coding region under the control of CaMV 35S promoter was introduced into the shoot apical meristems of immature embryo axes by particle acceleration. The explants were then cultured on a medium containing a high level of benzylaminopurine to induce shoot regeneration. Although the plantlets obtained were chimeric, the foreign gene was expressed in the recovered first generation offspring (McCabe et al 1988). In another work, using cotyledons of immature embryos of six varieties of soybeans, it was found that transient expression of the reporter gene driven by CaMV 35S promoter was dependent upon the stage of cotyledon development, with the younger stages being less responsive than the older ones, but the genotype had an overriding effect over the age of the embryo (Moore, Moore and Collins 1994). The development of a transformation system for cotton embryonic axes involved a protocol similar to that used for soybean and resulted in the production of transformants in which GUS gene was integrated and was transmitted to the progeny in a Mendelian fashion (McCabe and Martinell 1993). Stable transformation of embryos of Carica papaya (papaya) (Fitch et al 1990; Cabrera-Ponce, Vegas-Garcia and Herrera-Estrella 1995), bean (Aragao et al 1993), sunflower (Hunold et al 1995), Eucalyptus globulus (Rochange et al 1995; Serrano et al 1996), and Brassica juncea (Kost et al 1996) has been achieved following DNA delivery by particle bombardment, although regenerants were recovered only in papaya. In one exciting development in this line of investigation, the regeneration of transgenic papaya plants resistant to the papaya ring spot virus was possible by the bombardment of 2,4-Dtreated immature seed embryos with the gene for the ring spot virus coat protein (Fitch et al 1992). In another work, after particle bombardment of embryos, bean plants transgenic for GUS, BAR, and bean golden mosaic virus coat genes were characterized for over five generations of selfing without any loss of introduced genes (Russell et al 1993).
(ii) Other Methods of Embryo Transformation In a continuing search for methods to transform embryos of specific plants, other successful reports
530
may bear a closer examination. Soyfer (1980) found that injection of milky stage endosperm DNA of a nonwaxy mutant of barley into milky stage grains of a wx mutant caused normal starch production in the offspring. It may be that integration of the donor DNA into the proembryo occurred in this experiment; however, because of the reversion of the transformed state, the conditions for maintaining the stability of the transformants are in need of definition. Electroporation is another technique that has been used to introduce DNA into whole embryos. Fertile transgenic plants harboring the NPT11 gene were regenerated after electroporation both of immature maize embryos wounded either enzymatically or mechanically (D'Halluin et al 1992) and of mature half-embryos of rice (Xu and Li 1994). Gene transfer has been achieved into isolated rice embryos and into scutella of isolated wheat embryos electroporated without any pretreatment, although no transgenic plants have been regenerated by this method (Kloti et al 1993; Rao 1995). In its simplest form, the transient expression of chimeric genes is possible when dry embryos excised from cereals and legumes are imbibed in a solution containing DNA (Topfer et al 1989; Yoo and Jung 1995); the uptake of DNA is thought to occur due to changes in the physico-chemical properties of plant cell membranes when dehydrated embryos are hydrated. This method, however, does not seem to have caught on. In short, considering the results obtained with dicots and monocots, it can be said that the particle acceleration method of introducing new genes into embryos has proved to be an efficient method of producing transgenic plants. No doubt, the presence of meristems makes isolated embryos an interesting target system for microprojectile bombardment. The ability of embryos to grow into new plants in culture directly, or through the intervention of a callus, or by somatic embryogenesis, is also a unique property of isolated embryos that makes them suitable starting material for genetic engineering. 3. TRANSFORMATION OF EMBRYOGENIC CELLS AND SOMATIC EMBRYOS
Embryogenic cells that regenerate somatic embryos possess features characteristic of meristematic cells and hence have served as favorite targets for DNA delivery. In a few instances, somatic embryos have been used for the transformation of regenerable plants. Microprojectile bombardment
Applications
is the most exploited of the techniques for gene transfer into embryogenic cells and embryoids. This technique has scored over Agrobacteriummediated transfer and electroporation because of the relatively high transformation frequencies obtained. (i) Monocots and Dicots
Experiments on a few species such as maize, in which embryogenic suspension cells are bombarded with the BAR gene, illustrate the ease of obtaining fertile transgenic plants by this route. Like transgenic wheat plants described in the previous section, transgenic maize plants regenerated from embryogenic cells by organogenesis or somatic embryogenesis are protected against basta and other L-PPT-containing herbicides (Fromm et al 1990; Gordon-Kamm et al 1990; Weymann et al 1993). A similar protocol was used to obtain transformed lines by selecting for resistance to hygromycin and chlorsulfuron (Fromm et al 1990; Walters et al 1992). There is now an impressive list of plants in which transformants for resistance to herbicides or antibiotics have been selected after microprojectile bombardment of embryogenic cells. Among monocots, this includes oat (Somers et al 1992),rice(Cao et al 1992), and barley (Wan and Lemaux 1994) selected for herbicide resistance; rice (L. Li et al 1993; S. Zhang et al 1996), Festuca arundinacea, F. rubra (Poaceae) (Spangenberg et al 1995a), Lolium perenne (Spangenberg et al 1995b), and Musa sp. (Musaceae) (Sagi et al 1995a) selected for hygromycin resistance; and sugarcane (Bower and Birch 1992) and Triticum X Hordeum amphidiploid hybrid (Barcelo et al 1994) selected for geneticin resistance. In other monocots, such as Agrostis palustris (Poaceae) (Zhong et al 1993) and Cenchrus ciliaris (Poaceae) (Ross, Manners and Birch 1995), transgenic plants have been recovered without any selection pressures after bombardment of embryogenic calluses. It appears likely that the efficiency of regeneration in each case may be related to the state of the callus at the time of the bombardment effecting DNA transfer process. Alternative methods to obtain transgenic plants include the incorporation of plasmid DNA by electroporation into untreated embryogenic calluses of cereals or of calluses wounded either enzymatically or mechanically and the incorporation by direct uptake following laser microbeam puncture of cells (D'Halluin et al 1992; Guo, Liang and Berns 1995; Pescitelli and Sukhapinda 1995). Attempts to introduce DNA by particle bombardment into embryogenic calluses or embryoids
Genetic Transformation of Embryos
531
of dicotyledons are limited to a few species, such as of Picea abies (Robertson et al 1992; Yibrah et al 1994; Gossypium hirsutum (Finer and McMullen 1990), Clapham et al 1995); in the embryogenic callus, Glycine max (Sato et al 1993; Hadi, McMullen and somatic embryos, and seedlings of P. glauca and P. Finer 1996), Vitis sp. (Hebert et al 1993; Scorza et al marina (Bommineni et al 1993; Charest et al 1993; 1995; Kikkert et al 1996), Coffea sp. (van Boxtel et al Ellis et al 1993; Charest, Devantier and Lachance 1995), Lathyrus sativus (Barna and Mehta 1995), 1996); and in the embryogenic cells of Pinus radiata Carica papaya (Cabrera-Ponce, Vegas-Garcia and (Walter et al 1994). As other methods of gene delivHerrera-Estrella 1995), Citrus reticulata X C. paradisiery are tested with embryogenic cells, it should be (Yao et al 1996), and Manihot esculenta (Schopke possible to tailor one of them to generate a high freet al 1996). In V. vinifera, recovery of trans- quency of regeneration in gymnosperms. genic plants resistant to kanamycin was facilitated The formulation of a routine method to obtain when somatic embryos initially bombarded with stably transformed gymnosperms on a level comuncoated microprojectile particles were coculti- parable to that achieved in angiosperms is not on vated with Agrobacterium tumefaciens or A. rhizo-the horizon. Two major reasons can be cited for this genes housing GUS or NPT11 genes (Scorza et al pessimistic view. Although somatic embryos of 1995). Direct cocultivation with A. tumefaciens gymnosperms possess the capacity to regenerate containing genes with suitable expression signals whole plants, the process is relatively slow. has proved to be reliable and efficient with other Attempts to establish the inheritance of foreign embryogenic calluses or somatic embryos for genes have to be seen in terms of the number of regeneration of transgenic plants. Examples of trans- years required for plants to reach the cone-bearing genic plants obtained by this means whose trans- age. The limitation of performing controlled polliformation status has been established are Juglans nation experiments is also a strong reason for regia (McGranahan et al 1988, 1990), Carica papaya believing that many years of hard work lie ahead (Fitch et al 1993; Cabrera-Ponce, Vegas-Garcia on the way to reaching the goal of demonstrating and Herrera-Estrella 1996), Asparagus officinalis sexual inheritance of the introduced gene. (Delbreil, Guerche and Jullien 1993), V. rupestris (Mullins, Tang and Facciotti 1990), and V. berlandieri X V. rupestris (Le Gall et al 1994). However, trans- 4. GENERAL COMMENTS genic plants of V. vinifera recovered after cocultivaCritical insights into the fundamental processes tion of embryogenic callus with an engineered of sexual reproduction in angiosperms, combined strain of A. rhizogenes harboring both GUS and with the use of recombinant DNA techniques, have NPTII genes exhibited various phenotypes, includhelped to pioneer the production of transgenic ing normal plants and plants with wrinkled leaves plants by the introduction of external genetic mateand abundant roots (Nakano, Hoshino and Mii rial into the genotype of the embryo. The transfor1994). In the approach taken with Dendrathema granmation systems described in this chapter have the diflora (Asteraceae), leaf discs transformed with A. advantage that they will allow efficient gene transtumefaciens were induced to regenerate transgenic fer into germ cells, embryos, and embryoids, which somatic embryos and plants in a suitable medium can regenerate directly into transgenic plants. It has (Pavingerova' et al 1994). Although transformation not been convincingly established that naked DNA rates obtained by cocultivation with bacteria were can be taken up by egg cells or integrated into them somewhat low, this was compensated for by the through pollen tubes and that the altered phenohigh regenerative ability of the embryogenic cells types of the transformed offspring are heritable. To and by the large number of transformed somatic some extent this is compensated for by the rigorous embryos. approaches employed to transform pollen embryoids, zygotic embryos, embryogenic cells, and somatic embryos so that they can regenerate geneti(ii) Gymnosperms cally engineered phenotypes of some of our agriculThe development of protocols for somatic tural crops. When techniques for the transformation embryogenesis in gymnosperm genera has led to of the egg or zygote are perfected, they may provide the exploitation of transformation techniques to a strategic testing ground for the genetic engineering study the regulation of gene expression in stably of the plant phenotype beginning with the first cell transformed cells. Biolistic approaches have been of the sporophyte. Therefore, systems that may be used to monitor the expression of GUS or NPTII uniquely suited to this approach deserve to be genes in the embryogenic cells and somatic embryos explored widely.
References
Note: For a given author, all references with one or two coauthors are first listed alphabetically. All et al references are listed alphabetically after the last three-author reference for a given author.
162-169. New Delhi: Council of Scientific & Industrial Research. Aguirre, P.J., and Smith, A.G. 1993. Molecular characterization of a gene encoding a cysteine-rich protein preferentially expressed in anthers of Lycopersicon Aalen, R.B. 1995. The transcripts encoding two oleosin esculentum. Plant Mol. Biol. 23:477-487. isoforms are both present in the aleurone and in the Ahokas, H. 1978. Cytoplasmic male sterility in barley. embryo of barley (Hordeum vulgare L.) seeds. Plant II. Physiology and anther cytology of msm 1. Mol. Biol. 28:583-588. Hereditas 89:7-21. Aalen, R.B., Opsahl-Ferstad, H.-G., Linnestad, C, and Ahokas, H. 1980. Cytoplasmic male sterility in barley. Olsen, O.-A. 1994. Transcripts encoding an oleosin V. Physiological characterization of the msml-Rfmla and a dormancy-related protein are present in both system. Physiol. Plant. 48:231-238. the aleurone layer and the embryo of developing Ahokas, H. 1982a. Cytoplasmic male sterility in barley. barley (Hordeum vulgare L.) seeds. Plant ]. 5:385-396. VIII. Lipoxygenase activity and anther amino nitrogen in the msml-Rfmla system. Plant Physiol. Aarts, M.G.M., Dirkse, W.G., Stiekema, W.J., and Pereira, 69:268-272. A. 1993. Transposon tagging of male sterility gene in Arabidopsis. Nature 363:715-717. Ahokas, H. 1982b. Cytoplasmic male sterility in barley: Aarts, M.G.M., Keijzer, C.J., Stiekema, W.J., and Pereira, evidence for the involvement of cytokinins in A. 1995. Molecular characterization of the CER1 fertility restoration. Proc. Natl Acad. Sci. USA gene of Arabidopsis involved in epicuticular wax 79:7605-7608. biosynthesis and pollen fertility. Plant Cell Ai, Y, Kron, E., and Kao, T.-H. 1991. S-alleles are retained 7:2115-2127. and expressed in a self-compatible cultivar of Abad, A.R., Mehrtens, B.J., and Mackenzie, S.A. 1995. Petunia hybrida. Mol. Gen. Genet. 230:353-358. Specific expression in reproductive tissues and fate Ai, Y, Singh, A., Coleman, C.E., Ioerger, T.R., Kheyr-Pour, of a mitochondrial sterility-associated protein in A., and Kao, T.-H. 1990. Self-incompatibility in cytoplasmic male-sterile bean. Plant Cell 7:271-285. Petunia inflata: isolation and characterization of Abbott, A.G., Ainsworth, C.C., and Flavell, R.B. 1984. cDNAs encoding three S-allele-associated proteins. Characterization of anther differentiation in cytoSex. Plant Reprod. 3:130-138. plasmic male sterile maize using a specific isozyme Ainsworth, C, Crossley, S., Buchanan-Wollaston, V., system (esterase). Theor. Appt. Genet. 67:469-473. Thangavelu, M., and Parker, J. 1995. Male and Abdalla, M.M.F., and Hermsen, J.G.T. 1972. Unilateral female flowers of the dioecious plant sorrel show incompatibility: hypothesis, debate and its implicadifferent patterns of MADS box gene expression. tions for plant breeding. Euphytica 21:32—47. Plant Cell 7:1583-1598. Abe, T., and Futsuhara, Y. 1989. Selection of higher regen- Akatsuka, T., and Nelson, O.E. 1966. Starch granuleerative callus and change in isozyme pattern in rice bound adenosine diphosphate glucose-starch gluco(Oryza sativa L.). Theor. Appl. Genet. 78:648-652. syltransferase of maize seeds. /. Biol. Chem. Acevedo, A., and Scandalios, j.G. 1990. Expression of the 241:2280-2285. catalase and superoxide dismutase genes in mature Akhundova, G.G., Grinikh, L.I., and Shevchenko, V.V. pollen in maize. Theor. Appl. Genet. 80:705-711. 1978. The embryogenesis in Arabidopsis thaliana folAckerson, R.C. 1984a. Regulation of soybean embryogenlowing the y-irradiation of the plants in the generaesis by abscisic acid. /. Expt. Bot. 35:403-413. tive phase. Ontogenez 9:514-519. Ackerson, R.C. 1984b. Abscisic acid and precocious ger- Alam, S., and Sandal, PC. 1969. Electrophoretic analyses mination in soybeans. /. Expt. Bot. 35:414—421. of anther proteins from male-fertile and male-sterile Adler, K., and Miintz, K. 1983. Origin and development of Sudangrass, Sorghum vulgare var. sudanense (Piper). protein bodies in cotyledons of Vicia faba. Proposal Crop Sci. 9:157-159. for a uniform mechanism. Planta 157:401-410. Albani, D., Altosaar, I., Arnison, P.G., and Fabijanski, S.F. Agarwal, S. 1962. Embryology of Quinchamalium chilense 1991. A gene showing sequence similarity to pectin Lam. In Plant Embryology - A Symposium, pp. esterase is specifically expressed in developing 533
534 pollen of Brassica napus. Sequences in its 5 flanking region are conserved in other pollen-specific promoters. Plant Mol. Biol. 16:501-513. Albani, D., Robert, L.S., Donaldson, P.A., Altosaar, I., Amison, P.G., and Fabijanski, S.F. 1990. Characterization of a pollen-specific gene family from Brassica napus which is activated during early microspore development. Plant Mol. Biol. 15:605-622. Albani, D., Sardana, Rv Robert, L.S., Altosaar, I., Arnison, P.G., and Fabijanski, S.F. 1992. A Brassica napus gene family which shows sequence similarity to ascorbate oxidase is expressed in developing pollen. Molecular characterization and analysis of promoter activity in transgenic tobacco plants. Plant ]. 2:331-342. Albertini, L. 1965. fitude autoradiographique des syntheses d'acide ribonucl&que (RNA) au cours de la microsporogenese chez le Rhoeo discolor (Hance).
References
Allen, R.D., Trelease, R.N., and Thomas, T.L. 1988. Regulation of isocitrate lyase gene expression in sunflower. Plant Physiol. 86:527-532. Allen, R.D., Cohen, E.A., Vonder Haar, R.A., Adams, C.A., Ma, D.P., Nessler, C.L., and Thomas, T.L. 1987. Sequence and expression of a gene encoding an albumin storage protein in sunflower. Mol. Gen. Genet. 210:211-218. Allen, R.L., and Lonsdale, D.M. 1992. Sequence analysis of three members of the maize polygalacturonase gene family expressed during pollen development. Plant Mol. Biol. 20:343-345.
Allen, R.L., and Lonsdale, D.M. 1993. Molecular characterization of one of the maize polygalacturonase gene family members which are expressed during late pollen development. Plant J. 3:261-271. Allison, D.C., and Fisher, W.D. 1964. A dominant gene for male-sterility in upland cotton. Crop Sci. 4:548-549. Compt. Rend. Acad. Sci. Paris 260:651-653. Almoguera, C, Coca, M.C., and Jordano, J. 1995. Differential accumulation of sunflower tetraubiquiAlbertini, L. 1967. fitude autoradiographique des syntin mRNAs during zygotic embryogenesis and these de prot£ines au cours de la microsporogenese chez le Rhoeo discolor (Hance). Compt. Rend. Acad. Sci. developmental regulation of their heat-shock response. Plant Physiol. 107:765-773. Paris 264D:2773-2776. Albertini, L. 1971. Les acides nucl£iques et les prot£ines Almoguera, C, and Jordano, J. 1992. Developmental and au cours de la microsporogenese chez le Rhoeo disenvironmental concurrent expression of sunflower color Hance. Etude autoradiographique et cytophodry-seed-stored low-molecular-weight heat-shock tom<$trique. Rev. Cytol. Biol. Veget. 34:49-92. protein and Lea mRNAs. Plant Mol. Biol. 19:781-792. Albertini, L. 1975. fitude autoradiographique de l'incor- Alpi, A., Tognoni, F., and D'Amato, F. 1975. Growth reguporation du tryptophane-3H dans les microsporolator levels in embryo and suspensor of Phaseolus coccineus at two stages of development. Planta cytes et dans le tapis du Rhoeo discolor Hance. Caryologia 28:445-458. 127:153-162. Albertini, L., Grenet-Auberger, H., and Souvr<§, A. 1981. Alpi, A., Lorenzi, R., Cionini, P.G., Bennici, A., and fitude autoradiographique de l'incorporation de D'Amato, F. 1979. Identification of gibberellin Aj in pr^curseurs des polysaccharides et des lipides dans the embryo suspensor of Phaseolus coccineus. Planta les microsporocytes et le tapis du Rhoeo discolor 147:225-228. Hance. Caryologia 34:53-68. Altenbach, S.B., Kuo, C.-C, Staraci, L.C., Pearson, K.W., Albertini, L., Souvre, A., and Audran, J.C. 1987. Le tapis Wainwright, C, Georgescu, A., and Townsend, J. de l'anthere et ses relations avec les microsporocytes 1992. Accumulation of a Brazil nut albumin in seeds et les grains de pollen. Rev. Cytol. Biol. Veget.-Bot. of transgenic canola results in enhanced levels of 10:211-242. seed protein methionine. Plant Mol. Biol. 18:235-245. Albertsen, M.C., and Palmer, R.G. 1979. A comparative Altenbach, S.B., Pearson, K.W., Leung, F.W., and Sun, light- and electron-microscopic study of microsporoS.S.M. 1987. Cloning and sequence analysis of a genesis in male sterile (MS.) and male fertile soycDNA encoding a Brazil nut protein exceptionally beans (Glycine max (L.) Merr.). Am. J. Bot. 66:253-265. rich in methionine. Plant Mol. Biol. 8:239-250. Albertsen, M.C., and Phillips, R.L. 1981. Developmental Altenbach, S.B., Pearson, K.W., Meeker, G., Staraci, L.C., cytology of 13 genetic male sterile loci in maize. Can. and Sun, S.S.M. 1989. Enhancement of the methionine content of seed proteins by the expression of a J. Genet. Cytol. 23:195-208. chimeric gene encoding a methionine-rich protein in Alch6, J.D., Fernandez, M.C., and Rodriguez-Garcia, M.I. transgenic plants. Plant Mol. Biol. 13:513-522. 1994. Cytochemical features common to nucleoli and cytoplasmic nucleoloids of Olea europaea meiocytes: Altschuler, M., and Mascarenhas, J.P. 1982. Heat shock detection of rRNA by in situ hybridization. /. Cell Sci. proteins and effects of heat shock in plants. Plant 107:621-629. Mol. Biol. 1:103-115. Aldrich, H.C., and Vasil, I.K. 1970. infrastructure of the Altschuler, Y., Rosenberg, N., Harel, R., and Galili, G. 1993. The N- and C-terminal regions regulate the postmeiotic nuclear envelope in microspores of Podocarpus macrophyllus. J. Ultrastr. Res. 32:307-315. transport of wheat y-gliadin through the endoplasmic reticulum in Xenopus oocytes. Plant Cell Aleith, F., and Richter, G. 1990. Gene expression during 5:443^50. induction of somatic embryogenesis in carrot cell suspensions. Planta 183:17-24. Alvarez, M.R. 1968. Quantitative changes in nuclear DNA accompanying postgermination embryonic developAlexander, R., Alamillo, J.M., Salamini, F., and Bartels, D. ment in Vanda (Orchidaceae). Am.}. Bot. 55:1036-1041. 1994. A novel embryo-specific barley cDNA clone encodes a protein with homologies to bacterial glu- Alvarez, M.R., and Sagawa, Y. 1965a. A histochemical cose and ribitol dehydrogenase. Planta 192:519-525. study of embryo sac development in Vanda (Orchidaceae). Caryologia 18:241-249. Allen, R.D., Bernier, F., Lessard, P.A., and Beachy, R.N. 1989. Nuclear factors interact with a soybean p-con- Alvarez, M.R., and Sagawa, Y. 1965b. A histochemical glycinin enhancer. Plant Cell 1:623-631. study of embryo development in Vanda (Orchidaceae). Caryologia 18:251-261. Allen, R.D., Nessler, C.L., and Thomas, T.L. 1985. Developmental expression of sunflower US storage Amarasinghe, V., Dhami, R., and Carlson, J.E. 1996. protein genes. Plant Mol. Biol. 5:165-173. Polyamine biosynthesis during somatic embryogen-
References
535
two MADS box genes in wild-type and mutant esis in interior spruce (Picea glauca X P. engelmannii complex). Plant Cell Rep. 15:495-499. Petunia flowers. Plant Cell 4:983-993. Ambrose, M.J., Wang, T.L., Cook, S.K., and Hedley, C.L. Angenent, G.C., Franken, J., Busscher, M., Colombo, L., and van Tunen, A.J. 1993. Petal and stamen forma1987. An analysis of seed development in Pisum tion in Petunia is regulated by the homeotic gene sativum. IV. Cotyledon cell populations in vivo and in vitro. J. Expt. Bot. 38:1909-1920. jbpl. Plant /.4:101-112. Ammirato, P.V. 1974. The effects of abscisic acid on the Angenent, G.C., Franken, J., Busscher, M., van Dijken, A., development of somatic embryos from cells of carvan Went, J.L., Dons, H.J.M., and van Tunen, A.J. away (Carum carvi L.). Bot. Gaz. 135:328-337. 1995b. A novel class of MADS box genes is involved in ovule development in Petunia. Plant Cell Ammirato, P.V. 1977. Hormonal control of somatic 7:1569-1582. embryo development from cultured cells of caraway. Interactions of abscisic acid, zeatin, and gibberellic Angold, R.E. 1967. The ontogeny and fine structure of the acid. Plant Physiol. 59:579-586. pollen grain of Endymion non-scriptus. Rev. Palaeobot. Palynol. 3:205-212. Ammirato, P.V, and Steward, F.C. 1969. Indirect effects of irradiation: morphogenetic effects of irradiated Angold, R.E. 1968. The formation of the generative cell in the pollen grain of Endymion non-scriptus (L.). /. Cell sucrose. Devel Biol. 19:87-106. Sci. 3:573-578. An, Y.-Q., Huang, S., McDowell, J.M., McKinney, E.C., and Meagher, R.B. 1996. Conserved expression of the Anthony, J.L., Vonder Haar, R.A., and Hall, T.C. 1991. Arabidopsis ACT1 and ACT3 actin subclass in organ Nucleotide sequence of a genomic clone encoding primordia and mature pollen. Plant Cell 8:15-30. arcelin, a lectin-like seed protein from Phaseolus vulgaris. Plant Physiol. 97:839-840. Anand, V.V., Arekal, G.D., and Swamy, B.G.L. 1980. Chimeral embryoids of pollen origin in tobacco. Aoyagi, K., Bassham, J.A., and Greene, F.C. 1984. Curr. Sci. 49:603-604. Pyruvate orthophosphate dikinase gene expression Anderberg, R.J., and Walker-Simmons, M.K. 1992. in developing wheat seeds. Plant Physiol. 75:393-396. Isolation of a wheat cDNA clone for an abscisic acid- Aoyagi, K., and Chua, N.-H. 1988. Cell-specific expression of pyruvate, Pi dikinase. In situ mRNA inducible transcript with homology to protein hybridization and immunolocalization labeling of kinases. Proc. Natl Acad. Sci. USA 89:10183-10187. protein in wheat seed. Plant Physiol. 86:364-368. Anderson, J.M., and Barrett, S.C.H. 1986. Pollen tube growth in tristylous Pontederia cordata Appels, R., Bouchard, R.A., and Stem, H. 1982. cDNA (Pontederiaceae). Can. ]. Bot. 64:2602-2607. clones from meiotic-specific poly(A)+RNA in Liliunr. Anderson, M.A., Sandrin, M.S., and Clarke, A.E. 1984. A homology with sequences in wheat, rye, and maize. high proportion of hybridomas raised to a plant Chromosoma 85:591-602. extract secrete antibody to arabinose or galactose. Apuya, N.R., and Zimmerman, J.L. 1992. Heat shock gene Plant Physiol. 75:1013-1016. expression is controlled primarily at the translational level in carrot cells and somatic embryos. Anderson, M.A., Cornish, E.C., Mau, S.-L., Williams, E.G., Plant Cell 4:657-665. Hoggart, R., Atkinson, A., Bonig, I., Grego, B., Simpson, R., Roche, P.J., Haley, J.D., Penschow, J.D., Aragao, F.J.L., Grossi de Sa, M.-F., Davey, M.R., Brasileiro, Niall, H.D., Tregear, G.W., Coghlan, J.P., Crawford, A.C.M., Faria, J.C., and Rech, E.L. 1993. Factors influR.J., and Clarke, A.E. 1986. Cloning of cDNA for a encing transient gene expression in bean (Phaseolus stylar glycoprotein associated with expression of vulgaris L.) using an electrical particle acceleration self-incompatibility in Nicotiana alata. Nature device. Plant Cell Rep. 12:483-490. 321:38-44. Arahira, M., and Fukazawa, C. 1994. Ginkgo US seed storage protein family mRNA: unusual Asn-Asn linkage Anderson, M.A., Harris, P.J., Bonig, I., and Clarke, A.E. as post-transitional cleavage site. Plant Mol. Biol. 1987. Immuno-gold localization of a-L-arabinofura25:597-605. nosyl residues in pollen tubes of Nicotiana alata Link Arasu, N.T. 1968. Self-incompatibility in angiosperms: a et Otto. Planta 171:438-442. Anderson, M.A., McFadden, G.I., Bernatzky, R., Atkinson, review. Genetka 39:1-24. A., Orpin, T., Dedman, H., Tregear, G., Fernley, R., Arditti, J. 1979. Aspects of the physiology of orchids. Adv. and Clarke, A.E. 1989. Sequence variability of three Bot. Res. 7:421-655. alleles of the self-incompatibility gene of Nicotiana Arekal, G.D., and Ramaswamy, S.N. 1973. Embryology of alata. Plant Cell 1:483-491. Burmannia pusilla (Wall, ex Miers) THW. and its taxAnderson, O.D., Greene, F.C, Yip, R.E., Halford, N.G., onomic status. Beitr. Biol. Pflanz. 49:35^5. Shewry, PR., and Malpica-Romero, J.-M. 1989. Arnison, P.G., Donaldson, P., Ho, L.C.C., and Keller, W.A. Nucleotide sequences of the two high-molecular1990. The influence of various physical parameters weight glutenin genes from the D-genome of a hexaon anther culture of broccoli (Brassica oleracea var. ploid bread wheat, Triticum aestivum L. cv. italica). Plant Cell Tissue Organ Cult. 20:147-155. Cheyenne. Nud. Acids Res. 17:461-462. Arrigoni, O., de Gara, L., Tommasi, F., and Liso, R. 1992. Anderson, O.D., Litts, J.C., Gautier, M.-F., and Greene, Changes in the ascorbate system during seed develF.C. 1984. Nucleic acid sequence and chromosome opment of Viciafaba L. Plant Physiol. 99:235-238. assignment of a wheat storage protein gene. Nucl. Artschwager, E. 1947. Pollen degeneration in male-sterile Acids Res. 12:8129-8144. sugar beets, with special reference to the tapetal Angenent, G.C., Busscher, M., Franken, J., Dons, H.J.M., plasmodium. /. Agr. Res. 75:191-197. and van Tunen, A.J. 1995a. Functional interaction Aryan, A.P., An, G., and Okita, T.W. 1991. Structural and between the homeotic genes jbpl and pMADSl durfunctional analysis of promoter from gliadin, an ing Petunia floral organogenesis. Plant Cell endosperm-specific storage protein gene of Triticum 7:507-516. aestivum L. Mol. Gen. Genet. 225:65-71. Angenent, G.C., Busscher, M., Franken, J., Mol, J.N.M., Asahi, T., Kumashiro, T., and Kubo, T. 1988. Constitution of and van Tunen, A.J. 1992. Differential expression of mitochondrial and chloroplast genomes in male ster-
536
References
ile tobacco obtained by protoplast fusion of Nicotiana protein which is processed into five homologous tabacum and N. debneyi. Plant Cell Physiol. 29:43-49. inhibitors. Plant Cell 5:203-213. Ascher, P.D. 1966. A gene action model to explain game- Atkinson, B.G., Raizada, M., Bouchard, R.A., Frappier, tophytic self-incompatibility. Euphytica 15:179-183. J.R.H., and Walden, D.B. 1993. The independent Ascher, P.D. 1971. The influence of RNA-synthesis stage-specific expression of the 18-kDa heat shock inhibitors on in vivo pollen tube growth and the selfprotein genes during microsporogenesis in Zea mays incompatibility reaction in Lilium longiflorum Thunb. L. Devel Genet. 14:15-26. Theor. Appl. Genet. 41:75-78. Attia, M.S. 1950. The nature of incompatibility in cabbage. Ascher, P.D. 1975. Special stylar property required for Proc. Am. Soc. Hort. Sci. 56:369-371. compatible pollen-tube growth in Lilium longiflorum Auboiron, E., Carron, M.-P., and Michaux-Ferriere, N. Thunb. Bot. Gaz. 136:317-321. 1990. Influence of atmospheric gases, particularly Ascher, P.D. 1977. Localization of the self- and the interethylene, on somatic embryogenesis of Hevea specific-incompatibility reactions in style sections of brasiliensis. Plant Cell Tissue Organ Cult. 21:31-37. Lilium longiflorum. Plant Sci. Lett. 10:199-203. Audran, J.C., and Dicko-Zafimahova, L.D. 1992. Aspects Ascher, P.D., and Drewlow, L.W. 1975. The effect of preultrastructuraux et cytochimiques du tapis staminal pollination injection with stigmatic exudate on interchez Calotropis procera (Asclepiadaceae). Grana specific pollen tube growth in Lilium longiflorum 31:253-272. Thunb. styles. Plant Sci. Lett. 4:401^05. Aukerman, M.J., Schmidt, R.J., Burr, B., and Burr, F.A. Ascher, P.D., and Peloquin, S.J. 1966a. Effect of floral aging 1991. An arginine to lysine substitution in the bZIP on the growth of compatible and incompatible pollen domain of an opaque-2 mutant in maize abolishes tubes in Lilium longiflorum. Am. J. Bot. 53:99-102. specific DNA binding. Genes Devel. 5:310-320. Ascher, P.D., and Peloquin, S.J. 1966b. Influence of tem- Avanzi, S., Cionini, P.G., and D'Amato, F. 1970. perature on incompatible and compatible pollen Cytochemical and autoradiographic analyses on the tube growth in Lilium longiflorum. Can. ]. Genet. Cytol. embryo suspensor cells of Phaseolus coccineus. 8:661-664. Caryologia 23:605-638. Ascher, P.D., and Peloquin, S.J. 1968. Pollen tube growth Avanzi, S., Corsi, G., D'Amato, F., Floris, C, and Meletti, and incompatibility following intra- and inter-speP. 1967. The chromosome breaking effect of the cific pollinations in Lilium longiflorum. Am. ]. Bot. irradiated endosperm in water-soaked seeds of 55:1230-1234. durum wheat. Mutation Res. 4:704-707. Asghar, R., Fenton, R.D., DeMason, D.A., and Close, T.J. Avjioglu, A., and Knox, R.B. 1989. Storage lipid accumu1994. Nuclear and cytoplasmic localization of maize lation by zygotic and somatic embryos in culture. embryo and aleurone dehydrin. Protoplasma Ann. Bot. 63:409-420. 177:87-94. Ashihara, H., Fujimura, T., and Komamine, A. 1981. Ba, L.T., Cave", G., Henry, M., and Guignard, J.-L. 1978. Pyrimidine nucleotide biosynthesis during somatic Embryogenie des Potamogetonacees. fitude en embryogenesis in a carrot cell suspension culture. Z. microscopie electronique a balayage de 1'origine du Pflanzenphysiol. 104:129-137. cotyledon chez Potamogeton lucens L. Compt. Rend. Acad. Sci. Paris 286D:1351-1353. Ashley, T. 1972. Zygote shrinkage and subsequent development in some Hibiscus hybrids. Planta Baaziz, M., Aissam, F., Brakez, Z., Bendiab, K., El 108:303-317. Hadrami, I., and Cheikh, R. 1994. Electrophoretic patterns of acid soluble proteins and active isoAshley, T. 1975a. Fine structure of early endosperm develforms of peroxidase and polyphenoloxidase typifyopment in Hibiscus. Caiyologia 28:63-71. ing calli and somatic embryos of two reputed date Ashley, T. 1975b. Alterations in the fine structure of develpalm cultivars in Morocco. Euphytica 76:159-168. oping endosperm of Hibiscus hybrids. Caryologia 28:73-80. Babbar, S.B., and Gupta, S.C. 1980. Chilling induced Asker, S.E., and Jerling, L. 1992. Apomixis in Plants. Boca androgenesis in anthers of Petunia hybrida without Raton: CRC Press. any culture medium. Z. Pflanzenphysiol. 100:279-283. Aspart, L., Meyer, Y, Laroche, M., and Penon, P. 1984. Babbar, S.B., and Gupta, S.C. 1986. Putative role of ethylene in Datura metel microspore embryogenesis. Developmental regulation of the synthesis of proteins encoded by stored mRNA in radish embryos. Physiol. Plant. 68:141-144. Plant Physiol. 76:664-673. Bacic, A., Gell, A.C., and Clarke, A.E. 1988. Aspinall, CO., and Rosell, K.-G. 1978. Polysaccharide Arabinogalactan proteins from stigmas of Nicotiana alata. Phytochemistry 27:679-684. component in the stigmatic exudate from Lilium longiflorum. Phytochemistry 17:919-922. Backs-Hiisemann, D., and Reinert, J. 1970. EmbryoAspuria, E.T., Nagato, Y., and Uchimiya, H. 1994. bildung durch isolierte Einzelzellen aus Histochemical localization of a chimeric gene (rolCGewebekulturen von Daucus carota. Protoplasma GUS) expression in zygotic embryos of transgenic 70:49-60. tobacco plants. Ann. Bot. 73:465-469. Badenoch-Jones, J., Spencer, D., Higgins, T.J.V., and Millerd, A. 1981. The role of glycosylation in storageAstrom, H., Sorri, O., and Raudaskoski, M. 1995. Role of protein synthesis in developing pea seeds. Planta microtubules in the movement of the vegetative 153:201-209. nucleus and generative cell in tobacco pollen tubes. Sex. Plant Reprod. 8:61-69. Bae, J.M., Giroux, M., and Hannah, L. 1990. Cloning and Astrom, H., Virtanen, I., and Raudaskoski, M. 1991. Coldcharacterization of the brittle-2 gene of maize. stability in the pollen tube cytoskeleton. Protoplasma Maydica 35:317-322. Baerson, S.R., and Lamppa, G.K. 1993. Developmental 160:99-107. regulation of an acyl carrier protein gene promoter Atkinson, A.H., Heath, R.L., Simpson, R.J., Clarke, A.E., and in vegetative and reproductive tissues. Plant Mol. Anderson, M.A. 1993. Proteinase inhibitors in Biol. 22:255-267. Nicotiana alata stigmas are derived from a precursor
References
537
Baerson, S.R., Vander Heiden, M.G., and Lamppa, G.K. Bal, A.K., and De, D.N. 1961. Developmental changes in 1994. Identification of domains in an Arabidopsis acyl the submicroscopic morphology of the cytoplasmic carrier protein gene promoter required for maximal components during microsporogenesis in Tradescantia. organ-specific expression. Plant Mol. Biol. 26:1947-1959. Bevel Biol. 3:241-254. Bagga, Sv Adams, H v Kemp, J.D., and Sengupta-Gopalan, Balconi, C, Rizzi, E., Motto, M., Salamini, F., and C. 1995. Accumulation of 15-kilodalton zein in novel Thompson, R. 1993. The accumulation of zein protein bodies in transgenic tobacco. Plant Physiol. polypeptides and zein mRNA in cultured 107:13-23. endosperms of maize is modulated by nitrogen supply. Plant J. 3:325-334. Bagga, S., Sutton, D., Kemp, J.D., and Sengupta-Gopalan, C. 1992. Constitutive expression of the f3-phaseolin Baldan, B., Frattini, C, Guzzo, F., Branca, C, Terzi, M., gene in different tissues of transgenic alfalfa does Mariani, P., and Lo Schiavo, F. 1995. A stage-specific not ensure phaseolin accumulation in non-seed tisblock is produced in carrot somatic embryos by 1,2sue. Plant Mol. Biol. 19:951-958. benzisoxazole-3-acetic acid. Plant Sci. 108:85-92. Bagni, N., Adamo, P., and Serafini-Fracassini, D. 1981. Baldi, B.C., Franceschi, V.R., and Loewus, F.A. 1987. RNA, proteins and polyamines during tube growth Preparation and properties of pollen sporoplasts. in germinating apple pollen. Plant Physiol. Protoplasma 141:47-55. 68:727-730. Baldwin, T.C., Coen, E.S., and Dickinson, H.G. 1992. The Bai, S., and Sung, Z.R. 1995. The role of EMF1 in regulatptll gene expressed in the transmitting tissue of ing the vegetative and reproductive transition in Antirrhinum encodes an extensin-like protein. Plant j . Arabidopsis thaliana (Brassicaceae). Am. ]. Bot. 2:733-739. 82:1095-1103. Balsamo, R.A., Wang, J.-L., Eckard, K.J., Wang, C.-S., and Bailey, C.J., Cobb, A., and Boulter, D. 1970. A cotyledon Lord, E.M. 1995. Immunogold localization of a slice system for the electron autoradiographic study developmentally regulated, tapetal-specific, 15 kDa of the synthesis and intracellular transport of the lily anther protein. Protoplasma 189:17-25. seed storage protein of Viciafaba. Planta 95:103-118. Baltz, R., Domon, C, Pillay, D.T.N., and Steinmetz, A. Bailey, D.S., DeLuca, V., Diirr, M., Verma, D.P.S., and 1992a. Characterization of a pollen-specific cDNA Maclachlan, G.A. 1980. Involvement of lipid-linked from sunflower encoding a zinc finger protein. Plant oligosaccharides in synthesis of storage glycoproJ. 2:713-721. teins in soybean seeds. Plant Physiol. 66:1113-1118. Baltz, R., Evrard, J.-L., Domon, C, and Steinmetz, A. Bailey-Serres, J., Dixon, L.K., Liddell, A.D., and Leaver, 1992b. A LIM motif is present in a pollen-specific C.J. 1986a. Nuclear-mitochondrial interactions in protein. Plant Cell 4:1465-1466. cytoplasmic male-sterile Sorghum. Theor. Appl. Genet.Banas, M., Tirlapur, U.K., Charzyriska, M., and Cresti, M. 73:252-260. 1996. Some events of mitosis and cytokinesis in the Bailey-Serres, J., Hanson, D.K., Fox, T.D., and Leaver, C.J. generative cell of Ornithogatum virens L. Planta 1986b. Mitochondrial genome rearrangement leads 190:202-208. to extension and relocation of the cytochrome c oxi- Banerjee, U.C., Rowley, J.R., and Alessio, M. 1965. Exine dase subunit I gene in Sorghum. Cell 47:567-576. plasticity during pollen grain maturation. /. Palynol. Bajaj, Y.P.S. 1974a. Isolation and culture studies on pollen 1:70-89. tetrad and pollen mother-cell protoplasts. Plant Sci. Banga, S.S., Labana, K.S., and Banga, S.K. 1984. Male sterilLett. 3:93-99. ity in Indian mustard (Brassica juncea (L.) Coss.) - a Bajaj, Y.P.S. 1974b. Induction of repeated cell division in biochemical characterization. Theor. Appl. Genet. isolated pollen mother cells of Atropa belladonna. 67:515-519. Plant Sci. Lett. 3:309-312. Bapat, S.A., Rawal, S.K., and Mascarenhas, A.F. 1992. Bajaj, Y.P.S. 1979. Test-tube fertilization and development Isozyme profiles during ontogeny of somatic embryos of maize {Zea mays L.) plants. Indian J. Expt. Biol. in wheat (Triticum aestivum L.). Plant Sci. 82:235-242. 17-A75-478. Barcelo, P., Hagel, C, Becker, D., Martin, A., and Lorz, H. Bajaj, Y.P.S. 1983. In vitro production of haploids. In 1994. Transgenic cereal (Tritordeum) plants obtained Handbook of Plant Cell Culture, Vol. 1, ed. D.A. Evans, at high efficiency by microprojectile bombardment W.R. Sharp, P.V. Ammirato, and Y. Yamada, pp. of inflorescence tissue. Plant J. 5:583-592. 228-287. New York: Macmillan Publishing Co. Barker, S.J., Harada, J.J., and Goldberg, R.B. 1988. Cellular Bajaj, Y.P.S. 1990. In vitro production of haploids and their localization of soybean storage protein mRNA in use in cell genetics and plant breeding. In transformed tobacco seeds. Proc. Natl Acad. Sci. USA Biotechnology in Agriculture and Forestry 12. Haploids 85:458-462. in Crop Improvement I, ed. Y.P.S. Bajaj, pp. 3-44. Barna, K.S., and Mehta, S.L. 1995. Genetic transformation Berlin: Springer-Verlag. and somatic embryogenesis in Lathyrus sativus. Bajer, A. 1965. Cine micrographic analysis of cell plate for}. Plant Biochem. Biotech. 4:67-71. mation in endosperm. Expt. Cell Res. 37:376-398. Barnabas, B., Fransz, P.F., and Schel, J.H.N. 1987. Baker, H.G. 1966. The evolution, functioning and breakUltrastructural studies on pollen embryogenesis in down of heteromorphic incompatibility systems. I. maize {Zea mays L.). Plant Cell Rep. 6:212-215. The Plumbaginaceae. Evolution 20:349-368. Barnabas, B., and Fridvalszky, L. 1984. Adhesion and gerBaker, ]., Steele, C, and Dure, L., III. 1988. Sequence and mination of differentially treated maize pollen characterization of 6 Lea proteins and their genes grains on the stigma. Ada Bot. Hung. 30:329-332. from cotton. Plant Mol. Biol. 11:277-291. Barnabcis, B., and Kov^cs, G. 1992. In vitro pollen maturaBaker, S.S., Rugh, C.L., Whitmore, F.W., and Kamalay, J.C. tion and successful seed production in detached 1996. Genes encoding HS-globulin-like proteins are spikelet cultures in wheat (Triticum aestivum L). Sex. expressed in the megagametophyte soon after fertilPlant Reprod. 5:286-291. ization in eastern white pine (Pinus strobus L.). Int. ]. Barnes, S.H., and Blackmore, S. 1987. Preliminary obserPlant Sci. 157:453-461. vations on the formation of the male germ unit in
538
References
and germination in relation to protein synthesis. Catananche caerulea L. (Compositae: Lactuceae). Brief Plant Sci. Lett. 35:55-60. report. Protoplasma 138:187-189. Barratt, D.H.P. 1986a. Modulation by abscisic acid of stor- Bass, H.W., Webster, C, OBrian, G.R., Roberts, J.K.M., and Boston, R.S. 1992. A maize ribosome-inactivating age protein accumulation in Vicia faba L. cotyledons protein is controlled by the transcriptional activator cultured in vitro. Plant Sci. 46:159-167. opaque-2. Plant Cell 4:225-234. Barratt, D.H.P. 1986b. Regulation of storage protein accumulation by abscisic acid in Vicia faba L. cotyle- Bassett, C.L., Mothershed, C.P., and Galau, G.A. 1988. dons cultured in vitro. Ann. Bot. 57:245-256. Floral-specific polypeptides of the Japanese morning glory. Planta 175:221-228. Barratt, D.H.P., and Clark, J.A. 1991. Proteins arising during the late stages of embryogenesis in Pisum sativum Bassiiner, R., Baumlein, H., Huth, A., Jung, R., Wobus, U., L. Planta 184:14-23. Rapoport, T.A., Saalbach, G., and Miintz, K. 1988. Abundant embryonic mRNA in field bean (Vicia faba Barratt, D.H.P, and Clark, J.A. 1993. A stress-induced, L.) codes for a new class of seed proteins: cDNA developmentally regulated, highly polymorphic cloning and characterization of the primary translaprotein family in Pisum sativum L. Planta 191:7-17. tion product. Plant Mol. Biol. 11:321-334. Barratt, D.H.P., Domoney, C, and Wang, T.L. 1989. Purification and partial characterisation of two Bassuner, R., Manteuffel, R., Muntz, K., Puchel, M., abscisic-acid-responsive proteins induced in culSchmidt, P., and Weber, E. 1983. Analysis of in vivo tured embryos of Pisum sativum L. Planta 180:16-23. and in vitro globulin formation during cotyledon development of field beans (Vicia faba L. var. minor). Barratt, D.H.P, and Flavell, R.B. 1975. Alterations in mitoBiochem. Physiol. Pflanz. 178:665-684. chondria associated with cytoplasmic and nuclear genes concerned with male sterility in maize. Theor. Bastola, D.R., and Minocha, S.C. 1995. Increased Appl. Genet. 45:315-321. putrescine biosynthesis through transfer of mouse ornithine decarboxylase cDNA in carrot promotes Barratt, D.H.P, Whitford, P.N., Cook, S.K., Butcher, G., somatic embryogenesis. Plant Physiol. 109:63-71. and Wang, T.L. 1989. An analysis of seed development in Pisum sativum. VIII. Does abscisic acid pre- Baszczynski, C.L., and Fallis, L. 1990. Isolation and nucleotide sequence of a genomic clone encoding a vent precocious germination and control storage new Brassica napus napin gene. Plant Mol. Biol. protein synthesis? /. Expt. Bot. 40:1009-1014. 14:633-635. Barrett, S.C.H. 1977. The breeding system of Pontederia rotundifolia L., a tristylous species. New Phytol. Batchelder, C, Ross, J.H.E., and Murphy, D.J. 1994. Synthesis and targeting of Brassica napus oleosin in 78:209-220. transgenic tobacco. Plant Sci. 104:39-47. Barrett, S.C.H., ed. 1992. Evolution and Function of Heterostyly. Berlin: Springer-Verlag. Bateman, A.J. 1952. Self-incompatibility systems in angiosperms. I. Theory. Heredity 6:285—310. Bartalesi, A., Del Casino, C, Moscatelli, A., Cai, G., and Tiezzi, A. 1991. Confocal laser scanning microscopy Bateman, A.J. 1954. Self-incompatibility systems in angiosperms. II. Iberis amara. Heredity 8:305-332. of the microtubular system of dividing generative cell in Nicotiana tabacum pollen tube. Giorn. Bot. Hal. Bateman, A.J. 1955. Self-incompatibility systems in angiosperms. III. Cruciferae. Heredity 9:53-68. 125:21-28. Bartels, D., Singh, M., and Salamini, F. 1988. Onset of Batygina, T.B. 1969. On the possibility of separation of a new type of embryogenesis in angiosperms. Rev. desiccation tolerance during development of the Cytol. Biol. Veget. 32:335-341. barley embryo. Planta 175:485^92. Bartels, D., and Thompson, R.D. 1983. The characteriza- Baumgartner, B., Tokuyasu, K.T., and Chrispeels, M.J. 1980. Immunocytochemical localization of reserve tion of cDNA clones coding for wheat storage proprotein in the endoplasmic reticulum of developing teins. Nucl. Acids Res. 11:2961-2977. bean (Phaseolus vulgaris) cotyledons. Planta Bartels, D., and Thompson, R.D. 1986. Synthesis of 150:419^25. mRNAs coding for abundant endosperm proteins during wheat grain development. Plant Sci. Baumlein, H., Boerjan, W., Nagy, I., Bassuner, R., van Montagu, M., Inz6, D., and Wobus, U. 1991a. A novel 46:117-125. seed protein gene from Vicia faba is developmentally Bartels, D., Thompson, R.D., and Rothstein, S. 1985. regulated in transgenic tobacco and Arabidopsis Synthesis of a wheat storage protein subunit in plants. Mol. Gen. Genet. 225:459-467. Escherichia coli using novel expression vectors. Gene 35:159-167. Baumlein, H., Boerjan, W., Nagy, I., Panitz, R., Inz<5, D., and Wobus, U. 1991b. Upstream sequences regulatBartels, D., Altosaar, I., Harberd, N.P., Barker, R.F., and ing legumin gene expression in heterologous transThompson, R.D. 1986. Molecular analysis of ygenic plants. Mol. Gen. Genet. 225:121-128. gliadin gene families at the complex Gli-1 locus of bread wheat (T. aestivum L.). Theor. Appl. Genet. Baumlein, H., Mise"ra, S., LuerGen, H., Kolle, K., 72:845-853. Horstmann, C, Wobus, U., and Muller, A.J. 1994. The FUS3 gene of Arabidopsis thaliana is a regulator Bartels, D., Engelhardt, K., Roncarati, R., Schneider, K., of gene expression during late embryogenesis. Plant Rotter, M., and Salamini, F. 1991. An ABA and GA J. 6:379-387. modulated gene expressed in the barley embryo encodes an aldose reductase related protein. EMBO Baumlein, H., Muller, A.J., Schiemann, J., Helbing, D., Manteuffel, R., and Wobus, U. 1987. A legumin B J. 10:1037-1043. gene of Vicia faba is expressed in developing seeds of Barton, M.K., and Poethig, R.S. 1993. Formation of the transgenic tobacco. Biol. Zentral. 106:569-575. shoot apical meristem in Arabidopsis thaliana: an analysis of development in the wild type and in Baumlein, H., Muller, A.J., Schiemann, J., Helbing, D., Manteuffel, R., and Wobus, U. 1988. Expression of a the shoot meristemless mutant. Development Vicia faba legumin B gene in transgenic tobacco 119:823-831. plants: gene dosage-dependent protein accumulaBashe, D., and Mascarenhas, J.P. 1984. Changes in potastion. Biochem. Physiol. Pflanz. 183:205-210. sium ion concentrations during pollen dehydration
References
539
A.A.M. 1990. Organization of the actin cytoskeleton Baumlein, H., Nagy, I., Villarroel, R., Inz6, D., and Wobus, during megasporogenesis in Gasteria verrucosa visuU. 1992. Cis-analysis of a seed protein gene proalized with fluorescent-labelled phalloidin. Acta Bot. moter: the conservative RY repeat CATGCATG Neerl. 39:43-48. within the legumin box is essential for tissue-specific expression of a legumin gene. Plant J. 2:233-239. Bednarska, E. 1981. Autoradiographic studies of DNA and histone synthesis in successive differentiation Baumlein, H., Wobus, U., Pustell, J., and Kafatos, F.C. stages of pollen grain in Hyacinthus orientalis L. Acta 1986. The legumin gene family: structure of a B type Soc. Bot. Polon. 50:367-380. gene of Vicia faba and a possible legumin gene specific regulatory element. Nucl. Acids Res. Bednarska, E. 1984. Ultrastructural and metabolic transformations of differentiating Hyacinthus orientalis L. 14:2707-2720. pollen grain cells. I. RNA and protein synthesis. Acta Baus, A.D., Franzmann, L., and Meinke, D.W. 1986. Soc. Bot. Polon. 53:145-158. Growth in vitro of arrested embryos from lethal mutants of Arabidopsis thaliana. Theor. Appl. Genet. Bednarska, E. 1989a. Localization of calcium on the 72:577-586. stigma surface of Ruscus aculeatus L. Studies using chlorotetracycline and X-ray microanalysis. Planta Bawa, K.S., and Beach, J.H. 1983. Self-incompatibility systems in the Rubiaceae of a tropical lowland wet for179:11-16. est. Am.}. Bot. 70:1281-1288. Bednarska, E. 1989b. The effect of exogenous Ca2+ ions on pollen grain germination and pollen tube growth. Bayliss, M.W., and Dunn, S.D.M. 1979. Factors affecting Investigations with 45Ca2+ together with verapamil, callus formation from embryos of barley (Hordeum La3+, and ruthenium red. Sex. Plant Reprod. 2:53-58. vulgare). Plant Sci. Lett. 14:311-316. Beachy, R.N. 1980. In vitro synthesis of the a and a' sub- Bednarska, E. 1991. Calcium uptake from the stigma by units of the 7S storage proteins (conglycinins) of soygerminating pollen in Primula officinalis L. and Ruscus aculeatus L. Sex. Plant Reprod. 4:36-38. bean seeds. Plant Physiol. 65:990-994. Beachy, R.N., Jarvis, N.P., and Barton, K.A. 1981. Bednarska, E. 1993. Localization of calcium-dependent Biosynthesis of subunits of the soybean 7S storage ATPase in germinating pollen grain and pollen tube protein. /. Mol. Appl. Genet. 1:19-27. in Vicia faba. Folia Histochem. Cytobiol. 31:147-151. Beachy, R.N., Thompson. J.F., and Madison, J.T. 1978. Bednarska, E. 1995. Localization of membrane-associated calcium in unpollinated and pollinated pistil of Isolation of polyribosomes and messenger RNA Petunia hybrida Hort. Acta Soc. Bot. Polon. 64:19-24. active in in vitro synthesis of soybean seed proteins. Bednarska, E., and Butowt, R. 1994. Calcium in pollenPlant Physiol. 61:139-144. pistil interaction in Petunia hybrida Hort. I. Beachy, R.N., Chen, Z.-L., Horsch, R.B., Rogers, S.G., Localization of Ca2+ ions in mature pollen grain Hoffmann, N.J., and Fraley, R.T. 1985. Accumulation using pyroantimonate and autoradiographic methand assembly of soybean f}-conglycinin in seeds of ods. Folia Histochem. Cytobiol. 32:265-269. transformed Petunia plants. EMBO /. 4:3047-3053. Beadle, G.W. 1932. Genes in maize for pollen sterility. Bednarska, E., and Butowt, R. 1995a. Calcium in Genetics 17:413-431. pollen-pistil interaction in Petunia hybrida Hort. II. Localization of Ca2+ ions and Ca2+-ATPase in unpolBechtel, D.B., and Barnett, B.D. 1986. A freeze-fracture linated pistil. Folia Histochem. Cytobiol. 33:43-52. study of storage protein accumulation in unfixed wheat starchy endosperm. Cereal Chem. 63:232-240. Bednarska, E. and Butowt, R. 1995b. Calcium in pollen-pistil interaction in Petunia hybrida Hort. III. Bechtel, D.B., Gaines, R.L., and Pomeranz, Y. 1982. Early Localization of Ca2+ ions and Ca2+-ATPase in pollistages in wheat endosperm formation and protein nated pistil. Folia Histochem. Cytobiol. 33:125-132. body initiation. Ann. Bot. 50:507-518. Bechtel, D.B., and Juliano, B.0.1980. Formation of protein Bednarska, E., and Karbowska, J. 1990. Localization of bodies in the starchy endosperm of rice (Oryza sativa Ca2+ and Ca-ATPase in papillae of maturing stigma L.): a re-investigation. Ann. Bot. 45:503-509. in Pharbitis nil L. Phytomorphology 40:323-329. Bechtel, D.B., and Pomeranz, Y. 1977. Ultrastructure of the Bednarska, E., and Tretyn, A. 1989. Ultrastructural localmature ungerminated rice (Oryza sativa) caryopsis. ization of acetylcholinesterase activity in the stigma The caryopsis coat and the aleurone cells. Am. J. Bot. of Pharbitis nil. Cell Biol. Int. Rep. 13:275-281. Beerhues, L., Forkmann, G., Schopker, H., Stotz, G., and 64:966-973. Wiermann, R. 1989. Flavanone 3-hydroxylase and Bechtel, D.B., and Pomeranz, Y. 1981. Ultrastructure and dihydroflavonol oxygenase activities in anthers of cytochemistry of mature oat (Avena sativa L.) Tulipa. The significance of the tapetum fraction in endosperm. The aleurone layer and starchy flavonoid metabolism. /. Plant Physiol. 133:743-746. endosperm. Cereal Chem. 58:61-69. Becker, D., Brettschneider, R., and Lorz, H. 1994. Fertile Beevers, L., and Poulson, R. 1972. Protein synthesis in transgenic wheat from microprojectile bombardcotyledons of Pisum sativum L. I. Changes in cell-free ment of scutellar tissue. Plant ]. 5:299-307. amino acid incorporation capacity during seed development and maturation. Plant Physiol. Bedinger, P.A., and Edgerton, M.D. 1990. Developmental staging of maize microspores reveals a transition in 49:476-481. developing microspore proteins. Plant Physiol. Belanger, EC, and Kriz, A.L. 1989. Molecular characteri92:474-479. zation of the major maize embryo globulin encoded Bednara, J. 1977. Female gametophyte and pollen tube of by the Glbl gene. Plant Physiol. 91:636-643. Epilobium palustre L. Acta Soc. Bot. Polon. 46:603-615. Belanger, EC, and Kriz, A.L. 1991. Molecular basis for Bednara, J., van Lammeren, A.A.M., and Willemse, M.T.M. allelic polymorphism of the maize globulin-1 gene. 1988. Microtubular configurations during meiosis and Genetics 129:863-872. megasporogenesis in Gasteria verrucosa and Bell, J., and Hicks, G. 1976. Transmitting tissue of the Chamaenerion angustifolium. Sex. Plant Reprod. pistil of tobacco: light and electron microscopic 1:164-172. observations. Planta 131:187-200. Bednara, J., Willemse, M.T.M., and van Lammeren, Belliard, G., Vedel, F., and Pelletier, G. 1979.
540
References
Mitochondrial recombination in cytoplasmic sequence associated with self-incompatibility in hybrids of Nicotiana tabacum by protoplast fusion. Nicotiana alata has homology with mitochondrial Nature 281:401-403. DNA. Theor. Appl. Genet. 77:320-324. Benfey, P.N., and Chua, N.-H. 1989. Regulated genes in Bewley, J.D., and Black, M. 1994. Seeds. Physiology of transgenic plants. Science 244:174-181. Development and Germination, 2nd ed. New York: Plenum Press. Benfey, P.N., Ren, L., and Chua, N.-H. 1989. The CaMV 35S enhancer contains at least two domains which Bhadula, S.K., and Sawhney, V.K. 1987. Esterase activity can confer different developmental and tissue-speand isozymes during the ontogeny of stamens of cific expression patterns. EMBO J. 8:2195-2202. male fertile Lycopersicon esculentum Mill., a male sterBenfey, P.N., Ren, L., and Chua, N.-H. 1990. Tissue-speile stamenless-2 mutant and the low temperaturecific expression from CaMV 35S enhancer subdoreverted mutant. Plant Sci. 52:187-194. mains in early stages of plant development. EMBO J. Bhadula, S.K., and Sawhney, V.K. 1989a. Protein analysis 9:1677-1684. of floral organs of some members of Solanaceae. Bot. Mag. Tokyo 102:85-91. Benfey, P.N., Linstead, P.J., Roberts, K., Schiefelbein, J.W., Hauer, M.-T, and Aeschbacher, R.A. 1993. Root Bhadula, S.K., and Sawhney, V.K. 1989b. Amylolytic activdevelopment in Arabidopsis: four mutants with draity and carbohydrate levels during the stamen matically altered root morphogenesis. Development ontogeny of a male fertile, and a "gibberellin-sensi119:57-70. tive' male sterile mutant of tomato (Lycopersicon esculentum). ]. Expt. Bot. 40:789-794. Benito Moreno, R.M., Macke, F., Alwen, A., and HeberleBors, E. 1988. In-situ seed production after pollina- Bhadula, S.K., and Sawhney, V.K. 1991. Protein analysis tion with m-w'tro-matured, isolated pollen. Planta during the ontogeny of normal and male sterile sta176:145-148. menless-2 mutant stamens of tomato (Lycopersicon esculentum Mill.). Biochem. Genet. 29:29-41. Bennett, M.D., and Hughes, W.G. 1972. Additional mitosis in wheat pollen induced by ethrel. Nature Bhalla, P.L., Singh, M.B., and Malik, C.P. 1980a. Dark fixa240:566-568. tion of CO2 by embryo-suspensors of Nasturtium (Tropaeolum majus). Biochem. Physiol. Pflanz. Bennett, M.D., and Smith, J.B. 1976. The nuclear DNA 175:263-267. content of the egg, the zygote and young proembryo cells in Hordeum. Caryologia 29:435-446. Bhalla, PL., Singh, M.B., and Malik, C.P. 1980b. Changes in level of major biochemical constituents and some Bennett, M.D., Smith, J.B., and Barclay, I. 1975. Early seed enzymes of carbohydrate metabolism during development in the Triticeae. Phil. Trans. Roy. Soc. embryogeny in okra Abelmoschus esculentus (L.) Land. 272B:199-227. Moench. Indian ]. Expt. Biol. 18:686-689. Bennett, M.D., and Stern, H. 1975. The time and duration of female meiosis in Lilium. Proc. Roy. Soc. Lond. Bhalla, P.L., Singh, M.B., and Malik, C.P. 1981a. Studies on the comparative biosynthetic activities of 188B:459-475. embryo and suspensor in Tropaeolum majus L. Z. Bennett, M.D., Finch, R.A., Smith, J.B., and Rao, M X Pflanzenphysiol. 103:115-119. 1973a. The time and duration of female meiosis in wheat, rye and barley. Proc. Roy. Soc. Lond. Bhalla, PL., Singh, M.B., and Malik, C.P. 1981b. Characterization of pentose phosphate pathway in 183B:301-319. embryo suspensor of Tropaeolum majus. Biochem. Bennett, M.D., Rao, M.K., Smith, J.B., and Bayliss, M.W. Physiol. Pflanz. 176:789-792. 1973b. Cell development in the anther, the ovule, and the young seed of Triticum aestivum L. var. Bhandari, N.N. 1984. The microsporangium. In Embryology of Angiosperms, ed. B.M. Johri, pp. Chinese Spring. Phil. Trans. Roy. Soc. Lond. 266B:39-81. 53-121. Berlin: Springer-Verlag. Bennici, A., and Cionini, P.G. 1979. Cytokinins and in vitro Bhandari, N.N., and Bhargava, M. 1983. Wall ingrowths development of Phaseolus coccineus embryos. Planta in the antipodal cells in Papaver somniferum L. Beitr. 147:27-29. Biol. Pflanz. 58:53-58. Berger, C, and Erdelska", O. 1973. Ultrastructural aspects Bhandari, N.N., Bhargava, M., and Chitralekha, P. 1986. Cellularization of free-nuclear endosperm of Papaver of the embryo sac of Jasione montana L.: cell walls. somniferum L. Phytomorphology 36:357-366. Caryologia 25: Suppl. 109-120. Bergfeld, R., Kuhnl, T., and Schopfer, P. 1980. Formation Bhandari, N.N., Bhargava, M., and Geier, T. 1981. A persisting cellulosic wall of microspore mother cells of protein storage bodies during embryogenesis in during microsporogenesis in Allium tuberosum Rottl. cotyledons of Sinapis alba L. Planta 148:146-156. and Cyclamen persicum Mill. Ann. Bot. 48:425^131. Bergfeld, R., Hong, Y.-N., Kuhnl, T, and Schopfer, P. 1978. Formation of oleosomes (storage lipid bodies) dur- Bhandari, N.N., Kishori, R., and Natesh, S. 1976. Ontogeny, cytology, and histochemistry of anther ing embryogenesis and their breakdown during tapetum in relation to pollen development in Nigella seedling development in cotyledons of Sinapis alba L. damascena. Phytomorphology 26:46-59. Planta 143:297-307. Berleth, T., and Jurgens, G. 1993. The role of the Bhandari, N.N., and Sachdeva, A. 1983. Some aspects of organization and histochemistry of the embryo sac monopteros gene in organising the basal body region of Scilla sibirica Sato. Protoplasma 116:170-178. of the Arabidopsis embryo. Development 118:575-587. Bernard, S. 1971. Developpement d'embryons haploides a Bhandari, N.N., Soman, P., and Bhargava, M. 1980. Histochemical studies on the female gametophyte of partir d'antheres cultivees in vitro. Etude Argemone mexicana L. Cytologia 45:281-291. cytologique comparee chez le tabac et le Petunia. Rev. Bhaskaran, S., and Swaminathan, M.S. 1959. Stage of Cytol. Biol. Veget. 34:165-188. deoxyribonucleic acid synthesis during mitosis and Bernatzky, R., Anderson, M.A., and Clarke, A.E. 1988. meiosis. Curr. Sci. 28:335-336. Molecular genetics of self-incompatibility in flowerBhatnagar, S.P., and Kallarackal, J. 1980. Cytochemical ing plants. Devel Genet. 9:1-12. studies on the endosperm of Linaria bipartita (Vent.) Bernatzky, R., Mau, S.-L., and Clarke, A.E. 1989. A nuclear
541
References
Willd. with a note on the role of endosperm haustoria. Cytologia 45:247-256. Bhatnagar, S.P., and Sawhney, V. 1981. Endosperm - its morphology, ultrastructure, and histochemistry. Int. Rev. Cytol. 73:55-102. Bhattacharyya, M.K., Paiva, N.L., Dixon, R.A., Korth, K.L., and Stermer, B.A. 1995. Features of the hmg 1 subfamily of genes encoding HMG-CoA reductase in potato. Plant Mol. Biol. 28:1-15. Bhattacharyya, M.K., Smith, A.M., Ellis, T.H.N., Hedley, C, and Martin, C. 1990. The wrinkled-seed character of pea described by Mendel is caused by a transposon-like insertion in a gene encoding starch-branching enzyme. Cell 60:115-122. Bhave, M.R., Lawrence, S., Barton, C, and Hannah, L.C. 1990. Identification and molecular characterization of shrunken-2 cDNA clones of maize. Plant Cell 2:581-588. Bhojwani, S.S., and Cocking, E.C. 1972. Isolation of protoplasts from pollen tetrads. Nature New Biol. 239:29-30. Bhojwani, S.S., Dunwell, J.M., and Sunderland, N. 1973. Nucleic-acid and protein contents of embryogenic tobacco pollen. /. Expt. Bot. 24:863-871. Biddington, N.L., Sutherland, R.A., and Robinson, H.T. 1988. Silver nitrate increases embryo production in anther culture of Brussels sprouts. Ann. Bot. 62:181-185. Biddle, J.A. 1979. Anther and pollen development in garden pea and cultivated lentil. Can. J. Bot. 57:1883-1900. Bilodeau, P., Lafontaine, J.-G., and Bellemare, G. 1994. Far upstream activating promoter regions are responsible for expression of the BnCl cruciferin gene from
with desiccation tolerance in soybean. Plant Physiol. 96:868-874. Blechl, A.E., and Anderson, O.D. 1996. Expression of a novel high-molecular-weight glutenin subunit gene in transgenic wheat. Nature Biotech. 14:875-879. Blechl, A.E., Lorens, G.F., Greene, F.C., Mackey, B.E., and Anderson, O.D. 1994. A transient assay for promoter activity of wheat seed storage protein genes and other genes expressed in developing endosperm. Plant Sci. 102:69-80. Blundy, K.S., Blundy, M.A.C., and Crouch, M.L. 1991. Differential expression of members of the napin storage protein gene family during embryogenesis in Brassica napus. Plant Mol. Biol. 17:1099-1104.
Bobb, A.J., Eiben, H.G., and Bustos, M.M. 1995. PvAlf, an embryo-specific acidic transcriptional activator enhances gene expression from phaseolin and phytohemagglutinin promoters. Plant J. 8:331-343. Boblenz, K., Nothnagel, T., and Metzlaff, M. 1990. Paternal inheritance of plastids in the genus Daucus. Mol. Gen. Genet. 220:489-491.
Boeshore, M.L., Hanson, M.R., and Izhar, S. 1985. A variant mitochondrial DNA arrangement specific to Petunia stable sterile somatic hybrids. Plant Mol. Biol. 4:125-132. Boeshore, M.L., Lifshitz, I., Hanson, M.R., and Izhar, S. 1983. Novel composition of mitochondrial genomes in Petunia somatic hybrids derived from cytoplasmic male sterile and fertile plants. Mol. Gen. Genet. 190:459^67. Bogdanov, Y.F., Strokov, A.A., and Reznickova, S.A. 1973. Histone synthesis during meiotic prophase in Lilium. Chromosoma 43:237-245. Bogue, M.A., Vonder Haar, R.A., Nuccio, M.L., Griffing, Brassica napus. Plant Cell Rep. 14:125-130. L.R., and Thomas, T.L. 1990. Developmentally regulated expression of a sunflower IIS seed protein Bimal, R., Schel, J.H.N., Tavoletti, S., and Willemse, gene in transgenic tobacco. Mol. Gen. Genet. M.T.M. 1995. Localization of poly(A)+-containing 222:49-57. RNA during female gametophyte development in Medkago sativa and the diploid mutant Medicago Bohdanowicz, J. 1973. Karyological anatomy of the sussativa ssp. falcata using digoxigenin-labelled oligopensor in Alisma L. I. Alisma plantago-aquatica L. Acta dT probes. Sex. Plant Reprod. 8:18-26. Biol. Cracow. Ser. Bot. 16:235-246. Binarova, P., Straatman, K., Hause, B., Hause, G., and van Bohdanowicz, J. 1987. Alisma embryogenesis: the development and ultrastructure of the suspensor. Lammeren, A.A.M. 1993. Nuclear DNA synthesis Protoplasma 137:71-83. during the induction of embryogenesis in cultured microspores and pollen of Brassica napus L. Theor. Bohdanowicz, J., Ciampolini, F., and Cresti, M. 1995. Appl. Genet. 87:9-16. Striped projections of the outer membrane of the Bino, R.J. 1985a. Ultrastructural aspects of cytoplasmic generative cell in Convallaria majalis pollen. Sex. male sterility in Petunia hybrida. Protoplasma Plant. Reprod. 8:223-227. 127:230-240. Bohdanowicz, J., and Turala-Szybowska, K. 1985. Bino, R.J. 1985b. Histological aspects of microsporogeneUltrastructure of endopolyploid antipodals in sis in fertile, cytoplasmic male sterile and restored Aconitum vulparia Rchb. I. Antipodals in the mature fertile Petunia hybrida. Theor. Appl. Genet. 69:423-428. embryo sac. Protoplasma 127:163-170. Bino, R.J., van Tuyl, J.M., and de Vries, J.N. 1990. Flow cyto- Bohdanowicz, J., and Turala-Szybowska, K. 1987. metric determination of relative nuclear DNA contents Ultrastructure of endopolyploid antipodals in in bicellulate and tricellulate pollen. Ann. Bot. 65:3-8. Aconitum vulparia Rchb. II. Antipodals in the period of free nuclear endosperm. Protoplasma 140:13-21. Bird, J., Porter, E.K., and Dickinson, H.G. 1983. Events in the cytoplasm during male meiosis in Lilium. ]. Cell Bohn, G.W., and Principe, J.A. 1964. A second male-sterilSci. 59:27-42. ity gene in the muskmelon. /. Hered. 55:211-215. Blackbourn, H.D., and Jackson, A.P. 1996. Plant clathrin Bollini, R., and Chrispeels, M.J. 1978. Characterization heavy chain: sequence analysis and restricted localiand subcellular localization of vicilin and phytosation in growing pollen tubes. /. Cell Sci. hemagglutinin, the two major reserve proteins of 109:777-787. Phaseolus vulgaris L. Planta 142:291-298. Blackman, S.A., Obendorf, R.L., and Leopold, A.C. 1992. Bollini, R., and Chrispeels, M.J. 1979. The rough Maturation proteins and sugars in desiccation tolerendoplasmic reticulum is the site of reserve-protein ance of developing soybean seeds. Plant Physiol. synthesis in developing Phaseolus vulgaris cotyle100:225-230. dons. Planta 146:487-501. Blackman, S.A., Wettlaufer, S.H., Obendorf, R.L., and Bollini, R., van der Wilden, W., and Chrispeels, M.J. 1982. Leopold, A.C. 1991. Maturation proteins associated A precursor of the reserve-protein, phaseolin, is tran-
542
References
siently associated with the endoplasmic reticulum of developing Phaseolus vulgaris cotyledons. Physiol. Plant. 55:82-92. Bollini, R., Vitale, A., and Chrispeels, M.J. 1983. In vivo and in vitro processing of seed reserve protein in the endoplasmic reticulum: evidence for two glycosylation steps. /. Cell Biol. 96:999-1007. Bolz, G. 1961. Genetisch-zuchterische Untersuchungen bei Tagetes. II. Herstellung, Genetik und Verwendung rontgeninduzierter Mutanten in der Zuchtung. Z. PflanzenzUchtg 45:121-142. Bommineni, V.R., and Greyson, R.I. 1987. In vitro culture of ear shoots of Zea mays and the effect of kinetin on sex expression. Am. ]. Bot. 74:883-890. Bommineni, V.RV and Greyson, R.I. 1990. Regulation of flower development in cultured ears of maize (Zea mays L.). Sex. Plant Reprod. 3:109-115.
Bommineni, V.R., Atkinson, B.G., Greyson, R.I., and Walden, D.B. 1990. Polypeptides synthesized during the maturation of flower organs from tassel and ear inflorescences of Zea mays L. Maydica 35:195-201. Bommineni, V.R., Chibbar, R.N., Datla, R.S.S., and Tsang, E.W.T. 1993. Transformation of white spruce (Picea glauca) somatic embryos by microprojectile bombardment. Plant Cell Rep. 13:17-23. Bonhomme, S., Budar, R, Fdrault, M., and Pelletier, G. 1991. A 2.5 kb Ncol fragment of Ogura radish mitochondrial DNA is correlated with cytoplasmic male-sterility in Brassica cybrids. Curr. Genet. 19:121-127. Bonhomme, S., Budar, R, Lancelin, D., Small, I., Defrance, M.-C, and Pelletier, G. 1992. Sequence and transcript analysis of the Nco2.5 Ogura-specific fragment correlated with cytoplasmic male sterility in Brassica cybrids. Mol. Gen. Genet. 235:340-348. Boothe, J.G., and Walden, D.B. 1990. Gene expression in embryos and seedlings of maize. Maydica 35:187-194. Booy, G., Krens, F.A., and Bino, R.J. 1992. Analysis of pollen-tube growth in cultured maize silks. Sex. Plant Reprod. 5:227-231.
Booy, G., Krens, F.A., and Huizing, H.J. 1989. Attempted pollen-mediated transformation of maize. /. Plant Physiol. 135:319-324. Bopp-Hassenkamp, G. 1960. Elektronenmikroskopische Untersuchungen an Pollenschlauchen zweier Liliaceen. Z. Naturfors. 15b:91-94. Borisjuk, L., Weber, H., Panitz, R., Manteuffel, R., and Wobus, U. 1995. Embryogenesis of Viciafaba L.: histodifferentiation in relation to starch and storage protein synthesis. /. Plant Physiol. 147:203-218. Borkird, C., Choi, J.H., and Sung, Z.R. 1986. Effect of 2,4dichlorophenoxyacetic acid on the expression of embryogenic program in carrot. Plant Physiol. 81:1143-1146. Borkird, C, Choi, J.H., Jin, Z.-H., Franz, G., Hatzopoulos, P., Chorneau, R., Bonas, U., Pelegri, R, and Sung, Z.R. 1988. Developmental regulation of embryonic genes in plants. Proc. Natl Acad. Sci. USA 85:6399-6403. Boronat, A., Martinez, M.C., Reina, M, Puigdomenech, P., and Palau, J. 1986. Isolation and sequencing of a 28 kD glutelin-2 gene from maize. Common elements in the 5' flanking regions among zein and glutelin genes. Plant Sci. 47:95-102. Borroto, K., and Dure, L., III. 1987. The globulin seed storage proteins of flowering plants are derived from two ancestral genes. Plant Mol. Biol. 8:113-131. Borroto, K.E., and Dure, L., III. 1986. The expression of
chloroplast genes during cotton embryogenesis. Plant Mol. Biol. 7:105-113.
Borthwick, H.A. 1931. Development of the macrogametophyte and embryo of Daucus carota. Bot. Gaz. 92:23^4. Bosio, M.G. 1940. Ricerche sulla fecondazione intraovarica in Helleborus e Poeonia. Nuovo Giorn. Bot. Hal.
47:591-598. Bosnes, M., and Olsen, O.-A. 1992. The rate of nuclear gene transcription in barley endosperm syncytia increases sixfold before cell-wall formation. Planta 186:376-383. Bosnes, M., Weideman, R, and Olsen, O.-A. 1992. Endosperm differentiation in barley wild-type and sex mutants. Plant J. 2:661-674. Bosnes, M., Harris, E., Aigeltinger, L., and Olsen, O.-A. 1987. Morphology and ultrastructure of 11 barley shrunken endosperm mutants. Theor. Appl. Genet. 74:177-187. Boston, R.S., Becwar, M.R., Ryan, R.D., Goldsbrough, P.B., Larkins, B.A., and Hodges, T.K. 1987. Expression from heterologous promoters in electroporated carrot protoplasts. Plant Physiol. 83:742-746. Boston, R.S., Fontes, E.B.P., Shank, B.B., and Wrobel, R.L. 1991. Increased expression of the maize immunoglobulin binding protein homolog b-70 in three zein regulatory mutants. Plant Cell 3:497-505. Botti, C, and Vasil, I.K. 1983. Plant regeneration by somatic embryogenesis from parts of cultured mature embryos of Pennisetum americanum (L.) K. Schum. Z. Pflanzenphysiol. 111:319-325. Bouchard, R.A. 1990. Characterization of expressed meiotic prophase repeat transcript clones of Lilium: meiosis-specific expression, relatedness, and affinities to small heat shock protein genes. Genome 33:68-79. Bouchard, R.A., and Stern, H. 1980. DNA synthesized at pachytene in Lilium: a non-divergent subclass of moderately repetitive sequences. Chromosoma 81:349-363. Bouharmont, J. 1977. Cytology of microspores and calli after anther culture in Hordeum vulgare. Caryologia 30:351-360. Boulter, D., Evans, I.M., Ellis, J.R., Shirsat, A., Gatehouse, J.A., and Croy, R.R.D. 1987. Differential gene expression in the development of Pisum sativum. Plant Physiol. Biochem. 25:283-289.
Bouthyette, P.-Y., Spitsberg, V, and Gregory, P. 1985. Mitochondrial interaction with Helminthosporium maydis race T toxin: blocking by dicyclohexylcarbodiimide. /. Expt. Bot. 36:511-528. Boutilier, K.A., Gines, M.-J., DeMoor, J.M., Huang, B., Baszczynski, C.L., Iyer, V.N., and Miki, B.L. 1994. Expression of the BnmNAP subfamily of napin genes coincides with the induction of Brassica microspore embryogenesis. Plant Mol. Biol. 26:1711-1723. Boutry, M., and Briquet, M. 1982. Mitochondrial modifications associated with the cytoplasmic male sterility in faba beans. Eur. ]. Biochem. 127:129-135. Bower, R., and Birch, R.G. 1992. Transgenic sugarcane plants via microprojectile bombardment. Plant J. 2:409^16. Bowman, J.L., Drews, G.N., and Meyerowitz, E.M. 1991. Expression of the Arabidopsis floral homeotic gene AGAMOUS is restricted to specific cell types late in flower development. Plant Cell 3:749-758. Bowman, J.L., Smyth, D.R., and Meyerowitz, E.M. 1989.
References
543
Rasmussen, S.K. 1985. Primary structure of a Bl Genes directing flower development in Arabidopsis. hordein gene from barley. Carlsberg Res. Commun. Plant Cell 1:37-52. 50:333-345. Bowman, J.L., Smyth, D.R., and Meyerowitz, E.M. 1991. Genetic interactions among floral homeotic genes of Braun, C.J., Siedow, J.N., and Levings, C.S., III. 1990. Arabidopsis. Development 112:1-20. Fungal toxins bind to the URF13 protein in maize mitochondria and Escherichia coli. Plant Cell Bowman, J.L., Alvarez, J., Weigel, D., Meyerowitz, E.M., 2:153-161. and Smyth, D.R. 1993. Control of flower development in Arabidopsis thaliana by APETALA1 and inter-Braun, C.J., Siedow, J.N., Williams, M.E., and Levings, acting genes. Development 119:721-743. C.S., III. 1989. Mutations in the maize mitochondrial T-urfl3 gene eliminate sensitivity to a fungal pathoBowman, J.L., Sakai, H., Jack, T., Weigel, D., Mayer, U., toxin. Proc. Natl Acad. Sci. USA 86:4435-4439. and Meyerowitz, E.M. 1992. SUPERMAN, a regulator of floral homeotic genes in Arabidopsis. Brawley, S.H., Wetherell, D.F., and Robinson, K.R. 1984. Electrical polarity in embryos of wild carrot preDevelopment 114:599-615. cedes cotyledon differentiation. Proc. Natl Acad. Sci. Bown, D., Ellis, T.H.N., and Gatehouse, J.A. 1988. The USA 81:6064-6067. sequence of a gene encoding convicilin from pea {Pisum sativum L.) shows that convicilin differs from Bray, E.A., and Beachy, R.N. 1985. Regulation by ABA of vicilin by an insertion near the N-terminus. Biochem. P-conglycinin expression in cultured developing ]. 251:717-726. soybean cotyledons. Plant Physiol. 79:746-750. Bown, D., Levasseur, M., Croy, R.R.D., Boulter, D., and Bray, E.A., Naito, S., Pan, N.-S., Anderson, E., Dub£, P., and Beachy, R.N. 1987. Expression of the P-subunit Gatehouse, J.A. 1985. Sequence of a pseudogene in of p-conglycinin in seeds of transgenic plants. Planta the legumin gene family of pea (Pisum sativum L.). Nucl. Acids Res. 13:4527-4538. 172:364-370. Boyer, C, Hilbert, J.-L., and Vasseur, J. 1993. Bredemeijer, G.M.M. 1974. Peroxidase activity and peroxEmbryogenesis-related protein synthesis and accuidase isoenzyme composition in self-pollinated, mulation during early acquisition of somatic cross-pollinated and unpollinated styles of Nicotiana alata. Ada Bot. Need. 23:149-157. embryogenesis competence in Cichorium. Plant Sci. 93:41-53. Bredemeijer, G.M.M., and Blaas, J. 1975. A possible role of a stylar peroxidase gradient in the rejection of Boyes, D.C., and Nasrallah, J.B. 1993. Physical linkage of incompatible growing pollen tubes. Ada Bot. Neerl. the SLG and SRK genes at the self-incompatibility 24:37-48. locus of Brassica oleracea. Mol. Gen. Genet. 236:369-373. Bredemeijer, G.M.M., and Blaas, J. 1980. Do S allele-specific peroxidase isoenzymes exist in self-incompatiBoyes, D.C., and Nasrallah, J.B. 1995. An anther-specific ble Nicotiana alata? Theor. Appl. Genet. 57:119-123. gene encoded by an S locus haplotype of Brassica produces complementary and differentially regu- Bredemeijer, G.M.M., and Blaas, J. 1981. S-specific prolated transcripts. Plant Cell 7:1283-1294. teins in styles of self-incompatible Nicotiana alata. Theor. Appl. Genet. 59:185-190. Boyes, D.C., Chen, C.-H., Tantikanjana, T., Esch, J.J., and Nasrallah, J.B. 1991. Isolation of a second S-locus- Breen, J.P., and Crouch, M.L. 1992. Molecular analysis of a cruciferin storage protein gene family of Brassica related cDNA from Brassica oleracea: genetic relationnapus. Plant Mol. Biol. 19:1049-1055. ships between the S locus and two related loci. Genetics 127:221-228. Breiteneder, H., Pettenburger, K., Bito, A., Valenta, R., Kraft, D., Rumpold, H., Scheiner, O., and Boyle, S.A., and Yeung, E.C. 1983. Embryogeny of Breitenbach, M. 1989. The gene coding for the major Phaseolus: developmental pattern of lactate and alcobirch pollen allergen Betvl, is highly homologous to hol dehydrogenases. Phytochemistry 22:2413-2416. a pea disease resistance response gene. EMBO ]. Brace, J., Ockendon, D.J., and King, GJ. 1993. 8:1935-1938. Development of a method for the identification of S alleles in Brassica oleracea based on digestion of PCR- Breslavetz, L.P. 1935. Abnormal development of pollen in different races and grafts of hemp. Genetica amplified DNA with restriction endonucleases. Sex. 17:154-169. Plant Reprod. 6:133-138. Bradley, D., Carpenter, R., Sommer, H., Hartley, N., and Breton, A.M., and Sung, Z.R. 1982. Temperature-sensitive Coen, E. 1993. Complementary floral homeotic phecarrot variants impaired in somatic embryogenesis. notypes result from opposite orientations of a transDevel Biol. 90:58-66. poson at the plena locus of Antirrhinum. Cell Brettell, R.I.S., Thomas, E., and Ingram, D.S. 1980. 72:85-95. Reversion of Texas male-sterile cytoplasm maize in culture to give fertile, T-toxin resistant plants. Theor. Brady, T. 1973. Feulgen cytophotometric determination of Appl. Genet. 58:55-58. the DNA content of the embryo proper and suspensor cells of Phaseolus coccineus. Cell Diffn 2:65-75. Brewbaker, J.L. 1957. Pollen cytology and self-incompatibility systems in plants. /. Hered. 48:271-277. Brady, T., and Walthall, E.D. 1985. The effect of the suspensor and gibberellic acid on Phaseolus vulgaris Brewbaker, J.L. 1959. Biology of the angiosperm pollen grain. Indian }. Genet. Plant Breed. 19:121-133. embryo protein content. Devel Biol. 107:531-536. Brander, K.A., and Kuhlemeier, C. 1995. A pollen-specific Brewbaker, J.L. 1967. The distribution and phylogenetic DEAD-box protein related to translation initiation significance of binucleate and trinucleate pollen factor eIF-4A from tobacco. Plant Mol. Biol. grains in the angiosperms. Am. J. Bot. 54:1069-1083. Brewbaker, J.L., and Emery, G.C. 1962. Pollen radiob27:637-649. otany. Radiation Bot. 1:101-154. Brandt, A., and Ingversen, J. 1976. In vitro synthesis of barley endosperm proteins on wild type and mutant Brewbaker, J.L., and Gorrez, D.D. 1967. Genetics of selftemplates. Carlsberg Res. Commun. 41:311-320. incompatibility in the monocot genera, Ananas Brandt, A., Montembault, A., Cameron-Mills, V., and (pineapple) and Gasteria. Am. ]. Bot. 54:611-616.
544 Brewbaker, J.L., and Kwack, B.H. 1963. The essential role of calcium ion in pollen germination and pollen tube growth. Am. ]. Bot. 50:859-865. Brewbaker, J.L., and Majumder, S.K. 1961. Cultural studies of the pollen population effect and the selfincompatibility inhibition. Am. ]. Bot. 48:457-464. Brewbaker, J.L., and Natarajan, A.T. 1960. Centric fragments and pollen-part mutation of incompatibility alleles in Petunia. Genetics 45:699-704. Briarty, L.G. 1980. Stereological analysis of cotyledon cell development in Phaseolus. II. The developing cotyledon. /. Expt. Bot. 31:1387-1398. Briarty, L.G., Hughes, C.E., and Evers, A.D. 1979. The developing endosperm of wheat - a stereological analysis. Ann. Bot. 44:641-658. Briggs, C.L. 1992. A light and electron microscope study of the mature central cell and egg apparatus of Solatium nigrum L. (Solanaceae). Int. ]. Plant Sci. 153:40-48. Briggs C.L. 1995. The initiation, development and removal of embryo sac wall ingrowths in the developing seeds of Solanum nigrum L. An ultrastructural study. Ann. Bot. 76:429^39. Brighigna, L., Fiordi, A.C., and Palandri, M.R. 1981. Ultrastructural investigations on the two-nucleate pollen grain of Tillandsia caput-medusae Morr. (Bromeliaceae). Am. J. Bot. 68:1033-1041. Brighigna, L., and Papini, A. 1993. The ultrastructure of the tapetum of Tillandsia albida Mez et Purpus. Phytomorphology 43:261-274. Bright, S.W.J., Wood, E.A., and Miflin, B.J. 1978. The effect of aspartate-derived amino acids (lysine, threonine, methionine) on the growth of excised embryos of wheat and barley. Planta 139:113-117. Brinegar, A.C., Stevens, A., and Fox, J.E. 1985. Biosynthesis and degradation of a wheat embryo cytokinin-binding protein during embryogenesis and germination. Plant Physiol. 79:706-710. Brink, R.A., and Cooper, D.C. 1941. Incomplete seed failure as a result of somatoplastic sterility. Genetics 26:487-505. Brink, R.A., and Cooper, D.C. 1947a. Effect of the DE|7 allele on development of the maize caryopsis. Genetics 32:350-368. Brink, R.A., and Cooper, D.C. 1947b. The endosperm in seed development. Bot. Rev. 13:423-541. Brink, R.A., and MacGillivray, J.H. 1924. Segregation for the waxy character in maize pollen and differential development of the male gametophyte. Am. ]. Bot. 11:465-469. Brock, R.D. 1954. Fertility in Lilium hybrids. Heredity 8:409-420. Brooking, I.R. 1979. Male sterility in Sorghum bicolor (L.) Moench induced by low night temperature. II. Genotypic differences in sensitivity. Aust. J. Plant Physiol. 6:143-147. Brooks, ]., and Shaw, G. 1978. Sporopollenin: a review of its chemistry, palaeochemistry and geochemistry. Grana 17:91-97. Brooks, M.H. 1962. Comparative analyses of some of the free amino acids in anthers of fertile and genetic cytoplasmic male sterile Sorghum. Genetics 47:1629-1638. Brooks, M.H., Brooks, J.S., and Chien, L. 1966. The anther tapetum in cytoplasmic- genetic male sterile Sorghum. Am. ]. Bot. 53:902-908. Broothaerts, W., Janssens, G.A., Proost, P., and Broekaert,
References
W.F. 1995. cDNA cloning and molecular analysis of two self-incompatibility alleles from apple. Plant Mol. Biol. 27:499-511. Broothaerts, W., Vanvinckenroye, P., Decock, B., van Damme, J., and Vendrig, J. C. 1991. Petunia hybrida Sproteins: ribonuclease activity and the role of their glycan side chains in self-incompatibility. Sex. Plant Reprod. 4:258-266. Broothaerts, W.J., van Laere, A., Witters, R., Prt>aux, G., Decock, B., van Damme, J., and Vendrig, J.C. 1989. Purification and N-terminal sequencing of style glycoproteins associated with self-incompatibility in Petunia hybrida. Plant Mol. Biol. 14:93-102. Brown, D.C.W. 1988. Germplasm determination of in vitro somatic embryogenesis in alfalfa. HortScience 23:526-531. Brown, D.C.W., and Atanassov, A. 1985. Role of genetic background in somatic embryogenesis in Medicago. Plant Cell Tissue Organ Cult. 4:111-122. Brown, D.C.W., Finstad, K.I., and Watson, E.M. 1995. Somatic embryogenesis in herbaceous dicots. In In Vitro Embryogenesis in Plants, ed. T.A. Thorpe, pp. 345-415. Dordrecht: Kluwer Academic Publishers. Brown, J.W.S., Ersland, D.R., and Hall, T.C. 1982. Molecular aspects of storage protein synthesis during seed development. In The Physiology and Biochemistry of Seed Development, Dormancy and Germination, ed. A.A. Khan, pp. 3—42. Amsterdam: Elsevier Biomedical Press. Brown, J.W.S., Wandelt, C, and Feix, G. 1986. The upstream regions of zein genes: sequence analysis and expression in the unicellular green alga Acetabularia. Eur. J. Cell Biol. 42:161-170. Brown, R.C., and Lemmon, B.E. 1988. Microtubules associated with simultaneous cytokinesis of coenocytic microsporocytes. Am. J. Bot. 75:1848-1856. Brown, R.C., and Lemmon, B.E. 1989. Minispindles and cytoplasmic domains in microsprogenesis of orchids. Protoplasma 148:26-32. Brown, R.C., and Lemmon, B.E. 1991a. Pollen development in orchids. 1. Cytoskeleton and the control of division plane in irregular patterns of cytokinesis. Protoplasma 163:9-18. Brown, R.C., and Lemmon, B.E. 1991b. Pollen development in orchids. 2. The cytokinetic apparatus in simultaneous cytokinesis. Protoplasma 165:155-166. Brown, R.C., and Lemmon, B.E. 1991c. Pollen development in orchids. 3. A novel generative pole microtubule system predicts unequal pollen mitosis. /. Cell Sci. 99:273-281. Brown, R.C., and Lemmon, B.E. 1991d. Pollen development in orchids. 5. A generative cell domain involved in spatial control of the hemispherical cell plate. /. Cell Sci. 100:559-565. Brown, R.C., and Lemmon, B.E. 1992a. Control of division plane in normal and griseofulvin-treated microsporocytes of Magnolia. J. Cell Sci. 103:1031-1038. Brown, R.C., and Lemmon, B.E. 1992b. Pollen development in orchids. 4. Cytoskeleton and ultrastructure of the unequal pollen mitosis in Phalaenopsis. Protoplasma 167:183-192. Brown, R.C., and Lemmon, B.E. 1994. Pollen mitosis in the slipper orchid Cypripedium fasciculatum. Sex. Plant Reprod. 7:87-94. Brown, R.C., and Lemmon, B.E. 1996. Nuclear cytoplasmic domains, microtubules and organelles in microsporocytes of the slipper orchid Cypripedium
References
545
californicum A. Gray dividing by simultaneous Burgess, J. 1970a. The occurrence of cross-linked microtubules in a higher plant cell. Planta 92:25-28. cytokinesis. Sex. Plant Reprod. 9:145-152. Brown, R.C, Lemmon, B.E., and Olsen, O.-A. 1994. Burgess, J. 1970b. Cell shape and mitotic spindle formaEndosperm development in barley: microtubule tion in the generative cell of Endymion non-scriptus. involvement in the morphogenetic pathway. Plant Planta 95:72-85. Cell 6:1241-1252. Burgess, J. 1970c. Microtubules and cell division in the Brown, S.M., and Crouch, M.L. 1990. Characterization of microspore of Dactylorchis fuschii. Protoplasma a gene family abundantly expressed in Oenothera 69:253-264. organensis pollen that shows sequence similarity to Burns, J.A., and Wetzstein, H.Y. 1994. Storage reserves in polygalacturonase. Plant Cell 2:263-274. pecan somatic embryos derived from suspension culture. Plant Sci. 102:213-219. Brown, S.W. 1949. Endomitosis in the tapetum of tomato. Burow, M.D., Sen, P., Chlan, C.A., and Murai, N. 1992. Am. ]. Bot. 36:703-716. Developmental control of the (J-phaseolin gene Brown, S.W., and Cave, M.S. 1953. Induced dominant requires positive, negative, and temporal seed-spelethality in Lilium. Proc. Natl Acad. Sci. USA 39:97-102. cific transcriptional regulatory elements and a negaBrown, S.W., and Cave, M.S. 1954. The detection and tive element for stem and root expression. Plant J. nature of dominant lethals in Lilium. 1. Effects of X 2:537-548. rays on the heritable component and functional ability of the pollen grain. Am. J. Bot. 41:455-469. Burr, B., and Burr, F.A. 1976. Zein synthesis in maize Bruck, D.K., and Walker, D.B. 1985a. Cell determination endosperm by polyribosomes attached to protein during embryogenesis in Citrus jambhiri. I. Ontogeny bodies. Proc. Natl Acad. Sci. USA 73:515-519. of the epidermis. Bot. Gaz. 146:188-195. Burr, B., Burr, F.A., Rubenstein, I., and Simon, M.N. 1978. Bruck, D.K., and Walker, D.B. 1985b. Cell determination Purification and translation of zein messenger RNA during embryogenesis in Citrus jambhiri. II. from maize endosperm protein bodies. Proc. Natl Epidermal differentiation as a one-time event. Am. J. Acad. Sci. USA 75:696-700. Bot. 72:1602-1609. Burr, B., Burr, F.A., St. John, T.P., Thomas, M., and Davis, Brunori, A. 1967. Relationship between DNA synthesis R.W. 1982. Zein storage protein gene family of and water content during ripening of Viciafaba seed. maize. An assessment of heterogeneity with cloned Caryologia 20:333-338. messenger RNA sequences. /. Mol. Biol. 154:33-49. Brunori, A., and D'Amato, F. 1967. The DNA content of Burr, F.A., and Burr, B. 1982. Three mutations in Zea mays affecting zein accumulation: comparison of zein nuclei in the embryo of dry seeds of Pinus pinea and polypeptides, in vitro synthesis and processing, Lactuca sativa. Caryologia 20:153-161. mRNA levels, and genome organization. /. Cell Biol. Brunori, A., Forino, L.M.C., and Frediani, M. 1989. 94:201-206. Polyploid DNA contents in the starchy endosperm nuclei and seed weight in Triticum aestivum. J. Genet.Burton, R.A., Bewley, J.D., Smith, A.M., Bhattacharyya, Breed. 43:131-134. M.K., Tatge, H., Ring, S., Bull, V., Hamilton, W.D.O., and Martin, C. 1995. Starch branching enzymes Bruun, L. 1987. The mature embryo sac of the sugar beet, belonging to distinct enzyme families are differenBeta vulgaris: a structural investigation. Nord. J. Bot. tially expressed during pea embryo development. 7:543-551. Plant}. 7:3-15. Bruun, L., and Olesen, P. 1989. A structural investigation of the ovule in sugarbeet, Beta vulgaris: the micropy- Busch, M., Mayer, U., and Jiirgens, G. 1996. Molecular lar nucellus. Nord. }. Bot. 9:81-87. analysis of the Arabidopsis pattern formation gene Bryan, J.H.D. 1951. DNA-protein relations during GNOM: gene structure and intragenic complementamicrosporogenesis of Tradescantia. Chromosoma tion. Mol. Gen. Genet. 250:681-691. 4:369-392. Busri, N., Chapman, G.P., and Greenham, J. 1993. The Bucciaglia, P.A., and Smith, A.G. 1994. Cloning and charembellum: a newly defined structure in the grass acterization of Tag 1, a tobacco anther B-l,3-gluovule. Sex. Plant Reprod. 6:191-198. canase expressed during tetrad dissolution. Plant Bustos, M.M., Begum, D., Kalkan, F.A., Battraw, M.J., and Mol. Biol. 24:903-914. Hall, T.C. 1991. Positive and negative as-acting Bucher, M., Brander, K.A., Sbicego, S., Mandel, T, and DNA domains are required for spatial and temporal Kuhlemeier, C. 1995. Aerobic fermentation in regulation of gene expression by a seed storage protobacco pollen. Plant Mot. Biol. 28:739-750. tein promoter. EMBO ]. 10:1469-1479. Buckley, L.G., and Trigiano, R.N. 1994. Changes in ovule Bustos, M.M., Guiltinan, M.J., Jordano, ]., Begum, D., protein profiles associated with somatic embryogenKalkan, F.A., and Hall, T.C. 1989. Regulation of B-glucuronidase expression in transgenic tobacco plants by esis in Cercis canadensis (redbud). Plant Cell Rep. an A/T rich,ris-actingsequence found upstream of a 14:27-30. French bean p-phaseolin gene. Plant Cell 1:839-853. Budelier, K.A., Smith, A.G., and Gasser, C.S. 1990. Regulation of a stylar transmitting tissue-specific Bustos, M.M., Luckow, V.A., Griffing, L.R., Summers, M.D., and Hall, T.C. 1988. Expression, glycosylation gene in wild-type and transgenic tomato and and secretion of phaseolin in a baculovirus system. tobacco. Mol. Gen. Genet. 224:183-192. Plant Mol. Biol. 10:475-488. Buffard-Morel, J. 1968. Effets du glucose, du 16vulose, du maltose et du saccharose sur le d£veloppement des Butler, W.M., and Cuming, A.C. 1993. Differential molecular responses to abscisic acid and osmotic stress in embryons de palmier a huile (Elaeis guineensis Jacq. var. Dura Bee.) en culture in vitro. Compt. Rend. Acad. viviparous maize embryos. Planta 189:47-54. Button, J., Kochba, J., and Bornman, C.H. 1974. Fine strucSci. Paris 185D:185-188. ture of and embryoid development from embryoBufe, A., Spangfort, M.D., Kahlert, H., Schlaak, M., and genie ovular callus of ~Shamouti' orange (Citrus Becker, W.-M. 1996. The major birch pollen allergen, Bet v 1, shows ribonuclease activity. Planta 199:413—415. sinensis Osb.). /. Expt. Bot. 25:446-457.
546
References
Buttrose, M.S. 1963a. Infrastructure of the developing body formation in the developing barley wheat endosperm. Aust. J. Bio/. Sri. 16:305-317. endosperm. Carlsberg Res. Commun. 45:577-594. Buttrose, M.S. 1963b. Ultrastructure of the developing Campbell, R.J., and Ascher, P.D. 1975. Incorporation of aleurone cells of wheat grain. Aust. J. Bio/. Sci. radioactive label into nucleic acids of compatible 16:768-774. and incompatible pollen tubes of Lilium longiflorum Bystedt, P.-A. 1990. The transmitting tract in Trimezia fosThunb. Theor. Appl. Genet. 46:143-148. teriana (Iridaceae). I. Ultrastructure in the stigma, Campbell, W.P., Lee, J.W., O'Brien, T.P., and Smart, M.G. style and ovary. Nord. ]. Bot. 9:507-518. 1981. Endosperm morphology and protein body formation in developing wheat grain. Aust. ]. Plant Physiol. 8:5-19. Cabrera-Ponce, J.L., Vegas-Garcia, A., and HerreraEstrella, L. 1995. Herbicide resistant transgenic Campenot, M.K., Zhang, G., Cutler, A.J., and Cass, D.D. papaya plants produced by an efficient particle bom1992. Zea mays embryo sacs in culture. I. Plant regenbardment transformation method. Plant Cell Rep. eration from 1 day after pollination embryos. Am. J. 15:1-7. Bot. 79:1368-1373. Cabrera-Ponce, J.L., Vegas-Garcia, A., and Herrera- Canas, L., and Malmberg, R. 1992. Isolation and characterization of three monoclonal antibodies specific for Estrella, L. 1996. Regeneration of transgenic papaya pistil and stamen tissues of tobacco (Nicotiana plants via somatic embryogenesis induced by Agrobacterium rhizogenes. In Vitro Cell Devel Biol. tabacum L.). Plant Sci. 83:195-203. 32:86-90. Canas, L.A., Busscher, M., Angenent, G.C., Beltrdn, J.-P., Cadic, A., and Sangwan-Norreel, B.S. 1983. Modifications and van Tunen, A.J. 1994. Nuclear localization of the Petunia MADS box protein FBP1. Plant ]. 6:597-604. ultrastructurales provoqufies par des traitements promoteurs de l'androgenese chez le Datura innoxia Cao, J., Duan, X., McElroy, D., and Wu, R. 1992. Mill. Ann. Sci. Nat. Bot. Ser. XIII 5:97-114. Regeneration of herbicide resistant transgenic rice plants following microprojectile-mediated transforCai, G., Bartalesi, A., Del Casino, C, Moscatelli, A., Tiezzi, mation of suspension culture cells. Plant Cell Rep. A., and Cresti, M. 1993. The kinesin-immunoreactive 11:586-591. homologue from Nicotiana tabacum pollen tubes: biochemical properties and subcellular localization. Capkova, V., Hrabetova\ E., and Tupy, J. 1983. Reduction Planta 191:496-506. of leucine efflux and protein release from tobacco Cai, G., Moscatelli, A., Del Casino, C, and Cresti, M. 1996. pollen tubes in culture by actinomycin D in the Cytoplasmic motors and pollen tube growth. Sex. absence of calcium. Biochem. Physiol. Pflanz. Plant Reprod. 9:59-64. 178:521-527. Cai, X., Dong, Y., and Sodmergen. 1995. Actin organiza- Capkova, V., Hrabetova\ E., and Tupy, J. 1987. Protein tion and distribution during hydration, activation changes in tobacco pollen culture; a newly syntheand germination of pollen in Amaryllis vittata Ait. sized protein related to pollen tube growth. /. Plant using TRITC-phalloidin as a probe. Cytologia Physiol. 130:307-314. 60:133-139. Capkova', V., Hrabetova\ E., and Tupy, J. 1988. Protein synCaiola, M.G., Banas, M., and Canini, A. 1993. Ultrathesis in pollen tubes: preferential formation of new structure and germination percentage of Crocus species independent of transcription. Sex. Plant biflorus Miller subsp. biflorus (Iridaceae) pollen. Bot. Reprod. 1:150-155. Acta 106:488^95. Capkova, V., Zbrozek, J., and Tupy, J. 1994. Protein synCaligo, M.A., Nuti Ronchi, V., and Nozzolini, M. 1985. thesis in tobacco pollen tubes: preferential synthesis Proline and serine affect polarity and development of cell-wall 69-kDa and 66-kDa glycoproteins. Sex. of carrot somatic embryos. Cell Diffn 17:193-198. Plant Reprod. 7:57-66. Callis, J., and Bedinger, P. 1994. Developmentally regu- Capkova, V., Hrabetov^, E., Tupy, J., and Rihova, L. 1983. lated loss of ubiquitin and ubiquitinated proteins Amino acid uptake and protein synthesis in cultured during pollen maturation in maize. Proc. Natl Acad. tobacco pollen. Biochem. Physiol. Pflanz. 178:511-520. Sci. USA 91:6074-6077. Capkova-Balatkovd, V., Hrabetova", E., and Tupy, J. 1980. Callis, J., Raasch, J.A., and Vierstra, R.D. 1990. Ubiquitin Effects of cycloheximide on pollen of Nicotiana extension proteins of Arabidopsis thaliana. Structure, tabacum in culture. Biochem. Physiol. Pflanz. 175:412-420. localization, and expression of their promoters in transgenic tobacco. /. Biol. Chem. 265:12486-12493. Carasco, J.F., Croy, R., Derbyshire, E., and Boulter, D. 1978. The isolation and characterization of the major Calzoni, G.L., Speranza, A., Li, Y.Q., Ciampolini, R, and polypeptides of the seed globulin of cowpea (Vigna Cresti, M. 1993. Wall biosynthesis and wall polysacunguiculata L. Walp) and their sequential synthesis in charide composition in pollen tubes of Malus domestica growing at low rate after monensin treatment. developing seeds. /. Expt. Bot. 29:309-323. Acta Bot. Neerl. 42:473-480. Carlberg, I., Soderhall, K., Glimelius, K., and Eriksson, T. Camadro, E.L., and Masuelli, R.W. 1995. A genetic model 1984. Protease activities in non-embryogenic and for the endosperm balance number (EBN) in the embryogenic carrot cell strains during callus growth wild potato Solanum acaule Bitt. and two related and embryo formation. Physiol. Plant. 62:458-464. diploid species. Sex. Plant Reprod. 8:283-288. Carman, J.G. 1988. Improved somatic embryogenesis in Camefort, H., and Sangwan, R.S. 1979. Effets d'un choc wheat by partial simulation of the in-ovulo oxygen, thermique sur certaines ultrastructures des grains de growth-regulator and desiccation environments. pollen embryogenes du Datura metel L. Compt. Rend. Planta 175:417-424. Acad. Sci. Paris 288D:1383-1386. Carman, J.G., Crane, C.F., and Riera-Lizarazu, O. 1991. Comparative histology of cell walls during meiotic Cameron-Mills, V., and Brandt, A. 1988. A y-hordein gene. and apomeiotic megasporogenesis in two hexaploid Plant Mol. Biol. 11:449-461. Australasian Elymus species. Crop Sci. 31:1527-1532. Cameron-Mills, V., and von Wettstein, D. 1980. Protein
References
547
Megagametophyte development in Hordeum vulgare. Carmichael, J.S., and Friedman, W.E. 1995. Double fertil1. Early megagametogenesis and the nature of cell ization in Gnetum gnemon: the relationship between wall formation. Can. j. Bot. 63:2164-2171. the cell cycle and sexual reproduction. Plant Cell Cass, D.D., Peteya, D.J., and Robertson, B.L. 1986. 7:1975-1988. Megagametophyte development in Hordeum vulgare. Carmona, M.J., Hernandez-Lucas, C, San Martin, C, 2. Later stages of wall development and morphologGonzalez, P., and Garrfa-Olmedo, F. 1993. Subcellular ical aspects of megagametophyte cell differentiation. localization of type I thionins in the endosperms of Can. ]. Bot. 64:2327-2336. wheat and barley. Protoplasma 173:1-7. Carniel, K. 1952. Das Verhalten der Kerne im Tapetum der Castle, L.A., and Meinke, D.W. 1994. A FUSCA gene of Angiospermen mit besonderer Berucksichtigung Arabidopsis encodes a novel protein essential for von Endomitosen und sogenannten Endomitosen. plant development. Plant Cell 6:25-41. Osterr. Bot. Z. 99:318-362. Castle, L.A., Errampalli, D., Atherton, T.L., Franzmann, L.H., Yoon, E.S., and Meinke, D.W. 1993. Genetic and Carniel, K. 1961. Das Antherentapetum von Zea mays. molecular characterization of embryonic mutants Osterr. Bot. Z. 108:89-96. identified following seed transformation in Carniel, K. 1967a. Uber die Embryobildung in der Arabidopsis. Mol. Gen. Genet. 241:504-514. Gattung Paeonia. Osterr. Bot. Z. 114:4-19. Carniel, K. 1967b. Licht- und elektronenmikroskopische Cavallini, A., Natali, L., Balconi, C, Rizzi, E., Motto, M., Cionini, G., and D'Amato, F. 1995. Chromosome Untersuchung der Ubischkorperentwicklung in der endoreduplication in endosperm cells of two maize Gattung Oxalis. Osterr. Bot. Z. 114:490-501. genotypes and their progenies. Protoplasma Carpenter, J.L., Ploense, S.E., Snustad, D.P., and Silflow, 189:156-162. CD. 1992. Preferential expression of an a-tubulin gene of Arabidopsis in pollen. Plant Cell 4:557-571. Cave, M.S., Arnott, H.J., and Cook, S.A. 1961. Embryogeny in the California peonies with referCarpenter, R., and Coen, E.S. 1990. Floral homeotic mutaence to their taxonomic position. Am. ]. Bot. tions produced by transposon-mutagenesis in 48:397-404. Antirrhinum majus. Genes Devel. 4:1483-1493. Carraro, L., and Lombardo, G. 1976. Tapetal ultrastruc- Cave, M.S., and Brown, S.W. 1954. The detection and nature of dominant lethals in Lilium. II. Cytological tural changes during pollen development. II. Studies abnormalities in ovules after pollen irradiation. Am. on Pelargonium zonale and Kalanchoe obtusa. ]. Bot. 41:469^83. Caryologia 29:339-344. Carraro, L., Lombardo, G., and Gerola, F.M. 1985. Cebrat, J., and Zad^cka, A. 1978. Development of anthers in three male-sterile lines of rye (Secale cereale L.). Electron-cytochemical localization of peroxidase in Genet. Polon. 19:25-31. self- and cross-pollinated styles of Primula acaulis. Caryologia 38:83-94. Ceccarelli, N., Lorenzi, R., and Alpi, A. 1979. Kaurene and kaurenol biosynthesis in cell-free system of Phaseolus Carraro, L., Lombardo, G., and Gerola, F.M. 1986. Stylar coccineus suspensor. Phytochemistry 18:1657-1658. peroxidase and incompatibility reactions in Petunia hybrida. ]. Cell Sci. 82:1-10. Ceccarelli, N., Lorenzi, R., and Alpi, A. 1981a. Gibberellin Casas, A.M., Kononowicz, A.K., Zehr, U.B., Tomes, D.T., biosynthesis in Phaseolus coccineus suspensor. Z. Pflanzenphysiol. 102:37-44. Axtell, J.D., Butler, L.G., Bressan, R.A., and Hasegawa, P.M. 1993. Transgenic sorghum plants Ceccarelli, N., Lorenzi, R., and Alpi, A. 1981b. Kaurene metabolism in cell-free extracts of Phaseolus coccineus via microprojectile bombardment. Proc. Natl Acad. Sci. USA 90:11212-11216. suspensors. Plant Sci. Lett. 21:325-332. Casey, R., Domoney, C, and Ellis, N. 1986. Legume stor- Chamberland, S., Daigle, N., and Bernier, F. 1992. The legumin boxes and the 3 part of a soybean fi-conglycinin age proteins and their genes. Oxford Surv. Plant Mol. Cell Biol. 3:1-95. promoter are involved in seed gene expression in transgenic tobacco plants. Plant Mol. Biol. 19:937-949. Casey, R., Domoney, C, and Nielsen, N.C. 1985. Isolation of a cDNA clone for pea (Pisum sativum) seed lipoxy- Chamberlin, M.A., Homer, H.T., and Palmer, R.G. 1993a. Nutrition of ovule, embryo sac, and young embryo genase. Biochem. ]. 232:79-85. in soybean: an anatomical and autoradiographic Cass, D.D. 1972. Occurrence and development of a filistudy. Can. J. Bot. 71:1153-1168. form apparatus in the egg of Plumbago capensis. Am. ]. Bot. 59:279-283. Chamberlin, M.A., Homer, H.T., and Palmer, R.G. 1993b. Nuclear size and DNA content of the embryo and Cass, D.D. 1973. An ultrastructural and Nomarski-interendosperm during their initial stages of developference study of the sperms of barley. Can. ]. Bot. ment in Glycine max (Fabaceae). Am. ]. Bot. 51:601-605. 80:1209-1215. Cass, D.D., and Fabi, G.C. 1988. Structure and properties of sperm cells isolated from the pollen of Zea mays. Chan, M.-T., Chang, H.-H., Ho, S.-L., Tong, W.-F., and Yu, S.-M. 1993. Agrobacterium-mediated production of Can. ]. Bot. 66:819-825. transgenic rice plants expressing a chimeric oc-amyCass, D.D., and Jensen, W.A. 1970. Fertilization in barley. lase promoter/fj-glucuronidase gene. Plant Mol. Biol. Am. ]. Bot. 57:62-70. 22:491-506. Cass, D.D., and Karas, I. 1974. Ultrastructural organization of the egg of Plumbago zeylanica. ProtoplasmaChandlee, J.M., and Scandalios, J.G. 1983. Gene expression during early kernel development in Zea mays. 81:49-62. Devel Genet. 4:99-115. Cass, D.D., and Karas, I. 1975. Development of sperm Chandlee, J.M., and Scandalios, J.G. 1984. Regulation of cells in barley. Can. J. Bot. 53:1051-1062. Catl gene expression in the scutellum of maize durCass, D.D., and Peteya, D.J. 1979. Growth of barley pollen ing early sporophytic development. Proc. Natl Acad. tubes in vivo. I. Ultrastructural aspects of early tube Sci. USA 81:4903^907. growth in the stigmatic hair. Can. J. Bot. 57:386-396. Cass, D.D., Peteya, D.J., and Robertson, B.L. 1985. Chandler, P.M., Higgins, T.J.V., Randall, P.J., and Spencer,
548
References
D. 1983. Regulation of legumin levels in developing Charne, D.G., and Beversdorf, W.D. 1988. Improving pea seeds under conditions of sulfur deficiency. microspore culture as a rapeseed breeding tool: the Rates of legumin synthesis and levels of legumin use of auxins and cytokinins in an induction mRNA. Plant Physiol. 71:47-54. medium. Can.}. Bot. 66:1671-1675. Chandler, P.M., Spencer, D., Randall, P.J., and Higgins, Charzynska, M., Ciampolini, F., and Cresti, M. 1988. T.J.V. 1984. Influence of sulfur nutrition on developGenerative cell division and sperm cell formation in mental patterns of some major pea seed proteins and barley. Sex. Plant Reprod. 1:240-247. their mRNAs. Plant Physiol. 75:651-657. Charzynska, M., and Cresti, M. 1993. Early events in diviChandra, S., and Bhatnagar, S.P. 1974. Reproductive biolsion of the generative cell of Ornithogalum virens. ogy of Triticum. II. Pollen germination, pollen tube Protoplasma 172:77-83. growth, and its entry into the ovule. Phytomorphology Charzynska, M., and Lewandowska, E. 1990. Generative 24:11-217. cell division and sperm cell association in the pollen grain of Sambucus nigra. Ann. Bot. 65:685-689. Chandra Sekhar, K.N., and Heij, E.G. 1995. Changes in proteins and peroxidases induced by compatible Charzynska, M., and Maleszka, J. 1978. 3H-thymidine pollination in the ovary of Nicotiana tabacum L. incorporation into the microspores and pollen grains ahead of the advancing pollen tubes. Sex. Plant nuclei in excised Tradescantia stamens. Acta Soc. Bot. Reprod. 8:369-374. Polon. 47:163-171. Chandra Sekhar, K.N., and Williams, E.G. 1992. Charzynska, M., Murgia, M., and Cresti, M. 1989. Nonradioactive in situ localization of poly(A)+RNA Ultrastructure of the vegetative cell of Brassica napus during pollen development in anthers of tobacco pollen with particular reference to microbodies. {Nicotiana tabacum L.). Protoplasma 169:9-17. Protoplasma 152:22-28. Chang, M.T., and Neuffer, M.G. 1989. Maize microsporo- Charzynska, M., Murgia, M., and Cresti, M. 1990. genesis. Genome 32:232-244. Microspore of Secale cereale as a transfer cell type. Protoplasma 158:26-32. Chang, Y.C., and Walling, L.L. 1991. Abscisic acid negatively regulates expression of chlorophyll alb bind- Charzynska, M., and Pannenko, I. 1976. Inhibition of cytokinesis in the microspores of Tradescantia ing protein genes during soybean embryogeny. Plant Physiol. 97:1260-1264. bracteata Small, by caffeine. Acta Soc. Bot. Polon. 45:469^176. Chang, Y.C., and Walling, L.L. 1992. Chlorophyll a/b-binding protein genes are differentially expressed during Charzynska, M., Murgia, M., Milanesi, M., and Cresti, M. soybean development. Plant Mol. Biol. 19:217-230. 1989. Origin of sperm cell association in the "male Chao, C.-Y. 1971. A periodic acid-Schiff's substance germ unit" of Brassica pollen. Protoplasma 149:1-4. related to the directional growth of pollen tube into Chase, CD. 1994. Expression of CMS-unique and flankthe embryo sac in Paspalum ovules. Am. ]. Bot. ing mitochondrial DNA sequences in Phaseolus vul58:649-654. garis L. Curr. Genet. 25:245-251. Chao, C.-Y. 1977. Further cytological studies of a periodic Chase, CD., and Ortega, V.M. 1992. Organization of ATPA coding and 3' flanking sequences associated acid-Schiff's substance in the ovules of Paspalum with cytoplasmic male sterility in Phaseolus vulgaris orbiculare and P. longifolium. Am. J. Bot. 64:921-930. h. Curr. Genet. 22:147-153. Chapman, G.P. 1987. The tapetum. Int. Rev. Cytol. 107:111-125. Chaubal, R., and Reger, B.J. 1990. Relatively high calcium is localized in synergid cells of wheat ovaries. Sex. Chapman, G.P, and Busri, N. 1994. Apomixis in Pennisetum: an ultrastructural study. Int. ]. Plant Sci. Plant Reprod. 3:98-102. 155:492^97. Chaubal, R., and Reger, B.J. 1992a. Calcium in the synergids and other regions of pearl millet ovaries. Sex. Chappell, J., and Chrispeels, M.J. 1986. Transcriptional and Plant Reprod. 5:34-46. posttranscriptional control of phaseolin and phytohemagglutinin gene expression in developing cotyle- Chaubal, R., and Reger, B.J. 1992b. The dynamics of caldons of Phaseolus vulgaris. Plant Physiol. 81:50-54. cium distribution in the synergid cells of wheat after pollination. Sex. Plant Reprod. 5:206-213. Chardard, R. 1958. L'ultrastructure des grains de pollen Chaubal, R., and Reger, B.J. 1993. Prepollination degenerd'Orchidees. Rev. Cytol. Biol. Veget. 19:223-235. ation in mature synergids of pearl millet: an examiChardard, R. 1962. Recherches sur les cellules-meres des nation using antimonate fixation to localize calcium. microspores des Orchidees. Etude au microscope Sex. Plant Reprod. 6:225-238. electronique. Rev. Cytol. Biol. Veget. 24:1-148. Charest, P.J., Devantier, Y, and Lachance, D. 1996. Stable Chaubal, R., and Reger, B.J. 1994. Dynamics of antigenetic transformation of Picea marina (Black spruce) monate-precipitated calcium and degeneration in via particle bombardment. In Vitro Cell Devel Biol. unpollinated pearl millet synergids after maturity. 32:91-99. Sex. Plant Reprod. 7:122-134. Charest, P.J., Calero, N., Lachance, D., Datla, R.S.S., Chaudhuri, S., and Messing, J. 1994. Allele-specific Duchesne, L.C., and Tsang, E.W.T. 1993. parental imprinting of drzl, a posttranscriptional Microprojectile-DNA delivery in conifer species: facregulator of zein accumulation. Proc. Natl Acad. Sci. tors affecting assessment of transient gene expresUSA 91:4867-4871. sion using the (3-glucoronidase reporter gene. Plant Chaudhury, A.M., Craig, S., Bloemer, K.C., Farrell, L., and Cell Rep. 12:189-193. Dennis, E.S. 1992. Genetic control of male fertility in Charlton, W.L., Keen, C.L., Merriman, C, Lynch, P., higher plants. Aust. ]. Plant Physiol. 19:419^426. Greenland, A.J., and Dickinson, H.G. 1995. Chaudhury, A.M., Lavithis, M., Taylor, P.E., Craig, S., Endosperm development in Zea mays; implication of Singh, M.B., Signer, E.R., Knox, R.B., and Dennis, gametic imprinting and paternal excess in regulation E.S. 1994. Genetic control of male fertility in of transfer layer development. Development Arabidopsis thaliana: structural analysis of premeiotic 121:3089-3097. developmental mutants. Sex. Plant Reprod. 7:17-28.
References
549
Chen, Z., Schertz, K.F., Mullet, J.E., DuBell, A., and Hart, G.E. 1995. Characterization and expression of rpoCl in CMS and fertile lines of sorghum. Plant Mol. Biol. 28:799-809. Chen, Z.-K., Wang, F.-H., and Zhou, F. 1988. On the origin, development and ultrastructure of the orbicules and pollenkitt in the tapetum of Anemarrhena asphodeloides (Liliaceae). Grana 27:273-282. Chen, Z.-L., Pan, N.-S., and Beachy, R.N. 1988. A DNA sequence element that confers seed-specific enhancement to a constitutive promoter. EMBO J. 7:297-302. ration. Proc. Natl Acad. Sci. USA 92:1167-1171. Chen, Z.-L., Schuler, M.A., and Beachy, R.N. 1986. Functional analysis of regulatory elements in a plant Chay, C.H., Buehler, E.G., Thorn, J.M., Whelan, T.M., and embryo-specific gene. Proc. Natl Acad. Sci. USA Bedinger, P.A. 1992. Purification of maize pollen 83:8560-8564. exines and analysis of associated proteins. Plant Physiol. 100:756-761. Chen, Z.-L., Naito, S., Nakamura, I., and Beachy, R.N. 1989. Regulated expression of genes encoding soyChee, P.P., Jones, J.M., and Slightom, J.L. 1991. Expression of bean storage protein minigene in tobacco seeds. bean P-conglycinins in transgenic plants. Devel Introns are not required for seed specific expression. Genet. 10:112-122. /. Plant Physiol. 137:402-408. Chen, Z.-Y., Hsing, Y.-I.C, Lee, P.-F., and Chow, T.-Y. 1992. Nucleotide sequences of a soybean cDNA encoding Chee, P.P., Klassy, R.C., and Slightom, J.L. 1986. an 18 kilodalton late embryogenesis abundant proExpression of a bean storage protein "phaseolin tein. Plant Physiol. 99:773-774. minigene' in foreign plant tissues. Gene 41:47-57. Chen, C. 1977. In vitro development of plants from Cheng, P C , Greyson, R.I., and Walden, D.B. 1979. microspores of rice. In Vitro 13:484-489. Comparison of anther development in genie maleChen, C.C., Kasha, K.J., and Marsolais, A. 1984. sterile (mslO) and in male-fertile corn (Zen mays) Segmentation patterns and mechanisms of genome from light microscopy and scanning electron multiplication in cultured microspores of barley. microscopy. Can. ]. Bot. 57:578-596. Can. J. Genet. Cytol. 26:475^83. Cheng, W.-H., Taliercio, E.W., and Chourey, PS. 1996. The miniaturel seed locus of maize encodes a cell wall Chen, C.C., Howarth, M.J., Peterson, R.L., and Kasha, K.J. invertase required for normal development of 1984. Ultrastructure of androgenic microspores of endosperm and maternal cells in the pedicel. Plant barley during the early stages of anther culture. Can. Cell 8:971-983. ]. Genet. Cytol. 26:484-491. Chen, C.-G., Cornish, E.C., and Clarke, A.E. 1992. Specific Chern, M.-S., Eben, H.G., and Bustos, M.M. 1996. The developmentally regulated bZIP factor ROM1 modexpression of an extensin-like gene in the style of ulates transcription from lectin and storage protein Nicotiana data. Plant Cell 4:1053-1062. genes in bean embryos. Plant j . 10:135-148. Chen, C.-G., Mau, S.-L., and Clarke, A.E. 1993. Nucleotide sequence and style-specific expression of a novel Chernyshev, A.I., Davletova, S.K., Bashkirov, V.I., proline-rich protein gene from Nicotiana alata. Plant Shakhmanov, N.B., Mekhedov, S.L., and Anan'ev, E.V. 1989. Nucleotide sequence of the Bl hordein gene of Mol. Bid. 21:391-395. barley (Hordeum vulgare L.). Soviet Genet. 25:865-869. Chen, C.-H., and Nasrallah, J.B. 1990. A new class of S sequences defined by a pollen recessive self-incom- Chesnut, R.S., Shotwell, M.A., Boyer, S.K., and Larkins, patibility allele of Brassica oleracea. Mol. Gen. Genet. B.A. 1989. Analysis of avenin proteins and the 222:241-248. expression of their mRNAs in developing oat seeds. Plant Cell 1:913-924. Chen, K., Johal, S., and Wildman, S.G. 1977. Phenotypic markers for chloroplast DNA genes in higher plants Cheung, A.Y., Wang, H., and Wu, H.-M. 1995. A floral transand their use in biochemical genetics. In Nucleic Acids mitting tissue-specific glycoprotein attracts pollen and Protein Synthesis in Plants, ed. L. Bogorad and J. tubes and stimulates their growth. Cell 82:383-393. H. Weil, pp. 183-194. New York: Plenum Press. Cheung, A.Y., May, B., Kawata, E.E., Gu, Q., and Wu, H.M. 1993. Characterization of cDNAs for stylar transChen, K., and Meyer, V.G. 1979. Mutation in chloroplast mitting tissue-specific proline-rich proteins in DNA coding for the large subunit of fraction 1 protobacco. Plant J. 3:151-160. tein correlated with male sterility in cotton. /. Hered. 70:431-433. Cheung, A.Y., Zhan, X.-Y, Wang, H., and Wu, H.-M. 1996. Organ-specific and agamous-regulated expression Chen, L.-J., and Luthe, D.S. 1987. Analysis of proteins and glycosylation of a pollen tube growth-promotfrom embryogenic and non- embryogenic rice ing protein. Proc. Natl Acad. Sci. USA 93:3853-3858. (Oryza sativa L.) calli. Plant Sci. 48:181-188. Chen, R., Aguirre, P.J., and Smith, A.G. 1994. Chibbar, R.N., Kartha, K.K., Leung, N., Qureshi, J., and Characterization of an anther- and tapetum-specific Caswell, K. 1991. Transient expression of marker gene encoding a glycine-rich protein from tomato. /. genes in immature zygotic embryos of spring wheat Plant Physiol. 143:651-658. (Triticum aestivum) through microprojectile bombardment. Genome 34:453-460. Chen, R., and Smith, A.G. 1993. Nucleotide sequence of a stamen- and tapetum-specific gene from Lycopersicon Chibbar, R.N., Shyluk, J., Georges, F., Mallard, C.S., and esculentum. Plant Physiol. 101:1413. Constabel, F. 1988. Esterase isozymes as markers of Chen, Y.-C, and Chourey, PS. 1989. Spatial and temporal somatic embryogenesis in cultured carrot cells. /. expression of the two sucrose synthase genes in Plant Physiol. 133:367-370. maize: immunohistological evidence. Theor. Appl. Chibi, F., Angosto, T., Garrido, D., and Matilla, A. 1993. Requirement of polyamines for in-vitro maturation Genet. 78:553-559. Chauhan, S.V.S., and Kinoshita, T. 1979. Histochemical localization of histones, DNA and proteins in anthers of male-fertile and male-sterile plants, jap. J. Breed. 29:287-293. Chauhan, S.V.S., and Singh, S.P. 1966. Pollen abortion in male-sterile hexaploid wheat ('Norin') having Aegilops ovata L. cytoplasm. Crop Sci. 6:532-535. Chaumont, R, Bernier, B., Buxant, R., Williams, M.E., Levings, C.E., III, and Boutry, M. 1995. Targeting the maize 7-urfl3 product into tobacco mitochondria confers methomyl sensitivity to mitochondrial respi-
550
References of the mid-binucleate pollen of Nicotiana tabacum. ]. Plant Physiol. 142:452-456.
Chichiricco, G., Constantino, P., and Spano, L. 1992. Expression of the rolB oncogene from Agrobacterium rhizogenes during zygotic embryogenesis in tobacco. Plant Cell Physiol. 33:827-832.
Childers, W.R. 1952. Male sterility in Medicago sativa L. Sri. Agr. 32:351-364. Chitralekha, P., and Bhandari, N.N. 1991. Post-fertilization development of antipodal cells in Ranunculus sceleratus Linn. Phytomorphology 41:200-212.
Chitralekha, P., and Bhandari, N.N. 1992. Cellularization of endosperm in Asphodelus tenuifolius Cav. Phytomorphology 42:185-193. Chitralekha, P., and Bhandari, N.N. 1993. Cellularization of free-nuclear endosperm in Ranunculus sceleratus Linn. Phytomorphology 43:165-183. Chlan, C.A., and Dure, L., III. 1983. Plant seed embryogenesis as a tool for molecular biology. Mol. Cell. Biochem. 55:5-15. Chlan, C.A., Borroto, K., Kamalay, J.A., and Dure, L., III. 1987. Developmental biochemistry of cottonseed embryogenesis and germination. XIX. Sequences and genomic organization of the a-globulin (vicilin) genes of cottonseed. Plant Mol. Biol. 9:533-546. Chlan, C.A., Pyle, J.B., Legocki, A.B., and Dure, L., III. 1986. Developmental biochemistry of cottonseed embryogenesis and germination. XVIII. cDNA and amino acid sequences of members of the storage protein families. Plant Mol. Biol. 7:475^89. Cho, T.-J., Davies, C.S., Fischer, R.L., Turner, N.E., Goldberg, R.B., and Nielsen, N.C. 1989. Molecular characterization of an aberrant allele for the Cy3 glycinin gene: a chromosomal rearrangement. Plant Cell 1:339-350. Cho, U.-H., and Kasha, K.J. 1989. Ethylene production and embryogenesis from anther cultures of barley (Hordeum vulgare). Plant Cell Rep. 8:415-417.
transport of phytohemagglutinin in the protein bodies in bean cotyledons. Planta 158:140-151. Chrispeels, M.J. 1985. UDP-GlcNAcglycoprotein GlcNActransferase is located in the Golgi apparatus of developing bean cotyledons. Plant Physiol. 78:835-838. Chrispeels, M.J., Vitale, A., and Staswick, P. 1984. Gene expression and synthesis of phytohemagglutinin in the embryonic axes of developing Phaseolus vulgaris seeds. Plant Physiol. 76:791-796. Chrispeels, M.J., Higgins, T.J.V., Craig, S., and Spencer, D. 1982. Role of the endoplasmic reticulum in the synthesis of reserve proteins and the kinetics of their transport to protein bodies in developing pea cotyledons. /. Cell Biol. 93:5-14. Christ, B. 1959. Entwicklungsgeschichtliche und physiologische Untersuchungen iiber die Selbststerilitat von Cardamine pratensis L. Z. Bot. 47:88-112.
Christensen, J.E., and Horner, H.T., Jr. 1974. Pollen pore development and its spatial orientation during microsporogenesis in the grass Sorghum bicolor. Am. /. Bot. 61:604-623. Christensen, J.E., Horner, H.T., Jr., and Lersten, N.R. 1972. Pollen wall and tapetal orbicular wall development in Sorghum bicolor (Gramineae). Am. ]. Bot. 59:43-58. Christianson, M.L. 1986. Fate map of the organizing shoot apex in Gossypium. Am. J. Bot. 73:947-958.
Christou, P. 1995. Strategies for variety-independent genetic transformation of important cereals, legumes and woody species utilizing particle bombardment. Euphytica 85:13-27. Christou, P., Ford, T.L., and Kofron, M. 1991. Production of transgenic rice (Oryza sativa L.) plants from agronomically important indica and japonica varieties via electric discharge particle acceleration of exogenous DNA into immature zygotic embryos. Biotechnology 9:957-962.
Chu, C, Liu, H.-T., and Du, R.-H. 1982. Microphotometric determination of DNA contents of early developmental pollen grains in tobacco anther culture. Ada
Choi, J.H., Liu, L.-S., Borkird, C, and Sung, Z.R. 1987. Bot. Sinica 24:1-9. Cloning of genes developmentally regulated during plant embryogenesis. Proc. Natl Acad. Sci. USA Chung, I.-K., Ito, T., Tanaka, H., Ohta, A., Nan, H.G., and 84:1906-1910. Takagi, M. 1994. Molecular diversity of three S-allele Choinski, J.S., Jr., and Trelease, R.N. 1978. Control of enzyme cDNAs associated with gametophytic self-incomactivities in cotton cotyledons during maturation and gerpatibility in Lycopersicon peruvianum. Plant Mol. Biol. mination, n. Glyoxysomal enzyme development in 26:757-762. embryos. Plant Physiol. 62:141-145. Chung, Y.-Y., Kim, S.-R., Kang, H.-G., Noh, Y.-S., Park, Choinski, J.S., Jr., Trelease, R.N., and Doman, D.C. 1981. M.C., Finkel, D., and An, G. 1995. Characterization Control of enzyme activities in cotton cotyledons of two rice MADS box genes homologous to GLOBOSA. Plant Sci. 109:45-56. during maturation and germination. III. In vitro embryo development in the presence of abscisic Ciampolini, R, Cresti, M., and Kapil, R.N. 1983. Fine acid. Planta 152:428-435. structural and cytochemical characteristics of style and stigma in olive. Caryologia 36:211-230. Chojecki, A.J.S., Bayliss, M.W., and Gale, M.D. 1986. Cell production and DNA accumulation in the wheat Ciampolini, F., Moscatelli, A., and Cresti, M. 1988. Ultrastructural features of Aloe ciliaris pollen. I. endosperm, and their association with grain weight. Mature grain and its activation in vitro. Sex. Plant Ann. Bot. 58:809-817. Reprod. 1:88-96. Chourey, P.S. 1981. Genetic control of sucrose synthetase in maize endosperm. Mol. Gen. Genet. 184:372-376. Ciampolini, F., Shivanna, K.R., and Cresti, M. 1991. High humidity and heat stress causes dissociation of Chourey, P.S., and Nelson, O.E. 1976. The enzymatic defiendoplasmic reticulum in tobacco pollen. Bot. Ada ciency conditioned by the shrunken-1 mutations in 104:110-116. maize. Biochem. Genet. 14:1041-1055. Chourey, P.S., and Schwartz, D. 1971. Ethyl methanesul- Ciampolini, F., Cresti, M., Sarfatti, G., and Tiezzi, A. 1981. infrastructure of the stylar canal cells of Citrus limon fonate-induced mutations of the Shj protein in (Rutaceae). Plant Syst. Evol. 138:263-274. maize. Mutation Res. 12:151-157. Chowdhury, J.B., and Das, K. 1968. Cyto-morphological Ciarrocchi, G., Cella, R., and Nielsen, E. 1981. Release of nucleotide-cleaving acid phosphatase from carrot studies on male sterility in Brassica campestris L. cells grown in suspension culture. Physiol. Plant. Cytologia 33:195-199. 53:375-377. Chrispeels, M.J. 1983. The Golgi apparatus mediates the
References
551
ative cells of Solanum (Solanaceae). Am. J. Bot. Cionini, P.G., Bennici, A., Alpi, A., and D'Amato, F. 1976. 64:377-383. Suspensor, gibberellin and in vitro development of Phaseolus coccineus embryos. Planta 131:115-117. Clement, C, and Audran, J.C. 1995. Anther wall layers Clapham, D. 1971. In vitro development of callus from the control pollen sugar nutrition in Lilium. Protoplasma 187:172-181. pollen of Lolium and Hordeum. Z. Pflanzenzuchtg 65:285-292. Clowes, F.A.L. 1978a. Origin of the quiescent centre in Zea Clapham, D., Manders, G., Yibrah, H.S., and von Arnold, mays. New Phytol. 80:409-419. S. 1995. Enhancement of short- and medium-term Clowes, F.A.L. 1978b. Origin of quiescence at the root pole of pea embryos. Ann. Bot. 42:1237-1239. expression of transgenes in embryogenic suspensions of Picea abies (L.) Karst. /. Expt. Bot. 46:655-662.Clutter, M., Brady, T., Walbot, V., and Sussex, I. 1974. Macromolecular synthesis during plant embryoClapham, D.H., and Ostergren, G. 1984. Immunogeny. Cellular rates of RNA synthesis in diploid and cytochemistry of tubulin at meiosis in Tradescantia polytene cells in bean embryos. /. Cell Biol. by a protein-A gold method. Hereditas 101:137-142. 63:1097-1102. Clark, A.J., Higgins, P., Martin, H., and Bowles, D.J. 1991. An embryo-specific protein of barley (Hordeum vul- Cobb, B.G, and Hannah, L.C. 1983. Development of wild type, shrunken-1 and shrunken-2 maize kernels gare). Eur. ). Biochem. 199:115-121. grown in vitro. Theor. Appl. Genet. 65:47-51. Clark, E., Garni, Y., and Izhar, S. 1988. Loss of CMS-specific mitochondrial DNA arrangement in fertile segre- Coca, M.A., Almoguera, C, and Jordano, J. 1994. Expression of sunflower low-molecular-weight heatgants of Petunia hybrids. Plant Mol. Biol. 11:249-253. shock proteins during embryogenesis and persisClark, E.M., Izhar, S., and Hanson, M.R. 1985. tence after germination: localization and possible Independent segregation of the plastid genome and functional implications. Plant Mol. Biol. 25:479^92. cytoplasmic male sterility in Petunia somatic hybrids. Mol. Gen. Genet. 199:440-445. Cochrane, M.P. 1994. Observations on the germ aleurone of barley. Morphology and histochemistry. Ann. Bot. Clark, F.J. 1940. Cytogenetic studies of divergent meiotic 73:113-119. spindle formation in Zea mays. Am. J. Bot. 27:547-559. Clark, J.K., and Sheridan, W.F. 1986. Developmental pro- Cochrane, M.P, and Duffus, CM. 1980. The nucellar projection and modified aleurone in the crease region of files of the maize embryo-lethal mutants dek22 and developing caryopses of barley (Hordeum vulgare L. dek23. J. Hered. 77:83-92. var. distichum). Protoplasma 103:361-375. Clark, J.K., and Sheridan, W.F. 1988. Characterization of the two maize embryo-lethal defective kernel mutants Cochrane, M.P, and Duffus, CM. 1981. Endosperm cell number in barley. Nature 289:399-101. rgh*-1210 andfl*-1253B: effects on embryo and gametophyte development. Genetics 120:279-290. Cocucci, A., and Jensen, W.A. 1969a. Orchid embryology: the mature megagametophyte of Epidendrum scutella. Clark, J.K., and Sheridan, W.F. 1991. Isolation and characKurtziana 5:23-38. terization of 51 embryo-specific mutations of maize. Plant Cell 3:935-951. Cocucci, A., and Jensen, W.A. 1969b. Orchid embryology: pollen tetrads of Epidendrum scutella in the anther Clark, K.R., and Sims, T.L. 1994. The S-ribonuclease gene and on the stigma. Planta 84:215-229. of Petunia hybrida is expressed in nonstylar tissue, including immature anthers. Plant Physiol. Cocucci, A., and Jensen, W.A. 1969c. Orchid embryology: 106:25-36. megagametophyte of Epidendrum scutella following Clark, K.R., Okuley, J.J., Collins, P.D., and Sims, T.L. 1990. fertilization. Am. J. Bot. 56:629-640. Sequence variability and developmental expression Coen, E.S., and Meyerowitz, E.M. 1991. The war of the of S-alleles in self-incompatible and pseudo-selfcomwhorls: genetic interactions controlling flower patible Petunia. Plant Cell 2:815-826. development. Nature 353:31-37. Clark, S.E., Running, M.P., and Meyerowitz, E.M. 1995. Colasanti, J., Tyers, M., and Sundaresan, V. 1991. Isolation CLAVATA3 is a specific regulator of shoot and floral and characterization of cDNA clones encoding a meristem development affecting the same processes functional p34cdc2 homologue from Zea mays. Proc. as CLAVATA1. Development 121:2057-2067. Natl Acad. Sci. USA 88:3377-3381. Clarke, A., Gleeson, P., Harrison, S., and Knox, R.B. 1979. Cole, K. 1959. Inheritance of male sterility in green sproutPollen-stigma interactions: identification and ing broccoli. Can. }. Genet. Cytol. 1:203-207. characterization of surface components with recog- Coleman, C.E., and Kao, T.-H. 1992. The flanking regions of nition potential. Proc. Natl Acad. Sci. USA two Petunia inflata S alleles are heterogeneous 76:3358-3362. and contain repetitive sequences. Plant Mol. Biol. 18:725-737. Clarke, A.E., Abbot, A., Mandel, T.E., and Pettitt, J.M. 1980. Organization of the wall layers of the stigmatic Coleman, C.E., Lopes, M.A., Gillikin, J.W., Boston, R.S., and Larkins, B.A. 1995. A defective signal peptide in papillae of Gladiolus gandavensis: a freeze-fracture study. /. Ultrastr. Res. 73:269-281. the maize high-lysine mutant floury 2. Proc. Natl Clarke, A.E., Considine, J.A., Ward, R., and Knox, R.B. Acad. Sci. USA 92:6828-6831. 1977a. Mechanism of pollination in Gladiolus: roles of Colhoun, C.W., and Steer, M.W. 1981. Microsporogenesis the stigma and pollen-tube guide. Ann. Bot. 41:15-20. and the mechanism of cytoplasmic male sterility in Clarke, A.E., Harrison, S., Knox, R.B., Rapp, J., Smith, P., and maize. Ann. Bot. 48:417-424. Marchalonis, J.P. 1977b. Common antigens and male- Colhoun, C.W., and Steer, M.W. 1983. The cytological female recognition in plants. Nature 265:161-163. effects of the gametocides ethrel and RH-531 on microsporogenesis in barley (Hordeum vulgare L.). Clarke, E., and Steer, M.W. 1983. Cytoplasmic structure of Plant Cell Environ. 6:21-29. germinated and ungerminated pollen grains of Collada, C, Allona, I., Aragoncillo, P., and Aragoncillo, C. Tradescantia virginiana. Caryologia 36:299-305. Clauhs, R.P., and Grun, P. 1977. Changes in plastid and 1993. Development of protein bodies in cotyledons mitochondrion content during maturation of generof Fagus sylvatica. Physiol. Plant. 89:354-359.
552
References
Collins, G.B., and Grosser, J.W. 1984. Culture of embryos. In Conde, M.F., Pring, D.R., Schertz, K.F., and Ross, W.M. Cell Culture and Somatic Cell Genetics of Plants, Vol. 1, 1982. Correlation of mitochondrial DNA restriction ed. I.K. Vasil, pp. 241-257. Orlando: Academic Press. endonuclease patterns with sterility expression in six male-sterile Sorghum cytoplasms. Crop Sci. Collins, G.B., and Sunderland, N. 1974. Pollen-derived 22:536-539. haploids of Nicotiana knightiana, N. raimondii, and N. attenuata. J. Expt. Bot. 25:1030-1035. Condeelis, J.S. 1974. The identification of F actin in the Colombo, L., Franken, J., Koetje, E., van Went, J., Dons, pollen tube and protoplast of Amaryllis belladonna. Expt. Cell Res. 88:435-439. H.J.M., Angenent, G.C., and van Tunen, A.J. 1995. The Petunia MADS box gene FBP11 determines Conger, B.V., Hanning, G.E., Gray, D.J., and McDaniel, ovule identity. Plant Cell 7:1859-1868. J.K. 1983. Direct embryogenesis from mesophyll cells of orchardgrass. Science 221:850-851. Colorado, P., Nicola's, G., and Rodriguez, D. 1995. Unusual sequence and characteristics of a chick-pea Conley, C.A., and Hanson, M.R. 1994. Tissue-specific proseed protein which is regulated by abscisic acid and tein expression in plant mitochondria. Plant Cell is similar to late-embryogenesis-abundant proteins. 6:85-91. Planta 196:622-625. Conley, C.A., Parthasarathy, M.V., and Hanson, M.R. Colorado, P., Nicola's, C, Nicola's, G., and Rodriguez, D. 1994. Effects of Petunia cytoplasmic male sterile 1995. Expression of three ABA-regulated clones and (CMS) cytoplasm on the development of sterile and their relationship to maturation processes during the fertility-restored P. parodii anthers. Am. J. Bot. embryogenesis of chick-pea seeds. Physiol. Plant. 81:630-640. 94:1-6. Connett, M.B., and Hanson, M.R. 1990. Differential mitoColot, V., Bartels, D., Thompson, R., and Flavell, R. 1989. chondrial electron transport through the cyanideMolecular characterization of an active wheat LMW sensitive and cyanide-insensitive pathways in glutenin gene and its relation to other wheat and isonuclear lines of cytoplasmic male sterile, male ferbarley prolamin genes. Mol. Gen. Genet. 216:81-90. tile, and restored Petunia. Plant Physiol. 93:1634-1640. Colot, V., Robert, L.S., Kavanagh, T.A., Bevan, M.W., and Considine, J.A., and Knox, R.B. 1979. Development and histochemistry of the pistil of the grape, Vifis vinifera. Thompson, R.D. 1987. Localization of sequences in Ann. Bot. 43:11-22. wheat endosperm protein genes which confer tissuespecific expression in tobacco. EMBO J. 6:3559-3564. Cook, F.S., and Walden, D.B. 1967. The male gametophyte of Zea mays L. III. The influence of temperature and Comai, L., Baden, C.S., and Harada, J.J. 1989. Deduced calcium on pollen germination and tube growth. sequence of a malate synthase polypeptide encoded Can. ]. Bot. 45:605-613. by a subclass of the gene family. /. Biol. Chem. 264:2778-2782. Cooper, D.C., and Brink, R.A. 1945. Seed collapse following matings between diploid and tetraploid races of Comai, L., and Harada, J.J. 1990. Transcriptional activities in dry seed nuclei indicate the timing of the transiLycopersicon pimpinellifolium. Genetics 30:376-401. tion from embryogeny to germination. Proc. NatI Cooper, P., Ho, T.-H.D., and Hauptmann, R.M. 1984. Tissue specificity of the heat-shock response in Acad. Sci. USA 87:2671-2674. maize. Plant Physiol. 75:431-441. Comai, L., Dietrich, R.A., Maslyar, D.J., Baden, C.S., and Harada, J.J. 1989. Coordinate expression of tran- Cope, F.W. 1962. The mechanism of pollen incompatibilscriptionally regulated isocitrate lyase and malate ity in Theobroma cacao L. Heredity 17:157-182. synthase genes in Brassica napus L. Plant Cell Coppens, L., and Dewitte, D. 1990. Esterase and peroxi1:293-300. dase zymograms from barley (Hordeum vulgare L.) callus as a biochemical marker system of embryogeComai, L., Matsudaira, K.L., Heupel, R.C., Dietrich, R.A., nesis and organogenesis. Plant Sci. 67:97-105. and Harada, J.J. 1992. Expression of a Brassica napus malate synthase gene in transgenic tomato plants Cordewener, J., Booij, H., van der Zandt, H., van Engelen, during the transition from late embryogeny to gerF., van Kammen, A., and de Vries, S. 1991. mination. Plant Physiol. 98:53-61. Tunicamycin-inhibited carrot somatic embryogenesis can be restored by secreted cationic peroxidase Comeau, A., Nadeau, P., Plourde, A., Simard, R., Maes, O., isoenzymes. Planta 184:478^86. Kelly, S., Harper, L, Lettre, J., Landry, B., and St.Pierre, C.-A. 1992. Media for the in ovulo culture of Cordewener, J.H.G., Busink, R., Traas, J.A., Custers, proembryos of wheat and wheat-derived interspeJ.B.M., Dons, H.J.M., and van Lookeren Campagne, cific hybrids or haploids. Plant Sci. 81:117-125. M.M. 1994. Induction of microspore embryogenesis in Brassica napus L. is accompanied by specific Comstock, J.C., Martinson, C.A., and Gengenbach, B.G. changes in protein synthesis. Planta 195:50-56. 1973. Host specificity of a toxin from Phyllosticta maydis for Texas cytoplasmically male-sterile maize. Cordewener, J.H.G., Hause, G., Gorgen, E., Busink, R., Hause, B., Dons, H.J.M., van Lammeren, A.A.M., Phytopathology 63:1357-1361. van Lookeren Campagne, M.M., and Pechan, P. Concei^ao, A. dS., and Krebbers, E. 1994. A cotyledon reg1995. Changes in synthesis and localization of ulatory region is responsible for the different spatial members of the 70-kDa class of heat-shock proteins expression patterns of Arabidopsis 2S albumin genes. Plant ]. 5:493-505. accompany the induction of embryogenesis in Brassica napus L. microspores. Planta 196:7*17-755. Conceicao, A. dS., van Vliet, A., and Krebbers, E. 1994. Unexpectedly higher expression levels of a chimeric Corke, F.M.K., Hedley, C.L., and Wang, T.L. 1990a. An 2S albumin seed protein transgene from a tandem analysis of seed development in Pisum sativum L. XI. array construct. Plant Mol. Biol. 26:1001-1005. Cellular development and the deposition of storage protein in immature embryos grown in vivo and in Concha, I.I., Mansilla, J., Riveros, M., and Burzio, L.O. vitro. Protoplasma 155:127-135. 1995. Ul snRNP components are present in the vegetative and generative nuclei of the pollen grain. Sex. Corke, F.M.K., Hedley, C.L., and Wang, T.L. 1990b. An Plant Reprod. 8:339-344. analysis of seed development in Pisum sativum L. XII.
References
553
In vitro manipulation of embryo development using Craig, S., Goodchild, D.J., and Hardham, A.R. 1979. xenobiotic compounds. Protoplasma 155:136-143. Structural aspects of protein accumulation in develCornish, E.C., Anderson, M.A., and Clarke, A.E. 1988. oping pea cotyledons. I. Qualitative and quantitative Molecular aspects of fertilization in flowering changes in parenchyma cell vacuoles. Aust. J. Plant plants. Annu. Rev. Cell Biol. 4:209-228. Physiol. 6:81-98. Cornish, E.C., Pettitt, J.M., and Clarke, A.E. 1988. Self- Craig, S., Millerd, A., and Goodchild, D.J. 1980. Structural incompatibility genes in flowering plants. In aspects of protein accumulation in developing pea Temporal and Spatial Regulation of Plant Genes, ed. cotyledons. III. Immunocytochemical localization of D.P.S. Verma and R.B. Goldberg, pp. 117-130. New legumin and vicilin using antibodies shown to be York: Springer-Verlag. specific by the enzyme-linked immunosorbent assay (ELISA). Aust. J. Plant Physiol. 7:339-351. Cornish, E.C., Pettitt, J.M., Bonig, I., and Clarke, A.E. 1987. Developmentally controlled expression of a gene Cramer, J.H., Lea, K., and Slightom, J.L. 1985. Expression associated with self-incompatibility in Nicotiana of phaseolin cDNA genes in yeast under control of alata. Nature 326:99-102. natural plant DNA sequences. Proc. Natl Acad. Sci. USA 82:334-338. Corre, E, Henry, Y., Rode, A., and Hartmann, C. 1996. Em gene expression during somatic embryogenesis in Crang, R.E. 1966. A fine structural study of in vivo pollen the monocot Triticum aestivum L. Plant Sci. tube penetration in Lychnis alba. Trans. Am. Micros. 117:139-149. Soc. 85:564-570. Corriveau, J.L., and Coleman, A.W. 1988. Rapid screening Crang, R.E., and Miles, G.B. 1969. An electron microscope method to detect potential biparental inheritance of study of germinating Lychnis alba pollen. Am. }. Bot. plastid DNA and results for over 200 angiosperm 56:398-405. species. Am. ]. Bot. 75:1443-1458. Creech, R.G. 1965. Genetic control of carbohydrate synCorsi, G. 1972. The suspensor of Eruca sativa Miller thesis in maize endosperm. Genetics 52:1175-1186. (Cruciferae) during embryogenesis in vitro. Ciorn. Cremonini, R., and Cionini, P.G. 1977. Extra DNA syntheBot. Hal. 106:41-54. sis in embryo suspensor cells of Phaseolus coccineus. Corsi, G., and Renzoni, G.C. 1972. II genere AHium L. in Protoplasma 91:303-313. Italia. IV. Studio citofotometrico sul granulo Cresti, M., Ciampolini, F., and Kapil, R.N. 1983. pollinico di Allium chamaemoly L. Giorn. Bot. Ital. Ultrastructure of S3 S4 genotype pollen grains of 106:257-264. Oenothera organensis. Ada Bot. Neerl. 32:177-183. Corsi, G., Renzoni, G.C, and Viegi, L. 1973. A DNA Cresti, M., Ciampolini, F., and Kapil, R.N. 1984. Generative cells of some angiosperms with particucytophotometric investigation on the suspensor of lar emphasis on their microtubules. /. Submicros. Eruca sativa Miller. Caryologia 26:531-540. Cytol. 16:317-326. Corti, E.F., and Cecchi, A.F. 1970. The behaviour of the cytoplasm during the megasporogenesis in Cresti, M., Ciampolini, F., and Sansavini, S. 1980. Paphiopedilum spicerianum (Rchb. F.) Pfitzer. Ultrastructural and histochemical features of pistil Caryologia 23:715-727. oiMalus communis: the stylar transmitting tissue. Sci. Hort. 12:327-337. Corti, E.F., and Fiordi, A.C. 1973. Comportement d'une formation cytoplasmatique du me'gasporocyte en Cresti, M., Ciampolini, F., and Sarfatti, G. 1980. Bupleurum. Caryologia 25: Suppl. 43-58. Ultrastructural investigations on Lycopersicum peruCoutos-Thevenot, P., Jouanneau, J.P., Brown, S., Petiard, vianum pollen activation and pollen tube organizaV., and Guern, J. 1990. Embryogenic and nontion after self- and cross-pollination. Planta embryogenic cell lines of Daucus carota cloned from 150:211-217. meristematic cell clusters: relation with cell ploidy Cresti, M., Ciampolini, F., and Tiezzi, A. 1986. determined by flow cytometry. Plant Cell Rep. Ultrastructural studies on Nicotiana tabacum pollen 8:605-608. tubes grown in different culture medium (prelimiCoutos-Thevenot, P., Jouenne, T, Maes, O., Guerbette, F., nary results). Ada Bot. Neerl. 35:285-292. Grosbois, M., le Caer, J.P., Boulay, M., Deloire, A., Cresti, M., Ciampolini, F., and van Went, J.L. 1991. StripKader, J.C., and Guern J. 1993. Four 9-kDa proteins shaped projections at the cytoplasmic face of the excreted by somatic embryos of grapevine are isoouter membrane of the generative cell in Amaryllis forms of lipid transfer proteins. Eur. J. Biochem. belladonna. Ann. Bot. 68:105-107. 217:885-889. Cresti, M., and Keijzer, C.J. 1985. The structure of the Coutos-Thevenot, P., Maes, O., Jouenne, T., Mauro, M.C., endoplasmic reticulum in pollen grains and pollen Boulay, M., Deloire, A., and Guern J. 1992. tubes, after osmium tetroxide-potassium ferriExtracellular protein patterns of grapevine cell suscyanide staining. /. Submicros. Cytol. 17:615-620. pensions in embryogenic and non-embryogenic situ- Cresti, M., Lancelle, S.A., and Hepler, P.K. 1987. Structure ations. Plant Sci. 86:137-145. of the generative cell wall complex after freeze substitution in pollen tubes of Nicotiana and Impatiens. ]. Coxon, D.T., and Davies, D.R. 1982. The effect of the ra and rb loci on the lipid content of the seed of Pisum Cell Sci. 88:373-378. sativum. Theor. Appl. Genet. 64:47-50. Cresti, M., Murgia, M., and Theunis, C.H. 1990. Craig, S., and Goodchild, D.J. 1984a. Periodate-acid treatMicrotubule organization in sperm cells in the ment of sections permits on-grid immunogold localpollen tubes of Brassica oleracea L. Protoplasma 154:151-156. ization of pea seed vicilin in ER and Golgi. Cresti, M., and van Went, J.L. 1976. Callose deposition Protoplasma 122:35-44. and plug formation in Petunia pollen tubes in situ. Craig, S., and Goodchild, D.J. 1984b. Golgi-mediated Planta 133:35-40. vicilin accumulation in pea cotyledon cells is redirected by monensin and by nigericin. Protoplasma Cresti, M., Ciampolini, F., Mulcahy, D.L.M., and Mulcahy, G. 1985. Ultrastructure of Nicotiana alata pollen, its 122:91-97.
554
References
germination and early tube formation. Am. ]. Bot. 72:719-727. Cresti, M., Ciampolini, R, Pacini, E., and Sarfatti, G. 1978a. Phytoferritin in plastids of the style of Olea
Gatehouse, J.A., Shirsat, A.H., Kang, A., Ellis, J.R., Thompson, A., and Boulter, D. 1988. Expression of pea legumin sequences in pea, Nicotiana and yeast. Biochem. Physiol. Pflanz. 183:183-197.
Croy, R.R.D., Gatehouse, J.A., Evans, I.M., and Boulter, D. 1980a. Characterisation of the storage protein subCresti, M., Ciampolini, R, Pacini, E., Sarfatti, G., van units synthesised in vitro by polyribosomes and Went, J.L., and Willemse, M.T.M. 1979. RNA from developing pea (Pisum sativum L.). I. Ultrastructural differences between compatible and Legumin. Planta 148:49-56. incompatible pollen tubes in the stylar transmitting tissue of Petunia hybrida. ]. Submicros. Cytol. Croy, R.R.D., Gatehouse, J.A., Evans, I.M., and Boulter, D. 11:209-219. 1980b. Characterisation of the storage protein subunits synthesised in vitro by polyribosomes and Cresti, M., Ciampolini, R, Pacini, E., Sree Ramulu, K., and RNA from developing pea (Pisum sativum L.). II. Devreux, M. 1978b. Gamma irradiation of Prunus Vicilin. Planta 148:57-63. avium L. flower buds: effects on stylar development - an ultrastructural study. Ada Bot. Neerl. 27:97-106. Croy, R.R.D., Lycett, G.W., Gatehouse, J.A., Yarwood, J.N., and Boulter, D. 1982. Cloning and analysis of cDNAs Cresti, M., Ciampolini, R, van Went, J.L., and Wilms, H.J. encoding plant storage protein precursors. Nature 1982. Ultrastructure and histochemistry of Citrus 295:76-79. limon (L.) stigma. Planta 156:1-9. Cresti, M., Hepler, P.K., Tiezzi, A., and Ciampolini, R Cruz-Alvarez, M., Kirihara, J.A., and Messing, J. 1991. 1986a. Fibrillar structures in Nicotiana pollen: Post-transcriptional regulation of methionine conchanges in ultrastructure during pollen activation tent in maize kernels. Mol. Gen. Genet. 225:331-339. and tube emission. In Biotechnology and Ecology of Cui, X., Wise, R.P., and Schnable, P.S. 1996. The rfl nuclear Pollen, ed. D.L. Mulcahy, G.B. Mulcahy, and E. restorer gene of male-sterile T-cytoplasm maize. Ottaviano, pp. 283-288. New York: Springer-Verlag. Science 272:1334-1336. Cresti, M., Keijzer, C.J., Tiezzi, A., Ciampolini, R, and Cullis, C.A. 1976. Chromatin-bound DNA-dependent Pocardi, P. 1986b. Stigma of Nicotiana: ultrastructural RNA polymerase in developing pea cotyledons. and biochemical studies. Am. j . Bot. 73:1713-1722. Planta 131:293-298. Cresti, M, Milanesi, C, Salvatici, P., and van Aelst, A.C. Cullis, C.A. 1978. Chromatin-bound DNA-dependent RNA polymerase in developing pea cotyledons. II. 1990. Ultrastructural observations of Papaver rhoeas Polymerase activity and template availability under mature pollen grains. Bot. Ada 103:349-354. different growth conditions. Planta 144:57-62. Cresti, M., Milanesi, C, Tiezzi, A., Ciampolini, R, and Moscatelli, A. 1988. Ultrastructure of Linaria vulgaris Cuming, A.C. 1984. Developmental regulation of gene pollen grains. Ada Bot. Neerl. 37:379-386. expression in wheat embryos. Molecular cloning of a DNA sequence encoding the early-methionineCresti, M., Pacini, E., Ciampolini, P., and Sarfatti, G. 1977. labelled (E ) polypeptide. Eur. ]. Biochem. Germination and early tube development in vitro of Lycopersicum peruvianum pollen: ultrastructural fea145:351-357."" tures. Planta 136:239-247. Cummings, D.P., Green, C.E., and Stuthman, D.D. 1976. Cresti, M., van Went, J.L., Pacini, E., and Willemse, M.T.M. Callus induction and plant regeneration in oats. Crop 1976. Ultrastructure of transmitting tissue of Sci. 16:465-470. Lycopersicon peruvianum style: development and his- Cummins, I., and Murphy, D.J. 1992. cDNA sequence of a tochemistry. Planta 132:305-312. sunflower oleosin and transcript tissue specificity. Plant Mol. Biol. 19:873-876. Cre'te, P. 1963. Embryo. In Recent Advances in the Embryology of Angiosperms, ed. P. Maheshwari, pp. Cummins, I., Hills, M.J., Ross, J.H.E., Hobbs, D.H., 171-220. Delhi: International Society of Plant Watson, M.D., and Murphy, D.J. 1993. Differential, Morphologists. temporal and spatial expression of genes involved in storage oil and oleosin accumulation in developing Croissant-Sych, Y, and Okita, T.W. 1996. Identification of rapeseed embryos: implication for the role of positive and negative regulatory cis-elements of the oleosins and the mechanisms of oil-body formation. rice glutelin Gt3 promoter. Plant Sci. 116:27-35. Plant Mol. Biol. 23:1015-1027. Cross, J.W., and Adams, W.R., Jr. 1983. Embryo-specific globulins from Zea mays L. and their subunit compo- Curry, J., Morris, C.R, and Walker-Simmons, M.K. 1991. sition. /. Agr. Pood Chem. 31:534-538. Sequence analysis of a cDNA encoding a group 3 LEA mRNA inducible by ABA or dehydration stress Crossley, S.J., Greenland, A.J., and Dickinson, H.G. 1995. in wheat. Plant Mol. Biol. 16:1073-1076. The characterisation of taperum-specific cDNAs isolated from a Lilium henryi L. meiocyte subtractive Custers, J.B.M., and Bergervoet, J.H.W. 1990. In vitro culture of embryos of Cucumis spp.: heart-stage cDNA library. Planta 196:523-529. embryos have a higher ability of direct plant formaCrouch, M.L. 1982. Non-zygotic embryos of Brassica napus tion than advanced-stage embryos. Sex. Plant Reprod. L. contain embryo-specific storage proteins. Planta 3:152-159. 156:520-524. Crouch, M.L., and Sussex, I.M. 1981. Development and Custers, J.B.M., Cordewener, J.H.G., No'llen, Y, Dons, H.J.M., and van Lookeren Campagne, M.M. 1994. storage-protein synthesis in Brassica napus L. Temperature controls both gametophytic and sporoembryos in vivo and in vitro. Planta 153:64—74. phytic development in microspore cultures of Crouch, M.L., Tenbarge, K.M., Simon, A.E., and Ferl, R. Brassica napus. Plant Cell Rep. 13:267-271. 1983. cDNA clones for Brassica napus seed storage proteins: evidence from nucleotide sequence analy- Cyr, R.J., Bustos, M.M., Guiltinan, M.J., and Posket, D.E. 1987. Developmental modulation of rubulin protein sis that both subunits of napin are cleaved from a and mRNA levels during somatic embryogenesis in precursor polypeptide. /. Mol. Appl. Genet. 2:273-283. cultured carrot cells. Planta 171:365-376. Croy, R.R.D., Evans, I.M., Yarwood, J.N., Harris, N., europaea L. Ada Bot. Neerl. 27:417-423.
555
References
Czak6, M., Jang, J.-C, Herr, J.M., Jr., and Marton, L. 1992. Davey, J.E., and van Staden, J. 1978. Ultrastructural Differential manifestation of seed mortality induced aspects of reserve protein deposition during cotyleby seed-specific expression of the gene for diphthedonary cell development in Lupinus albus. 2. ria toxin A chain in Arabidopsis and tobacco. Mol. Pflanzenphysiol. 89:259-271. Gen. Genet. 235:33-40. Davies, D.R. 1980. The ra locus and legumin synthesis in Pisum sativum. Biochem. Genet. 18:1207-1219.
D'Alascio-Deschamps, R. 1972. Le sac embryonnaire du lin apres la fecondation. Botaniste 50:273-288. D'Alascio-Deschamps, R. 1973. Organisation du sac embryonnaire du Linum catharticum L., espece recoltee en station naturelle; 6tude ultrastructurale.
Davies, D.R., and Bedford, I.D. 1982. Abscisic acid and storage protein accumulation in Pisum sativum embryos grown in vitro. Plant Sci. Lett. 27:337-343. Davies, D.R., and Brewster, V. 1975. Studies of seed development in Pisum sativum. II. Ribosomal RNA conBull. Soc. Bot. Fr. 120:189-200. tents in reciprocal crosses. Planta 124:303-309. D'Alascio-Deschamps, R. 1981. Embryologie du Linum Davies, H.M., and Delmer, D.P. 1981. Two kinds of protein catharticum L. Le zygote: £tude ultrastructurale. Bull. glycosylation in a cell-free preparation from developing cotyledons of Phaseolus vulgaris. Plant Physiol. Soc. Bot. Fr. Lett. Bot. 128:269-278. D'Amato, F. 1984. Role of polyploidy in reproductive 68:284-291. organs and tissues. In Embryology of Angiosperms, ed. Davies, M.D., and Dickinson, D.B. 1971. Effects of freezeB.M. Johri, pp. 519-566. Berlin: Springer-Verlag. drying on permeability and respiration of germinatD'Amato, F., Devreux, M., and Scarascia Mugnozza, G.T. ing lily pollen. Physiol. Plant. 24:5-9. 1965. The DNA content of the nuclei of the pollen Davies, M.D., and Dickinson, D.B. 1972. Properties of urigrains in tobacco and barley. Caryologia 18:377-382. dine diphosphoglucose dehydrogenase from pollen of Lilium longiflorum. Arch. Biochem. Biophys. Dahlgren, K.V.O. 1934. Die Embryologie von Impatiens 152:53-61. roylei. Svensk. Bot. Tidskr. 28:103-125. Dahmer, M.L., Hildebrand, D.F., and Collins, G.B. 1992. Davis, B.D. 1983. Growth of excised pea embryonic axes Comparative protein accumulation patterns in soyon different sugars. Am. J. Bot. 70:816-820. bean somatic and zygotic embryos. In Vitro Cell Davis, R.W., Smith, J.D., and Cobb, B.G. 1990. A light and Devel Biol. 28P:106-114. electron microscope investigation of the transfer cell region of maize caryopses. Can. ]. Bot. 68:471-479. Dale, P.J. 1975. Pollen dimorphism and anther culture in Dawe, R.K., and Freeling, M. 1990. Clonal analysis of the barley. Planta 127:213-220. cell lineages in the male flower of maize. Devel Biol. Dale, P.J., and Deambrogio, E. 1979. A comparison of cal142:233-245. lus induction and plant regeneration from different explants of Hordeum vulgare. Z. Pflanzenphysiol. Dawe, R.K., and Freeling, M. 1992. The role of initial cells in maize anther morphogenesis. Development 94:65-77. 116:1077-1085. Dannenhoffer, J.M., Bostwick, D.E., Or, E., and Larkins, B.A. 1995. Opaque-15, a maize mutation with prop- Dawe, R.K., Lachmansingh, A.R., and Freeling, M. 1993. Transposon-mediated mutations in the untranslated erties of a defective opaque-2 modifier. Proc. Natl Acad. Set USA 92:1931-1935. leader of maize Adhl that increase and decrease pollen-specific gene expression. Plant Cell 5:311-319. Das, N.K. 1965. Inactivation of the nucleolar apparatus during meiotic prophase in corn anthers. Expt. Cell Dawe, R.K., Sedat, J.W., Agard, D.A., and Cande, W.Z. Res. 40:360-364. 1994. Meiotic chromosome pairing in maize is associated with a novel chromatin organization. Cell Das, O.P., and Messing, J.W. 1987. Allelic variation and 76:901-912. differential expression at the 27-kilodalton zein locus in maize. Mol. Cell. Biol. 7:4490^497. Dawson, J., Wilson, Z.A., Aarts, M.G.M., Braithwaite, A.F., Briarty, L.G., and Mulligan, B.J. 1993. Dasgupta, J., and Bewley, J.D. 1982. Desiccation of axes of Microspore and pollen development in six malePhaseolus vulgaris during development of a switch sterile mutants of Arabidopsis thaliana. Can. J. Bot. from a development pattern of protein synthesis to a 71:629-638. germination pattern. Plant Physiol. 70:1224-1227. Dashek, W.V., Erickson, S.S., Hayward, D.M., Lindbeck, De, D.N. 1961. Autoradiographic studies of nucleoprotein G., and Mills, R.R. 1979. Peroxidase in cytoplasm metabolism during the division cycle. Nucleus and cell wall of germinating lily pollen. Bot. Gaz. 4:1-24. 140:261-265. de Barros, E.G., Takasaki, K., Kirleis, A.W., and Larkins, Dashek, W.V., and Harwood, H.I. 1974. Proline, hydroxB.A. 1991. Nucleotide sequence of a cDNA clone yproline, and lily pollen tube elongation. Ann. Bot. encoding y-kafirin protein from Sorghum bicolor. 38:947-959. Plant Physiol. 97:1606-1607. Dashek, W.V., and Rosen, W.G. 1966. Electron microscop- de Block, M., and Debrouwer, D. 1993. Engineered fertilical localization of chemical components in the ity control in transgenic Brassica napus L.: histochemgrowth zone of lily pollen tubes. Protoplasma ical analysis of anther development. Planta 61:192-204. 189:218-225. Dashek, W.V., Thomas, H.R., and Rosen, W.G. 1971. de Boer-de Jeu, M.J. 1978. Megasporogenesis. A comparaSecretory cells of lily pistils. II. Electron microscope tive study of the ultrastructural aspects of megascytochemistry of canal cells. Am. }. Bot. 58:909-920. porogenesis in Lilium, Allium and Impatiens. Meded. Landboumhoges. Wageningen 78-16:1-127. Datta, S.K. 1987. Plant regeneration by pollen embryogenesis from cultured whole spikes of barley (Hordeum de Castro, L.A.B., Lacerda, Z., Aramayo, R.A., Sampaio, vulgare). Theor. Appl. Genet. 74:121-124. M.J.A.M., and Gander, E.S. 1987. Evidence for a preDauphin-Guerin, B., Teller, G., and Durand, D. 1980. cursor molecule of Brazil nut 2S seed proteins from Different endogenous cytokinins between male and biosynthesis and cDNA analysis. Mol. Gen. Genet. female Mercurialis annua L. Planta 148:124-129. 206:338-343.
556
References
de Clercq, A., Vandewiele, M., de Rycke, R., van Damme, de Silva, J., Loader, N.M., Jarman, C, Windust, J.H.C., J., van Montagu, M., Krebbers, E., and Hughes, S.G., and Safford, R. 1990. The isolation and Vandekerckhove, J. 1990a. Expression and processsequence analysis of two seed-expressed acyl carrier ing of an Arabidopsis 2S albumin in transgenic protein genes from Brassica napus. Plant Mol. Biol. tobacco. Plant Physiol. 92:899-907. 14:537-548. de Clercq, A., Vandewiele, M., van Damme, J., Guerche, de Vries, A.P., and Ie, T.S. 1970. Electron-microscopy on P., van Montagu, M., Vandekerckhove, J., and anther tissue and pollen of male sterile and fertile Krebbers, E. 1990b. Stable accumulation of modified wheat {Triticum aestivum L.). Euphytica 19:103-120. 2S albumin seed storage proteins with higher de Vries, S.C, Booij, H., Janssens, R., Vogels, R., Saris, L., methionine contents in transgenic plants. Plant Lo Schiavo, F., Terzi, M., and van Kammen, A. 1988a. Physiol. 94:970-979. Carrot somatic embryogenesis depends on the phytohormone-controlled presence of correctly glycosyde Freitas, F.A., Yunes, J.A., da Silva, M.J., Arruda, P., and lated extracellular proteins. Genes Devel. 2:462-476. Leite, A. 1994. Structural characterization and promoter activity analysis of the y-kafirin gene from de Vries, S.C, Booij, H., Meyerink, P., Huisman, G., Wilde, sorghum. Mol. Gen. Genet. 245:177-186. H.D., Thomas, T.L., and van Kammen, A. 1988b. Acquisition of embryogenic potential in carrot de Jong, A.J., Schmidt, E.D.L., and de Vries, S.C. 1993. cell-suspension cultures. Planta 176:196-204. Early events in higher-plant embryogenesis. Plant Mol. Biol. 22:367-377. de Wet, J.M.J., Bergquist, R.R., Harlan, J.R., Brink, D.E., de Jong, A.J., Cordewener, J., Lo Schiavo, R, Terzi, M., Cohen, C.E., Newell, C.A., and de Wet, A.E. 1985. Vandekerckhove, J., van Kammen, A., and de Vries, Exogenous gene transfer in maize (Zea mays) using S.C. 1992. A carrot somatic embryo mutant is resDNA-treated pollen. In The Experimental Manipulation of Ovule Tissues, ed. G.P. Chapman, S.H. cued by chitinase. Plant Cell 4:425^33. Mantell, and R.W. Daniels, pp. 197-209. New York: de Jong, A.J., Heidstra, R., Spaink, H.P., Hartog, M.V., Longman. Meijer, E.A., Hendriks, T., Lo Schiavo, F., Terzi, M., Bisseling, T., van Kammen, A., and de Vries, S.C. Degenhardt, J., Fiebig, C, and Link, G. 1991. Chloroplast 1993. Rhizobium lipooligosaccharides rescue a carrot and nuclear transcripts for plastid proteins in somatic embryo mutant. Plant Cell 5:615-620. Arabidopsis thaliana: tissue distribution in mature plants and during seedling development and de Jong, A.J., Hendriks, T., Meijer, E.J., Penning, M., Lo embryogenesis. Bot. Ada 104:455-463. Schiavo, E, Terzi, M., van Kammen, A., and de Vries, S.C. 1995. Transient reduction in secreted 32 kD chiti- Del Casino, C, Li, Y.-Q., Moscatelli, A., Scali, M., Tiezzi, nase prevents somatic embryogenesis in the carrot A., and Cresti, M. 1993. Distribution of microtubules (Daucus carota L.) variant tsll. Bevel Genet. 16:332-343. during the growth of tobacco pollen tubes. Biol. Cell. 79:125-132. de la Pefia, A. 1986. 7« vitro' culture of isolated meiocytes of rye, Secale cereale L. Environ. Expt. Bot. 26:17-23. Del Casino, C, Tiezzi, A., Wagner, V.T., and Cresti, M. de la Pefia, A., Lorz, H., and Schell, J. 1987. Transgenic rye 1992. The organization of the cytoskeleton in the plants obtained by injecting DNA into young floral generative cell and sperms of Hyacinthus orientalis. tillers. Nature 325:274-276. Protoplasma 168:41-50. de Mey, J., Lambert, A.M., Bajer, A.S., Moeremans, M., Del Rosario, A.G., and de Guzman, E.V. 1976. The growth of coconut "Makapuno" embryos in vitro as affected and de Brabander, M. 1982. Visualization of microtubules in interphase and mitotic plant cells of by mineral composition and sugar level of the Haemanthus endosperm with immuno-gold staining medium during the liquid and solid cultures. Philipp. J. Sci. 105:215-222. method. Proc. Natl Acad. Sci. USA 79:1898-1902. de Nettancourt, D. 1977. Incompatibility in Angiosperms. Delannay, X. 1979. Evolution of male sterility mechanisms in gynodioecious and dioecious species of Cirsium Berlin: Springer-Verlag. (Cynareae, Compositae). Plant Syst. Evol. de Nettancourt, D., Devreux, M., Bozzini, A., Cresti, M., 132:327-332. Pacini, E., and Sarfatti, G. 1973a. Ultrastructural aspects of the self-incompatibility mechanism in Delannay, X., and Palmer, R.G. 1982. Genetics and cytolLycopersicum peruvianum Mill. /. Cell Sci. 12:403—419. ogy of the mst male-sterile soybean. /. Hered. 73:219-223. de Nettancourt, D., Devreux, M., Laneri, U., Cresti, M., Pacini, E., and Sarfatti, G. 1974. Genetical and ultra- Delbreil, B., Guerche, P., and Jullien, M. 1993. Agrobacterium-mediated transformation of Asparagus structural aspects of self and cross incompatibility in interspecific hybrids between self-compatible officinalis L. long-term embryogenic callus and Lycopersicum esculentum and self-incompatible L. regeneration of transgenic plants. Plant Cell Rep. peruvianum. Theor. Appl. Genet. 44:278-288. 12:129-132. de Nettancourt, D., Devreux, M., Laneri, U., Pacini, E., DeLisle, A.J., and Crouch, M.L. 1989. Seed storage protein transcription and mRNA levels in Brassicfa napus Cresti, M., and Sarfatti, G. 1973b. Ultrastructural during development and in response to exogenous aspects of unilateral interspecific incompatibility abscisic acid. Plant Physiol. 91:617-623. between Lycopersicum peruvianum and L. esculentum. Caryologia 25: Suppl. 207-217. DeLong, A., Calderon-Urrea, A., and Dellaporta, S.L. 1993. Sex determination gene TASSELSEED2 of de Paepe, R., Forchioni, A., Chetrit, P., and Vedel, F. 1993. Specific mitochondrial proteins in pollen: presence maize encodes a short-chain alcohol dehydrogenase of an additional ATP synthase (3-subunit. Proc. Natl required for stage-specific floral organ abortion. Cell Acad. Sci. USA 90:5934-5938. 74:757-768. de Pater, S., Pham, K., Chua, N.-H., Memelink, J., and Delorme, V, Giranton, J.-L., Hatzfeld, Y, Friry, A., Kijne, J. 1993. A 22-bp fragment of the pea lectin proHeizmann, P., Ariza, M.J., Dumas, C, Gaude, T., and moter containing essential TGAC-like motifs confers Cock, J.M. 1995. Characterization of the S locus seed-specific gene expression. Plant Cell 5:877-886. genes, SLG and SRK, of the Brassica S3 haplotype:
References
557
Cappadocia, M. 1994. Molecular cloning of two identification of a membrane-localized protein Solanum chacoense S-alleles and a hypothesis conencoded by the S locus receptor kinase gene. Plant ]. 7:429-440. cerning their evolution. Sex. Plant Reprod. 7:169-176. Delvallee, I., and Dumas, C. 1988. Anther development in Desprez, B., Chupeau, M.-C, Vermeulen, A., Delbreil, B., Zea mays: changes in protein, peroxidase, and Chupeau, Y., and Bourgin, J.-P. 1995. Regeneration esterase patterns. /. Plant Physiol. 132:210-217. and characterization of plants produced from mature tobacco pollen protoplasts via gametosoDeMason, D.A. 1986. Endosperm structure and storage matic hybridization. Plant Cell Rep. 14:204-209. reserve histochemistry in the palm, Washingtonia filifera. Am. J. Bot. 73:1332-1340. Detchepare, S., Heizmann, P., and Dumas, C. 1989. Changes in protein patterns and protein synthesis DeMason, D.A., and Chandra Sekhar, K.N. 1988. The during anther development in Brassica oleracea. j . breeding system in the date palm (Phoenix dactylifera Plant Physiol. 135:129-137. L.) and its recognition by early cultivators. Adv. Econ. Bot. 6:20-35. Deurenberg, J.J.M. 1976. In vitro protein synthesis with DeMason, D.A., Chandra Sekhar, K.N., and Harris, M. polysomes from unpollinated, cross- and self-polli1989. Endosperm development in the date palm nated Petunia ovaries. Planta 128:29-33. {Phoenix dactylifera) (Arecaceae). Am. J. Bot. Deurenberg, J.J.M. 1977. Differentiated protein synthesis 76:1255-1265. with polysomes from Petunia ovaries before fertilizaDeMason, D.A., Sexton, R., and Grant Reid, J.S. 1983. tion. Planta 133:201-206. Structure, composition and physiological state of the DeVerno, L.L., Charest, P.J., and Bonen, L. 1994. endosperm of Phoenix dactylifera L. Ann. Bot. Mitochondrial DNA variation in somatic embryo52:71-80. genie cultures of Larix. Theor. Appl. Genet. 88:727-732. Demerec, M. 1923. Heritable characters of maize. XV. Devi, P.M. 1964. Heterostyly in Biophytum sensitivum DC. Germless seeds. /. Hered. 14:297-300. /. Genet. 59:41-48. Demerec, M. 1924. A case of pollen dimorphism in maize. Devic, M., Albert, S., and Delseny, M. 1996. Induction and Am. ]. Bot. 11:461-464. expression of seed-specific promoters in Arabidopsis Deng, W., Grayburn, W.S., Hamilton-Kemp, T.R., Collins, embryo-defective mutants. Plant J. 9:205-215. G.B., and Hildebrand, D.F. 1992. Expression of soy- Devreux, M. 1963. Effets de l'irradiation gamma bean-embryo lipoxygenase 2 in transgenic tobacco chronique sur l'embryogenese de Capsella bursa-pastissue. Planta 187:203-208. toris Moench. In L'Energia Nucleare in Agricoltura, VI Denis, M., Delourme, R., Gourret, J.-P., Mariani, C, and Congr. Nud. (Roma), pp. 199-217. Vallecchi: Comitato Renard, M. 1993. Expression of engineered nuclear Nazionale Energia Nucleare. male sterility in Brassica napus. Plant Physiol. Devreux, M., and Scarascia Mugnozza, G.T. 1962. Action 101:1295-1304. des rayons gamma sur les premiers stades de develDepigny-This, D., Raynal, M., Aspart, L., Delseny, M., and oppement de l'embryon de Nicotiana rustica L. Caryologia 15:279-291. Grellet, F. 1992. The cruciferin gene family in radish. Plant Mol. Bid. 20:467-479. Dewey, R.E., Levings, C.S., III, and Timothy, D.H. 1986. Novel recombinations in the maize mitochondrial Derksen, J., Pierson, E.S., and Traas, J.A. 1985. genome produce a unique transcriptional unit in the Microtubules in vegetative and generative cells of Texas male-sterile cytoplasm. Cell 44:439-449. pollen tubes. Eur. J. Cell Biol. 38:142-148. Derksen, J., Rutten, T., van Amstel, T., de Win, A., Doris, Dewey, R.E., Timothy, D.H., and Levings, C.S., III. 1987. A F., and Steer, M. 1995. Regulation of pollen tube mitochondrial protein associated with cytoplasmic growth. Ada Bot. Neerl. 44:93-119. male sterility in the T cytoplasm of maize. Proc. Natl Acad. Sci. USA 84:5374-5378. DeRocher, A., and Vierling, E. 1995. Cytoplasmic HSP70 homologues of pea: differential expression in vege- Dewey, R.E., Siedow, J.N., Timothy, D.H., and Levings, tative and embryonic organs. Plant Mol. Biol. C.S., III. 1988. A 13-kilodalton maize mitochondrial 27-M1-A56. protein in E. coli confers sensitivity to Bipolaris mayDeRocher, A.E., and Vierling, E. 1994. Developmental dis toxin. Science 239:293-295. control of small heat shock protein expression dur- DeWitt, N.D., Harper, J.F., and Sussman, M.R. 1991. ing pea seed maturation. Plant J. 5:93-102. Evidence for a plasma membrane proton pump in phloem cells of higher plants. Plant ]. 1:121-128. DeRose, R.T., Ma, D.-P., Kwon, l.-S., Hasnain, S.E., Klassy, R.C., and Hall, T.C. 1989. Characterization of the Dexheimer, J. 1965. Sur les structures cytoplasmiques dans les grains de pollen de Lobelia erinus. Compt. kafirin gene family from sorghum reveals extensive homology with zein from maize. Plant Mol. Biol. Rend. Acad. Sci. Paris 260:6963-6965. 12:245-256. Dexheimer, J. 1968. Sur la synthese d'acide ribonucleique Desborough, S., and Peloquin, SJ. 1968. Disc-elecpar les tubes polliniques en croissance. Compt. Rend. Acad. Sci. Paris 267D:2126-2128. trophoresis of proteins and enzymes from styles, pollen and pollen tubes of self-incompatible culti- Dexheimer, J. 1970. Recherches cytophysiologiques sur les vars of Lilium longiflorum. Theor. Appl. Genet. grains de pollen. Rev. Cytol. Biol. Veget. 33:169-233. 38:327-331. Dexheimer, J. 1972. Etude experimental des grains de Deschamps, R. 1969. Premiers stades du developpement pollen des angiospermes. Rev. Cytol. Biol. Veyet. de l'embryon et de l'albumen du lin: etude au micro35:17-39. scope electronique. Rev. Cytol. Biol. Veget. Dhaliwal, A.S., and Malik, C.P. 1982. Localization of some 32:379-390. hydrolases in the pollen and stigma of Brassica Deshusses, ]., Gumber, S.C, and Loewus, F.A. 1981. Sugar campestris following self- and cross-pollination. uptake in lily pollen. A proton symport. Plant Phytomorphology 32:37-41. Physiol. 67:793-796. Dhaliwal, A.S., Malik, C.P, and Singh, M.B. 1981. Despres, C, Saba-El-Leil, M., Rivard, S.R., Morse, D., and Overcoming incompatibility in Brassica campestris L.
558
References
by carbon dioxide, and dark fixation of the gas by Dickinson, H.G. 1981. The structure and chemistry of self- and cross-pollinated pistils. Ann. Bot. plastid differentiation during male meiosis in Lilium 48:227-233. henryi. J. Cell Sci. 52:223-241. D'Halluin, K., Bonne, E., Bossut, M., de Beuckeleer, M., Dickinson, H.G. 1990. Self-incompatibility in flowering and Leemans, J. 1992. Transgenic maize plants by tisplants. Bioessays 12:155-161. sue electroporation. Plant Cell 4:1495-1505. Dickinson, H.G., and Andrews, L. 1977. The r61e of memDhillon, S.S., and Miksche, J.P. 1983. DNA, RNA, protein brane-bound cytoplasmic inclusions during gametoand heterochromatin changes during embryo develgenesis in Lilium longiflorum Thumb. Planta opment and germination of soybean (Glycine max 134:229-240. L.). Histochem. J. 15:21-37. Dickinson, H.G., and Bell, PR. 1970. Nucleocytoplasmic interaction at the nuclear envelope in post meiotic di Fonzo, N., Gentinetta, E., Salamini, R, and Soave, C. microspores of Pinus banksiana. ]. Ultrastr. Res. 1979. Action of the opaque-7 mutation on the accu33:356-359. mulation of storage products in maize endosperm. Plant Sci. Lett. 14:345-354. Dickinson, H.G., and Bell, PR. 1972a. The role of the tapeturn in the formation of sporopollenin-containing di Fonzo, N., Hartings, H., Brembilla, M., Motto, M., structures during microsporogenesis in Pinus Soave, C, Navarro, E., Palau, J., Rhode, W., and banksiana. Planta 107:205-215. Salamini, F. 1988. The b-32 protein from maize endosperm, an albumin regulated by the 02 locus: Dickinson, H.G., and Bell, PR. 1972b. Structures resemnucleic acid (cDNA) and amino acid sequences. Mol. bling nuclear pores at the orifice of nuclear invagiGen. Genet. 212:481-487. nations in developing microspores of Pinus banksiana. Devel Biol. 27:425-429. di Fonzo, N., Manzocchi, L., Salamini, F., and Soave, C. 1986. Purification and properties of an endospermic Dickinson, H.G., and Bell, PR. 1976. The changes in the protein of maize associated with the opaque-2 and tapetum of Pinus banksiana accompanying formation opaque-6 genes. Planta 167:587-594. and maturation of the pollen. Ann. Bot. 40:1101-1109. Diboll, A.G. 1968. Fine structural development of the Dickinson, H.G., and Elleman. C.J. 1985. Structural changes in the pollen grain of Brassica oleracea during megagametophyte of Zea mays following fertilizadehydration in the anther and development on the tion. Am. ]. Bot. 55:797-806. stigma as revealed by anhydrous fixation techDiboll, A.G., and Larson, D.A. 1966. An electron microniques. Micron Micros. Ada 16:255-270. scopic study of the mature megagametophyte in Zea mays. Am. J. Bot. 53:391^02. Dickinson, H.G., and Heslop-Harrison, J. 1968. Common mode of deposition for the sporopollenin of sexine Dickinson, CD., Evans, R.P., and Nielsen, N.C. 1988. RY and nexine. Nature 220:926-927. repeats are conserved in the 5 -flanking regions of legume seed-protein genes. Nucl. Acids Res. 16:371. Dickinson, H.G., and Heslop-Harrison, J. 1970a. The behaviour of plastids during meiosis in the Dickinson, CD., Hussein, E.H.A., and Nielsen, N.C. 1989. microsporocyte of Lilium longiflorum Thunb. Cytobios Role of posttranslational cleavage in glycinin assem2:103-118. bly. Plant Cell 1:459-469. Dickinson, D.B. 1965. Germination of lily pollen: respira- Dickinson, H.G., and Heslop-Harrison, J. 1970b. The ribosome cycle, nucleoli, and cytoplasmic nucleoloids in tion and tube growth. Science 150:1818-1819. the meiocytes of Lilium. Protoplasma 69:187-200. Dickinson, D.B. 1966. Inhibition of pollen respiration by Dickinson, H.G., and Heslop-Harrison, J. 1971. The mode oligomycin. Nature 210:1362-1363. of growth of the inner layer of the pollen-grain exine Dickinson, D.B. 1967. Permeability and respiratory propin Lilium. Cytobios 4:233-243. erties of germinating pollen. Physiol. Plant. Dickinson, H.G., and Heslop-Harrison, J. 1977. 20:118-127. Ribosomes, membranes and organelles during meioDickinson, D.B., and Davies, M.D. 1971. Nucleoside sis in angiosperms. Phil. Trans. Roy. Soc. Lond. diphosphate kinase from lily pollen. Plant Cell 277B:327-342. Physiol. 12:157-160. Dickinson, H.G. 1970a. Ultrastructural aspects of primex- Dickinson, H.G., and Lawson, J. 1975. Pollen tube growth ine formation in the microspore tetrad of Lilium in the stigma of Oenothera organensis following comlongiflorum. Cytobiologie 1:437-449. patible and incompatible intraspecific pollinations. Dickinson, H.G. 1970b. Membrane investing the Proc. Roy. Soc. Lond. 188B:327-344. microsporangium of Pinus banksiana. New Phytol. Dickinson, H.G., and Lewis, D. 1973a. Cytochemical and 69:1065-1068. ultrastructural differences between intraspecific Dickinson, H.G. 1971a. The role played by sporopollenin compatible and incompatible pollinations in Raphanus. Proc. Roy. Soc. Lond. 183B:21-38. in the development of pollen in Pinus banksiana. In Sporopollenin, ed. J. Brooks, PR. Grant, M. Muir, P. Dickinson, H.G., and Lewis, D. 1973b. The formation of the tryphine coating the pollen grains of Raphanus, van Gijzel, and G. Shaw, pp. 31-67. London: and its properties relating to the self-incompatibility Academic Press. system. Proc. Roy. Soc. Lond. 184B:149-165. Dickinson, H.G. 1971b. Nucleo-cytoplasmic interaction following meiosis in the young microspores of Dickinson, H.G., and Lewis, D. 1974. Changes in the pollen grain wall of Linum grandiflorum following Lilium longiflorum; events at the nuclear envelope. compatible and incompatible intraspecific pollinaGrana 11:117-127. tions. Ann. Bot. 38:23-29. Dickinson, H.G. 1973. The role of plastids in the formation Dickinson, H.G., Moriarty, J., and Lawson, J. 1982. of pollen grain coatings. Cytobios 8:25-40. Pollen-pistil interaction in Lilium longiflorum: the Dickinson, H.G. 1976. Common factors in exine deposirole of the pistil in controlling pollen tube growth tion. In The Evolutionary Significance of the Exine, ed. following cross- and self-pollinations. Proc. Roy. Soc. I.K. Ferguson and J. Muller, pp. 67-89. London: Lond. 215B:45-62. Academic Press.
References
559
Dickinson, H.G., and Potter, U. 1975. Post meiotic expression and cytoplasmic male sterility in Sorghum. Plant Mol. Biol. 2:89-102. nucleo-cytoplasmic interaction in Pinus banksiana: the secretion of RNA by the nucleus. Planta Dixon, L.K., Leaver, C.J., Brettell, R.I.S., and Gengenbach, 122:99-104. B.G. 1982. Mitochondrial sensitivity to Drechslera maydis T-toxin and the synthesis of a variant mitoDickinson, H.G., and Potter, U. 1976. The development of chondrial polypeptide in plants derived from maize patterning in the alveolar sexine of Cosmos bipinnatissue cultures with Texas male-sterile cytoplasm. tus. New Phytol. 76:543-550. Theor. Appl. Genet. 63:75-80. Dickinson, H.G., and Potter, U. 1978. Cytoplasmic changes accompanying the female meiosis in Lilium Dobrofsky, S., and Grant, W.F. 1980. Electrophoretic evilongiflorum Thunb. /. Cell Sci. 29:147-169. dence supporting self-incompatibility in Lotus corniculatus. Can. }. Bot. 58:712-716. Dickinson, H.G., and Potter, U. 1979. Post-meiotic nucleo-cytoplasmic interaction in Cosmos bipinnatus. Dodds, K.S., and Simmonds, N.W. 1946. A cytological basis of sterility in Tripsacum laxum. Ann. Bot. Early events at the nuclear envelope. Planta 10:109-116. 145:449-457. Dickinson, H.G., and Sheldon, J.M. 1984. A radial system Dodds, P.N., Clarke, A.E., and Newbigin, E. 1996. A molecular perspective on pollination in flowering of microtubules extending between the nuclear plants. Cell 85:141-144. envelope and the plasma membrane during early male haplophase in flowering plants. Planta Dodds, P.N., Bonig, I., Du, H., Rodin, J., Anderson, M.A., 161:86-90. Newbigin, E., and Clarke, A.E. 1993. S-RNase gene of Nicotiana alata is expressed in developing pollen. Dickinson, H.G., and Willson, C. 1983. Two stages in the redifferentiation of amyloplasts in the microspores Plant Cell 5:1771-1782. of Lilium. Ann. Bot. 52:803-810. Dolfini, S.F., and Sparvoli, F. 1988. Cytological characteriDickinson, H.G., and Willson, C. 1985. Behaviour of zation of the embryo-lethal mutant dek-1 of maize. nucleoli and cytoplasmic nucleoloids during meiotic Protoplasma 144:142-148. divisions in Lilium henryi. Cytobios 43:349-365. Dolfini, S.F., Landoni, M., Tonelli, C, Bernard, L., and Dieckert, J.W., and Dieckert, M.C. 1976. The chemistry Viotti, A. 1992. Spatial regulation in the expression of and cell biology of the vacuolar proteins of seeds. /. structural and regulatory storage-protein genes in Food Sci. 41:475-482. Zea mays endosperm. Devel Genet. 13:264-276. Dierks-Ventling, C. 1981. Storage proteins in Zea mays (L.): Domon, C, and Steinmetz, A. 1994. Exon shuffling in anther-specific genes from sunflower. Mol. Gen. interrelationship of albumins, globulins and zeins in Genet. 244:312-317. the opaque-2 mutation. Eur. ]. Biochem. 120:177-182. Diers, L. 1963. Elektronenmikroskopische Beobachtungen Domon, C, Evrard, J.-L., Herdenberger, F., Pillay, D.T.N., an der generativen Zelle von Oenothera hookeri Torr. and Steinmetz, A. 1990. Nucleotide sequence of two et Gray. Z. Naturfors. 18b:562-566. anther-specific cDNAs from sunflower (Helianthus Dieterich, K. 1924. Uber Kultur von Embryonen annuus L.). Plant Mol. Biol. 15:643-646. aufierhalb des Samens. Flora 117:379^17. Domon, C, Evrard, J.-L., Pillay, D.T.N., and Steinmetz, A. Dietrich, P.S., Bouchard, R.A., Casey, E.S., and Sinibaldi, 1991. A 2.6 kb intron separates the signal peptide R.M. 1991. Isolation and characterization of a small coding sequence of an anther-specific protein from heat shock protein gene from maize. Plant Physiol. the rest of the gene in sunflower. Mol. Geu. Genet. 96:1268-1276. 229:238-244. Dietrich, R.A., Radke, S.E., and Harada, J.J. 1992. Domon, J.-M., Dumas, B., Laine, E., Meyer, Y., David, A., Downstream DNA sequences are required to actiand David, H. 1995. Three glycosylated polypepvate a gene expressed in the root cortex of embryos tides secreted by several embryogenic cell cultures and seedlings. Plant Cell 4:1371-1382. of pine show highly specific serological affinity to antibodies directed against the wheat germin Dietrich, R.A., Maslyar, D.J., Heupel, R.C., and Harada, apoprotein monomer. Plant Physiol. 108:141-148. J.J. 1989. Spatial patterns of gene expression in Brassica napus seedlings: identification of a cortex- Domon, j.-M., Meyer, Y, Faye, L., David, A., and David, specific gene and localization of mRNAs encoding H. 1994. Extracellular (glyco)proteins in embryoisocitrate lyase and a polypeptide homologous to genic and non-embryogenic cell lines of Carribbean proteinases. Plant Cell 1:73-80. pine. Comparison between phenotypes of stage one Dieu, P., and Dunwell, J.M. 1988. Anther culture with difsomatic embryos. Plant Physiol. Biochem. 32:137-147. ferent genotypes of opium poppy (Papaver som- Domoney, C, and Casey, R. 1983. Cloning and characteriniferum L.): effect of cold treatment. Plant Cell Tissue zation of complementary DNA for convicilin, a Organ Cult. 12:263-271. major seed storage protein in Pisum sativum L. Planta Dijak, M., and Simmonds, D.H. 1988. Microtubule organi159:446^53. zation during early direct embryogenesis from mes- Domoney, C, and Casey, R. 1985. Measurement of gene ophyll protoplasts of Medicago sativa L. Plant Sci. number for seed storage proteins in Pisum. Nucl. 58:183-191. Acids Res. 13:687-699. Dijak, M., Smith, D.L., Wilson, T.J., and Brown, D.C.W. Domoney, C, and Casey, R. 1987. Changes in legumin 1986. Stimulation of direct embryogenesis from mesmessenger RNAs throughout seed development in ophyll protoplasts of Medicago sativa. Plant Cell Rep. Pisum sativum L. Planta 170:562-566. 5:468-470. Domoney, C, Davies, D.R., and Casey, R. 1980. The initiaDinis, A.M., and Mesquita, J.F. 1993. The F-actin distribution of legumin synthesis in immature embryos of tion during microsporogenesis in Magnolia Pisum sativum L. grown in vivo and in vitro. Planta soulangeana Soul. (Magnoliaceae). Sex. Plant Reprod. 149:454-460. 6:57-63. Domoney, C, Ellis, N., Turner, L., and Casey, R. 1991. A Dixon, L.K., and Leaver, C.J. 1982. Mitochondrial gene developmentally regulated early-embryogenesis
560
References
protein in pea {Pisum sativum L.) is related to the Drews, G.N., Beals, T.P., Bui, A.Q., and Goldberg, R.B. heat-shock protein (HSP70) gene family. Planta 1992. Regional and cell-specific gene expression pat184:350-355. terns during petal development. Plant Cell Domoney, C, Firmin, J.L., Sidebottom, C, Ealing, P.M., 4:1383-1404. Slabas, A., and Casey, R. 1990. Lipoxygenase hetero- Driscoll, CJ. 1986. Nuclear male sterility systems in seed geneity in Pisum sativum. Planta 181:35-43. production of hybrid varieties. Crit. Rev. Plant Sci. Dong, J., and Yang, H.-Y. 1989. An ultrastructural study of 3:227-256. embryo sac in Oryza sativa L. Ada Bot. Sinica 31:81-88. Dryanovska, O.A. 1981. Cytochemical and autoradiDong, J.G., Satoh, S., and Fujii, T. 1988. Variation in endoographic studies of meiosis and microsporanogeneplasmic-reticulum-associated glycoproteins of carrot sis in Tradescantia paludosa. Compt. Rend. Acad. Sci. cells cultured in vitro. Planta 173:419-423. Bulg. 34:1169-1172. Dong, J.-Z., and Dunstan, D.I. 1994. Growth parameters, Du, H., Simpson, R.J., Clarke, A.E., and Bacic, A. 1996. protein and DNA synthesis of an embryogenic susMolecular characterization of a stigma-specific gene pension culture of white spruce (Picea glauca). ]. encoding an arabinogalactan-protein (AGP) from Plant Physiol. 144:201-208.
Nicotiana alata. Plant J. 9:313-323.
Dong, J.-Z., and Dunstan, D.I. 1996. Expression of abun- Du, H., Simpson, R.J., Moritz, R.L., Clarke, A.E., and dant mRNAs during somatic embryogenesis of Bacic, A. 1994. Isolation of the protein backbone of white spruce [Picea glauca (Moench) Voss]. Planta an arabinogalactan-protein from the styles of 199:459-466. Nicotiana alata and characterization of a corresponding cDNA. Plant Cell 6:1643-1653. Donini, B., and Hussain, S. 1968. Development of the embryo in Triticum durum following irradiation of Dubald, M., Barakate, A., Mandaron, P., and Mache, R. male or female gamete. Radiation Bot. 8:289-295. 1993. The ubiquitous presence of exopolygalacturDooner, H.K. 1985. Viviparous-1 mutation in maize condionase in maize suggests a fundamental cellular functions pleiotropic enzyme deficiencies in the aleution for this enzyme. Plant J. 4:781-791. rone. Plant Physiol. 77:486-488. Dubey, D.K., and Singh, S.P. 1965. Mechanism of pollen Dooner, H.K., and Nelson, O.E. 1979. Interaction among abortion in three male sterile lines of flax (Linum usiC, R and Vp in the control of the Bz glucosyltranstatissimum L.). Crop Sci. 5:121-124. ferase during endosperm development in maize. Dubois, T., Guedira, M., Dubois, J., and Vasseur, J. 1990. Genetics 91:309-315. Direct somatic embryogenesis in roots of Cichorium: Dos Santos, A.V.P., Cutter, E.G., and Davey, M.R. 1983. is callose an early marker? Ann. Bot. 65:539-545. Origin and development of somatic embryos in Dubois, T., Guedira, M., Dubois, J., and Vasseur, J. 1991. Medicago sativa L. (alfalfa). Protoplasma 117:107-115. Direct somatic embryogenesis in leaves of Cichorium. Dougall, D.K., and Verma, D.C. 1978. Growth and embryo A histological and SEM study of early stages. formation in wild-carrot suspension cultures with Protoplasma 162:120-127. ammonium ion as a sole nitrogen source. In Vitro Duchenne, M., Lejeune, B., Fouillard, P., and Quetier, F. 14:180-182. 1989. Comparison of the organization and expression Doughty, J., Hedderson, F., McCubbin, A., and Dickinson, of mtDNA of fertile and male-sterile sugar beet variH. 1993. Interaction between a coating-borne pepeties (Beta vulgaris L.). Theor. Appl. Genet. 78:633-640. tide of the Brassica pollen grain and stigmatic S (self- Duck, N., McCormick, S., and Winter, J. 1989. Heat shock incompatibility)-locus-specific glycoproteins. Proc. protein hsp70 cognate gene expression in vegetative Natl Acad. Sci. USA 90:467-471. and reproductive organs of Lycopersicon esculentum. Proc. Natl Acad. Sci. USA 86:3674-3678. Dover, G.A. 1972. The organization and polarity of pollen mother cells of Triticum aestivum. ]. Cell Sci. Duck, N.B., and Folk, W.R. 1994. Hsp70 heat shock pro11:699-711. tein cognate is expressed and stored in developing tomato pollen. Plant Mol. Biol. 26:1031-1039. Dow, D.A., and Mascarenhas, J.P. 1991a. Optimization of conditions for in situ hybridization and determina- Ducker, S.C., and Knox, R.B. 1976. Submarine pollination tion of the relative number of ribosomes in the cells in seagrasses. Nature 263:705-706. of the mature embryo sac of maize. Sex. Plant Reprod. Ducrocq, C, Sangwan, R.S., and Sangwan-Norreel, B.S. 4:244-249. 1994. Production of Agrobaderium-mediated transgenic fertile plants by direct somatic embryogenesis Dow, D.A., and Mascarenhas, J.P. 1991b. Synthesis and from immature zygotic embryos of Datura innoxia. accumulation of ribosomes in individual cells of the female gametophyte of maize during its developPlant Mol. Biol. 25:995-1009. ment. Sex. Plant Reprod. 4:250-253. Dudareva, N., Evrard, J.-L., Pillay, D.T.N., and Steinmetz, A. 1994. Nucleotide sequence of a pollen-specific Dowrick, V.P.J. 1956. Heterostyly and homostyly in cDNA from Helianthus annuus L. encoding a highly Primula obconica. Heredity 10:219-236. basic protein. Plant Physiol. 106:403-404. Doyle, J.J., Schuler, M.A., Godette, W.D., Zenger, V., and Beachy, R.N. 1986. The glycosylated seed storage Dudits, D., Bogre, L. and Gyorgyey, J. 1991. Molecular and cellular approaches to the analysis of plant proteins of Glycine max and Phaseolus vulgaris. embryos development from somatic cells in vitro. J. Structural homologies of genes and proteins. /. Biol. Cell Sci. 99:473^82. Chem. 261:9228-9238. Dresselhaus, T., Lorz, H., and Kranz, E. 1994. Dudits, D., Gyorgyey, J., Bogre, L., and Bako, L. 1995. Molecular biology of somatic embryogenesis. In In Representative cDNA libraries from few plant cells. Vitro Embryogenesis in Plants, ed. T.A. Thorpe, pp. Plant ]. 5:605-610. 267-308. Dordrecht: Kluwer Academic Publishers. Drews, G.N., Bowman, J.L., and Meyerowitz, E.M. 1991. Negative regulation of the Arabidopsis homeotic gene Duffus, CM., and Rosie, R. 1975. Biochemical changes during embryogeny in Hordeum distichum. AGAMOUS by the APETALA2 product. Cell Phytochemistry 14:319-323. 65:991-1002.
References
561
Dulberger, R. 1970. Tristyly in Lythrum junceum. New male-sterile and fertile lines of pigeon pea (Cajanus Phytol. 69:751-759. cajan (L.) Millsp.). Euphytica 30:431-435. Dulberger, R. 1973. Distyly in Linum pubescens and L. Dundas, I.S., Saxena, K.B., and Byth, D.E. 1982. Pollen mother cell and anther wall development in a phomucronatum. Bot. }. Linn. Soc. 66:117-126. toperiod-insensitive male-sterile mutant of pigeon Dulberger, R. 1974. Structural dimorphism of stigmatic pea {Cajanus cajan (L.) Millsp.). Euphytica 31:371-375. papillae in distylous Linum species. Am. ]. Bot. 61:238-243. Dunstan, D.I., Tautorus, T.E., and Thorpe, T.A. 1995. Dulberger, R. 1975. Intermorph structural differences Somatic embryogenesis in woody plants. In In Vitro Embryogenesis in Plants, ed. T.A. Thorpe, pp. 471-538. between stigmatic papillae and pollen grains in relaDordrecht: Kluwer Academic Publishers. tion to incompatibility in Plumbaginaceae. Proc. Roy. Soc. Land. 188B:257-274. Dunwell, J.M. 1979. Anther culture of Nicotiana tabacum: the role of the culture vessel atmosphere in pollen Dulberger, R. 1981. Dimorphic exine sculpturing in three distylous species of Linum (Linaceae). Plant Syst. embryo induction and growth. /. Expt. Bot. 30:419^128. Evol. 139:113-119. Dulberger, R. 1987. Fine structure and cytochemistry of Dunwell, J.M. 1981. Influence of genotype and environment on growth of barley embryos in vitro. Ann. Bot. the stigma surface and incompatibility in some dis48:535-542. tylous Linum species. Ann. Bot. 59:203-217. Dulberger, R. 1989. The apertural wall in pollen of Linum Dunwell, J.M., Cornish, M., and de Courcel, A.G.L. 1985. grandiflorum. Ann. Bot. 63:421-431. Influence of genotype, plant growth temperature and anther incubation temperature on microspore Dulberger, R. 1990. Release of proteins from the pollen wall of Linum grandiflorum. Sex. Plant Reprod. 3:18-22. embryo production in Brassica napus spp. oleifera. ]. Expt. Bot. 36:679-689. Dulieu, H.L. 1966. Pollination of excised ovaries and culture of ovules of Nicotiana tabacum L. Phytomorphology Dunwell, J.M., and Sunderland, N. 1974a. Pollen ultra16:69-75. structure in anther cultures of Nicotiana tabacum. I. Dumas, C. 1973. Contribution a l'6tude cyto-physiEarly stages of culture. /. Expt. Bot. 25:352-361. ologique du stigmate. III. Evolution et role du r£tic- Dunwell, J.M., and Sunderland, N. 1974b. Pollen ultraulum endoplasmique au cours de la secretion chez structure in anther cultures of Nicotiana tabacum. II. Forsythia intermedia Z.; etude cytochimique. Z. Changes associated with embryogenesis. /. Expt. Bot. Pflanzenphysiol. 70:119-130. 25:363-373. Dumas, C. 1974a. Contribution a l'etude cyto-physi- Dunwell, J.M., and Sunderland, N. 1975. Pollen ultraologique du stigmate. VIII. Les associations r6ticustructure in anther cultures of Nicotiana tabacum. III. lum endoplasmique-plaste et la s6cr6tion The first sporophytic division. /. Expt. Bot. stigmatique. Botaniste 56:81-102. 26:240-252. Dumas, C. 1974b. Some aspects of stigmatic secretion in Dunwell, J.M., and Sunderland, N. 1976. Pollen ultraForsythia. In Fertilization in Higher Plants, ed. H.F. structure in anther cultures of Datura innoxia. I. Linskens, pp. 119-126. Amsterdam: North-Holland Division of the presumptive vegetative cell. /. Cell Publishing Co. Sci. 22:469-480. Dumas, C. 1977a. Lipochemistry of the progamic stage of Dupuis, F. 1972. Formation de la paroi de separation entre a self-incompatible species: neutral lipids and fatty la cellule geneiatrice et la cellule vegetative dans le acids of the secretory stigma during its glandular pollen d'Impatiens balsamina L. Bull. Soc. Bot. Fr. activity, and of the solid style, the ovary and the 119:41-50. anther in Forsythia intermedia Zab. (heterostylic Dupuis, I., and Dumas, C. 1989. In vitro pollination as a species). Planta 137:177-184. model for studying fertilization in maize (Zea mays Dumas, C. 1977b. fitablissement d'un modele de la cineL.). Sex. Plant Reprod. 2:265-269. tique de la secretion lipo-polyph£nolique du stig- Dupuis, I., and Dumas, C. 1990a. Biochemical markers of mate de Forsythia intermedia Zabel. Compt. Rend. female receptivity in maize (Zea mays L.) assessed Acad. Sci. Paris 284D: 1777-1779. using in vitro fertilization. Plant Sci. 70:11-19. Dumas, C. 1978. Stigmates s£cr6teurs et lipides neutres Dupuis, I., and Dumas, C. 1990b. Influence of temperasecretes. Bull. Soc. Bot. Fr. Actual. Bot. 125:61-68. ture stress on in vitro fertilization and heat shock Dumas, C. and Gaude, T. 1981. Stigma-pollen recognition protein synthesis in maize (Zea mays L.) reproducand pollen hydration. Phytomorphology 31:191-201. tive tissues. Plant Physiol. 94:665-670. Dumas, C, Knox, R.B., and Gaude, T. 1985. The spatial Dupuis, I., and Pace, G.M. 1993. Gene transfer to maize association of the sperm cells and vegetative nucleus male reproductive structure by particle bombardin the pollen grain of Brassica. Protoplasma ment of tassel primordia. Plant Cell Rep. 12:607-611. 124:168-174. Dupuis, I., Roeckel, P., Matthys-Rochon, E., and Dumas, Dumas, C, Knox, R.B., McConchie, C.A., and Russell, C. 1987. Procedure to isolate viable sperm cells from S.D. 1984. Emerging physiological concepts in fertilcorn (Zea mays L.) pollen grains. Plant Physiol. ization. Wlrat's Neio in Plant Physiol. 15:17-20. 85:876-878. Dumas, C, Rougier, M., Zandonella, P., Ciampolini, F., Durand, B. 1966. Action d'une kineline sur les caracteres Cresti, M., and Pacini, E. 1978. The secretory stigma sexuels de Mercurialis annua L. (2» = 16). Compt. in Lycopersicum peruvianum Mill.: ontogenesis and Rend. Acad. Sci. Paris 263D:1309-1311. glandular activity. Protoplasma 96:173-187. Durante, M., Cionini, P.G., Avanzi, S., Cremonini, R., and Dunbar, A. 1973. Pollen ontogeny in some species of D'Amato, F. 1977. Cytological localization of the Campanulaceae. A study by electron microscopy. genes for the four classes of ribosomal RNA (25S, Bot. Not. 126:277-315. 18S, 5.8S and 5S) in polytene chromosomes of Dundas, I.S., Saxena, K.B., and Byth, D.E. 1981. Phaseolus coccineus. Chromosoma 60:269-282. Microsporogenesis and anther wall development in Durante, M., Cremonini, R., Tagliasacchi, A.M., Forino,
562 L.M.C., and Cionini, P.G. 1987. Characterization and chromosomal localization of fast renaturing and satellite DNA sequences in Phaseolus coccineus. Protoplasma 137:100-108. Dure, L., III. 1985. Embryogenesis and gene expression during seed formation. Oxford Surv. Plant Mol. Cell Biol. 2:179-197. Dure, L., III. 1993. A repeating 11-mer amino acid motif and plant desiccation. Plant J. 3:363-369. Dure, L., III., and Chlan, C. 1981. Developmental biochemistry of cottonseed embryogenesis and germination. XII. Purification and properties of principal storage proteins. Plant Physiol. 68:180-186. Dure, L., III., and Galau, G.A. 1981. Developmental biochemistry of cotton seed embryogenesis and germination. XIII. Regulation of biosynthesis of principal storage proteins. Plant Physiol. 68:187-194. Dure, L., HI., Greenway, S.C., and Galau, G.A. 1981. Developmental biochemistry of cottonseed embryogenesis and germination: changing messenger ribonucleic acid populations as shown by in vitro and in vivo protein synthesis. Biochemistry 20:4162-4168. Dure, L., Ill, Crouch, M., Harada, J., Ho, T.-H.D., Mundy, J., Quatrano, R., Thomas, T., and Sung, Z.R. 1989. Common amino acid sequence domains among the LEA proteins of higher plants. Plant Mol. Biol. 12:475-486. Dure, L., Ill, Pyle, J.B., Chlan, C.A., Baker, J.C, and Galau, G.A., 1983. Developmental biochemistry of cottonseed embryogenesis and germination. XVII. Developmental expression of genes for the principal storage proteins. Plant Mol. Biol. 2:199-206. Dure, L.S., and Jensen, W.A. 1957. The influence of gibberellic acid and indoleacetic acid on cotton embryos cultured in vitro. Bot. Gaz. 118:254-261. Dute, R.R., and Peterson, CM. 1992. Early endosperm development in ovules of soybean, Glycine max (L.) Merr. (Fabaceae). Ann. Bot. 69:263-271. Dute, R.R., Peterson, CM., and Rushing, A.E. 1989. Ultrastructural changes of the egg apparatus associated with fertilization and proembryo development of soybean, Glycine max (Fabaceae). Ann. Bot. 64:123-135. Dutta, PC, and Appelqvist, L.-A. 1989. The effects of different cultural conditions on the accumulation of depot lipids notably petroselinic acid during somatic embryogenesis in Daucus carota L. Plant Sci. 64:167-177. Dutta, PC, Appelqvist, L.-A., Gunnarsson, S., and von Hofsten, A. 1991. Lipid bodies in tissue culture, somatic and zygotic embryo of Daucus carota L.: a qualitative and quantitative study. Plant Sci. 78:259-267. Dwyer, K.G., Balent, M.A., Nasrallah, J.B., and Nasrallah, M.E. 1991. DNA sequences of self-incompatibility
References
Dwyer, K.G., Lalonde, B.A., Nasrallah, J.B., and Nasrallah, M.E. 1992. Structure and expression of AtSl, an Arabidopsis thaliana gene homologous to the S-locus related genes of Brassica. Mol. Gen. Genet. 231:442-448 Dzelskalns, V.A., Nasrallah, J.B., and Nasrallah, M.E. 1992. Cell-cell communication in plants: self incompatibility in flower development. Devel Biol. 153:70-82. Dzelskalns, V.A., Thorsness, M.K., Dwyer, K.G., Baxter, J.S., Balent, M.A., Nasrallah, M.E. and Nasrallah, J.B. 1993. Distinct ris-acting elements direct pistilspecific and pollen-specific activity of the Brassica S locus glycoprotein gene promoter. Plant Cell 5:855-863. Eady, C, Lindsey, K., and Twell, D. 1994. Differential activation and conserved vegetative cell-specific activity of a late pollen promoter in species with bicellular and tricellular pollen. Plant J. 5:543-550. Eady, C, Lindsey, K., and Twell, D. 1995. The significance of microspore division and division symmetry for vegetative cell-specific transcription and generative cell differentiation. Plant Cell 7:65-74. East, E.M. 1934. Norms of pollen-tube growth in incompatible matings of self-sterile plants. Proc. Natl Acad. Sci. USA 20:225-230. East, E.M., and Mangelsdorf, A.J. 1925. A new interpretation of the hereditary behavior of self-sterile plants. Proc. Natl Acad. Sci. USA 11:166-171.
Ebert, P.R., Anderson, M.A., Bematzky, R., Altschuler, M., and Clarke, A.E. 1989. Genetic polymorphism of self-incompatibility in flowering plants. Cell 56:255-262. Echlin, P. 1972. The ultrastructure and ontogeny of pollen in Helleborus foetidus L. IV. Pollen grain maturation. /. Cell Sci. 11:111-129. Echlin, P., and Godwin, H. 1968a. The ultrastructure and ontogeny of pollen in Helleborus foetidus L. I. The development of the tapetum and Ubisch bodies. /. Cell Sci. 3:161-174. Echlin, P., and Godwin, H. 1968b. The ultrastructure and ontogeny of pollen in Helleborus foetidus L. II. Pollen grain development through the callose special wall stage. /. Cell Sci. 3:175-186. Echt, C.S., and Schwartz, D. 1981. Evidence for the inclusion of controlling elements within the structural gene at the waxy locus in maize. Genetics 99:275-284. Edwards, M., Scott, C, Gidley, M.J., and Grant Reid, J.S. 1992. Control of mannose/galactose ratio during galactomannan formation in developing legume seeds. Planta 187:67-74. Edwardson, J.R. 1956. Cytoplasmic male-sterility. Bot. Rev. 22:696-738. Edwardson, J.R. 1970. Cytoplasmic male sterility. Bot. Rev. 36:341-420. Edwardson, J.R., Bond, D.A., and Christie, R.G. 1976. Cytoplasmic sterility factors in Vicia faba L. Genetics genes from Brassica campestris and B. oleracea: poly82:443-449. morphism predating speciation. Plant Mol. Biol. 16:481^186. Edwardson, J.R., and Warmke, H.E. 1967. Fertility restoration in cytoplasmic male-sterile Petunia. ]. Hered. Dwyer, K.G., Chao, A., Cheng, B., Chen, C.H., and 58:195-196. Nasrallah, J.B. 1989. The Brassica self-incompatibility Egertsdotter, U., Mo, L.H., and von Arnold, S. 1993. multigene family. Genome 31:969-972. Extracellular proteins in embryogenic suspension Dwyer, K.G., Kandasamy, M.K., Mahosky, D.I., Acciai, ]., cultures of Norway spruce (Picea abies). Physiol. Kudish, B.I., Miller, J.E., Nasrallah, M.E., and Plant. 88:315-321. Nasrallah, J.B. 1994. A superfamily of S locus-related sequences in Arabidopsis: diverse structures and Egertsdotter, U., and von Arnold, S. 1995. Importance of arabinogalactan proteins for the development of expression patterns. Plant Cell 6:1829-1843.
References somatic embryos of Norway spruce {Picea abies). Physiol. Plant. 93:334-345. Ehlenfeldt, M.K., and Ortiz, R. 1995. Evidence on the nature and origins of endosperm dosage requirements in Solarium and other angiosperm genera. Sex. Plant Reprod. 8:189-196. Eisenberg, A.J., and Mascarenhas, J.P. 1985. Abscisic acid and the regulation of synthesis of specific seed proteins and their messenger RNAs during culture of soybean embryos. Planta 166:505-514. El-Ghazaly, G., and Jensen, W. 1986a. Studies of the development of wheat (Triticum aestivum) pollen: formation of the pollen aperture. Can. J. hot. 64:3141-3154. El-Ghazaly, G., and Jensen, W.A. 1986b. Studies of the development of wheat (Triticum aestivum) pollen. I. Formation of the pollen wall and Ubisch bodies. Grana 25:1-29. El-Ghazaly, G.A., and Jensen, W.A. 1990. Development of wheat (Triticum aestivum) pollen wall before and after effect of a gametocide. Can. J. Bot. 68:2509-2516. El-Ghazaly, G.A., and Nilsson, S. 1991. Development of tapetum and orbicules of Catharanthus roseus (Apocynaceae). In Pollen and Spores, ed. S. Blackmore and S.H. Barnes, pp. 317-329. Oxford: Clarendon Press. El Hadrami, I., Michaux-Ferriere, N., Carron, M.-P., and d'Auzac, J. 1989. Les polyamines, facteur limitant possible de l'embryogenese somatique chez YHevea brasiliensis. Compt. Rend. Acad. Sci. Paris Ser. HI 308:205-211. Elleman, C.J., and Dickinson, H.G. 1986. Pollen-stigma interactions in Brassica. IV. Structural reorganization in the pollen grains during hydration. /. Cell Sci. 80:141-157. Elleman, C.J., and Dickinson, H.G. 1990. The role of the exine coating in pollen-stigma interactions in Brassica oleracea L. New Phytol. 114:511-518. Elleman, C.J., and Dickinson, H.G. 1996. Identification of pollen components regulating pollination-specific responses in the stigmatic papillae of Brassica oleracea. New Phytol. 133:197-205. Elleman, C.J., Franklin-Tong, V., and Dickinson, H.G. 1992. Pollination in species with dry stigmas: the nature of the early stigmatic response and the pathway taken by pollen tubes. New Phytol. 121:413-424. Elleman, C.J., Willson, C.E., and Dickinson, H.G. 1987. Fixation of Brassica oleracea pollen during hydration; a comparative study. Pollen Spores 29:273-290. Elleman, C.J., Willson, C.E., Sarker, R.H., and Dickinson, H.G. 1988. Interaction between the pollen tube and stigmatic cell wall following pollination in Brassica oleracea. New Phytol. 109:111-117. Elliott, R.C., Betzner, A.S., Huttner, E., Oakes, M.P., Tucker, W.Q.J., Gerentes, D., Perez, P., and Smyth, D.R. 1996. AINTEGUMENTA, an APEL4LA2-like gene of Arabidopsis with pleiotropic roles in ovule development and floral organ growth. Plant Cell 8:155-168. Ellis, D.D., McCabe, D.E., Mclnnis, S., Ramachandran, R., Russell, D.R., Wallace, K.M., Martinell, B.J., Roberts, D.R., Raffa, K.L., and McCown, B.H. 1993. Stable transformation of Picea glauca by particle acceleration. Biotechnology 11:84-89. Ellis, J.R., Gates, P.J., and Boulter, D. 1987. Storage-protein deposition in the developing rice caryopsis in relation to the transport tissues. Ann. Bot. 60:663-670. Ellis, J.R., Shirsat, A.H., Hepher, A., Yarwood, J.N., Gatehouse, J.A., Croy, R.R.D., and Boulter, D. 1988. Tissue-specific expression of a pea legumin gene in
563 seeds of Nicotiana plumbaginifolia. Plant Mol. Biol. 10:203-214. Emons, A.M.C. 1994. Somatic embryogenesis: cell biological aspects. Acta Bot. Neerl. 43:1-14. Emons, A.M.C, and Kieft, H. 1990. Comparison of embryogenic and non-embryogenic suspension cells of maize by means of freeze-fracturing. Micron Micros. Acta 21:255-256. Emons, A.M.C, Mulder, M.M., and Kieft, H. 1993. Pyrolysis mass spectrometry of developmental stages of maize embryos. Acta Bot. Neerl. 42:319-339. Emons, A.M.C, Vos, J.W., and Kieft, H. 1992. A freeze fracture analysis of the surface of embryogenic and non-embryogenic suspension cells of Daucus carota. Plant Sci. 87:85-97. Engell, K. 1989. Embryology of barley: time course and analysis of controlled fertilization and early embryo formation based on serial sections. Nord. ]. Bot. 9:265-280. Engell, K. 1994. Embryology of barley. IV. Ultrastructure of the antipodal cells of Hordeum vulgare L. cv. Bomi before and after fertilization of the egg cell. Sex. Plant Reprod. 7:333-346. Engels, F.M. 1973. Function of Golgi vesicles in relation to cell wall synthesis in germinating Petunia pollen. I. Isolation of Golgi vesicles. Acta Bot. Neerl. 22:6-13. Engels, F.M. 1974a. Function of Golgi vesicles in relation to cell wall synthesis in germinating Petunia pollen. II. Chemical composition of Golgi vesicles and pollen tube wall. Acta Bot. Neerl. 23:81-89. Engels, F.M. 1974b. Function of Golgi vesicles in relation to cell wall synthesis in germinating Petunia pollen. III. The ultrastructure of the tube wall. Acta Bot. Neerl. 23:201-207. Engels, F.M. 1974c. Function of Golgi vesicles in relation to cell wall synthesis in germinating Petunia pollen. IV. Identification of cellulose in pollen tube walls and Golgi vesicles by X-ray diffraction. Acta Bot. Neerl. 23:209-215. Engvild, K.C, Linde-Laursen, I., and Lundqvist, A. 1972. Anther cultures of Datura innoxia: flower bud stage and embryoid level of ploidy. Hereditas 72:331-332. Enjuto, M., Lumbreras, V., Marin, C , and Boronat, A. 1995. Expression of the Arabidopsis HMG2 gene, encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase, is restricted to meristematic and floral tissues. Plant Cell 7:517-527. Entwistle, J. 1988. Primary structure of a C-hordein gene from barley. Carlsberg Res. Commun. 53:247-258. Erbrich, P. 1965. Uber Endopolyploidie und Kernstrukturen in Endospermhaustorien. Osterr. Bot. Z. 112:197-262. Erdelska, O. 1966. Einfluss der Befruchtung auf die Entwicklung der Antipodenkerne der Gerste. Bidlogia 21:857-864. Erdelska, 0.1980. Some structural aspects of flax embryo nutrition. Biologia 35:243-249. Erdelska', O. 1985. Dynamics of the development of embryo and endosperm. II. (Linum usitatissimum, Capsella bursa-pastoris). Biologia 40:849-857. Erdelska, O. 1986. Cytolysis of the endosperm in different types of correlation in endosperm and endosperm development. Acta Bot. Neerl. 35:437-441. Erdelska, O., and Klasova, A. 1978. La region micropylaire du sac embryonnaire de Jasione montana L. avant et apres la fecondation. Bull. Soc. Bot. Fr. Actual. Bot. 125:249-252.
564
References
Ereken-Tumer, N., Richter, J.D., and Nielsen, N.C. 1982. ABA and IAA contents in somatic and zygotic Structural characterization of the glycinin precurembryos of Hevea brasiliensis (Mull. Arg.) during sors. /. Biol. Chem. 257:4016-4018. ontogenesis. Plant Sci. 92:111-119. Erickson, L., Grant, I., and Beversdorf, W. 1986a. Ettinger, W.F., and Harada, J.J. 1990. Translational or postCytoplasmic male sterility in rapeseed (Brassica translational processes affect differentially the accunapus L.). 1. Restriction patterns of chloroplast and mulation of isocitrate lyase and malate synthase mitochondrial DNA. Theor. Appl. Genet. 72:145-150. proteins and enzyme activities in embryos and seedlings of Brassica napus. Arch. Biochem. Biophys. Erickson, L., Grant, I., and Beversdorf, W. 1986b. 281:139-143. Cytoplasmic male sterility in rapeseed (Brassica napus L.). 2. The role of a mitochondrial plasmid. Eunus, A.M. 1955. The effects of X-rays on the embryonal Theor. Appl. Genet. 72:151-157. growth and development of Hordeum vulgare L. /. Erickson, R.O. 1948. Cytological and growth correlations Expt. Bot. 6:409-421. in the flower bud and anther of Lilium longiflorum. Evans, D.E., Rothnie, N.E., Palmer, M.V., Burke, D.G., Am. ]. Bot. 35:729-739. Sang, J.P., Knox, R.B., Williams, E.G., Hilliard, E.P., Ericson, M.L., Muren, E., Gustavsson, H.-O., Josefsson, L.and Salisbury, P.A. 1987. Comparative analysis of G., and Rask, L. 1991. Analysis of the promoter fatty acids in pollen and seed of rapeseed. region of napin genes from Brassica napus demonPhytochemistry 26:1895-1897. strates binding of nuclear protein in vitro to a con- Evans, D.E., Sang, J.P., Cominos, X., Rothnie, N.E., and served sequence motif. Eur. }. Biochem. 197:741-746. Knox, R.B. 1990. A study of phospholipids and galactolipids in pollen of two lines of Brassica napus Ericson, M.L., Rodin, }., Lenman, M., Glimelius, K., L. (rapeseed) with different ratios of linoleic to Josefsson, L.-G., and Rask, L. 1986. Structure of the linolenic acid. Plant Physiol. 92:418-424. rapeseed 1.7S storage protein, napin, and its precursor. /. Biol. Chem. 261:14576-14581. Evans, D.E., Taylor, P.E., Singh, M.B., and Knox, R.B. 1991. Errampalli, D., Patton, D., Castle, L., Mickelson, L., Quantitative analysis of lipids and protein from the Hansen, K., Schnall, }., Feldmann, K., and Meinke, pollen of Brassica napus L. Plant Sci. 73:117-126. D. 1991. Embryonic lethals and T-DNA insertional Evans, D.E., Taylor, P.E., Singh, M.B., and Knox, R.B. 1992. mutagenesis in Arabidopsis. Plant Cell 3:149-157. The interrelationship between the accumulation of Eschrich, W. 1961. Untersuchungen iiber den Ab- und lipids, protein and the level of acyl carrier protein Aufbau der Callose. (HI. Mitteilungiiber Callose). Z. during the development of Brassica napus L. pollen. Bot. 49:153-218. Planta 186:343-354. Esen, A., and Soost, R.K. 1973. Seed development in Citrus Evans, I.M., Bown, D., Lycett, G.W., Croy, R.R.D., Boulter, with special reference to 2X X 4X crosses. Am. }. Bot. D., and Gatehouse, J.A. 1985. Transcription of a legu60:448-462. min gene from pea (Pisum sativum L.) in vitro. Planta Espelie, K.E., Loewus, F.A., Pugmire, R.J., Woolfenden, 165:554-560. W.R., Baldi, B.G., and Given, P.H. 1989. Structural Evans, I.M., Croy, R.R.D., Hutchinson, P., Boulter, D., analysis of Lilium longiflorum sporopollenin by 13C Payne, PL, and Gordon, M.E. 1979. Cell free syntheNMR spectroscopy. Phytochemistry 28:751-753. sis of some storage protein subunits by polyribosomes and RNA isolated from developing seeds of Espelund, M., de Bedout, J.A., Outlaw, W.H., Jr., and pea (Pisum sativum L.). Planta 144:455-462. Jakobsen, K.S. 1995. Environmental and hormonal regulation of barley late-embryogenesis-abundant Evans, I.M., Gatehouse, J.A., Croy, R.R.D., and Boulter, D. (Lea) mRNAs is via different signal transduction 1984. Regulation of the transcription of storage-propathways. Plant Cell Environ. 18:943-949. tein mRNA in nuclei isolated from developing pea (Pisum sativum L.) cotyledons. Planta 160:559-568. Espelund, M., Sseboe-Larssen, S., Hughes, D.W., Galau, G.A., Larsen, R, and Jakobsen, K.S. 1992. Late Evans, P.T., Holaway, B.L., and Malmberg, R.L. 1988. Biochemical differentiation in the tobacco flower embryogenesis-abundant genes encoding proteins probed with monoclonal antibodies. Planta with different numbers of hydrophilic repeats are 175:259-269. regulated differentially by abscisic acid and osmotic Evans, P.T., and Malmberg, R.L. 1989. Alternative pathstress. Plant J. 2:241-252. ways of tobacco placental development: time of Esser, K. 1953. Genomverdopplung und Pollencommitment and analysis of a mutant. Devel Biol. schlauchwachstum bei Heterostylen. Z. Indukt. Abstamm. Vererbungs. 85:28-50. 136:273-283. Estelle, M.A., and Somerville, C. 1987. Auxin-resistant Everett, N.P., Wach, M.J., and Ashworth, D.J. 1985. mutants of Arabidopsis thaliana with an altered morBiochemical markers of embryogenesis in tissue culphology. Mol. Gen. Genet. 206:200-206. tures of the maize inbred B73. Plant Sci. 41:133-140. Estruch, J.J., Kadwell, S., Merlin, E., and Crossland, L. Evers, A.D. 1970. Development of the endosperm of 1994. Cloning and characterization of a maize wheat. Ann. Bot. 34:547-555. pollen-specific calcium-dependent calmodulin-inde- Evrard, J.-L., Jako, C, Saint-Guily, A., Weil, J.-H., and pendent protein kinase. Proc. Natl Acad. Sci. USA Kuntz, M. 1991. Anther-specific, developmentally 91:8837-8841. regulated expression of genes encoding a new class of proline-rich proteins in sunflower. Plant Mol. Biol. Etienne, H., Sotta, B., Montoro, P., Miginiac, E., and 16:271-281. Carron, M.-P. 1993a. Relations between exogenous growth regulators and endogenous indole-3-acetic Eyal, Y., Curie, C, and McCormick, S. 1995. Pollen speciacid and abscisic acid in the expression of somatic ficity elements reside in 30 bp of the proximal proembryogenesis in Hevea brasiliensis (Mull. Arg.). moters of two pollen-expressed genes. Plant Cell Plant Sci. 88:91-96. 7:373-384. Etienne, H., Sotta, B., Montoro, P., Miginiac, E., and Eyster, W.H. 1931. Heritable characters of maize. XLII. Reduced endosperm. /. Hered. 22:250-252. Carron, M.-P. 1993b. Comparison of endogenous
References
565
shrunken-endosperm mutant of barley. Can. ]. Bot. Faberg<§, A.C. 1937. The cytology of the male sterile 65:943-948. Lathyrus odoratus. Genetica 19:423-430. Fabijanski, S., and Altosaar, I. 1985. Evidence for transla- Fennell, A., and Hauptmann, R. 1992. Electroporation and PEG delivery of DNA into maize microspores. Plant tional control of storage protein biosynthesis during Cell Rep. 11:567-570. embryogenesis of Avena sativa L. (oat endosperm). Plant Mol. Biol. 4:211-218. Fernandez, D.E., Turner, F.R., and Crouch, M.L. 1991. In situ localization of storage protein mRNAs in develFabijanski, S., Matlashewski, G.J., and Altosaar, I. 1985. oping meristems of Brassica napus embryos. Characterization of developing oat seed mRNA: eviDevelopment 111:299-313. dence for many globulin mRNAs. Plant Mol. Biol. 4:205-210. Fernando, D.D., and Cass, D.D. 1994. Plasmodial tapetum and pollen wall development in Butomus umbellatus Falk, A., Ek, B., and Rask, L. 1995. Characterization of a (Butomaceae). Am. ]. Bot. 81:1592-1600. new myrosinase from Brassica napus. Plant Mol. Biol. 27:863-874. Fernando, D.D., and Cass, D.D. 1996. Development and structure of ovule, embryo sac, embryo, and Fan, Z., Armstrong, K.C., and Keller, W.A. 1988. endosperm in Butomus umbellatus (Butomaceae). Int. Development of microspores in vivo and in vitro in J. Plant Sci. 157:269-279. Brassica napus L. Protopksma 147:191-199. Faure, J.-E., Digonnet, C, and Dumas, C. 1994. An in vitro Ferrant, V., and Bouharmont J. 1994. Origin of gynogenetic embryos of Beta vulgaris L. Sex. Plant Reprod. system for adhesion and fusion of maize gametes. 7:12-16. Science 263:1598-1600. Faure, J.-E., Mogensen, H.L., Dumas, C, Lorz, H., and Ferrari, T.E., Brans, D., and Wallace, D.H. 1981. Isolation of a plant glycoprotein involved with control of Kranz, E. 1993. Karyogamy after electrofusion of sinintercellular recognition. Plant Physiol. 6"?'-.270-277. gle egg and sperm cell protoplasts from maize: cytological evidence and time course. Plant Cell 5:747-755. Ferrari, T.E., and Wallace, D.H. 1975. Germination of Brassica pollen and expression of incompatibility in Faure, J.-E., Mogensen, H.L., Kranz, E., Digonnet, C, and vitro. Euphytica 24:757-765. Dumas, C. 1992. Ultrastructural characterization and three-dimensional reconstruction of isolated maize Ferrari, T.E., and Wallace, D.H. 1976. Pollen protein syn(Zea mays L.) egg cell protoplasts. Protoplasma thesis and control of incompatibility in Brassica. 171:97-103. Theor. Appl. Genet. 48:243-249. Faure, O., and Nougarede, A. 1993. Nuclear DNA content Ferreira, P.C.G., Hemerly, A.S., de Almeida Engler, J., van Montagu, M., Engler, G., and Inze, D. 1994. of somatic and zygotic embryos of Vitis vinifera cv. Developmental expression of the Arabidopsis cyclin Grenache noir at the torpedo stage. Flow cytomegene cyclAt. Plant Cell 6:1763-1774. try and in situ DNA microspectrophotometry. Protoplasma 176:145-150. Ferrie, A.M.R., Palmer, C.E., and Keller, W.A. 1995. Haploid embryogenesis. In In Vitro Embryogenesis in Faure, O., Mengoli, M., Nougarede, A., and Bagni, N. Plants, ed. T.A. Thorpe, pp. 309-344. Dordrecht: 1991. Polyamine pattern and biosynthesis in zygotic Kluwer Academic Publishers. and somatic embryo stages of Vitis vinifera. ]. Plant Physiol. 138:545-549. Fett, W.F., Paxton, J.D., and Dickinson, D.B. 1976. Studies on the self-incompatibility response of Lilium longiFauron, C.M.-R., Havlik, M., and Brettell, R.I.S. 1990. The florum. Am. J. Bot. 63:1104-1108. mitochondrial genome organization of a maize fertile cmsT revertant line is generated through recom- Feys, B.J.F., Benedetti, C.E., Penfold, C.N., and Turner, J.G. bination between two sets of repeats. Genetics 1994. Arabidopsis mutants selected for resistance to 124:423-428. the phytotoxin coronatine are male sterile, insensitive to methyl jasmonate, and resistant to a bacterial Fauron, C.M.-R., Casper, M., Gesteland, R., and Albertsen, pathogen. Plant Cell 6:751-759. M. 1992. A multi-recombination model for the mtDNA rearrangements seen in maize cmsT regen- Fiedler, U., Filistein, R., Wobus, U., and Baumlein, H. erated plants. Plant ]. 2:949-958. 1993. A complex ensemble of c/s-regulatory elements Favre-Duchartre, M. 1978. Oogeneses chez les angiospercontrols the expression of a Vicia faba non-storage seed protein gene. Plant Mol. Biol. 22:669-679. mes et autres plantes ovul£es. Rev. Cytol. Biol. Veget.Fienberg, A.A., Choi, J.H., Lubich, W.P., and Sung, Z.R. Bot. 1:79-95. Feirer, R.P., Conkey, J.H., and Verhagen, S.A. 1989. 1984. Developmental regulation of polyamine Triglycerides in embryogenic conifer calli: a comparmetabolism in growth and differentiation of carrot ison with zygotic embryos. Plant Cell Rep. 8:207-209. culture. Planta 162:532-539. Feirer, R.P., Mignon, G., and Litvay, J.D. 1984. Arginine Filion, W.G., and Christie, B.R. 1966. The mechanism of decarboxylase and polyamines required for embryomale sterility in a clone of orchardgrass (Dactylis genesis in the wild carrot. Science 223:1433-1435. glomerata L.). Crop Sci. 6:345-347. Felker, F.C. 1987. Ultrastructure of maize endosperm sus- Filippini, F., Terzi, M., Cozzani, F., Vallone, D., and Lo pension cultures. Am. ]. Bot. 74:1912-1920. Schiavo, F. 1992. Modulation of auxin-binding proteins in cell suspensions. II. Isolation and initial charFelker, F.C, Peterson, D.M., and Nelson, O.E. 1984. acterization of carrot cell variants impaired in somatic Development of tannin vacuoles in chalaza and seed embryogenesis. Theor. Appl. Genet. 84:430-434. coat of barley in relation to early chalazal necrosis in the segl mutant. Planta 161:540-549. Finer, J.J., and McMullen, M.D. 1990. Transformation of Felker, F.C, Peterson, D.M., and Nelson, O.E. 1985. cotton (Gossypium hirsutum L.) via particle bombardment. Plant Cell Rep. 8:586-589. Anatomy of immature grains of eight maternal effect shrunken endosperm barley mutants. Am. }. Bot. Fineran, B.A., Wild, D.J.C, and Ingerfeld, M. 1982. Initial wall formation in the endosperm of wheat, Triticum 72:248-256. aestivum: a reevaluation. Can. J. Bot. 60:1776-1795. Felker, F.C, Peterson, D.M., and Nelson, O.E. 1987. Early grain development of the seg2 maternal-effect Finkelstein, R., and Somerville, C. 1989. Abscisic acid or
566
References
high osmoticum promote accumulation of longchain fatty acids in developing embryos of Brassica napus. Plant Sci. 61:213-217.
Finkelstein, R.R. 1993. Abscisic acid-insensitive mutations provide evidence for stage-specific signal pathways regulating expression of an Arabidopsis late embryogenesis-abundant (lea) gene. Mol. Gen. Genet. 238:401-408. Finkelstein, R.R., and Crouch, M.L. 1984. Precociously germinating rapeseed embryos retain characteristics of embryogeny. Planta 162:125-131. Finkelstein, R.R., and Crouch, M.L. 1986. Rapeseed embryo development in culture on high osmoticum is similar to that in seeds. Plant Physiol. 81:907-912. Finkelstein, R.R., and Somerville, C.R. 1990. Three classes of abscisic acid (ABA)-insensitive mutations of Arabidopsis define genes that control overlapping subsets of ABA responses. Plant Physiol. 94:1172-1179. Finkelstein, R.R., Tenbarge, K.M., Shumway, J.E., and Crouch, M.L. 1985. Role of ABA in maturation of rapeseed embryos. Plant Physiol. 78:630-636. Fischer, C, and Neuhaus, G. 1995. In vitro development of globular zygotic wheat embryos. Plant Cell Rep. 15:186-191. Fischer, C, and Neuhaus, G. 1996. Influence of auxin on the establishment of bilateral symmetry in monocots. Plant ]. 9:659-669. Fischer, H., Haake, V., Horstmann, C, and Jensen, U. 1995. Characterization and evolutionary relationships of Magnolia legumin-encoding cDNAs representing two divergent gene subfamilies. Eur. J. Biochem. 229:645-650. Fischer, R.L., and Goldberg, R.B. 1982. Structure and flanking regions of soybean seed protein genes. Cell 29:651-660. Fischer, W., Bergfeld, R., and Schopfer, P. 1987. Induction of storage protein synthesis in embryos of mature plant seeds. Naturwissenschaften 74:86-88. Fischer, W., Bergfeld, R., Plachy, C, Schafer, R., and Schopfer, P. 1988. Accumulation of storage materials, precocious germination and development of desiccation tolerance during seed maturation in mustard
protein gene of papaya ringspot virus. Biotechnology 10:1466-1472. Fitzgerald, M.A., and Knox, R.B. 1995. Initiation of primexine in freeze-substituted microspores of Brassica campestris. Sex. Plant Reprod. 8:99-104.
Fitzgerald, M.A., Barnes, S.H., Blackmore, S., Calder, D.M., and Knox, R.B. 1994. Exine formation in the pollinium of Dendrobium. Protoplasma 179:121-130. Flamand, M.-C, Goblet, J.-R, Due, G., Briquet, M., and Boutry, M. 1992. Sequence and transcription analysis of mitochondrial plasmids isolated from cytoplasmic male-sterile lines of Vicia faba. Plant Mol. Biol.
19:913-923. Flanagan, C.A., and Ma, H. 1994. Spatially and temporally regulated expression of the MADS-box gene AGL2 in wild-type and mutant Arabidopsis flowers. Plant Mol. Biol. 26:581-595.
Flavell, R. 1974. A model for the mechanism of cytoplasmic male sterility in plants, with special reference to maize. Plant Sci. Lett. 3:259-263. Fleming, A.J., and Hanke, D.E. 1993. The regulation of napin gene expression in secondary embryos of Brassica napus. Physiol. Plant. 87:396^02.
Flinn, B.S., Roberts, D.R., and Taylor, I.E.P. 1991. Evaluation of somatic embryos of interior spruce. Characterization and developmental regulation of storage proteins. Physiol. Plant. 82:624-632. Flinn, B.S., Roberts, D.R., Newton, C.H., Cyr, D.R., Webster, F.B., and Taylor, I.E.P. 1993. Storage protein gene expression in zygotic and somatic embryos of interior spruce. Physiol. Plant. 89:719-730. Flinn, B.S., Roberts, D.R., Webb, D.T., and Sutton, B.C.S. 1991. Storage protein changes during zygotic embryogenesis in interior spruce. Tree Physiol. 8:71-81. Flint, D., Ayers, G.S., and Ries, S.K. 1975. Synthesis of endosperm proteins in wheat seed during maturation. Plant Physiol. 56:381-384. Floris, C, Meletti, P., and D'Amato, F. 1970. Further observations on embryo-endosperm relations in irradiated water soaked seeds of durum wheat. Mutation Res. 10:253-255. Folsom, M.W., and Cass, D.D. 1986. Changes in transfer (Sinapis alba L.). Bot. Ada 101:344-354. cell distribution in the ovule of soybean after fertilization. Can. ]. Bot. 64:965-972. Fisher, D.B., and Jensen, W.A. 1969. Cotton embryogenesis: the identification, as nuclei, of the X-bodies in the Folsom, M.W., and Cass, D.D. 1988. The characteristics and fate of the soybean inner nucellus. Ada Bot. degenerated synergid. Planta 84:122-133. Need. 37:387-393. Fisher, D.B., and Jensen, W.A. 1972. Nuclear and cytoplasmic DNA synthesis in cotton embryos: a correlated Folsom, M.W., and Cass, D.D. 1989. Embryo sac development in soybean: ultrastructure of megasporogenelight and electron microscope autoradiographic study. sis and early megagametogenesis. Can. J. Bot. Histochemie 32:1-22. 67:2841-2849. Fisher, D.B., Jensen, W.A., and Ashton, M.E. 1968. Histochemical studies of pollen: storage pockets in the Folsom, M.W., and Cass, D.D. 1990. Embryo sac development in soybean: cellularization and egg apparatus endoplasmic reticulum (ER). Histochemie 13:169-182. expansion. Can. ]. Bot. 68:2135-2147. Fisher, R.H., Barton, M.K., Cohen, J.D., and Cooke, T.J. 1996. Hormonal studies of fass, an Arabidopsis Folsom, M.W., and Cass, D.D. 1992. Embryo sac development in soybean: the central cell and aspects of fermutant that is altered in organ elongation. Plant tilization. Am. J. Bot. 79:1407-1417. Physiol. 110:1109-1121. Fitch, M.M.M., Manshardt, R.M., Gonsalves, D., and Folsom, M.W., and Peterson, CM. 1984. Ultrastructural aspects of the mature embryo sac of soybean, Glycine Slightom, J.L. 1993. Transgenic papaya plants from max (L.) Merr. Bot. Gaz. 145:1-10. Agrobacterium-mediated transformation of somatic embryos. Plant Cell Rep. 12:245-249. Fong, F., Smith, J.D., and Koehler, D.E. 1983. Early events in maize seed development. l-Methyl-3-phenyl-5-(3Fitch, M.M.M., Manshardt, R.M., Gonsalves, D., Slightom, [trifluoromethyl]phenyl)-4-(lH )-pyridinone inducJ.L., and Sanford, J.C. 1990. Stable transformation of tion of vivipary. Plant Physiol. 73:899-901. papaya via microprojectile bombardment. Plant Cell Fontana, G.S., Santini, L., Caretto, S., Frugis, G., and Rep. 9:189-194. Mariotti, D. 1993. Genetic transformation in the Fitch, M.M.M., Manshardt, R.M., Gonsalves, D., Slightom, grain legume Cicer arietinum L. (chickpea). Plant Cell J.L., and Sanford, J.C. 1992. Virus resistant papaya Rep. 12:194-198. plants derived from tissues bombarded with the coat
References
567
forme et revolution cytoplasmique des synergides Fontes, E.B.P., Shank, B.B., Wrobel, R.L., Moose, S.P., du sac embryonnaire d'Aquilegia vulgaris. Compt. OBrian, G.R., Wurtzel, E.T., and Boston, R.S. 1991. Rend. Acad. Sci. Paris 280D:2445-2447. Characterization of an immunoglobulin binding protein homolog in the maize floury-2 endosperm Fowke, L.C, Attree, S.M., Wang, H., and Dunstan, D.I. 1990. Microtubule organization and cell division in mutant. Plant Cell 3:483-496. embryogenic protoplast cultures of white spruce Foote, H.C.C., Ride, J.P., Franklin-Tong, V.E., Walker, E.A., (Picea glauca). Protoplasma 158:86-94. Lawrence, M.J., and Franklin, F.C.H. 1994. Cloning and expression of a distinctive class of self-incom- Franchi, G.G., and Pacini, E. 1980. Wall projections in the patibility (S) gene from Palaver rhoeas L. Proc. Natl vegetative cell of Parietariaofficinalis L. pollen. Acad. Sci. USA 91:2265-2269. Protoplasma 104:67-74. Forde, B.G., and Leaver, C.J. 1980. Nuclear and cytoplas- Francis, R.R., and Bemis, W.P. 1970. A cytomorphological mic genes controlling synthesis of variant mitochonstudy of male sterility in a mutant of Cucurbita maxdrial polypeptides in male-sterile maize. Proc. Natl ima Dutch. Econ. Bot. 24:325-332. Acad. Sci. USA 77:418-422. Franke, W.W., Herth, W., VanDerWoude, W.J., and Morr6, D.J. 1972. Tubular and filamentous strucForde, B.G., Oliver, R.J.C., and Leaver, C.J. 1978. Variation in mitochondrial translation products associated tures in pollen tubes: possible involvement as with male-sterile cytoplasms in maize. Proc. Natl guide elements in protoplasmic streaming and vectorial migration of secretory vesicles. Planta Acad. Sci. USA 75:3841-3845. 105:317-341. Forde, B.G., Heyworth, A., Pywell, J., and Kreis, M. 1985a. Nucleotide sequence of a Bl hordein gene Frankel, O.H. 1940. Studies in Hebe. II. The significance of and the identification of possible upstream regulamale sterility in the genetic system. /. Genet. tory elements in endosperm storage protein genes 40:171-184. from barley, wheat and maize. Nucl. Acids Res. Frankel, R., and Galun, E. 1977. Pollination Mechanisms, 13:7327-7339. Reproduction and Plant Breeding. Berlin: SpringerVerlag. Forde, B.G., Kreis, M., Williamson, M.S., Fry, R.P., Pywell, J., Shewry, P.R., Bunce, N., and Miflin, B.J. 1985b. Frankel, R., Izhar, S., and Nitsan, J. 1969. Timing of callase activity and cytoplasmic male sterility in Petunia. Short tandem repeats shared by B- and C-hordein cDNAs suggest a common evolutionary origin for Biochem. Genet. 3:451-455. two groups of cereal storage protein genes. EM BO J. Frankel, R., Scowcroft, W.R., and Whitfeld, PR. 1979. 4:9-15. Chloroplast DNA variation in isonuclear male-sterile lines of Nicotiana. Mol. Gen. Genet. 169:129-135. Forde, B.G., Oliver, R.J.C., Leaver, C.J., Gunn, R.E., and Kemble, R.J. 1980. Classification of normal and male- Frankis, R., and Mascarenhas, J.P. 1980. Messenger RNA sterile cytoplasms in maize. I. Electrophoretic analyin the ungerminated pollen grain: a direct demonsis of variation in mitochondrially synthesized stration of its presence. Ann. Bot. 45:595-599. proteins. Genetics 95:443-450. Frankis, R.C., Jr. 1990. RNA and protein synthesis in gerForde, J., Forde, B.G., Fry, R.P., Kreis, M., Shewry, P.R., and minating pine pollen. /. Expt. Bot. 41:1469-1473. Miflin, B.J. 1983. Identification of barley and wheat Frankis, R.C., Jr., and Grayson, G.K. 1990. Heat-shock cDNA clones related to the high-Mr polypeptides of response in germinating pine pollen. Sex. Plant wheat gluten. FEBS Lett. 162:360-366. Reprod. 3:195-199. Forde, J., Malpica, J.-M., Halford, N.G., Shewry, PR., Franklin, F.C.H., Lawrence, M.J., and Franklin-Tong, V.E. 1995. Cell and molecular biology of self-incompatiAnderson, O.D., Greene, F.C., and Miflin, B.J. 1985. bility in flowering plants. Int. Rev. Cytol. 158:1-64. The nucleotide sequence of a HMW glutenin subunit gene located on chromosome 1A of wheat (Triticum Franklin-Tong, V.E., and Franklin, F.C.H. 1992. aestivum L.). Nucl. Acids Res. 13:6817-6832. Gametophytic self-incompatibility in Papaver rhoeas L. Sex. Plant Reprod. 5:1-7. Forino, L.M.C., Tagliasacchi, A.M., Cavallini, A., Cionini, G., Giraldi, E., and Cionini, P.G. 1992. RNA synthesis Franklin-Tong, V.E., and Franklin, F.C.H. 1993. Gametophytic self-incompatibility: contrasting in the embryo suspensor of Phaseolus coccineus at two mechanisms for Nicotiana and Papaver. Trends Cell stages of embryogenesis, and the effect of supplied Biol. 3:340-345. gibberellic acid. Protoplasma 167:152-158. Forman, M., and Jensen, W.A. 1965. Respiration and Franklin-Tong, V.E., Lawrence, M.J., and Franklin, F.C.H. embryogenesis in cotton. Plant Physiol. 40:765-769. 1988. An in vitro bioassay for the stigmatic product of the self-incompatibility gene in Papaver rhoeas L. New Forster, B.P., and Dale, J.E. 1983. A comparative study of Phytol. 110:109-118. early seed development in genotypes of barley and Franklin-Tong, V.E., Lawrence, M.J., and Franklin, F.C.H. rye. Ann. Bot. 52:603-612. 1990. Self-incompatibility in Papaver rhoeas L.: inhibiFosket, D.E., and Morejohn, L.C. 1992. Structural and tion of incompatible pollen tube growth is dependent functional organization of tubulin. Annu. Rev. Plant Physiol. Plant Mol. Biol. 43:201-240. on pollen gene expression. New Phytol. 116:319-324. Foster, G.D., Robinson, S.W., Blundell, R.P., Roberts, M.R., Franklin-Tong, V.E., Ride, J.P, and Franklin, F.C.H. 1995. Hodge, R., Draper, J., and Scott, R.J. 1992. A Brassica Recombinant stigmatic self-incompatibility (S-) protein elicits a Ca2+ transient in pollen of Papaver rhoeas. napus mRNA encoding a protein homologous to Plant J. 8:299-307. phospholipid transfer proteins, is expressed specifically in the tapetum and developing microspores. Franklin-Tong, V.E., Atwal, K.K., Howell, E.C., Lawrence, M.J., and Franklin, F.C.H. 1991. Self-incompatibility Plant Sci. 84:187-192. Foster, T.S., and Stern, H. 1959. The accumulation of soluin Papaver rhoeas: there is no evidence for the ble deoxyribosidic compounds in relation to nuclear involvement of stigmatic ribonuclease activity. Plant division in anthers of Lilium longiflorum. ]. Biophys. Cell Environ. 14:423-429. Franklin-Tong, V.E., Ride, J.P, Read, N.D., Trewavas, A.J., Biochem. Cytol. 5:187-192. and Franklin, F.C.H. 1993. The self-incompatibility Fougere-Rifot, M. 1975. L'edification de l'appareil fili-
568
References
response in Papaver rhoeas is mediated by cytosolic Fromm, M.E., Morrish, F., Armstrong, C, Williams, R., free calcium. Plant J. 4:163-177. Thomas, J., and Klein, T.M. 1990. Inheritance and expression of chimeric genes in the progeny of transFranklin-Tong, V.E., Ruuth, E., Marmey, P., Lawrence, genic maize plants. Biotechnology 8:833-839. M.JV and Franklin, F.C.H. 1989. Characterization of a stigmatic component from Papaver rhoeas L. which Frova, C. 1990. Analysis of gene expression in exhibits the specific activity of a self-incompatibility microspores, pollen, and silks of Zea mays L. Sex. (S-) gene product. New Phytol. 112:307-315. Plant Reprod. 3:200-206. Franssen-Verheijen, M.A.W., and Willemse, M.T.M. 1990. Frova, C, Taramino, G., and Binelli, G. 1989. Heat-shock The ovule of Gasteria verrucosa at receptivity of the proteins during pollen development in maize. Devel stigma: an ultrastructural study. Ada Bot. Neerl. Genet. 10:324-332. 39:53-63. Frova, C, Taramino, G., and Ottaviano, E. 1991. Franssen-Verheijen, M.A.W., and Willemse, M.T.M. 1993. Sporophytic and gametophytic heat shock protein Micropylar exudate in Gasteria (Aloaceae) and its synthesis in Sorghum bicolor. Plant Sci. 73:35-44. possible function in pollen tube growth. Am. J. Bot. Frova, C, Sari Gorla, M., Ottaviano, E., and Pella, C. 1983. 80:253-262. Haplodiploid gene expression in maize and its detection. Biochem. Genet. 21:923-931. Fransz, P.F., de Ruijter, N.C.A., and Schel, J.H.N. 1989. Isozymes as biochemical and cytochemical markers Fujii, N., and Uchimiya, H. 1991. Conditions favorable for in embryogenic callus cultures of maize (Zea mays the somatic embryogenesis in carrot cell culture L). Plant Cell Rep. 8:67-70. enhance expression of the rolC promoter-GUS fusion gene. Plant Physiol. 95:238-241. Franz, Gv Hatzopoulos, P., Jones, T.J., Krauss, M., and Sung, Z.R. 1989. Molecular and genetic analysis of Fujii, N., Yokoyama, R., and Uchimiya, H. 1994. Analysis of the rolC promoter region involved in somatic an embryonic gene, DC 8, from Daucus carota L. Mol. embryogenesis-related activation in carrot cell culGen. Genet. 218:143-151. tures. Plant Physiol. 104:1151-1157. Franzmann, L., Patton, D.A., and Meinke, D.W. 1989. In vitro morphogenesis of arrested embryos from lethal Fujimura, T., and Komamine, A. 1975. Effects of various mutants of Arabidopsis thaliana. Theor. Appl. Genet. growth regulators on the embryogenesis in a carrot cell suspension culture. Plant Sci. Lett. 5:359-364. 77:609-616. Frediani, M., Cremonini, R., Salvi, G., Caprari, C, Fujimura, T., and Komamine, A. 1979a. Involvement of endogenous auxin in somatic embryogenesis in a Desiderio, A., D'Ovidio, R., Cervone, F., and de carrot cell suspension culture. Z. Pflanzenphysiol. Lorenzo, G. 1993. Cytological localization of the 95:13-19. PG1P genes in the embryo suspensor cells of Phaseolus vulgaris L. Theor. Appl. Genet. 87:369-373. Fujimura, T., and Komamine, A. 1979b. Synchronization of somatic embryogenesis in a carrot cell suspension Frediani, M., Forino, L.M.C., Tagliasacchi, A.M., Cionini, culture. Plant Physiol. 64:162-164. P.G., Durante, M., and Avanzi, S. 1986a. Functional heterogeneity, during early embryogenesis, of Fujimura, T, and Komamine, A. 1980a. Mode of action of Phaseolus coccineus ribosomal cistrons in polytene 2,4-D and zeatin on somatic embryogenesis in a carchromosomes of embryo suspensor. Protoplasma rot cell suspension culture. Z. Pflanzenphysiol. 99:1-8. Fujimura, T, and Komamine, A. 1980b. The serial obser132:51-57. vation of embryogenesis in a carrot cell suspension Frediani, M., Tagliasacchi, A.M., Durante, M., and Avanzi, culture. New Phytol. 86:213-218. S. 1986b. Distribution of 5-methylcytosine-rich regions in the polytene chromosomes of Phaseolus Fujimura, T, Komamine, A., and Matsumoto, H. 1980. Aspects of DNA, RNA and protein synthesis during coccineus embryo suspensor as shown by the somatic embryogenesis in a carrot cell suspension immunoperoxidase technique. Expt. Cell Res. culture. Physiol. Plant. 49:255-260. 167:337-342. Freeling, M. 1976. Intragenic recombination in maize: Fujimura, T., Komamine, A., and Matsumoto, H. 1981. pollen analysis methods and the effect of parental Changes in chromosomal proteins during early AdhV isoalleles. Genetics 83:701-717. stages of synchronized embryogenesis in a carrot cell suspension culture. Z. Pflanzenphysiol. Frey-Wyssling, A., Grieshaber, E., and Miihlethaler, K. 102:293-298. 1963. Origin of spherosomes in plant cells. /. Ultrastr. Res. 8:506-516. Fujiwara, T, and Beachy, R.N. 1994. Tissue-specific and temporal regulation of a (3-conglycinin gene: roles of Friedman, W.E. 1990. Double fertilization in Ephedra, a the RY repeat and other cis-acting elements. Plant nonflowering seed plant: its bearing on the origin of Mol. Biol. 24:261-272. angiosperms. Science 247:951-954. Friedman, W.E. 1991. Double fertilization in Ephedra tri- Fujiwara, T., Lessard, P.A., and Beachy, R.N. 1992. Seedspecific repression of GUS activity in tobacco plants furca, a non-flowering seed plant: the relationship by antisense RNA. Plant Mol. Biol. 20:1059-1069. between fertilization events and the cell cycle. Fujiwara, T, Hirai, M.Y., Chino, M., Komeda, Y., and Protoplasma 165:106-120. Naito, S. 1992. Effects of sulfur nutrition on expresFriedman, W.E. 1992. Evidence of a pre-angiosperm orision of the soybean seed storage protein genes in gin of endosperm: implications for the evolution of transgenic Petunia. Plant Physiol. 99:263-268. flowering plants. Science 255:336-339. Frisch, D.A., van der Geest, A.H.M., Dias, K., and Hall, Fukasawa, H. 1954. On the free amino acids in anthers of male-sterile wheat and maize. Jap. }. Genet. T.C. 1995. Chromosomal integration is required for 29:135-137. spatial regulation of expression from the P-phaseolin promoter. Plant J. 7:503-512. Fukuda, Y. 1927. Cytological studies on the development of the pollen-grain in different races of Solanum Fritz, N.K., and Hanneman, R.E., Jr. 1989. Interspecific tuberosum L., with special reference to sterility. Bot. incompatibility due to stylar barriers in tuber-bearMag. Tokyo 41:459-476. ing and closely related non-tuber-bearing Solanums. Fukuoka, H., Ogawa, T., Minami, H., Yano, H., and Sex. Plant Reprod. 2:184-192.
569
References
nesis-abundant proteins. Plant Physiol. 101:695-696. Ohkawa, Y. 1996. Developmental stage-specific and nitrate-independent regulation of nitrate reductase Galili, G. 1989. Heterologous expression of a wheat high molecular weight glutenin gene in Escherichia coli. gene expression in rapeseed. Plant Physiol. 111:39-47. Proc. Natl Acad. Sci. USA 86:7756-7760.
Gabara, B. 1974. A possible role for the endoplasmic reticulum in exine formation. Grana 14:16-22. Gaget, Mv Said, C, Dumas, C., and Knox, R.B. 1984. Pollen-pistil interactions in interspecific crosses of Populus (sections Aigeiros and Leuce): pollen adhesion, hydration and callose responses. /. Cell Sci. 72:173-184. Gaiser, J.C., Robinson-Beers, K., and Gasser, C.S. 1995. The Arabidopsis SUPERMAN gene mediates asymmetric growth of the outer integument of ovules. Plant Cell 7:333-345. Galdn-Cano, J., Risuefio, M.C., and Gimenez-Martin, G. 1975. Nucleolar ultrastructure in megaspore mother cells of Pisum sativum L. Cytobiologie 10:479-484.
Galante, E., Vitale, A., Manzocchi, L., Soave, C, and Salamini, F. 1983. Genetic control of a membrane component and zein deposition in maize endosperm. Mol. Gen. Genet. 192:316-321. Galau, G.A., Bijaisoradat, N., and Hughes, D.W. 1987. Accumulation kinetics of cotton late embryogenesisabundant mRNAs and storage protein mRNAs: coordinate regulation during embryogenesis and the role of abscisic acid. Devel Biol. 123:198-212. Galau, G.A., Chlan, C.A., and Dure, L., III. 1983. Developmental biochemistry of cottonseed embryogenesis and germination. XVI. Analysis of the principal cotton storage protein gene family with cloned cDNA probes. Plant Mol. Biol. 2:189-198. Galau, G.A., and Close, T.J. 1992. Sequences of the cotton group 2 LEA/RAB/dehydrin proteins encoded by Lea3 cDNAs. Plant Physiol. 98:1523-1525. Galau, G.A., and Dure, L., III. 1981. Developmental biochemistry of cottonseed embryogenesis and germination: changing messenger ribonucleic acid populations as shown by reciprocal heterologous complementary deoxyribonucleic acid-messenger ribonucleic acid hybridization. Biochemistry 20:4169-4178. Galau, G.A., and Hughes, D.W. 1987. Coordinate accumulation of homeologous transcripts of seven cotton Lea gene families during embryogenesis and germination. Devel Biol. 123:213-221. Galau, G.A., Hughes, D.W., and Dure, L., HI. 1986. Abscisic acid induction of cloned cotton late embryogenesis-abundant (Lea) mRNAs. Plant Mol. Biol. 7:155-170. Galau, G.A., Jakobsen, K.S., and Hughes, W.D. 1991. The controls of late dicot embryogenesis and early germination. Physiol. Plant. 81:280-288. Galau, G.A., Wang, H.Y.-C, and Hughes, D.W. 1991. Sequence of the Gossypium hirsutum D-genome alloallele of legumin A and its mRNA. Plant Physiol. 97:1268-1270. Galau, G.A., Wang, H.Y.-C, and Hughes, D.W. 1992a. Cotton Mat5-A (C164) gene and Mat5-D cDNAs encoding methionine-rich 2S albumin storage proteins. Plant Physiol. 99:779-782. Galau, G.A., Wang, H.Y.-C, and Hughes, D.W. 1992b. Cotton Leal (D19) and LeaAl (D132) group 1 Lea genes encoding water stress-related proteins containing a 20-amino acid motif. Plant Physiol. 99:783-788. Galau, G.A., Wang, H.Y.-C, and Hughes, D.W. 1993. Cotton LeaS and Leali encode atypical late embryoge-
Galitz, D.S., and Howell, R.W. 1965. Measurement of ribonucleic acids and total free nucleotides of developing soybean seeds. Physiol. Plant. 18:1018-1021. Gallardo, D., Reina, M., Rigau, J., Boronat, A., and Palau, J. 1988. Genomic organization of the 28 kDa glutelin2 gene from maize. Plant Sci. 54:211-218. Galli, M.G., Bracale, M, Falavigna, A., and Soave, C. 1988. Sexual differentiation in Asparagus officinalis L. I. DNA characterization and mRNA activities in male and female flowers. Sex. Plant Reprod. 1:202-207. Gallusci, P., Salamini, F., and Thompson, R.D. 1994. Differences in cell type-specific expression of the gene opaque 2 in maize and transgenic tobacco. Mol. Gen. Genet. 244:391^00. Galun, E., Jung, Y, and Lang, A. 1963. Morphogenesis of floral buds of cucumber cultured in vitro. Devel Biol. 6:370-387. Gander, E.S., Holmstroem, K.-O., de Paiva, G.R., de Castro, L.A.B., Carneiro, M., and Grossi de Sci, M.-F. 1991. Isolation, characterization and expression of a gene coding for a 2S albumin from Bertholletia excelsa (Brazil nut). Plant Mol. Biol. 16:437-448. Gane, A.M., Weinhandl, J.A., Bacic, A., and Harris, P.J. 1994. Histochemistry and composition of the cell walls of styles of Nicotiana alata Link et Otto. Planta 195:217-225. Gao, X., Francis, D., Ormrod, J.C, and Bennett, M.D. 1992a. An electron microscopic study of double fertilization in allohexaploid wheat Triticum aestivum L. Ann. Bot. 70:561-568. Gao, X., Francis, D., Ormrod, J.C, and Bennett, M.D. 1992b. Changes in cell number and cell division activity during endosperm development in allohexaploid wheat, Triticum aestivum L. ]. Expt. Bot. 43:1603-1609. Garcia-Maroto, F., Salamini, F., and Rohde, W. 1993. Molecular cloning and expression patterns of three alleles of the de/iriens-homologous gene st-deficiens from Solanum tuberosum. Plant J. 4:771-780.
Garcia-Mas, J., Messeguer, R., Arus, P., and Puigdomenech, P. 1995. Molecular characterization of cDNAs corresponding to genes expressed during almond (Prunus amygdalus Batsch) seed development. Plant Mol. Biol. 27:205-210. Garcia-Mas, J., Messeguer, R., Ariis, P., and Puigdomenech, P. 1996. Accumulation of specific mRNAs during almond fruit development. Plant Sci. 113:185-192. Garrido, D., Chibi, F., and Matilla, A. 1995. Polyamines in the induction of Nicotiana tabacum pollen embryogenesis by starvation. /. Plant Physiol. 145:731-735. Garrido, D., Eller, N., Heberle-Bors, E., and Vicente, O. 1993. De novo transcription of specific mRNAs during the induction of tobacco pollen embryogenesis. Sex. Plant Reprod. 6:40-45.
Garrido, D., Vicente, O., Heberle-Bors, E., and Rodriguez-Garci'a, M.I. 1995. Cellular changes during the acquisition of embryogenic potential in isolated pollen grains of Nicotiana tabacum. Protoplasma 186:220-230. Gartner, P.-J., and Nagl, W. 1980. Acid phosphatase activity in plastids (plastolysomes) of senescing embryosuspensor cells. Planta 149:341-349. Gasser, C.S., Budelier, K.A., Smith, A.G., Shah, D.M., and
570
References
Geitmann, A., Li, Y.Q., and Cresti, M. 1996. The role of the cytoskeleton and dictyosome activity in the pulsatory growth of Nicotiana tabacum and Petunia hybrida pollen tubes. Bot. Acta. 109:102-109. Geitmann, A., Huddk, J., Vennigerholz, F., and Walles, B. sativum L.). Nucl. Acids Res. 15:7642. 1995. Immunogold localization of pectin and callose in pollen grains and pollen tubes of Brugmansia Gatehouse, J.A., Bown, D., Gilroy, J., Levasseur, M., suaveolens - implications for the self-incompatibility Castleton, J., and Ellis, T.H.N. 1988. Two genes reaction. /. Plant Physiol. 147:225-235. encoding 'minor' legumin polypeptides in pea (Pisum sativum L.). Characterization and complete Geli, M.I., Torrent, M., and Ludevid, D. 1994. Two structural sequence of the leg] gene. Biochem. J. 250:15-24. domains mediate two sequential events in y-zein targeting: protein endoplasmic reticulum retention and Gatehouse, J.A., Croy, R.R.D., Morton, H., Tyler, M., and protein body formation. Plant Cell 6:1911-1922. Boulter, D. 1981. Characterisation and subunit structures of the vicilin storage proteins of pea (Pisum Gell, A.C., Bade, A., and Clarke, A.E. 1986. Arabinosativum L.). Eur. J. Biochem. 118:627-633. galactan-proteins of the female sexual tissue of Nicotiana alata. I. Changes during flower developGatehouse, J.A., Evans, I.M., Bown, D., Croy, R.R.D., and ment and pollination. Plant Physiol. 82:885-889. Boulter, D. 1982. Control of storage-protein synthesis during seed development in pea (Pisum sativum L.). Geltz, N.R., and Russell, S.D. 1988. Two-dimensional elecBiochem. J. 208:119-127. trophoretic studies of the proteins and polypeptides in mature pollen grains and the male germ unit of Gatehouse, J.A., Evans, I.M., Croy, R.R.D., and Boulter, D. Plumbago zeylanica. Plant Physiol. 88:764-769. 1986. Differential expression of genes during legume seed development. Phil. Trans. Roy. Soc. Lond. Geneves, L. 1966. Evolution des infrastructures du cyto314B:367-384. plasme dans le tissu sporifere des antheres de Ribes rubrum L. (Grossulariacees). Compt. Rend. Acad. Sci. Gatehouse, J.A., Lycett, G.W., Delauney, A.J., Croy, R.R.D., Paris 262D:72-75. and Boulter, D. 1983. Sequence specificity of the post-translational proteolytic cleavage of vicilin, a Gengenbach, B.G. 1977a. Development of maize caryseed storage protein of pea (Pisum sativum L.). opses resulting from in-vitro pollination. Planta Biochem. J. 212:427-432. 134:91-93. Gaubier, P., Raynal, M., Hull, G., Huestis, G.M., Grellet, E, Gengenbach, B.G. 1977b. Genotypic influences on in vitro fertilization and kernel development of maize. Crop Arenas, C. Pages, M., and Delseny, M. 1993. Two difSci. 17:489-492. ferent Em-like genes are expressed in Arabidopsis thaliana seeds during maturation. Mol. Gen. Genet.Gengenbach, B.G., Green, C.E., and Donovan, CM. 1977. 238:409-418. Inheritance of selected pathotoxin resistance in Gaude, T., and Dumas, C. 1984. A membrane-like strucmaize plants regenerated from cell cultures. Proc. ture on the pollen wall surface in Brassica. Ann. Bot. Natl Acad. Sci. USA 74:5113-5117. 54:821-825. Gengenbach, B.G., Connelly, J.A., Pring, D.R., and Conde, Gaude, T., and Dumas, C. 1986. Organization of stigma M.F. 1981. Mitochondrial DNA variation in maize surface components in Brassica: a cytochemical plants regenerated during tissue culture selection. study. /. Cell Sci. 82:203-216. Theor. Appl. Genet. 59:161-167. Gaude, T., and Dumas, C. 1987. Molecular and cellular Gengenbach, B.G., Miller, R.J., Koeppe, D.E., and Arntzen, events of self-incompatibility. Int. Rev. Cytol. C.J. 1973. The effect of toxin from Helminthosporium 107:333-366. maydis (race T) on isolated corn mitochondria: swelling. Can. J. Bot. 51:2119-2125. Gaude, T., Friry, A., Heizmann, P., Mariac, C, Rougier, M., Fobis, I., and Dumas, C. 1993. Expression of a self- Genovesi, A.D., and Collins, G.B. 1982. In vitro production incompatibility gene in a self-compatible line of of haploid plants of corn via anther culture. Crop Sci. 22:1137-1144. Brassica oleracea. Plant Cell 5:75-86. Gaude, T., Rougier, M., Heizmann, P., Ockendon, D.J., and George, L., and Narayanaswamy, S. 1973. Haploid Capsicum through experimental androgenesis. Protoplasma Dumas, C. 1995. Expression level of the SLG gene is not correlated with the self-incompatibility pheno78:467-470. type in the class II S haplotypes of Brassica oleracea. Geraghty, D., Peifer, M.A., Rubenstein, I., and Messing, J. Plant Mol. Biol. 27:1003-1014. 1981. The primary structure of a plant storage protein: zein. Nucl. Acids Res. 9:5163-5174. Gavish, H., Vardi, A., and Fluhr, R. 1991. Extracellular proteins and early embryo development in Citrus Geraghty, D.E., Messing, J., and Rubenstein, I. 1982. Sequence analysis and comparison of cDNAs of the nucellar cell cultures. Physiol. Plant. 82:606-616. zein multigene family. EMBO ]. 1:1329-1335. Gavish, H., Vardi, A., and Fluhr, R. 1992. Suppression of somatic embryogenesis in Citrus cell cultures by Gerassimova-Navashina, G. 1960. A contribution to the cytology of fertilization in flowering plants. Nucleus extracellular proteins. Planta 186:511-517. 3:111-120. Gayler, K.R., and Sykes, G.E. 1981. p-Conglycinins in developing soybean seeds. Plant Physiol. 67:958-961. Gerlach-Cruse, D. 1969. Embryo- und Endospermentwicklung nach einer Rontgenbestrahlung der Fruchtknoten von Geetha, K.B., Lending, C.R., Lopes, M.A., Wallace, J.C., Arabidopsis thaliana (L.) Heynh. Radiation Bot. and Larkins, B.A. 1991. opaque-2 modifiers increase y9:433-442. zein synthesis and alter its spatial distribution in maize endosperm. Plant Cell 3:1207-1219. Gerster, ]., Allard, S., and Robert, L.S. 1996. Molecular characterization of two Brassica napus pollenGeitmann, A., Li, Y.-Q., and Cresti, M. 1995. expressed genes encoding putative arabinogalactan Ultrastructural immunolocalization of periodic proteins. Plant Physiol. 110:1231-1237. pectin depositions in the cell wall of Nicotiana tabacum pollen tubes. Protoplasma 187:168-171. Ghosh, B., Perry, M.P., and Marsh, D.G. 1991. Cloning the Fraley, R.T. 1989. Isolation of tissue-specific cDNAs from tomato pistils. Plant Cell 1:15-24. Gatehouse, J.A., Bown, D., Evans, I.M., Gatehouse, L.N., Jobes, D., Preston, P., and Croy, R.R.D. 1987. Sequence of the seed lectin gene from pea {Pisum
References
571
cDNA encoding the AmbtV allergen from giant rag- Giuliano, G., Rosellini, D., and Terzi, M. 1983. A new method for the purification of the different stages of weed (.Ambrosia trifida) pollen. Gene 101:231-238. carrot embryoids. Plant Cell Rep. 2:216-218. Ghosh, S., and Shivanna, K.R. 1980a. Pollen-pistil interaction in Linum grandiflorum. Scanning electron micro- Glab, N., Wise, R.P., Pring, D.R., Jacq, C, and Slonimski, P. 1990. Expression in Saccharomyces cerevisiae of a scopic observations and proteins of the stigma gene associated with cytoplasmic male sterility from surface. Planta 149:257-261. maize: respiratory dysfunction and uncoupling of Ghosh, S., and Shivanna, K.R. 1980b. Pollen-pistil interyeast mitochondria. Mol. Gen. Genet. 223:24-32. action in Linum grandiflorum: stigma-surface proteins and stigma receptivity. Proc. Indian Natl Sci. Acad. Glavin, T.L., Goring, D.R., Schafer, U., and Rothstein, S.J. 46B:177-183. 1994. Features of the extracellular domain of the Slocus receptor kinase from Brassica. Mol. Gen. Genet. Ghosh, S., and Shivanna, K.R. 1982a. Studies on 244:630-637. pollen-pistil interaction in Linum grandiflorum. Phytomorphology 32:385-395. Gleeson, P.A., and Clarke, A.E. 1979. Structural studies on the major component of Gladiolus style mucilage, Ghosh, S., and Shivanna, K.R. 1982b. Anatomical and arabinogalactan-protein. Biochem. ]. 181:607-621. cytochemical studies on the stigma and style in some Gleeson, P.A., and Clarke, A.E. 1980a. Comparison of the legumes. Bot. Gaz. 143:311-318. structures of the major components of the stigma Gibbs, P.E., and Bianchi, M. 1993. Post-pollination events and style secretions of Gladiolus: the arabino-3,6in species of Chorisia (Bombacaceae) and Tabebuia galactans. Carbohydr. Res. 83:187-192. (Bignoniaceae) with late-acting self-incompatibility. Gleeson, P.A., and Clarke, A.E. 1980b. Antigenic determiBot. Ada 106:64-71. nants of a plant proteoglycan, the Gladiolus style araGiese, H. 1992. Replication of DNA during barley binogalactan-protein. Biochem. J. 191:437-447. endosperm development. Can. ]. Bot. 70:313-318. Giese, H., Andersen, B., and Doll, H. 1983. Synthesis of Gleeson, P.A., and Clarke, A.E. 1980c. Arabinogalactans of sexual and somatic tissues of Gladiolus and Lilium. the major storage protein, hordein, in barley. PulsePhytochemistry 19:1777-1782. labeling study of grain filling in liquid-cultured detached spikes. Planta 159:60-65. Glenk, H.-O., Schimmer, O., and Wagner, W. 1970. Die Calcium-Verteilung in Oenof/iera-Pflanzen und ihr Giese, H., and Hejgaard, J. 1984. Synthesis of salt-soluble moglicher Einflufi auf den Chemotropismus der proteins in barley. Pulse-labeling study of grain filling Pollenschlauche und auf die Befruchtung. Phyton in liquid-cultured detached spikes. Planta 161:172-177. Austria 14:97-111. Gifford, D.J. 1988. An electrophoretic analysis of the seed proteins from Pinus monticola and eight other species Glover, D.E., and Barrett, S.C.H. 1983. Trimorphic incompatibility in Mexican populations of Pontederia sagitof pine. Can. J. Bot. 66:1808-1812. tata Presl. (Pontederiaceae). New Phytol. 95:439-455. Gifford, D.J., Greenwood, J.S., and Bewley, J.D. 1982. Deposition of matrix and crystalloid storage pro- Goblet, J.-P., Flamand, M.-C, and Briquet, M. 1985. A mitochondrial plasmid specifically associated with teins during protein body development in the male sterility and its relation with other mitochonendosperm of Ricinus communis L. cv. Hale seeds. drial plasmids in Viciafaba L. Curr. Genet. 9:423-426. Plant Physiol. 69:1471-1478. Giles, K.L., Bassett, H.C.M., and Eastin, J.D. 1975. The Goblet, J.-P, Boutry, M., Due, G., and Briquet, M. 1983. structure and ontogeny of the hilum region in Mitochondrial plasmid-like molecules in fertile and male sterile Viciafaba L. Plant Mol. Biol. 2:305-309. Sorghum bicolor. Aust. ]. Bot. 23:795-802. Gilissen, L.J.W. 1977. The influence of relative humidity Goday, A., Jensen, A.B., Culi^nez-Macia, F.A., Mar Alba, on the swelling of pollen grains in vitro. Planta M., Figueras, M., Serratosa, J., Torrent, M., and Pages, M. 1994. The maize abscisic acid-responsive protein 137:299-301. Rabl7 is located in the nucleus and interacts with Gillissen, L.J.W., and Brantjes, N.B.M. 1978. Function of nuclear localization signals. Plant Cell 6:351-360. the pollen coat in different stages of the fertilization process. Ada Bot. Neerl. 27:205-212. Goday, A., Sanchez-Martinez, D., G6mez, J., Puigdomenech, P., and Pages, M. 1988. Gene expresGiorgetti, L., Vergara, M.R., Evangelista, M., Lo Schiavo, F., Terzi, M., and Nuti Ronchi, V. 1995. On the occursion in developing Zea mays embryos: regulation by rence of somatic meiosis in embryogenic carrot cell abscisic acid of a highly phosphorylated 23- to 25-kD group of proteins. Plant Physiol. 88:564-569. cultures. Mol. Gen. Genet. 246:657-662. Giovinazzo, G., Manzocchi, L.A., Bianchi, M.W., Godfrey, C.A., and Linskens, H.F. 1968. Nucleic acid estiCoraggio, I., and Viotti, A. 1992. Functional analysis mations in pollinated styles of Petunia hybrida. Planta of the regulatory region of a zein gene in transiently 80:185-190. transformed protoplasts. Plant Mol. Biol. 19:257-263. Godineau, J.-C. 1966. Infrastructure du sac embryonnaire Giranton, J.-L., Ariza, M.J., Dumas, C., Cock, J.M., and du Crepis tectorum L.: les cellules du p61e micropylaire. Compt. Rend. Acad. Sci. Paris 263D:852-855. Gaude, T. 1995. The S locus receptor kinase gene encodes a soluble glycoprotein corresponding to the Godineau, J.-C. 1969. Ultra-structure des synergides chez quelques composers. Rev. Cytol. Biol. Veget. SRK extracellular domain in Brassica oleracea. Plant]. 32:209-226. 8:827-834. Giraudat, J., Hauge, B.M., Valon, C, Smalle, J., Parcy, E, Godwin, H. 1968. The origin of the exine. New Phytol. and Goodman, H.M. 1992. Isolation of the 67:667-676. Arabidopsis ABB gene by positional cloning. Plant Godwin, H., Echlin, P., and Chapman, B. 1967. The develCell 4:1251-1261. opment of the pollen grain wall in Ipomoea purpurea (L.) Roth. Rev. Palaeobot. Palynol. 3:181-195. Giuliano, G., Lo Schiavo, F., and Terzi, M. 1984. Isolation and developmental characterization of temperature- Goffner, D., This, P., and Delseny, M. 1990. Effects of abscisic sensitive carrot cell variants. Theor. Appl. Genet. acid and osmotica on helianthinin gene expression in 67:179-183. sunflower cotyledons in vitro. Plant Sci. 66:211-219.
572 Goldberg, R.B., Barker, S.J., and Perez-Grau, L. 1989. Regulation of gene expression during plant embryogenesis. Cell 56:149-160. Goldberg, R.B., de Paiva, G., and Yadegari, R. 1994. Plant embryogenesis: zygote to seed. Science 266:605-614. Goldberg, R.B., Hoschek, G., Ditta, G.S., and Breidenbach, R.W. 1981a. Developmental regulation of cloned superabundant embryo mRNAs in soybean. Devel Biol. 83:218-231. Goldberg, R.B., Hoschek, G., Tarn, S.H., Ditta, G.S., and Breidenbach, R.W. 1981b. Abundance, diversity, and regulation of mRNA sequence sets in soybean embryogenesis. Devel Biol. 83:201-217. Goldberg, R.B., Hoschek, G., and Vodkin, L.O. 1983. An insertion sequence blocks the expression of a soybean lectin gene. Cell 33:465-475. Goldman, M.H.S., Goldberg, R.B., and Mariani, C. 1994. Female sterile tobacco plants are produced by stigma-specific cell ablation. EMBO ]. 13:2976-2984. Goldman, M.H.S., Pezzotti, M., Seurinck, ]., and Mariani, C. 1992. Developmental expression of tobacco pistilspecific genes encoding novel extensin-like proteins. Plant Cell 4:1041-1051. Goldsbrough, P.B., Gelvin, S.B., and Larkins, B.A. 1986. Expression of maize zein genes in transformed sunflower cells. Mol. Gen. Genet. 202:374-381. Golubovskaya, I., Avalkina, N.A., and Sheridan, W.F. 1992. Effects of several meiotic mutations on female meiosis in maize. Devel Genet. 13:411-424. Golubovskaya, I.N. 1989. Meiosis in maize: mei genes and conception of genetic control of meiosis. Adv. Genet. 26:149-192. Golynskaya, E.L., Bashkirova, N.V., and Tomchuk, N.N. 1976. Phytohemagglutinins of the pistil in Primula as possible proteins of generative incompatibility. Soviet Plant Physiol. 23:69-76. Golz, J.F., Clarke, A.E., and Newbigin, E. 1995. Selfincompatibility in flowering plants. Curr. Op. Genet. Devel. 5:640-645. G6mez, ]., Sdnchez-Martinez, D., Stiefel, V., Rigau, ]., Puigdomenech, P., and Pages, M. 1988. A gene induced by the plant hormone abscisic acid in response to water stress encodes a glycine-rich protein. Nature 334:262-264. Gong, Z., Shen, W., Zhou, G., Huang,)., and Qian, S. 1988. Introducing exogenous DNA into plants after pollination - DNA traverse pathway of pollen tube to reach embryonic sac. Sci. Sinica 31B.-1080-1084. Gonzdlez-Melendi, P., Testillano, P.S., Ahmadian, P., Fad6n, B., Vicente, O., and Risuefio, M.C. 1995. In situ characterization of the late vacuolate microspore as a convenient stage to induce embryogenesis in Capsicum. Protoplasma 187:60-71. Gordon-Kamm, W.J., Michael-Spencer, T., Mangano, M.L., Adams, T.R., Daines, R.J., Start, W.G., O'Brien, J.V., Chambers, S.A., Adams, W.R., Jr., Willetts, N.G., Rice, T.B., Mackey, C.J., Krueger, R.W., Kausch, A.P., and Lemaux, P.G. 1990. Transformation of maize cells and regeneration of fertile transgenic plants. Plant Cell 2:603-618. Gori, P. 1976. Protein bodies in the nucellus of Euphorbia helioscopia. }. Ultrastr. Res. 54:53-58. Gori, P. 1977. Wall ingrowths in the embryo sac of Euphorbia helioscopia. Israel ]. Bot. 26:202-208. Gori, P. 1987. The fine structure of the developing Euphorbia dulcis endosperm. Ann. Bot. 60:563-569. Goring, D.R., and Rothstein, S.J. 1992. The S-locus recep-
References
tor kinase gene in a self-incompatible Brassica napus line encodes a functional serine/threonine kinase. Plant Cell 4:1273-1281. Goring, D.R., Banks, P., Beversdorf, W.D., and Rothstein, S.J. 1992a. Use of the polymerase chain reaction to isolate an S-locus glycoprotein cDNA introgressed from Brassica campestris into B. napus ssp. oleifera. Mol. Gen. Genet. 234:185-192. Goring, D.R., Banks, P., Fallis, L., Baszczynski, C.L., Beversdorf, W.D., and Rothstein, S.J. 1992b. Identification of an S-locus glycoprotein allele introgressed from B. napus ssp. rapifera to B. napus ssp. oleifera. Plant J. 2:983-989. Goring, D.R., Glavin, T.L., Schafer, U., and Rothstein, S.J. 1993. An S receptor kinase gene in self-compatible Brassica napus has a 1-bp deletion. Plant Cell 5:531-539. G6rska-Brylass, A. 1967a. Temporary callose wall in the generative cell of pollen grains. Naturwissenschaften 54:230-231. Gorska-Brylass, A. 1967b. Transitory callose envelope surrounding the generative cell in pollen grains. Ada Soc. Bot. Polon. 36:419-422. G6rska-Brylass, A. 1970. The "callose stage" of the generative cells in pollen grains. Grana 10:21-30. G6rska-Brylass, A., Bednarska, E., and Swierzowicz, I. 1986. At which morphological maturation stage of pollen grain takes place DNA replication? Acta Univ. N. Copernici 28:13-19. Gorst, ]., Overall, R.L., and Wernicke, W. 1987. Ionic currents traversing cell clusters from carrot suspension cultures reveal perpetuation of morphogenetic potential as distinct from induction of embryogenesis. Cell Viffn 21:101-109. Goto, K., and Meyerowitz, E.M. 1994. Function and regulation of the Arabidopsis floral homeotic gene P1STILLATA. Genes Devel. 8:1548-1560. Gottschalk, W., and Kaul, M.L.H. 1974. The genetic control of microsporogenesis in higher plants. Nucleus 17:133-166. Gottschalk, W., and Klein, H.D. 1976. The influence of mutated genes on sporogenesis. A survey on the genetic control of meiosis in Pisum sativum. Theor. Appl. Genet. 48:23-34. Gould, K.S., and Lord, E.M. 1988. Growth of anthers in Lilium longiflorum. A kinematic analysis. Planta 173:161-171. Goupil, P., Hatzopoulos, P., Franz, G., Hempel, F.D., You, R., and Sung, Z.R. 1992. Transcriptional regulation of a seed-specific carrot gene, DC8. Plant Mol. Biol. 18:1049-1063. Grafi, G., and Larkins, B.A. 1995. Endoreduplication in maize endosperm: involvement of M phase-promoting factor inhibition and induction of S phaserelated kinases. Science 269:1262-1264. Grafl, I. 1941. Ober das Wachstum der Antipodenkerne von Caltha palustris. Chromosoma 2:1-11. Graham, J.S.D., Jennings, A.C., Morton, R.K., Palk, B.A., and Raison, J.K. 1962. Protein bodies and protein synthesis in developing wheat endosperm. Nature 196:967-969. Graham, T.A., and Gunning, B.E.S. 1970. Localization of legumin and vicilin in bean cotyledon cells using fluorescent antibodies. Nature 228:81-82. Grambow, H.J., Kao, K.N., Miller, R.A., and Gamborg, O.L. 1972. Cell division and plant development from protoplasts of carrot cell suspension cultures. Planta 103:348-355.
573
References
in the genie male-sterile (ms 9) mutant of corn Zea Grant, I., Beversdorf, W.D., and Peterson, R.L. 1986. A mays. Can. ]. Genet. Cytol. 22:153-166. comparative light and electron microscopic study of microspore and tapetal development in male fertile Griffith, I.J., Smith, P.M., Pollock, J., Theerakulpisut, P., Avjioglu, A., Davies, S., Hough, T., Singh, M.B., and cytoplasmic male sterile oilseed rape (Brassica Simpson, R.J., Ward, L.D., and Knox, R.B. 1991. napus). Can. J. Bot. 64:1055-1068. Cloning and sequencing of Lol pi, the major allerGrant, M., and Bevan, M.W. 1994. Asparaginase gene genic protein of rye-grass pollen. FEBS Lett. expression is regulated in a complex spatial and 279:210-215. temporal pattern in nitrogen-sink tissues. Plant J. 5:695-704. Grill, L.K., and Garger, S.J. 1981. Identification and characterization of double-stranded RNA associated Gray, J.E., McClure, B.A., Bonig, I., Anderson, M.A., and with cytoplasmic male sterility in Vicia faba. Proc. Clarke, A.E. 1991. Action of the style product of the Natl Acad. Sci. USA 78:7043-7046. self-incompatibility gene of Nicotiana alata (S-RNase) on inOTfro-grownpollen tubes. Plant Cell 3:271-283. Grote, M., Swoboda, I., Meagher, R.B., and Valenta, R. 1995. Localization of profilin- and actin-like immunoreacGraybosch, R.A., and Palmer, R.G. 1985a. Male sterility in tivity in in uitro-germinated tobacco pollen tubes by soybean (Glycine max). I. Phenotypic expression of electron microscopy after special water-free fixation the ms2 mutant. Am. ]. Bot. 72:1738-1750. techniques. Sex. Plant Reprod. 8:180-186. Graybosch, R.A., and Palmer, R.G. 1985b. Male sterility in soybean (Glycine max). II. Phenotypic expression of Grun, P., and Aubertin, M. 1966. The inheritance and expression of unilateral incompatibility in Solanum. the ms4 mutant. Am. ]. Bot. 72:1751-1764. Heredity 21:131-138. Graybosch, R.A., Bernard, R.L., Cremeens, C.R., and Palmer, R.G. 1984. Genetic and cytological studies of Grun, P., and Radlow, A. 1961. Evolution of barriers to crossing of self-incompatible with self-compatible a male-sterile, female-fertile soybean mutant. A new species of Solanum. Heredity 16:137-143. male-sterile gene (ms2) in Glycine max (L.) Merr. /. Hexed. 75:383-388. Gu, Q., Kawata, E.E., Morse, M.-J., Wu, H.-M., and Cheung, A.Y. 1992. A flower-specific cDNA encodGreen, C.E., and Donovan, CM. 1980. Effect of aspartateing a novel thionin in tobacco. Mol. Gen. Genet. derived amino acids and aminoethyl cysteine on 234:89-96. growth of excised mature embryos of maize. Crop Sci. 20:358-362. Gu, S.-R., Gui, Y.-L., and Xu, T.-Y. 1984. Effects of physical Green, C.E., and Phillips, R.L. 1975. Plant regeneration and chemical factors on induction frequency of the from tissue cultures of maize. Crop Sci. 15:417-421. pollen plantlet and changes in starch formation in anthers. Ada Bot. Sinica 26:156-162. Greene, EC. 1981. In vitro synthesis of wheat (Triticum aestivum L.) storage proteins. Plant Physiol. 68:778-783. Guerche, P., de Almeida, E.R.P., Schwarztein, M.A., Gander, E., Krebbers, E., and Pelletier, G. 1990a. Greene, F.C. 1983. Expression of storage protein genes in Expression of the 2S albumin from Bertholletia excelsa developing wheat (Triticumaestivum L.) seeds. Plant in Brassica napus. Mol. Gen. Genet. 221:306-314. Physiol. 71:40-46. Greenwood, J.S., and Chrispeels, M.J. 1985a. Correct tar- Guerche, P., Tire, C, Grossi de Sa, F., de Clercq, A., van Montagu, M., and Krebbers, E. 1990b. Differential geting of the bean storage protein phaseolin in the expression of the Arabidopsis 2S albumin genes and seeds of transformed tobacco. Plant Physiol. 79:65-71. the effect of increasing gene family size. Plant Cell Greenwood, J.S., and Chrispeels, M.J. 1985b. 2:469-478. Immunocytochemical localization of phaseolin and phytohemagglutinin in the endoplasmic reticulum Guerrero, F.D., Crossland, L., Smutzer, G.S., Hamilton, and Golgi complex of developing bean cotyledons. D.A., and Mascarenhas, J.P. 1990. Promoter sequences from a maize pollen-specific gene direct Planta 164:295-302. tissue-specific transcription in tobacco. Mol. Gen. Gregor, D., Reinert, J., and Matsumoto, H. 1974. Changes Genet. 224:161-168. in chromosomal proteins from embryo induced carGuha, S., and Maheshwari, S.C. 1964. In vitro production rot cells. Plant Cell Physiol. 15:875-881. of embryos from anthers of Datura. Nature 204:497. Gregory, P., Gracen, V.E., Yoder, O.C., and Steinkraus, N.A. 1977. Differential effects of digitonin on mito- Guha, S., and Maheshwari, S.C. 1966. Cell division and chondria isolated from male-sterile and non maledifferentiation of embryos in the pollen grains of sterile cytoplasms of corn. Plant Sci. Lett. 9:17-21. Datura in vitro. Nature 212:97-98. Greinwald, R., and Czygan, F.-C. 1991. Regeneration of Guha-Mukherjee, S. 1973. Genotypic differences in the HI vitro formation of embryoids from rice pollen. /. plantlets from callus cultures of Chamaecytisus purpureus and Chamaecytisus austriacus (Leguminosae). Expt. Bot. 24:139-144. Guignard, J.-L. 1984. The development of cotyledon and Bot. Ada 104:64-67. shoot apex in monocotyledons. Can. ]. Bot. Grelon, M., Budar, F., Bonhomme, S., and Pelletier, G. 62:1316-1318. 1994. Ogura cytoplasmic male-sterility (CMS)-associated orfl38 is translated into a mitochondrial mem- Guignard, L. 1899. Sur les anth^rozoi'des et le double copulation sexuelle chez les v£g6taux angiospermes. brane polypeptide in male-sterile Brassica cybrids. Mol. Gen. Genet. 243:540-547.
Rev. Gen. Bot. 11:129-135.
Gresshoff, P.M., and Doy, C.H. 1972a. Development and Guilford, W.J., Schneider, D.M., Labovitz, J., and Opella, S.J. 1988. High resolution solid state 13C NMR specdifferentiation of haploid Lycopersicon esculenlum troscopy of sporopollenins from different plant taxa. (tomato). Planta 107:161-170. Plant Physiol. 86:134-136. Gresshoff, P.M., and Doy, C.H. 1972b. Haploid Arabidopsis thaliana callus and plants from anther culture. Aust. Guilluy, C.-M., Trick, M., Heizmann, P., and Dumas, C. }. Bid. Sci. 25:259-264. 1991. PCR detection of transcripts homologous to Greyson, R.I., Walden, D.B., and Cheng, P.C. 1980. LM, the self-incompatibility gene in anthers of Brassica. TEM and SEM observations of anther development Theor. Appl. Genet. 82:466-472.
574 Guiltinan, M.J., Marcotte, W.R., Jr., and Quatrano, R.S. 1990. A plant leucine zipper protein that recognizes an abscisic acid responsive element. Science 250:267-271. Guldager, P. 1978. Immunoelectrophoretic analysis of seed proteins from Pisum sativum L. Theor. Appl. Genet. 53:241-250. Guo, F.L., and Hu, S.Y. 1995. Cytological evidence of biparental inheritance of plastids and mitochondria in Pelargonium. Protoplasma 186:201-207. Guo, Yv Liang, H., and Berns, M.W. 1995. Laser-mediated gene transfer in rice. Physiol. Plant. 93:19-24. Guo, Z.-S., Sun, A.-C., Wang, Y.-Y., Gui, Y.-L., Gu, S.-R., and Miao, S.-H. 1978. Studies on induction of pollen plants and androgenesis in maize. Ada Bot. Sinica 20:204-209. Gupta, M., Chourey, P.S., Burr, B., and Still, P.E. 1988. cDNAs of two non-allelic sucrose synthase genes in maize: cloning, expression, characterization and molecular mapping of the sucrose synthase-2 gene. Plant Mol. Biol. 10:215-224. Gupta, S.C., and Babbar, S.B. 1980. Enhancement of plantlet formation in anther cultures of Datura metel L. by pre-chilling of buds. Z. Pflanzenphysiol. 96:465-470. Gustafson-Brown, C., Savidge, B., and Yanofsky, M.F. 1994. Regulation of the Arabidopsisfloralhomeotic gene APETALA1. Cell 76:131-143. Gustavsson, H.-O., Ellerstrom, M, Stalberg, K., Ezcurra, I., Koman, A., Hoglund, A.-S., Rask, L., and Josefsson, L.-G. 1991. Distinct sequence elements in a napin promoter interact in vitro with DNA-binding proteins from Brassica napus. Physiol. Plant. 82:205-212. Guzzo, R, Baldan, B., Levi, M., Sparvoli, E., Lo Schiavo, F., Terzi, M., and Mariani, P. 1995. Early cellular events during induction of carrot explants with 2,4-D. Protoplasma 185:28-36. Guzzo, F., Baldan, B., Mariani, P., Lo Schiavo, R, and Terzi, M. 1994. Studies on the origin of totipotent cells in explants of Daucus carota L. /. Expt. Bot. 45:1427-1432. Gw6zdz, E.A., and Deckert, J.E. 1989. The formation and translational activity of polysomes from developing lupin seeds. Physiol. Plant. 75:208-214. Gyorgyey, ]., Gartner, A., Nemeth, K., Magyar, Z., Hirt, H., Heberle-Bors, E., and Dudits, D. 1991. Alfalfa heat shock genes are differentially expressed during somatic embryogenesis. Plant Mol. Biol. 16:999-1007.
References
labeling in anther tapetum during microsporogenesis. Plant Physiol. 95:861-870. Hackett, R.M., Lawrence, M.J., and Franklin, F.C.H. 1992. A Brassica S-locus related gene promoter directs expression in both pollen and pistil of tobacco. Plant ]. 2:613-617. Hadi, M.Z., McMullen, M.D., and Finer, J.J. 1996. Transformation of 12 different plasmids into soybean via particle bombardment. Plant Cell Rep. 15:500-505. Hagemann, R., and Schroder, M.-B. 1989. The cytological basis of the plastid inheritance in angiosperms. Protoplasma 152:57-64. Hager, K.-P, and Dank, N. 1996. Seed storage proteins of Cupressaceae are homologous to legumins from angiosperms: molecular characterization of cDNAs from incense cedar (Calocedrus decurrens [Torr.] Florin). Plant Sci. 116:85-96. Hager, K.-P., Braun, H., Czihal, A., Muller, B., and Baumlein, H. 1995. Evolution of seed storage protein genes: legumin genes of Ginkgo biloba. ]. Mol. Evol. 41:457-466. Hahne, G., Mayer, J.E., and Lorz, H. 1988. Embryogenic and callus-specific proteins in somatic embryogenesis of the grass, Dactylis glomerata L. Plant Sci. 55:267-279. Haig, D. 1990. New perspectives on the angiosperm female gametophyte. Bot. Rev. 56:236-274. Hakansson, A. 1957. Notes on the giant chromosomes of Allium nutans. Bot. Not. 110:196-200. Hakansson, G., Glimelius, K., and Bonnett, H.T. 1990. Respiration in cells and mitochondria of male-fertile and male-sterile Nicotiana spp. Plant Physiol. 93:367-373. Hakman, I., Fowke, L.C., von Arnold, S., and Eriksson, T. 1985. The development of somatic embryos in tissue cultures initiated from immature embryos of Picea abies (Norway spruce). Plant Sci. 38:53-59. Hakman, I., Stabel, P., Engstrom, P., and Eriksson, T. 1990. Storage protein accumulation during zygotic and somatic embryo development in Picea abies (Norway spruce). Physiol. Plant. 80:441-445. Halford, N.G., Forde, }., Anderson, O.D., Greene, F.C., and Shewry, PR. 1987. The nucleoride and deduced amino acid sequences of an HMW glutenin subunit gene from chromosome IB of bread wheat (Triticum aestivum L.) and comparison with those genes from chromosomes 1A and ID. Theor. Appl. Genet. 75:117-126. Habben, J.E., Kirleis, A.W., and Larkins, B.A. 1993. The Halford, N.G., Forde, ]., Shewry, PR., and Kreis, M. 1989. Functional analysis of the upstream regions of a origin of lysine-containing proteins in opaque-2 silent and an expressed member of a family of wheat maize endosperm. Plant Mol. Biol. 23:825-838. seed protein genes in transgenic tobacco. Plant Sci. Habben, J.E., Moro, G.L., Hunter, B.G., Hamaker, B.R., 62:207-216. and Larkins, B.A. 1995. Elongation factor 1 concentration is highly correlated with the lysine content of Hall, T.C., McLeester, R.C., and Bliss, F.A. 1972. maize endosperm. Proc. Natl Acad. Sci. USA Electrophoretic analysis of protein changes during 92:8640-8644. the development of the French bean fruit. Phytochemistry 11:647-649. Haccius, B. 1978. Question of unicellular origin of nonzygotic embryos in callus cultures. Phytomorphology Hall, T.C., Ma, Y, Buchbinder, B.U., Pyne, J.W., Sun, S.M., 28:74-81. and Bliss, F.A. 1978. Messenger RNA for Gl protein of French bean seeds: cell-free translation and prodHaccius, B., and Reichert, H. 1964. Restitutionserscheinungen uct characterization. Proc. Natl Acad. Sci. USA an pflanzlichen Meristemen nach Rontgenbestrahlung. 75:3196-3200. II. Advetiv-Embryonie nach Samenbestrahlung von Eranthis hiemalis. Planta 62:355-372. Halperin, W. 1966. Alternative morphogenetic events in Hack, E., Lin, C, Yang, H., and Homer, H.T. 1991. Tcell suspensions. Am. J. Bot. 53:443^153. URF13 protein from mitochondria of Texas male- Halperin, W., and Jensen, W.A. 1967. Ultrastructural changes during growth and embryogenesis in carrot sterile maize (Zea mays L.). Its purification and cell cultures. /. Ultrastr. Res. 18:428-443. submitochondrial localization, and immunogold
References
Halperin, W., and Wetherell, D.F. 1965. Ammonium requirement for embryogenesis in vitro. Nature 205:519-520. Hamilton, D.A., Bashe, D.M., Stinson, J.R., and Mascarenhas, J.P. 1989. Characterization of a pollenspecific genomic clone from maize. Sex. Plant Reprod. 2:208-212. Hamilton, D.A., Roy, M., Rueda, J., Sindhu, R.K., Sanford, J., and Mascarenhas, J.P. 1992. Dissection of a pollenspecific promoter from maize by transient transformation assays. Plant Mol. Biol. 18:211-218. Hammond-Kosack, M.C.U., Holdsworth, M.J., and Bevan, M.W. 1993. In vivo footprinting of a low molecular weight glutenin gene (LMWG-1D1) in wheat endosperm. EMBO J. 12:545-554. Hannah, L.C., and Nelson, O.E., Jr. 1976. Characterization of ADP-glucose pyrophosphorylase from shrunken-2 and brittle-2 mutants of maize. Biochem. Genet. 14:547-560. Hannah, L.C., Tuschall, D.M., and Mans, R.J. 1980. Multiple forms of maize endosperm ADP-glucose pyrophosphorylase and their control by shrunken-2 and brittle-2. Genetics 95:961-970. Hannig, E. 1904. Zur Physiologie pflanzlicher Embryonen. Ueber die Cultur von Cruciferen-Embryonen ausserhalb der Embryosacks. Bot. Ztg 62:45-80. Hansen, D.J., Bellman, S.K., and Sacher, R.M. 1976. Gibberellic acid-controlled sex expression of corn tassels. Crop. Sci. 16:371-374. Hansen, G., Estruch, J.J., Sommer, H., and Spena, A. 1993. NTGLO: a tobacco homologue of the GLOBOSA floral homeotic gene of Antirrhinum majus: cDNA sequence and expression pattern. Mol. Gen. Genet. 239:310-312. Hanson, D.D., Hamilton, D.A., Travis, J.L., Bashe, D.M., and Mascarenhas, J.P. 1989. Characterization of a pollen-specific cDNA clone from "Lea mays and its expression. Plant Cell 1:173-179. Hanson, M.R., and Conde, M.F. 1985. Functioning and variation of cytoplasmic genomes: lessons from cytoplasmic-nuclear interactions affecting male fertility in plants. Int. Rev. Cytol. 94:213-267. Hara-Nishimura, I., Nishimura, M., and Akazawa, T. 1985. Biosynthesis and intracellular transport of US globulin in developing pumpkin cotyledons. Plant Physiol. 77:747-752. Harada, J.J., Baden, C.S., and Comai, L. 1988. Spatially regulated genes expressed during seed germination and postgerminative development are activated during embryogeny. Mol. Gen. Genet. 212:466^173. Harada, J.J., Barker, S.J., and Goldberg, R.B. 1989. Soybean (3-conglycinin genes are clustered in several DNA regions and are regulated by transcriptional and posttranscriptional processes. Plant Cell 1:415^125. Harada, J.J., DeLisle, A.J., Baden, C.S., and Crouch, M.L. 1989. Unusual sequence of an abscisic acid-inducible mRNA which accumulates late in Brassica napus seed development. Plant Mol. Biol. 12:395-401. Hardenack, S., Ye, D., Saedler, H., and Grant, S. 1994. Comparison of MADS box gene expression in developing male and female flowers of the dioecious plant white campion. Plant Cell 6:1775-1787. Hardham, A.R. 1976. Structural aspects of the pathways of nutrient flow to the developing embryo and cotyledons of Pisum sativum L. Aust. J. Bot. 24:711-721. Hari, V. 1980. Effect of cell density changes and condi-
575 tioned media on carrot cell embryogenesis. Z. Pflanzenphysiol. 96:227-231. Harikrishna, K., Jampates-Beale, R., Milligan, S.B., and Gasser, C.S. 1996. An endochitinase gene expressed at high levels in the stylar transmitting tissue of tomatoes. Plant Mol. Biol. 30:899-911. Haring, V., Gray, J.E., McClure, B.A., Anderson, M.A., and Clarke, A.E. 1990. Self-incompatibility: a self-recognition system in plants. Science 250:937-941. Harris, N. 1979. Endoplasmic reticulum in developing seeds of Vicia faba. A high voltage electron microscope study. Planta 146:63-69. Harris, N., and Boulter, D. 1976. Protein body formation in cotyledons of developing cowpea (Vigna unguiculata) seeds. Ann. Bot. 40:739-744. Harris, N., and Chaffey, N.J. 1986. Plasmarubules - real modifications of the plasmalemma. Nord. ]. Bot. 6:599-607. Harris, N., and Croy, R.R.D. 1986. Localization of mRNA for pea legumin: in situ hybridization using a biotinylated cDNA probe. Protoplasma 130:57-67. Harris, N., and Juliano, B.O. 1977. infrastructure of endosperm protein bodies in developing rice grains differing in protein content. Ann. Bot. 41:1-5. Harris, N., Grindley, H. Mulchrone, J., and Croy, R.R.D. 1989. Correlated in situ hybridisation and immunochemical studies of legumin storage protein deposition in pea (Pisum sativum L.). Cell Biol. Int. Rep. 13:23-35. Harris, P.J., Weinhandl, J.A., and Clarke, A.E. 1989. Effect on in vitro pollen growth of an isolated style glycoprotein associated with self-incompatibility in Nicotiana alata. Plant Physiol. 89:360-367. Harris, P.J., Freed, K., Anderson, M.A., Weinhandl, J.A., and Clarke, A.E. 1987. An enzyme-linked immunosorbent assay (ELISA) for in vitro pollen growth based on binding of a monoclonal antibody to pollen tube surface. Plant Physiol. 84:851-855. Hartings, H., Lazzaroni, N., Marsan, P.A., Aragay, A., Thompson, R., Salamini, F., di Fonzo, N., Palau, J., and Motto, M. 1990. The b-32 protein from maize endosperm: characterization of genomic sequences encoding two alternative central domains. Plant Mol. Biol. 14:1031-1040. Hartings, H., Maddaloni, M., Lazzaroni, N., di Fonzo, N., Motto, M., Salamini, F., and Thompson, R. 1989. The 02 gene which regulates zein deposition in maize endosperm encodes a protein with structural homologies to transcriptional activators. EMBO ]. 8:2795-2801. Harwood, J.L., Sodja, A., Stumpf, P.K., and Spurr, A.R. 1971. On the origin of oil droplets in maturing castor bean seeds, Ricinus communis. Lipids 6:851-854. Hasenkampf, C, Qureshi, M., Horsch, A., and Riggs, CD. 1992. Temporal and spatial distribution of meiotin-1 in anthers of Lilium longiflorum. Devel Genet. 13:425-434. Hasitschka, G. 1956. Bildung von Chromosomenbiindeln nach Art der Speicheldriisenchromosomen, spiralisierte Ruhekernchromosomen und andere Struktureigentumlichkeiten in den endopolyploiden Riesenkernen der Antipoden von Papaver rhoeas. Chromosoma 8:87-113. Hasitschka-Jenschke, G. 1957. Die Entwicklung der Samenanlage von Allium ursinum mit besonderer Beriicksichtigung der endopolyploiden Kerne in Synergiden und Antipoden. Osterr. Bot. Z. 104:1-24.
576
References
Hasitschka-Jenschke, G. 1958. Zur Karyologie der Microtubular and actin filament configurations durSamenanlage dreier Allium-Aiten. Osterr. Bot. Z. ing microspore and pollen development in Brassica 105:71-82. napus cv. Topas. Can. ]. Bot. 70:1369-1376. Hasitschka-Jenschke, G. 1959. Vergleichende karyologis- Hause, G., and Schroder, M.-B. 1986. Reproduction in che Untersuchungen an Antipoden. Chromosoma Triticale. I. The structure of the mature embryo sac. 10:229-267. Biol. Zentral. 105:511-517. Hasitschka-Jenschke, G. 1962. Notizen tiber endopoly- Hause, G., and Schroder, M.-B. 1987. Reproduction in ploide Kerne im Bereich der Samenanlage von Triticale. 2. Karyogamy. Protoplasma 139:100-104. Angiospermen. Osterr. Bot. Z. 109:125-137. Hause, G., Cordewener, J.H.G., Ehrmanova, M., Hause, B., Binarova, P., van Lookeren Campagne, M.M., and Haskell, D.W., and Rogers, O.M. 1985. RNA synthesis by van Lammeren, A.A.M. 1995. Cell cycle dependent vegetative and sperm nuclei of trinucleate pollen. distribution of phosphorylated proteins in Cytologia 50:805-809. microspores and pollen of Brassica napus L., detected Hatton, D., Sablowski, R., Yung, M.-H., Smith, C, Schuch, by the monoclonal antibody MPM-2. Protoplasma W., and Bevan, M. 1995. Two classes of cis sequences 187:117-126. contribute to tissue-specific expression of a PAL2 promoter in transgenic tobacco. Plant ]. 7:859-876. Haufier, I., Herth, W., and Reiss, H.-D. 1984. Calmodulin in tip-growing plant cells, visualized by fluorescing Hattori, T., Terada, T., and Hamasuna, S.T. 1994. Sequence calmodulin-binding phenothiazines. Planta 162:33-39. and functional analyses of the rice gene homologous to the maize Vpl. Plant Mol. Biol. 24:805-810. Hauxwell, A.J., Corke, F.M.K., Hedley, C.L., and Wang, Hattori, T., Terada, T., and Hamasuna, S. 1995. Regulation T.L. 1990. Storage protein gene expression is of Osem gene by abscisic acid and the transcriptional localised to regions lacking mitotic activity in develactivator VP1: analysis of cis-acting promoter eleoping pea embryos. An analysis of seed developments required for regulation by abscisic acid and ment in Pisum sativum XIV. Development 110:283-289. VP1. Plant]. 7:913-925. Havel, L., and NovSk, F.J. 1981. In vitro pollination of maize (Zea mays L.) - proof of double fertilization. Hattori, T., Vasil, V, Rosenkrans, L., Hannah, L.C., Plant Cell Rep. 1:26-28. McCarty, D.R., and Vasil, I.K. 1992. The viviparous-1 gene and abscisic acid activate the Cl regulatory Hayman, D.L. 1956. The genetical control of incompatibilgene for anthocyanin biosynthesis during seed matity in Phalaris coerulescens Desf. Aust. ]. Biol. Sci. 9:321-331. uration in maize. Genes Devel. 6:609-618. Hatzopoulos, P., Fong, R, and Sung, Z.R. 1990. Abscisic He, C, Tirlapur, U., Cresti, M., Peja, M., Crone, D.E., and acid regulation of DC8, a carrot embryonic gene. Mascarenhas, J.P. 1996. An Arabidopsis mutant showPlant Physiol. 94:690-695. ing aberrations in male meiosis. Sex. Plant Reprod. 9:54-57. Hatzopoulos, P., Franz, G., Choy, L., and Sung, R.Z. 1990. Interaction of nuclear factors with upstream He, C.-P, and Yang, H.-Y. 1992. Ultracytochemical localization of calcium in the embryo sac of sunflower. sequences of a lipid body membrane protein gene from carrot. Plant Cell 2:457-467. Chinese ]. Bot. 4:99-106. Hauffe, K.D., Paszkowski, U., Schulze-Lefert, P., He, S., Lyznik, A., and Mackenzie, S. 1995. Pollen fertility restoration by nuclear gene Fr in CMS bean: nuclearHahlbrock, K., Dangl, J.L., and Douglas, C.J. 1991. A parsley 4CL-1 promoter fragment specifies complex directed alteration of a mitochondrial population. Genetics 139:955-962. expression patterns in transgenic tobacco. Plant Cell 3:435-443. Hearn, M.J., Franklin, F.C.H., and Ride, J.P. 1996. Identification of a membrane glycoprotein in pollen Haughn, G.W., Schultz, E.A., and Martinez-Zapater, J.M. of Papaver rhoeas which binds stigmatic self-incom1995. The regulation of flowering in Arabidopsis patibility (S-) proteins. Plant ]. 9:467-475. thaliana: meristems, morphogenesis and mutants. Heath, J.D., Weldon, R., Monnot, C, and Meinke, D.W. Can.]. Bot. 73:959-981. 1986. Analysis of storage proteins in normal and Haughn, G.W., and Somerville, C.R. 1988. Genetic control aborted seeds from embryo-lethal mutants of of morphogenesis in Arabidopsis. Devel Genet. Arabidopsis thaliana. Planta 169:304-312. 9:73-89. Hauptli, H., and Williams, S. 1988. Maize in vitro pollina- Heberle-Bors, E. 1982a. In vitro pollen embryogenesis in Nicotiana tabacum L. and its relation to pollen steriltion with single pollen grains. Plant Sci. 58:231-237. ity, sex balance, and floral induction of the pollen Hause, B., Hause, G., Pechan, P., and van Lammeren, donor plants. Planta 156:396-401. A.A.M. 1993. Cytoskeletal changes and induction of embryogenesis in microspore and pollen cultures of Heberle-Bors, E. 1982b. On the time of embryogenic Brassica napus L. Cell Biol. Int. 17:153-168. pollen grain induction during sexual development of Nicotiana tabacum L. plants. Planta 156:402-406. Hause, B., van Veenendaal, W.L.H., Hause, G., and van Lammeren, A.A.M. 1994. Expression of polarity dur- Heberle-Bors, E. 1983. Induction of embryogenic pollen grains in situ and subsequent in vitro pollen embryoing early development of microspore-derived and genesis in Nicotiana tabacum by treatments of the zygotic embryos of Brassica napus L. cv. Topas. Bot. Ada 107:407-415. pollen donor plants with feminizing agents. Physiol. Plant. 59:67-72. Hause, G. 1986. Organelle distribution during pollen development of Pisum sativum L. Biol. Zentral. Heberle-Bors, E. 1989. Isolated pollen culture in tobacco: 105:283-288. plant reproductive development in a nutshell. Sex. Plant Reprod. 2:1-10. Hause, G. 1991. Ultrastructural investigations of mature embryo sacs of Daucus carota, D. aureus and D. muri-Heberle-Bors, E., and Odenbach, W. 1985. In vitro pollen embryogenesis and cytoplasmic male sterility in catus - possible cytological explanations of paternal Triticum aestivum. Z. Pflanzenzuchtg 95:14-22. plastid inheritance. Sex. Plant Reprod. 4:288-292. Hause, G., Hause, B., and van Lammeren, A.A.M. 1992. Heberle-Bors, E., and Reinert, J. 1979. Androgenesis in
References
577
isolated pollen cultures of Nicotiana tabacum: depen- Hemerly, A.S., Ferreira, P., de Almeida Engler, J., van Montagu, M., Engler, G., and Inze, D. 1993. cdcla dence upon pollen development. Protoplasma expression in Arabidopsis is linked with competence 99:237-245. for cell division. Plant Cell 5:1711-1723. Heberle-Bors, E., and Reinert, J. 1980. Isolated pollen cultures and pollen dimorphism. Naturwissenschaften Hensgens, L.A.M., de Bakker, E.P.H.M., van Os-Ruygrok, E.P., Rueb, S., van de Mark, R, van der Maas, H., van 67:311. der Veen, S., Kooman-Gersmann, M., 't Hart, L., and Heberle-Bors, E., and Reinert, J. 1981. Environmental conSchilperoort, R.A. 1993. Transient and stable exprestrol and evidence for predetermination of pollen sion of gusA fusions with rice genes in rice, barley embryogenesis in Nicotiana tabacum pollen. and perennial ryegrass. Plant Mol. Biol. 23:643-669. Protoplasma 109:249-255. Hubert, D., Kikkert, J.R., Smith, F.D., and Reisch, B.I. 1993. Hepher, A., and Boulter, M.E. 1987. Pollen tube growth and fertilization efficiency in Salpiglossis sinuata: Optimization of biolistic transformation of embryoimplications for the involvement of chemotropic facgenie grape cell suspensions. Plant Cell Rep. tors. Ann. Bot. 60:595-601. 12:585-589. Hecht, N.B., and Stern, H. 1971. A late replicating DNA Hepler, P.K., and Jackson, W.T. 1968. Microtubules and early stages of cell-plate formation in the endosperm of protein complex from cells in meiotic prophase. Haemanthus katlierinae Baker. /. Cell Biol. 38:437-446. Expt. Cell Res. 69:1-10. Heck, G.R., Perry, S.E., Nichols, K.W., and Fernandez, Herd, Y.R., and Beadle, D.J. 1980. The site of the selfincompatibility mechanism in Tradescantia pallida. D.E. 1995. AGL15, a MADS domain protein Ann. Bot. 45:251-256. expressed in developing embryos. Plant Cell Herd, Y.R., and Steer, M.W. 1984. Microsporogenesis in 7:1271-1282. genie male-sterile lines of barley (Hordeum vulgare). Hedley, C.L., Lloyd, J.R., Ambrose, M.J., and Wang, T.L. Can. J. Bot. 62:1127-1135. 1994. An analysis of seed development in Pisum sativum. XVII. The effect of the rb locus alone and in Herdt, E., Siitfeld, R., and Wiermann, R. 1978. The occurrence of enzymes involved in phenylpropanoid combination with r on the growth and development metabolism in the tapetum fraction of anthers. Eur. ]. of the seed. Ann. Bot. 74:365-371. Cell Biol. 17:433-^41. Hedley, C.L., Smith, CM., Ambrose, M.J., Cook, S., and Wang, T.L. 1986. An analysis of seed development in Herman, E.M. 1987. Immunogold-localization and synPisum sativum. II. The effect of the r-locus on the thesis of an oil-body membrane protein in developgrowth and development of the seed. Ann. Bot. ing soybean seeds. Planta 172:336-345. 58:371-379. Hernandez-Fernandez, M.M., and Christie, B.R. 1989. Inheritance of somatic embryogenesis in alfalfa Heidecker, G., and Messing, J. 1983. Sequence analysis of {Medicago sativa L.). Genome 32:318-321. zein cDNAs obtained by an efficient mRNA cloning method. Nucl. Acids Res. 11:4891-4906. Hernould, M., Suharsono, S., Litvak, S., Araya, A., and Mouras, A. 1993. Male-sterility induction in transHeidecker, G., and Messing, J. 1986. Structural analysis of genic tobacco plants with an unedited atp9 mitoplant genes. Annu. Rev. Plant Physiol. y7-A2S-Aii6. chondrial gene from wheat. Proc. Natl Acad. Sci. USA Heim, U., Baumlein, H., and Wobus, U. 1994. The legumin 90:2370-2374. gene family: A reconstructed Viciafaba legumin gene encoding a high-molecular-weight subunit is related HSrouart, D., van Montagu, M., and Inze', D. 1994. to type B genes. Plant Mol. Biol. 25:131-135. Developmental and environmental regulation of the Nicotiana plumbaginifolia cytosolic Cu/Zn-superoxHeim, U., Manteuffel, R., Baumlein, H., Steinbiss, H.-H., ide dismutase promoter in transgenic tobacco. Plant and Wobus, U. 1995. Transient expression of a lysinePhysiol. 104:873-880. rich vicilin gene of Vicia faba in barley endosperm detected by immunological tissue printing after par- Herrero, M., and Dickinson, H.G. 1979. Pollen-pistil ticle bombardment. Plant Cell Rep. 15:125-138. incompatibility in Petunia hybrida: changes in the pisHeinlein, M., and Starlinger, R 1989. Tissue- and cell-spetil following compatible and incompatible intraspecific expression of the two sucrose synthase isoencific crosses. /. Cell Sci. 36:1-18. zymes in developing maize kernels. Mol. Gen. Genet. Herrero, M., and Dickinson, H.G. 1980. Ultrastructural 215:441-446. and physiological differences between buds and mature flowers of Petunia hybrida prior to and folHelleboid, S., Couillerot, J.-R, Hilbert, J.-L, and Vasseur, J. lowing pollination. Planta 148:138-145. 1995. Inhibition of direct somatic embryogenesis by a-difluoromethylarginine in a Cichorium hybrid: Herrero, M., and Dickinson, H.G. 1981. Pollen tube development in Petunia hybrida following compatible and effects on polyamine content and protein patterns. Planta 196:571-576. incompatible intraspecific matings. /. Cell Sci. 47:365-383. Helm, K.W., and Abernethy, R.H. 1990. Heat shock proteins and their mRNAs in dry and early imbibing Herth, W. 1978. Ionophore A 23187 stops tip growth, but embryos of wheat. Plant Physiol. 93:1626-1633. not cytoplasmic streaming, in pollen tubes of Lilium longiflorum. Protoplasma 96:275-282. Helsper, J.P.F.G. 1979. The possible role of lipid intermediates in the synthesis of p-glucans by a membrane Herth, W., Franke, W.W., Bittiger, H., Kuppel, A., and Keilich, G. 1974. Alkali-resistant fibrils of (3-1,3- and fraction from pollen tubes of Petunia hybrida. Planta 144:443-450. (3-1,4-glucans: structural polysaccharides in the Helsper, J.P.F.G., Linskens, H.F., and Jackson, J.F. 1984. pollen tube wall of Lilium longiflorum. Cytobiologie Phytate metabolism in Petunia pollen. Phytochemistry 9:344-367. Herz, M., and Brunori, A. 1985. Nuclear DNA content in 23:1841-1845. the endosperm of developing grain of hexaploid Helsper, J.P.F.G., Veerkamp, J.H., and Sassen, M.M.A. Triticales and parental species. Z. Pflanzenziichtg 1977. p-Glucan synthetase activity in Golgi vesicles of Petunia hybrida. Planta 133:303-308. 95:336-341.
578 Heslop-Harrison, J. 1957. The experimental modification of sex expression in flowering plants. Biol. Rev. 32:38-90. Heslop-Harrison, J. 1959. Photoperiod and fertility in
References
Heslop-Harrison, J. 1979a. Aspects of the structure, cytochemistry and germination of the pollen of rye (Secale cereale L.). Ann. Bot. 44: Suppl. 1,1-47.
Heslop-Harrison, J. 1979b. Pollen-stigma interaction in Rottboellia exaltata L.f. Ann. Bot. 23:345-349. grasses: a brief review. New Zealand J. Bot. Heslop-Harrison, J. 1962. Origin of exine. Nature 17:537-546. 195:1069-1071. Heslop-Harrison, J. 1979c. An interpretation of the hydrodynamics of pollen. Am. ]. Bot. 66:737-743. Heslop-Harrison, J. 1963a. An ultrastructural study of pollen wall ontogeny in Silene pendula. Grana Palynol. Heslop-Harrison, J. 1982. Pollen-stigma interaction and cross-incompatibility in the grasses. Science 4:1-24. 215:1358-1364. Heslop-Harrison, J. 1963b. Ultrastructural aspects of differentiation in sporogenous tissue. Symp. Soc. Expt. Heslop-Harrison, J. 1987. Pollen germination and pollentube growth. Int. Rev. Cytol. 107:1-78. Biol. 17:315-340. Heslop-Harrison, J. 1964. Cell walls, cell membranes and Heslop-Harrison, J., and Dickinson, H.G. 1967. A cycle of spherosome aggregation and disaggregation correprotoplasmic connections during meiosis and pollen lated with the meiotic divisions in Lilium. development. In Pollen Physiology and Fertilization, ed. H.F. Linskens, pp. 39-47. Amsterdam: NorthPhytomorphology 17:195-199. Holland Publishing Co. Heslop-Harrison, J., and Dickinson, H.G. 1969. Time relationships of sporopollenin synthesis associated with Heslop-Harrison, J. 1966a. Cytoplasmic continuities durtapetum and microspores in Lilium. Planta ing spore formation in flowering plants. Endeavour 84:199-214. 25:65-72. Heslop-Harrison, J. 1966b. Cytoplasmic connexions Heslop-Harrison, J., Heslop-Harrison, J.S., and HeslopHarrison, Y. 1986. The comportment of the vegetabetween angiosperm meiocytes. Ann. Bot. 30:221-230. tive nucleus and generative cell in the pollen and Heslop-Harrison, J. 1968a. Ribosome sites and S gene pollen tubes of Helleborus foetidus L. Ann. Bot. action. Nature 218:90-91. 58:1-12. Heslop-Harrison, J. 1968b. Pollen wall development. Science 161:230-237. Heslop-Harrison, J., and Heslop-Harrison, Y. 1973. Pollen-wall proteins: "gametophytic' and "sporoHeslop-Harrison, J. 1968c. Some fine structural features of phytic' fractions in the pollen walls of the intine growth in the young microspore of Lilium henMalvaceae. Ann. Bot. 37:403-412. ryi. Portug. Ada Biol. 10A:235-246. Heslop-Harrison, J. 1968d. The formation of the genera- Heslop-Harrison, J., and Heslop-Harrison, Y. 1975. Enzymic removal of the proteinaceous pellicle of the tive cell in massulate orchids. /. Cell Sci. 3:457-466. stigma papilla prevents pollen tube entry in the Heslop-Harrison, J. 1968e. Anther carotenoids and the Caryophyllaceae. Ann. Bot. 39:163-165. synthesis of sporopollenin. Nature 220:605. Heslop-Harrison, j . 1968f. Tapetal origin of pollen-coat Heslop-Harrison, J., and Heslop-Harrison, Y. 1980a. Cytochemistry and function of the Zwischenkorper substances in Lilium. New Phytol. 67:779-786. in grass pollens. Pollen Spores 22:5-10. Heslop-Harrison, J. 1968g. Wall development within the microspore tetrad of Lilium longiflorum. Can. J. Bot.Heslop-Harrison, J., and Heslop-Harrison, Y. 1980b. The pollen-stigma interaction in the grasses. I. Fine46:1185-1192. structure and cytochemistry of the stigmas of Heslop-Harrison, J. 1969a. The origin of surface features Hordeum and Secale. Ada Bot. Need. 29:261-276. of the pollen wall of Tagetes patula as observed by Heslop-Harrison, J., and Heslop-Harrison, Y. 1981. The scanning electron microscopy. Cytobios 2:177-186. pollen-stigma interaction in the grasses. 2. PollenHeslop-Harrison, J. 1969b. An acetolysis-resistant memtube penetration and the stigma response in Secale. brane investing tapetum and sporogenous tissue in Ada Bot. Neerl. 30:289-307. the anthers of certain Compositae. Can. J. Bot. Heslop-Harrison, J., and Heslop-Harrison, Y. 1982a The 47:541-542. growth of the grass pollen tube: 1. Characteristics of Heslop-Harrison, J. 1971a. Sporopollenin in the biological the polysaccharide particles ("P-particles") associcontext. In Sporopollenin, ed. J. Brooks, PR. Grant, M. ated with apical growth. Protoplasma 112:71-80. Muir, P. van Gijzel and G. Shaw, pp. 1-30. London: Heslop-Harrison, J., and Heslop-Harrison, Y. 1982b. The Academic Press. pollen-stigma interaction in the grasses. 4. An interHeslop-Harrison, J. 1971b. Wall pattern formation in pretation of the self-incompatibility response. Ada angiosperm microsporogenesis. Symp. Soc. Expt. Biol. 25:277-300. Bot. Neerl. 31:429-439. Heslop-Harrison, J. 1972. Sexuality of angiosperms. In Heslop-Harrison, J., and Heslop-Harrison, Y. 1982c. Pollen-stigma interaction in the Leguminosae: conPlant Physiology. A Treatise, Vol. VIC, ed. F.C. stituents of the stylar fluid and stigma secretion of Steward, pp. 133-289. New York: Academic Press. Trifolium pratense L. Ann. Bot. 49:729-735. Heslop-Harrison, J. 1975a. The physiology of the pollen grain surface. Proc. Roy. Soc. Lond. 190B:275-299. Heslop-Harrison, J., and Heslop-Harrison, Y. 1984. The disposition of gamete and vegetative-cell nuclei in Heslop-Harrison, J. 1975b. Incompatibility and the the extending pollen tubes of a grass species, pollen-stigma interaction. Annu. Rev. Plant Physiol. Alopecurus pratensis L. Ada Bot. Neerl. 33:131-134. 26:403-^125. Heslop-Harrison, J. 1976. The adaptive significance of the Heslop-Harrison, J., and Heslop-Harrison, Y. 1985. Germination of stress-tolerant Eucalyptus pollen. /. exine. In The Evolutionary Significance of the Exine, ed. Cell Sci. 73:135-157. I.K. Ferguson and J. Muller, pp. 27-37. London: Heslop-Harrison, J., and Heslop-Harrison, Y. 1987. An Academic Press. analysis of gamete and organelle movement in the Heslop-Harrison, J. 1978. Genetics and physiology of pollen tube of Secale cereale L. Plant Sci. 51:203-213. angiosperm incompatibility systems. Proc. Roy. Soc. Heslop-Harrison, J., and Heslop-Harrison, Y. 1988a. Lond. 202B:73-92.
References
579
Organelle movement and fibrillar elements of the Heslop-Harrison, J., Heslop-Harrison, Y, Cresti, M., Tiezzi, A., and Ciampolini, F. 1986. Actin during cytoskeleton in the angiosperm pollen tube. Sex. pollen germination. /. Cell Sci. 86:1-8. Plant Reprod. 1:16-24. Heslop-Harrison,J., and Heslop-Harrison, Y. 1988b. Some Heslop-Harrison, J., Heslop-Harrison, Y, Cresti, M., Tiezzi, A., and Moscatelli, A. 1988. Cytoskeletal elepermeability properties of angiosperm pollen ments, cell shaping and movement in the grains, pollen tubes and generative cells. Sex. Plant angiosperm pollen tube. /. Cell Sci. 91:49-60. Reprod. 1:65-73. Heslop-Harrison, J., and Heslop-Harrison, Y. 1988c. Heslop-Harrison, J., Heslop-Harrison, Y, Knox, R.B., and Howlett, B. 1973. Pollen-wall proteins: "gametoTubulin and male-gamete interconnections in the phytic' and 'sporophytic' fractions in the pollen pollen tubes of the grass Alppecurus pratensis. Ann. walls of the Malvaceae. Ann. Bot. 37:403-412. Bot. 61:249-254. Heslop-Harrison, J., and Heslop-Harrison, Y. 1988d. Sites Heslop-Harrison, J., f, R.B., Heslop-Harrison, Y, and Mattsson, O. 1975. Pollen-wall proteins: emission of origin of the peripheral microtubule system of the and role in incompatibility responses. In The Biology vegetative cell of the angiosperm pollen tube. Ann. of the Male Gamete, ed. J.G. Duckett and PA. Racey, Bot. 62:455-461. pp. 189-202. London: Academic Press. Heslop-Harrison, J., and Heslop-Harrison, Y. 1989a. Cytochalasin effects on structure and movement in Heslop-Harrison, J.S., Heslop-Harrison, J., HeslopHarrison, Y, and Reger, B.J. 1985. The distribution of the pollen tube of Iris. Sex. Plant Reprod. 2:27-37. calcium in the grass pollen tube. Proc. Roy. Soc. Lond. Heslop-Harrison, J., and Heslop-Harrison, Y. 1989b. 225B:315-327. Actomyosin and movement in the angiosperm pollen tube: an interpretation of some recent results. Heslop-Harrison, Y. 1976. Localisation of concanavalin A binding sites on the stigma surface of a grass species. Sex. Plant Reprod. 2:199-207. Micron 7:33-36. Heslop-Harrison, J., and Heslop-Harrison, Y. 1989c. Conformation and movement of the vegetative Heslop-Harrison, Y. 1977. The pollen-stigma interaction: pollen-tube penetration in Crocus. Ann. Bot. nucleus of the angiosperm pollen tube: association 41:913-922. with the actin cytoskeleton. /. Cell Sci. 93:299-308. Heslop-Harrison, J., and Heslop-Harrison, Y. 1989d. Heslop-Harrison, Y. 1981. Stigma characteristics and angiosperm taxonomy. Nord. ]. Bot. 1:401-420. Myosin associated with the surfaces of organelles, vegetative nuclei and generative cells in angiosperm Heslop-Harrison, Y, and Heslop-Harrison, J. 1982a. Pollen-stigma interactions in the Leguminosae: the pollen grains and tubes. /. Cell Sci. 94:319-325. secretory system of the style in Trifolium pratense L. Heslop-Harrison, J., and Heslop-Harrison, Y. 1990. Ann. Bot. 50:635-645. Dynamic aspects of apical zonation in the angiosperm pollen tube. Sex. Plant Reprod. 3:187-194. Heslop-Harrison, Y, and Heslop-Harrison, J. 1982b. The micro-fibrillar component of the pollen intine: some Heslop-Harrison, J., and Heslop-Harrison, Y. 1991a. The structural features. Ann. Bot. 50:831-842. actin cytoskeleton in unfixed pollen tubes following microwave-accelerated DMSO-permeabilisation and Heslop-Harrison, Y, and Heslop-Harrison, J. 1992. TRITC-phalloidin staining. Sex. Plant Reprod. 4:6-11. Germination of monocolpate angiosperm pollen: evolution of the actin cytoskeleton and wall during Heslop-Harrison, J., and Heslop-Harrison, Y. 1991b. hydration, activation and tube emergence. Ann. Bot. Structural and functional variation in pollen intines. 69:385-394. In Pollen and Spores, ed. S. Blackmore and S.H. Barnes, pp. 331-343. Oxford: Clarendon Press. Heslop-Harrison, Y, Heslop-Harrison, J., and Reger, B.J. 1985. The pollen-stigma interaction in the grasses. 7. Heslop-Harrison, J., and Heslop-Harrison, Y. 1992a. Pollen-tube guidance and the regulation of tube Cyclical transformations of the actin cytoskeleton of number in Zea mays L. Ada Bot. Neerl. 34:193-211. hyacinth pollen subjected to recurrent vapor-phase hydration and dehydration. Biol. Cell. 75:245-252. Heslop-Harrison, Y, Heslop-Harrison, J., and Shivanna, Heslop-Harrison, }., and Heslop-Harrison, Y. 1992b. K.R. 1981. Heterostyly in Primula. 1. Fine-structural Intracellular motility, the actin cytoskeleton and gerand cytochemical features of the stigma and style in minability in the pollen of wheat {Triticum aestivum Primula vulgaris Huds. Protoplasma 107:171-187. L.). Sex. Plant Reprod. 5:247-255. Heslop-Harrison, Y, Heslop-Harrison, J.S., and HeslopHeslop-Harrison, J., and Heslop-Harrison, Y. 1992c. Harrison, J. 1986. Germination of Corylus avellana L. Germination of monocolpate angiosperm pollen: (hazel) pollen: hydration and the function of the effects of inhibitory factors and the Ca2+-channel oncus. Ada Bot. Neerl. 35:265-284. blocker, nifedipine. Ann. Bot. 69:395-403. Heslop-Harrison, Y, and Reger, B.J. 1988. Tissue organisation, pollen receptivity and pollen tube guidance in Heslop-Harrison, J., Heslop-Harrison, Y, and Barber, J. normal and mutant stigmas of the grass Pemrisetum 1975. The stigma surface in incompatibility typhoides (Burm.) Stapf et Hubb. Sex. Plant Reprod. responses. Proc. Roy. Soc. Lond. 188B:287-297. 1:182-193. Heslop-Harrison, J., Knox, R. B., and Heslop-Harrison, Y. 1974. Pollen-wall proteins: exine-held fractions asso- Heslop-Harrison, Y, Reger, B.J., and Heslop-Harrison, J. ciated with the incompatibility response in 1984a. The pollen-stigma interaction in the grasses. 5. Tissue organisation and cytochemistry of the Cruciferae. Theor. Appl. Genet. 44:133-137. Heslop-Harrison, J., and Mackenzie, A. 1967. stigma ("silk") of Zea mays L. Ada Bot. Neerl. Autoradiography of soluble [2-14C]thymidine deriv33:81-99. atives during meiosis and microsporogenesis in Heslop-Harrison, Y, Reger, B.J., and Heslop-Harrison, J. Lilium anthers. /. Cell Sci. 2:387-400. 1984b. The pollen-stigma interaction in the grasses. Heslop-Harrison, J., Heslop-Harrison, Y, Cresti, M., and 6. The stigma ('silk') of Zea mays L. as host to the polCiampolini, F. 1991. Ultrastructural features of lens of Sorghum bicolor (L.) Moench and Pemiisetwn pollen tubes of Endymion non-scriptus modified by americanum (L.) Leeke. Ada Bot. Neerl. 33:205-227. cytochalasin D. Sex. Plant Reprod. 4:73-80. Heslop-Harrison, Y, and Shivanna, K.R. 1977. The recep-
580
References
ulation of the synthesis of a sulfur-rich protein in pea tive surface of the angiosperm stigma. Ann. Bot. seeds. /. Biol. Chem. 261:11124-11130. 41:1233-1258. Hess, D. 1980. Investigations on the intra- and interspe- Higgins, T.J.V., Newbigin, E.J., Spencer, D., Llewellyn, D.J., and Craig, S. 1988. The sequence of a pea vicilin cific transfer of anthocyanin genes using pollen as gene and its expression in transgenic tobacco plants. vectors. Z. Pflanzenphysiol. 98:321-337. Plant Mol. Biol. 11:683-695. Hess, D., and Dressier, K. 1989. Tumor transformation of Petunia hybrida via pollen co-cultured with Hilbert, J.-L., Dubois, T, and Vasseur, J. 1992. Detection of embryogenesis-related proteins during somatic Agrobacterium tumefaciens. Bot. Ada 102:202-207. embryo formation in Cichorium. Plant Physiol. Hess, D., Dressier, K., and Nimmrichter, R. 1990. Biochem. 30:733-741. Transformation experiments by pipetting Agrobacterium into the spikelets of wheat (Triticum Hildebrand, D.F., Versluys, R.T., and Collins, G.B. 1991. Changes in lipoxygenase isozyme levels during soyaestivum L.). Plant Sci. 72:233-244. bean embryo development. Plant Sci. 75:1-8. Hess, J.R., and Carman, J.G. 1993. Normalizing development of cultured Triticum aestivum L. embryos. I. Hill, J.E., and Breidenbach, R.W. 1974. Proteins of soyLow oxygen tensions and exogenous ABA. /. Expt. beans seeds. II. Accumulation of the major protein Bot. 44:1067-1073. components during seed development and maturation. Plant Physiol. 53:747-751. Hess, M.W. 1993. Cell-wall development in freeze-fixed pollen: intine formation of Ledebouria socialis Hill, J.P., and Lord, E.M. 1987. Dynamics of pollen tube (Hyacinthaceae). Planta 189:139-149. growth in the wild radish Raphanus raphanistrum (Brassicaceae). II. Morphology, cytochemistry and Hess, M.W., and Frosch, A. 1994. Subunits of forming ultrastructure of transmitting tissues, and path of pollen exine and Ubisch bodies as seen in freeze subpollen tube growth. Am. }. Bot. 74:988-997. stituted Ledebouria socialis Roth (Hyacinthaceae). Protoplasma 182:10-14. Hill, J.P., and Lord, E.M. 1989. Floral development in Hess, M.W., and Hesse, M. 1994. Ultrastructural observaArabidopsis thaliana: a comparison of the wild type tions on anther development of freeze-fixed and the homeotic pistillata mutant. Can. J. Bot. Ledebouria socialis Roth (Hyacinthaceae). Planta 67:2922-2936. 192:421-130. Hilson, P., de Froidmont, D., Lejour, C, Hirai, S.-I., Jacquemin, J.-M., and Yaniv, M. 1990. Fos and Jun Hesse, M., Pacini, E., and Willemse, M., ed. 1993. The oncogenes transactivate chimeric or native promotTapetum. Cytology, Function, Biochemistry and ers containing AP1/GCN4 binding sites in plant Evolution. Wien: Springer-Verlag. cells. Plant Cell 2:651-658. Hetherington, P.R., and Fry, S.C. 1993. Xyloglucan endotransglycosylase activity in carrot cell suspensions Hinata, K., and Nishio, T. 1978. S-allele specificity of during cell elongation and somatic embryogenesis. stigma proteins in Brassica oleracea and B. campestris. Plant Physiol. 103:987-992. Heredity 41:93-100. Hicks, G.S. 1975. Carpelloids on tobacco stamen primor- Hinata, K., Nishio, T., and Kimura, J. 1982. Comparative dia in vitro. Can. J. Bot. 53:77-81. studies on S-glycoproteins purified from different Sgenotypes in self-incompatible Brassica species. II. Hicks, G.S., Browne, R., and Sand, S.A. 1981. Immunological specificities. Genetics 100:649-657. Organogenesis from cultured floral meristems of a male sterile tobacco hybrid. Can. J. Bot. 59:1665-1670. Hinata, K., Watanabe, M., Toriyama, K., and Isogai, A. 1993. A review of recent studies on homomorphic Hicks, G.S., and Sand, S.A. 1977. In vitro culture of the staself-incompatibility. Int. Rev. Cytol. 143:257-296. men primordia from a male sterile tobacco. Plant Sci. Lett. 10:257-263. Hinz, G., Hoh, B., Hohl, I., and Robinson, D.G. 1995. Hicks, G.S., and Sussex, I.M. 1970. Development in vitro of Stratification of storage proteins in the protein storexcised flower primordia of Nicotiana tabacum. Can. ]. age vacuole of developing cotyledons of Pisum Bot. 48:133-139. sativum L. J. Plant Physiol. 145:437-442. Hiei, Y, Ohta, S., Komari, T, and Kumashiro, T. 1994. Hirai, M.Y., Fujiwara, T., Goto, K., Komeda, Y., Chino, M., Efficient transformation of rice (Oryza sativa L.) and Naito, S. 1994. Differential regulation of soymediated by Agrobacterium and sequence analysis of bean seed storage protein gene promoter-GUS the boundaries of the T-DNA. Plant J. 6:271-282. fusions by exogenously applied methionine in transgenic Arabidopsis thaliana. Plant Cell Physiol. Higgins, P., and Bowles, D.J. 1990. Comparative analysis 35:927-934. of translatable mRNA populations in zygotic and pollen-derived embryos of barley (Hordeum vulgare Hirano, H.-Y, and Sano, Y. 1991. Molecular characterizaL.). Plant Sci. 69:239-247. tion of the waxy locus of rice (Oryza sativa). Plant Cell Physiol. 32:989-997. Higgins, R.K., and Petolino, J.F. 1988. In vitro pollination fertilization of maize: influence of explant factors on ker- Hirano, H.-Y., Tabayashi, N., Matsumura, T, Tanida, M., nel development. Plant Cell Tissue Organ Cult. Komeda, Y, and Sano, Y. 1995. Tissue-dependent 12:21-30. expression of the rice wx* gene promoter in transgenic rice and Petunia. Plant Cell Physiol. 36:37-44. Higgins, T.J.V. 1984. Synthesis and regulation of major proteins in seeds. Annu. Rev. Plant Physiol. Hird, D.L., Worrall, D., Hodge, R., Smartt, S., Paul, W., 35:191-221. and Scott, R. 1993. The anther-specific protein Higgins, T.J.V., and Spencer, D. 1981. Precursor forms of encoded by the Brassica napus and Arabidopsis pea vicilin subunits. Modification by microsomal thaliana A6 gene displays similarity to P-l,3-glumembranes during cell-free translation. Plant canases. Plant J. 4:1023-1033. Physiol. 67:205-211. Hirt, H., Pay, A., Gyorgyey, J., Bako, L., Nemeth, K., Higgins, T.J.V., Chandler, P.M., Randall, P.J., Spencer, D., Bogre, L., Schweyen, R.J., Heberle-Bors, E., and Beach, L.R., Blagrove, R.J., Kortt, A.A., and Inglis, Dudits, D. 1991. Complementation of a yeast cell A.S. 1986. Gene structure, protein structure, and regcycle mutant by an alfalfa cDNA encoding a protein
References
581
two higher plant species. Protoplasma 138:126-136. kinase homologous to p34cdc2. Proc. Natl Acad. Sci. USA 88:1636-1640. Hogenboom, N.G. 1973. A model for incongruity in intimate partner relationships. Euphytica 22:219-233. Hiscock, S.J., Doughty, J., and Dickinson, H.G. 1995. Synthesis and phosphorylation of pollen proteins Hoggart, R.M., and Clarke, A.E. 1984. Arabinogalactans during the pollen-stigma interaction in self-compatare common components of angiosperm styles. Phytochemistry 23:1571-1573. ible Brassica napus L. and self-incompatible Brassica oleracea L. Sex. Plant Reprod. 8:345-353. Hoglund, A.-S., Lenman, M., Falk, A., and Rask, L. 1991. Distribution of myrosinase in rapeseed tissues. Plant Hiscock, S.J., Dewey, F.M., Coleman, J.O.D., and Physiol. 95:213-221. Dickinson, H.G. 1994. Identification and localization of an active cutinase in the pollen of Brassica napus L. Hoglund, A.-S., Rodin, J., Larsson, E., and Rask, L. 1992. Planta 193:377-384. Distribution of napin and cruciferin in developing rape seed embryos. Plant Physiol. 98:509-515. Hiscock, S.J., Doughty, J., Willis, A.C., and Dickinson, H.G. 1995. A 7-kDa pollen coating-borne peptide Hohler, B., and Borner, T. 1980. Vergleichende from Brassica napus interacts with S-locus glycoproUntersuchungen von Isoenzymen des Antherentein and S-locus-related glycoprotein. Planta gewebes fertiler und cytoplasmatisch mannlich ster196:367-374. iler Weizenpflanzen. Biochem. Physiol. Pflanz. 175:562-569. Hodgkin, T., and Lyon, G.D. 1984. Pollen germination inhibitors in extracts of Brassica oleracea stigmas. NewHolbrook, L.A., van Rooijen, G.J.H., Wilen, R.W., and Phytol. 96:293-298. Moloney, M.M. 1991. Oilbody proteins in microspore-derived embryos of Brassica napus. Hodgkin, T., Lyon, G.D., and Dickinson, H.G. 1988. Hormonal, osmotic, and developmental regulation Recognition in flowering plants: a comparison of the of synthesis. Plant Physiol. 97:1051-1058. Brassica self-incompatibility system and plant pathogen interactions. New Phytol. 110:557-569. Holden, M.J., and Sze, H. 1984. Helminthosporium maydis T Hoecker, U., Vasil, I.K., and McCarty, D.R. 1995. toxin increased membrane permeability to Ca2+ in Integrated control of seed maturation and germinasusceptible corn mitochondria. Plant Physiol. tion programs by activator and repressor functions 75:235-237. of viviparous-1 of maize. Genes Devel. 9:2459-2469. Holden, M.J., and Sze, H. 1987. Dissipation of the memHoefert, L.L. 1969a. Ultrastructure of Beta pollen. I. brane potential in susceptible corn mitochondria by Cytoplasmic constituents. Am. ]. Bot. 56:363—368. the toxin of Helminthosporium maydis, Race T, and toxin analogs. Plant Physiol. 84:670-676. Hoefert, L.L. 1969b. Fine structure of sperm cells in pollen grains of Beta. Brief report. Protoplasma 68:237-240. Holdsworth, M.J., Munoz-Blanco, J., Hammond-Kosack, M., Colot, V., Schuch, W., and Bevan, M.W. 1995. The Hoekstra, F.A. 1979. Mitochondrial development and maize transcription factor opaque-2 activates a activity of binucleate and trinucleate pollen during wheat germ glutenin promoter in plant and yeast germination in vitro. Planta 145:25-36. cells. Plant Mol. Biol. 29:711-720. Hoekstra, F.A., and Bruinsma, J. 1975a. Viability of Compositae pollen: germination in vitro and influ- Hollung, K., Espelund, M., and Jakobsen, K.S. 1994. ences of climatic conditions during dehiscence. Z. Another Lea B19 gene (group 1 Lea) from barley conPflanzenphysiol. 76:36-43. taining a single 20 amino acid hydrophilic motif. Plant Mol. Biol. 25:559-564. Hoekstra, F.A., and Bruinsma, J. 1975b. Respiration and vitality of binucleate and trinucleate pollen. Physiol. Holm, P.B. 1977. The premeiotic DNA replication of euchromatin and heterochromatin in Lilium longifloPlant. 34:221-225. rum (Thunb.). Carlsberg Res. Cotnmun. 42:249-281. Hoekstra, F.A., and Bruinsma, J. 1978. Reduced independence of the male gametophyte in angiosperm evo- Holm, P.B, Knudsen, S., Mouritzen, P., Negri, D., Olsen, lution. Ann. Bot. 42:759-762. F.L., and Rou6, C. 1994. Regeneration of fertile barley plants from mechanically isolated protoplasts of the Hoekstra, F.A., and Bruinsma, J. 1979. Protein synthesis of fertilized egg cell. Plant Cell 6:531-543. binucleate and trinucleate pollen and its relationship to tube emergence and growth. Planta 146:559-566. Holm, P.B., Knudsen, S., Mouritzen, P., Negri, D., Olsen, Hoekstra, F.A., and Bruinsma, J. 1980. Control of respiraF.L., and Roue, C. 1995. Regeneration of the barley tion of binucleate and trinucleate pollen under zygote in ovule culture. Sear. Plant Reprod. 8:49-59. humid conditions. Physiol. Plant. 48:71-77. Hong, B., Barg, R., and Ho, T.-H.D. 1992. Developmental and organ-specific expression of an ABA- and stressHoffman, L.M., and Donaldson, D.D. 1985. Characterinduced protein in barley. Plant Mol. Biol. 18:663-674. ization of two Phaseolus vulgaris phytohemagglutinin genes closely linked on the chromosome. Hong, S.K., Aoki, T., Kitano, H., Satoh, H., and Nagato, Y. EMBO ]. 4:883-889. 1995a. Temperature-sensitive mutation, embryoless 1, affects both embryo and endosperm development in Hoffman, L.M., Donaldson, D.D., and Herman, E.M. 1988. rice. Plant Sci. 108:165-172. A modified storage protein is synthesized, processed, and degraded in the seeds of transgenic Hong, S.K., Aoki, T., Kitano, H., Satoh, H., and Nagato, Y. 1995b. Phenotypic diversity of 188 rice embryo plants. Plant Mol. Biol. 11:717-729. mutants. Devel Genet. 16:298-310. Hoffman, L.M., Ma, Y., and Barker, R.F. 1982. Molecular cloning of Phaseolus vulgaris lectin mRNA and use of Hong, S.K., Kitano, H., Satoh, H., and Nagato, Y. 1996. cDNA as a probe to estimate lectin transcript levels How is embryo size genetically regulated in rice? in various tissues. Nucl. Acids Res. 10:7819-7828. Development 122:2051-2058. Hoffman, L.M., Donaldson, D.D., Bookland, R., Rashka, Hong-Qi, Z., Croes, A.F., and Linskens, H.F. 1984. K., and Herman, E.M. 1987. Synthesis and protein Qualitative changes in protein synthesis in germibody deposition of maize 15-kd zein in transgenic nating pollen of Lilium longiflorum after a heat shock. tobacco seeds. EMBO /. 6:3213-3221. Plant Cell Environ. 7:689-691. Hogan, C.J. 1987. Microtubule patterns during meiosis in Hopf, N., Plesofsky-Vig, N., and Brambl, R. 1992. The heat
582
References
shock response of pollen and other tissues of maize. of deoxyribosides and regulation of deoxyribonuPlant Mol. Biol. 19:623-630. cleic acid synthesis. /. Biophys. Biochem. Cytol. 11:311-319. Hopper, J.E., Ascher, P.D., and Peloquin, SJ. 1967. Inactivation of self-incompatibility following tem- Hotta, Y, and Stern, H. 1963a. Synthesis of messenger-like perature pretreatments of styles of Lilium longifloribonucleic acid and protein during meiosis in isorum. Euphytica 16:215-220. lated cells of Trillium erectum. J. Cell Biol. 19:45-58. Hopping, M.E., and Jerram, E.M. 1979. Pollination of Hotta, Y, and Stern, H. 1963b. Inhibition of protein synkiwifruit (Actinidia chinensis Planch.): stigma-style thesis during meiosis and its bearing on intracellular structure and pollen tube growth. Neiv Zealand J. Bot. regulation. /. Cell Biol. 16:259-279. 17:233-240. Hotta, Y, and Stern, H. 1963c. Molecular facets of mitotic Horn, R., Kohler, R.H., and Zetsche, K. 1991. A mitochonregulation, I. Synthesis of thymidine kinase. Proc. drial 16 kDa protein is associated with cytoplasmic Natl Acad. Sci. USA 49:648-654. male sterility in sunflower. Plant Mol. Biol. 17:29-36. Hotta, Y, and Stern, H. 1963d. Molecular facets of mitotic regulation, II. Factors underlying the removal of Horn, R., Hustedt, J.E.G., Horstmeyer, A., Hahnen, J., thymidine kinase. Proc. Natl Acad. Sci. USA Zetsche, K., and Friedt, W. 1996. The CMS-associated 16 kDa protein encoded by orfH522 in the PET1 cyto49:861-865. plasm is also present in other male-sterile cyto- Hotta, Y, and Stern, H. 1965a. Inducibility of thymidine plasms of sunflower. Plant Mol. Biol. 30: 523-538. kinase by thymidine as a function of interphase stage. /. Cell Biol. 25:99-108. Horner, H.T., Jr. 1977. A comparative light- and electronmicroscopic study of microsporogenesis in male-fer- Hotta, Y, and Stern, H. 1965b. Polymerase and kinase tile and cytoplasmic male-sterile sunflower activities in relation to RNA synthesis during meio(Helianthus annum). Am. ]. Bot. 64:745-759. sis. Protoplasma 60:218-232. Horner, H.T., Jr., and Lersten, N.R. 1971. Microsporogenesis Hotta, Y, and Stern, H. 1971a. A DNA-binding protein in in Citrus limon (Rutaceae). Am. ]. Bot. 58:72-79. meiotic cells of Lilium. Bevel Biol. 26:87-99. Horner, H.T., Jr., and Pearson, C.B. 1978. Pollen wall and Hotta, Y, and Stern, H. 1971b. Analysis of DNA synthesis aperture development in Helianthus annuus during meiotic prophase in Lilium. J. Mol. Biol. (Compositae: Heliantheae). Am. J. Bot. 65:293-309. 55:337-355. Horner, H.T., Jr., and Rogers, M.A. 1974. A comparative Hotta, Y, and Stern, H. 1974. DNA scission and repair light and electron microscopic study of microsporoduring pachytene in Lilium. Chromosoma 46:279-296. genesis in male-fertile and cytoplasmic male-sterile Hotta, Y, and Stern, H. 1976. Persistent discontinuities in pepper (Capsicum annuum). Can. ]. Bot. 52:435—441. late replicating DNA during meiosis in Lilium. Horner, M., and Mott, R.L. 1979. The frequency of Chromosoma 55:171-182. embryogenic pollen grains is not increased by in Hotta, Y, and Stern, H. 1978. DNA unwinding protein from vitro anther culture in Nicotiana tabacum L. Planta meiotic cells of Lilium. Biochemistry 17:1872-1880. 147:156-158. Hotta, Y, and Stern, H. 1979. The effect of dephosphoryHorner, M., and Pratt, M.L. 1979. Amino acid analysis of lation on the properties of a helix-destabilizing proin vivo and androgenic anthers of Nicotiana tabacum. tein from meiotic cells and its partial reversal by a Protoplasma 98:279-282. protein kinase. Eur. J. Biochem. 95:31-38. Horner, M., and Street, H.E. 1978. Pollen dimorphism - Hotta, Y, and Stern, H. 1981. Small nuclear RNA moleorigin and significance in pollen plant formation by cules that regulate nuclease accessibility in specific anther culture. Ann. Bot. 42:763-771. chromatin regions of meiotic cells. Cell 27:309-319. Horovitz, A., Galil, J., and Portnoy, L. 1972. Soluble sugars Hotta, Y, and Stern, H. 1984. The organization of DNA in the stigmatic exudate of Yucca aloifolia L. Phyton segments undergoing repair synthesis during pachytene. Chromosoma 89:127-137. Argentina 29:43-46. Horowitz, J. 1985. A Phaseolus mutation results in a Hotta, Y, Tabata, S., and Stern, H. 1984. Replication and reduced level of lectin mRNA. Mol. Gen. Genet. nicking of zygotene DNA sequences. Control by a 198:482-485. meiosis-specific protein. Chromosoma 90:243-253. Hoshikawa, K. 1976. Histology of endosperm develop- Hotta, Y, Bennett, M.D., Toledo, L.A., and Stern, H. 1979. ment and reserve substance accumulation in cereal Regulation of R-protein and endonuclease activities grains. In Gamma Field Symposium, No. 13, pp. 1-15. in meiocytes by homologous chromosome pairing. Japan: Institute of Radiation Breeding. Chromosoma 72:191-201. Hotta, Y, and Hecht, N. 1971. Methylation of Lilium DNA Hotta, Y, Tabata, S., Bouchard, R.A., Pifion, R., and Stern, during the meiotic cycle. Biochim. Biophys. Ada H. 1985a. General recombination mechanisms in 238:50-59. extracts of meiotic cells. Chromosoma 93:140-151. Hotta, Y, Ito, M., and Stern, H. 1966. Synthesis of DNA Hotta, Y, Tabata, S., Stubbs, L., and Stern, H. 1985b. during meiosis. Proc. Natl Acad. Sci. USA Meiosis-specific transcripts of a DNA component 56:1184-1191. replicated during chromosome pairing: homology across the phylogenetic spectrum. Cell 40:785-793. Hotta, Y, Parchman, L.G., and Stern, H. 1968. Protein synthesis during meiosis. Proc. Natl Acad. Sci. USA Hough, T, Singh, M.B., Smart, I.J., and Knox, R.B. 1986. Immunofluorescent screening of monoclonal anti60:575-582. bodies to surface antigens of animal and plant cells Hotta, Y, and Shepard. J. 1973. Biochemical aspects of bound to polycarbonate membranes. /. Immuno. colchicine action on meiotic cells. Mol. Gen. Genet. 122:243-260. Meth. 92:103-107. Hotta, Y, and Stern, H. 1961a. Deamination of deoxycyti- Howarth, C. 1990. Heat shock proteins in Sorghum bicolor dine and 5-methyldeoxycytidine in developing and Pennisetum americanum. II. Stored mRNA in anthers of Lilium longiflorum (vat. Croft). /. Biophys. sorghum seed and its relationship to heat shock protein synthesis during germination. Plant Cell Biochem. Cytol. 9:279-284. Hotta, Y., and Stern, H. 1961b. Transient phosphorylation Environ. 13:57-64.
References
583
Howell, S.H., and Hecht, N.B. 1971. The appearance of Huang, B. 1992. Genetic manipulation of microspores and microspore-derived embryos. In Vitro Cell Devel Biol. polynucleotide ligase and DNA polymerase during 28P:53-58. the synchronous mitotic cycle in Lilium microspores. Biochim. Biophys. Acta 240:343-352. Huang, B., and Sunderland, N. 1982. Temperature-stress pretreatment in barley anther culture. Ann. Bot. Howell, S.H., and Stern, H. 1971. The appearance of DNA 49:77-88. breakage and repair activities in the synchronous Huang, B.-Q., and Russell, S.D. 1990. Isolation of fixed meiotic cycle of Lilium. J. Mol. Bid. 55:357-378. and viable eggs, central cells, and embryo sacs from Howlett, B.J., Knox, R.B., and Heslop-Harrison, J. 1973. ovules of Plumbago zeylanica. Plant Physiol. 90:9-12. Pollen-wall proteins: release of the allergen antigen E from intine and exine sites in pollen grains of rag- Huang, B.Q., and Russell, S.D. 1992. Synergid degeneraweed and Cosmos. ]. Cell Sci. 13:603-619. tion in Nicotiana: a quantitative, fluorochromatic and chlorotetracycline study. Sex. Plant Reprod. 5:151-155. Howlett, B.J., Knox, R.B., Paxton, J.D., and HeslopHarrison, J. 1975. Pollen-wall proteins: physico- Huang, B.Q., and Russell, S.D. 1993. Polarity of nuclear and organellar DNA during megasporogenesis and chemical characterization and role in megagametogenesis in Plumbago zeylanica. Sex. Plant self-incompatibility in Cosmos bipinnatus. Proc. Roy. Soc. Lond. 188B:167-182. Reprod. 6:205-211. Hsing, Y.-I.C, Chen, Z.-Y., and Chow, T.-Y. 1992. Huang, B.-Q., and Russell, S.D. 1994. Fertilization in Nicotiana tabacum: cytoskeletal modifications in the Nucleotide sequences of a soybean complementary embryo sac during synergid degeneration. A DNA encoding a 50-kilodalton late embryogenesis hypothesis for short-distance transport of sperm abundant protein. Plant Physiol. 99:354-355. cells prior to gamete fusion. Planta 194:200-214. Hsing, Y.-I.C, and Wu, S.-J. 1992. Cloning and characterization of cDNA clones encoding soybean seed matu- Huang, B.-Q., and Sheridan, W.F. 1994. Female gametophyte development in maize: microtubular organiration polypeptides. Bot. Bull. Acad. Sinica 33:191-199. zation and embryo sac polarity. Plant Cell 6:845-861. Hsing, Y.-I.C, Rinne, R.W., Hepburn, A.G., and Zielinski, R.E. 1990. Expression of maturation-specific genes in Huang, B.-Q., Strout, G.W., and Russell, S.D. 1993. Fertilization in Nicotiana tabacum: ultrastructural soybean seeds. Crop Sci. 30:1343-1350. organization of propane-jet-frozen embryo sacs in Hsu, F.C. 1979. Abscisic acid accumulation in developing vivo. Planta 191:256-264. seeds of Phaseolus vulgaris L. Plant Physiol. Huang, B.-Q., Pierson, E.S., Russell, S.D., Tiezzi, A., and 63:552-556. Cresti, M. 1992. Video microscopic observations of Hu, N.-T., Peifer, M.A., Heidecker, G., Messing, J., and living, isolated embryo sacs of Nicotiana and their Rubenstein, I. 1982. Primary structure of a genomic component cells. Sex. Plant Reprod. 5:156-162. zein sequence of maize. EMBO /. 1:1337-1342. Hu, S.Y. 1964. Morphological and cytological observa- Huang, B.-Q., Pierson, E.S., Russell, S.D., Tiezzi, A., and Cresti, M. 1993. Cytoskeletal organisation and modtions on the process of fertilization of the wheat. Sci. ification during pollen tube arrival, gamete delivery Sinica 13:925-936. and fertilisation in Plumbago zeylanica. Zygote Hu, S.Y, and Chu, Z.C. 1964. A histochemical study of 1:143-154. DNA in sexual cells of some angiosperms. Ada Bot. Sinica 12:333-345. Huang, B.-Q., Russell, S.D., Strout, G.W., and Mao, L.-J. Hu, S.-Y, Li, L.-G., and Zhou, C. 1985. Isolation of viable 1990. Organization of isolated embryo sacs and eggs embryo sacs and their protoplasts of Nicotiana of Plumbago zeylanica (Plumbaginaceae) before and tabacum. Ada Bot. Sinica 27:337-344. after fertilization. Am. J. Bot. 77:1401-1410. Hu, S.-Y, Wang, Y, and Yuan, Z.-F. 1993. The regional Huang, H., Mizukami, Y, Hu, Y, and Ma, H. 1993. aggregation of plastids and mitochondria in Isolation and characterization of the binding microsporocyte and microspore of Gossypium hirsusequences for the product of the Arabidopsis floral tum. Chinese J. Bot. 5:11-17. homeotic gene AGAMOUS. Nucl. Acids Res. Hu, S.-Y, and Yu, H.-S. 1988. Preliminary observations on 21:4769-4776. the formation of the male germ unit in pollen tubes Huang, H., Tudor, M., Weiss, C.A., and Ma, H. 1995. The of Cyphomandra betacea Sendt. Pwtoplasma 147:55-63. Arabidopsis MADS-box gene AGL3 is widely expressed and encodes a sequence-specific DNAHu, S.-Y, and Zhu, C 1979. The fusion of male and female nuclei in fertilization of higher plants. Ada Bot. binding protein. Plant Mol. Biol. 28:549-567. Sinica 21:1-10. Huang, J., Lee, S.-H., Lin, C, Medici, R., Hack, E., and Myers, A.M. 1990. Expression in yeast of the THu, S.-Y, Zhu, C, and Xu, L.-Y. 1981. Ultrastructure of URF13 protein from Texas male-sterile maize mitomale gametophyte in wheat. II. Formation and chondria confers sensitivity to methomyl and to development of sperm cell. Ada Bot. Sinica 23:85-91. Texas-cytoplasm-specific fungal toxins. EMBO ]. Hu, S.-Y, Zhu, C, and Zee, S.Y. 1983. Transfer cells in sus9:339-347. pensor and endosperm during early embryogeny of Huang, S., Lee, H.-S., Karunanandaa, B., and Kao, T.-H. Vigna sinensis. Ada Bot. Sinica 25:1-7. 1994. Ribonuclease activity of Petunia inflata S proHu, S.-Y, Zhu, C, Xu, L.-Y, Li, X.-R., and Shen, J.-H. 1979. Ultrastructure of male gametophyte in wheat. I. The teins is essential for rejection of self-pollen. Plant Cell formation of generative and vegetative cells. Ada 6:1021-1028. Bot. Sinica 21:208-214. HudSk, J., Walles, B., and Vennigerholz, F. 1993. The transHuala, E., and Sussex, I.M. 1992. LEAFY interacts with flomitting tissue in Brugmansia suaveolens L.: ultrastrucral homeotic genes to regulate Arabidopsis floral ture of the stylar transmitting tissue. Ann. Bot. development. Plant Cell 4:901-913. 71:177-186. Huang, B. 1986. Ultrastructural aspects of pollen embryo- Hueros, G., Varotto, S., Salamini, F., and Thompson, R.D. genesis in Hordeum, Triticum and Paeonia. In Haploids 1995. Molecular characterization of BETJ, a gene expressed in the endosperm transfer cells of maize. in Higher Plants, ed. H. Hu and H. Yang, pp. 91-117. Beijing: China Academic Publishers. Plant Cell 7:747-757.
584 Hughes, D.W., and Galau, G.A. 1987. Translation efficiency of Lea mRNAs in cotton embryos: minor changes during embryogenesis and germination. Plant Mol. Biol. 9:301-313. Hughes, D.W., and Galau, G.A. 1989. Temporally modular gene expression during cotyledon development. Genes Devel. 3:358-369. Hughes, D.W., and Galau, G.A. 1991. Developmental and environmental induction of Lea and LeaA mRNAs and the postabscission program during embryo culture. Plant Cell 3:605-618. Hughes, D.W., Wang, H.Y.-C, and Galau, G.A. 1993. Cotton (Gossypium hirsutum) MatP6 and MtP7 oleosin genes. Plant Physiol. 101:697-698. Hull, G.A., Bies, N., Twell, D., and Delseny, M. 1996. Analysis of the promoter of an abscisic acid responsive late embryogenesis abundant gene of Arabidopsis thaliana. Plant Sci. 114:181-192. Hull, G.A., Halford, N.G., Kreis, M., and Shewry, P.R. 1991. Isolation and characterisation of genes encoding rye prolamins containing a highly repetitive sequence motif. Plant Mol. Biol. 17:1111-1115. Hulskamp, M., Schneitz, K., and Pruitt, R.E. 1995. Genetic evidence for a long-range activity that directs pollen tube guidance in Arabidopsis. Plant Cell 7:57-64. Hiilskamp, M., Kopczak, S.D., Horejsi, T.F., Kihl, B.K., and Pruitt, R.E. 1995. Identification of genes required for pollen-stigma recognition in Arabidopsis thaliana. Plant}. 8:703-714. Hunold, R., Burrus, M, Bronner, R., Duret, J.-R, and Hahne, G. 1995. Transient gene expression in sunflower (Helianthus annuus L.) following microprojectile bombardment. Plant Sci. 105:95-109. Hurkman, W.J., Smith, L.D., Richter, J., and Larkins, B.A. 1981. Subcellular compartmentalization of maize storage proteins in Xenopus oocytes injected with zein messenger RNAs. /. Cell Biol. 89:292-299. Hussey, P.J., Lloyd, C.W., and Gull, K. 1988. Differential and developmental expression of (i-tubulins in a higher plant. /. Biol. Chem. 263:5474-5479. Huxley, J. 1955. Morphism and evolution. Heredity 9:1-52.
References
Ioerger, T.R., Gohlke, J.R., Xu, B., and Kao, T.-H. 1991. Primary structural features of the self-incompatibility protein in Solanaceae. Sex. Plant Reprod. 4:81-87. Irish, E.E., Langdale, J.A., and Nelson, T.M. 1994. Interactions between Tassel seed genes and other sex determining genes in maize. Devel Genet. 15:155-171. Irish, E.E., and Nelson, T.M. 1993. Development of tassel seed 2 inflorescences in maize. Am. J. Bot. 80:292-299. Irish, V.F., and Sussex, I.M. 1990. Function of the apetala-1 gene during Arabidopsis floral development. Plant Cell 2:741-753. Isaac, P.G., Jones, V.P., and Leaver, C.J. 1985. The maize cytochrome c oxidase subunit I gene: sequence, expression and rearrangement in cytoplasmic male sterile plants. EMBO J. 4:1617-1623. Ishibashi, N., and Minamikawa, T. 1990. Molecular cloning and characterization of stored mRNA in cotyledons of Vigna unguiculata. Plant Cell Physiol. 31-39-U. Ishibashi, N., Yamauchi, D., and Minamikawa, T. 1990. Stored mRNA in cotyledons of Vigna unguiculata seeds: nucleotide sequence of cloned cDNA for a stored mRNA and induction of its synthesis by precocious germination. Plant Mol. Biol. 15:59-64. Ishida, Y., Saito, H., Ohta, S., Hiei, Y, Komari, T, and Kumashiro, T. 1996. High efficiency transformation of maize (Zea mays L.) mediated by Agrobacterium tumefaciens. Nature Biotech. 14:745-750. Isogai, A., Takayama, S., Shiozawa, H., Tsukamoto, C, Kanbara, T, Hinata, K., Okazaki, K., and Suzuki, A. 1988. Existence of a common glycoprotein homologous to S-glycoproteins in two self-incompatible homozygotes of Brassica campestris. Plant Cell Physiol. 29:1331-1336. Isogai, A., Takayama, S., Tsukamoto, C, Ueda, Y, Shiozawa, H., Hinata, K., Okazaki, K., and Suzuki, A. 1987. S-locus-specific glycoproteins associated with self-incompatibility in Brassica campestris. Plant Cell Physiol. 28:1279-1291. Isogai, A., Yamakawa, S., Shiozawa, H., Takayama, S., Tanaka, H., Kono, T, Watanabe, M., Hinata, K., and Suzuki, A. 1991. The cDNA sequence of NS, glycoprotein of Brassica campestris and its homology to SIdzikowska, K. 1981. Mitochondria during androgenesis locus-related glycoproteins of B. oleracea. Plant Mol. in Hordeum vulgare. Ada Soc. Bot. Polon. 50:359-366. Biol. 17:269-271. Idzikowska, K., Ponitka, A., and Mtodzianowski, F. 1982. Pollen dimorphism and androgenesis in Hordeum Israel, H.W., and Sagawa, Y. 1964. Post-pollination ovule vulgare. Ada Soc. Bot. Polon. 51:153-156. development in Dendrobium orchids. II. Fine structure of the nucellar and archesporial phases. Iglesias, V.A., Gisel, A., Bilang, R., Ledue, N., Potrykus, I., Caryologia 17:301-316. and Sautter, C. 1994. Transient expression of visible marker genes in meristem cells of wheat embryos Ito, M. 1973. Studies on the behavior of meiotic protoafter ballistic micro-targeting. Planta 192:84-91. plasts. I. Isolation from microsporocytes of liliaceous plants. Bot. Mag. Tokyo 86:133-141. Ihle, J.N., and Dure, L., III. 1970. Hormonal regulation of translation inhibition requiring RNA synthesis. Ito, M., and Hotta, Y. 1973. Radioautography of incorpoBiochem. Biophys. Res. Commun. 38:995-1001. rated 3H-thymidine and its metabolism during meiotic prophase in microsporocytes of Lilium. Ihle, J.N., and Dure, L.S., III. 1972. The developmental bioChromosoma 43:391-398. chemistry of cottonseed embryogenesis and germination. III. Regulation of the biosynthesis of Ito, M., Hotta, Y, and Stern, H. 1967. Studies of meiosis in enzymes utilized in germination. /. Biol. Chem. vitro. II. Effect of inhibiting DNA synthesis during 247:5048-5055. meiotic prophase on chromosome structure and behavior. Devel Biol. 16:54-77. Iida, A., Nagasawa, A., and Oeda, K. 1995. Positive and negative cis-regulatory regions in the soybean Ito, M., and Maeda, M. 1973. Fusion of meiotic protoplasts in liliaceous plants. Expt. Cell Res. 80:453-456. glycinin promoter identified by quantitative tranIto, M., and Stern, H. 1967. Studies of meiosis in vitro. I. In sient gene expression. Plant Cell Rep. 14:539-544. vitro culture of meiotic cells. Devel Biol. 16:36-53. Ioerger, T.R., Clark, A.G., and Kao, T.-H. 1990. Polymorphism at the self-incompatibility locus in Ito, M., and Takegami, M.H. 1982. Commitment of mitotic cells to meiosis during the G2 phase of premeiosis. Solanaceae predates speciation. Proc. Natl Acad. Sci. Plant Cell Physiol. 23:943-952. USA 87:9732-9735.
References
585
gibberellin deficiency of the gib-1 mutant of tomato. Plant Physiol. 97:409-414. Jacobsen, S.E., Shi, L., Xin, Z., and Olszewski, N.E. 1994. Gibberellin-induced changes in the translatable mRNA populations of stamens and shoots of gibberellin-deficient tomato. Planta 192:372-378. Jaffe, L.A., Weisenseel, M.H., and Jaffe, L.F. 1975. Calcium accumulations within the growing tips of pollen tubes. /. Cell Biol. 67:488-492. Jahne, A., Becker, D., and Lorz, H. 1995. Genetic engineering of cereal crop plants: a review. Euphytica 85:35-44. Jahne, A., Becker, D., Brettschneider, R., and Lorz, H. 1994. Regeneration of transgenic, microsporederived fertile, barley. Theor. Appl. Genet. 89:525-533. Jahnen, W., Lush, W.M., and Clarke, A.E. 1989. Inhibition of in vitro pollen tube growth by isolated S-glycoproteins of Nicotiana alata. Plant Cell 1:501-510. Jahnen, W., Batterham, M.P., Clarke, A.E., Moritz, R.L., and Simpson, R.J. 1989. Identification, isolation, and N-terminal sequencing of style glycoproteins associated with self-incompatibility in Nicotiana alata. Plant Cell 1:493-499. Jain, S.M., Gupta, P.K., and Newton, R.J., ed. 1995. Somatic Embryogenesis in Woody Plants, Vol. 3. Gymnosperms. Dordrecht: Kluwer Academic Publishers. Jakobsen, K., Klemsdal, S.S., Aalen, R.B., Bosnes, M., Alexander, D., and Olsen, O.-A. 1989. Barley aleurone cell development: molecular cloning of aleurone-specific cDNAs from immature grains. Plant Mol. Biol. 12:285-293. Jakobsen, K.S., Hughes, D.W., and Galau, G.A. 1994. Simultaneous induction of postabscission and germination mRNAs in cultured dicotyledonous embryos. Planta 192:384-394. Jalouzot, M.-F. 1971. Aspects ultrastructuraux de la megasporogenese d'Oenothera lamarckiana en rapport avec les depots callosiques observes. Ann. Univ. A.R.E.R.S. 9:36-45. Jalouzot, M.-F. 1975. Aspects ultrastructuraux du sac embryonnaire d'Oenothera lamarckiana. Compt. Rend. Acad. Sci. Paris 281D:1305-1308. Jalouzot, R. 1969a. Differenciation nucleaire et cytoplasmique du grain de pollen de Lilium candidum. Expt. Cell Res. 55:1-8. Jack, T, Brockman, L.L., and Meyerowitz, E.M. 1992. The Jalouzot, R. 1969b. Aspects cytochimiques des deux celhomeotic gene APETALA3 of Arabidopsis thaliana lules du grain de pollen de Crocus longiflorus. Rev. encodes a MADS box and is expressed in petals and Cytol. Biol. Veget. 32:115-120. stamens. Cell 68:683-697. James, M.G., Robertson, D.S., and Myers, A.M. 1995. Characterization of the maize gene sugaryl, a deterJack, T, Fox, G.L., and Meyerowitz, E.M. 1994. Arabidopsis minant of starch composition in kernels. Plant Cell homeotic gene APETALA3 ectopic expression: tran7:417-429. scriptional and posttranscriptional regulation determine floral organ identity. Cell 76:703-716. Janaki-Ammal, E.K. 1941. The breakdown of meiosis in a male-sterile Saccharum. Ann. Bot. 5:83-87. Jackson, J.F., Jones, G., and Linskens, H.F. 1982. Phytic Jansen, M.A.K., Booij, H., Schel, J.H.N., and de Vries, S.C. acid in pollen. Phytochemistry 21:1255-1258. 1990. Calcium increases the yield of somatic Jackson, J.F., Kamboj, R.K., and Linskens, H.F. 1983. embryos in carrot embryogenic suspension cultures. Localization of phytic acid in the floral structure of Plant Cell Rep. 9:221-223. Petunia hybrida and relation to the incompatibility Janska, H., and Mackenzie, S.A. 1993. Unusual mitochongenes. Theor. Appl. Genet. 64:259-262. drial genome organization in cytoplasmic male sterJackson, J.F., and Linskens, H.F. 1982. Phytic acid in ile common bean and the nature of cytoplasmic Petunia hybrida pollen is hydrolysed during germireversion to fertility. Genetics 135:869-879. nation by a phytase. Acta Bot. Neerl. 31:441-447. Jacobsen, J.V., Knox, R.B., and Pyliotis, N.A. 1971. The Janson, J. 1993. Placental pollination in Lilium longiflorum Thunb. Plant Sci. 90:105-115. structure and composition of aleurone grains in the Janson, J., and Willemse, M.T.M. 1995. Pollen tube penebarley aleurone layer. Planta 101:189-209. tration and fertilization in Lilium longiflorum Jacobsen, S.E., and Olszewski, N.E. 1991. Characterization (Liliaceae). Am. J. Bot. 82:186-196. of the arrest in anther development associated with Itoh, Y, Kitamura, Y, Arahira, M., and Fukazawa, C. 1993. Gs-acting regulatory regions of the soybean seed storage IIS globulin gene and their interactions with seed embryo factors. Plant Mol. Biol. 21:973-984. Iwabuchi, M., Kyozuka, J., and Shimamoto, K. 1993. Processing followed by complete editing of an altered mitochondrial atp6 RNA restores fertility of cytoplasmic male sterile rice. EMBO J. 12:1437-1446. Iwanami, Y. 1953. Physiological researches of pollen. (V). On the inductive tissue and the growth of the pollen tube in the style. Bot. Mag. Tokyo 66:189-196. Iwanami, Y 1956. Protoplasmic movement in pollen grains and tubes. Phytomorphology 6:288-295. Iwanami, Y 1959. Physiological studies of pollen. /. Yokohama Municip. Univ. (C34 Biol. 13) 116:1-137. Iyer, R.D., and Raina, S.K. 1972. The early ontogeny of embryoids and callus from pollen and subsequent organogenesis in anther cultures of Datura metel and rice. Planta 104:146-156. Izawa, T., Foster, R., Nakajima, M, Shimamoto, K., and Chua, N.-H. 1994. The rice bZIP transcriptional activator RITA-1 is highly expressed during seed development. Plant Cell 6:1277-1287. Izhar, S. 1975. The timing of temperature effect on microsporogenesis in cytoplasmic male-sterile Petunia. J. Hered. 66:313-314. Izhar, S. 1978. Cytoplasmic male sterility in Petunia. III. Genetic control of microsporogenesis and male fertility restoration. /. Hered. 69:22-26. Izhar, S., and Frankel, R. 1971. Mechanism of male sterility in Petunia: the relationship between pH, callase activity in the anthers, and the breakdown of the microsporogenesis. Theor. Appl. Genet. 41:104-108. Izhar, S., and Frankel, R. 1973a. Mechanism of male sterility in Petunia. II. Free amino acids in male fertile and male sterile anthers during microsporogenesis. Theor. Appl. Genet. 43:13-17. Izhar, S., and Frankel, R. 1973b. Duration of meiosis in Petunia anthers in vivo and in floral bud culture. Ada Bot. Need. 22:14-22. Izhar, S., Schlicter, M., and Swartzberg, D. 1983. Sorting out of cytoplasmic elements in somatic hybrids of Petunia and the prevalence of the heteroplasmon through several meiotic cycles. Mol. Gen. Genet. 190:468-474.
586 Jardinaud, M.-R, Souvre, A., and Alibert, G. 1993. Transient GUS gene expression in Brassica napus electroporated microspores. Plant Sci. 93:177-184. Jardinaud, M.F., SouvrS, A., Alibert, G., and Beckert, M. 1995. uidA gene transfer and expression in maize microspores using the biolistic method. Protoplasma 187:138-143. Jarvi, A.J., and Eslick, R.F. 1975. Shrunken endosperm mutants in barley. Crop Sci. 15:363-366. Jauh, G.Y., and Lord, E.M. 1995. Movement of the tube cell in the lily style in the absence of the pollen grain and the spent pollen tube. Sex. Plant Reprod. 8:168-172. Jauh, G.Y., and Lord, E.M. 1996. Localization of pectins and arabinogalactan-proteins in lily (Lilium longiflorum L.) pollen tube and style, and their possible roles in pollination. Planta 199:251-261. Jayakaran, M. 1972. Suppression of stamens in Capsicum annuum by a morphactin (EMD-IT 7839). Curr. Sci. 41:849-850. Jegla, D.E., and Sussex, I.M. 1989. Cell lineage patterns in the shoot meristem of the sunflower embryo in the dry seed. Devel Bid. 131:215-225. Jensen, W.A. 1963. Cell development during plant embryogenesis. In Meristems and Differentiation. Brookhaven Symp. Biol. 16:179-202. Jensen, W.A. 1964. Observations on the fusion of nuclei in plants. /. Cell Biol. 23:669-672. Jensen, W.A. 1965a. The composition and ultrastructure of the nucellus in cotton. /. Ultrastr. Res. 13:112-128. Jensen, W.A. 1965b. The ultrastructure and histochemistry of the synergids of cotton. Am. J. Bot. 52:238-256. Jensen, W.A. 1965c. The ultrastructure and composition of the egg and central cell of cotton. Am. J. Bot. 52:781-797. Jensen, W.A. 1968. Cotton embryogenesis: the zygote. Planta 79:346-366. Jensen, W.A. 1973. Fertilization in flowering plants.
References
Ji, L.-H., and Langridge, P. 1994. An early meiosis cDNA clone from wheat. Mol. Gen. Genet. 243:17-23. Jiang, L., Abrams, S.R., and Kermode, A.R. 1996. Vicilin and napin storage-protein gene promoters are responsive to abscisic acid in developing transgenic tobacco seed but lose sensitivity following premature desiccation. Plant Physiol. 110:1135-1144. Jobson, S., Knox, R.B., Kenrick, J., and Dumas, C. 1983. Plastid development and ferritin content of stigmas of the legumes Acacia, Lotus and Trifolium. Protoplasma 116:213-218. Joersbo, M., Andersen, J.M., Okkels, F.T., and Rajagopal, R. 1989. Isoperoxidases as markers of somatic embryogenesis in carrot cell suspension cultures. Physiol. Plant. 76:10-16.
Jofuku, K.D., and Goldberg, R.B. 1989. Kunitz trypsin inhibitor genes are differentially expressed during the soybean life cycle and in transformed tobacco plants. Plant Cell 1:1079-1093. Jofuku, K.D., Okamuro, J.K., and Goldberg, R.B. 1987. Interaction of an embryo DNA binding protein with a soybean lectin gene upstream region. Nature 328:734-737. Jofuku, K.D., Schipper, R.D., and Goldberg, R.B. 1989. A frameshift mutation prevents Kunitz trypsin inhibitor mRNA accumulation in soybean embryos. Plant Cell 1:427-435. Jofuku, K.D., den Boer, B.G.W., van Montagu, M., and Okamuro, J.K. 1994. Control of Arabidopsis flower and seed development by the homeotic gene APETALA2. Plant Cell 6:1211-1225. Johansen, D.A. 1950. Plant Embryology. Embryogeny of the Spermatophyta. Waltham: Chronica Botanica Co. Johansson, L. 1983. Effects of activated charcoal in anther cultures. Physiol. Plant. 59:397^03. Johansson, M., and Walles, B. 1993a. Functional anatomy of the ovule in broad bean (Vicia faba L.). I. Histogenesis prior to and after pollination. Int. ]. BioScience 23:21-27. Plant Sci. 154:80-89. Jensen, W.A., Ashton, M., and Heckard, L.R. 1974. Johansson, M., and Walles, B. 1993b. Functional anatomy Ultrastructural studies of the pollen of subtribe of the ovule in broad bean, Vicia faba L. II. Castilleiinae, family Scrophulariaceae. Bot. Gaz. Ultrastructural development up to early embryogenesis. Int. ]. Plant Sci. 154:535-549. 135:210-218. Jensen, W.A., Ashton, M.E., and Beasley, C.A. 1983. Pollen Johansson, M., and Walles, B. 1994. Functional anatomy of the ovule in broad bean (Vicia faba L.): ultrastructural tube-embryo sac interactions in cotton. In Pollen: seed development and nutrient pathways. Ann. Bot. Biology and Implications for Plant Breeding, ed. D.L. 74:233-244. Mulcahy and E. Ottaviano, pp. 67-72. New York: Johari, R.P., Mehta, S.L., and Naik, M.S. 1977. Protein Elsevier. synthesis and changes in nucleic acids during Jensen, W.A., and Fisher, D.B. 1967. Cotton embryogenegrain development of Sorghum. Phytochemistry sis: double fertilization. Phytomorphology 17:261-269. 16:19-24. Jensen, W.A., and Fisher, D.B. 1968a. Cotton embryogenesis: the entrance and discharge of the pollen tube in John, M.E., and Petersen, M.W. 1994. Cotton (Gossypium hirsutum L.) pollen-specific polygalacturonase mRNA: the embryo sac. Planta 78:158-183. tissue and temporal specificity of its promoter in transJensen, W.A., and Fisher, D.B. 1968b. Cotton embryogengenic tobacco. Plant Mol. Biol. 26:1989-1993. esis: the sperm. Protoplasma 65:277-286. Jensen, W.A., and Fisher, D.B. 1969. Cotton embryogene- Johns, C, Lu, M., Lyznik, A., and Mackenzie, S. 1992. A mitochondrial DNA sequence is associated with sis: the tissues of the stigma and style and their relaabnormal pollen development in cytoplasmic male tion to the pollen tube. Planta 84:97-121. sterile bean plants. Plant Cell 4:435-449. Jensen, W.A., and Fisher, D.B. 1970. Cotton embryogenesis: the pollen tube in the stigma and style. Johns, C, Nickels, R., Mclntosh, L., and Mackenzie, S. 1993. The expression of alternative oxidase and alterProtoplasma 69:215-235. native respiratory capacity in cytoplasmic male sterJensen, W.A., Fisher, D.B., and Ashton, M.E. 1968. Cotton ile common bean. Sex. Plant Reprod. 6:257-265. embryogenesis: the pollen cytoplasm. Planta 81:206-228. Johnson, CM., Mulcahy, D.L., and Galinat, W.C. 1976. Male gametophyte in maize: influences of the gameJensen, W.A., Schulz, P., and Ashton, M.E. 1977. An ultratophytic genotype. Theor. Appl. Genet. 48:299-303. structural study of early endosperm development and synergid changes in unfertilized cotton ovules. Johnson, E.D., Knight, J., and Gayler, K.R. 1985. Biosynthesis and processing of legumin-like storage Planta 133:179-189.
References
587
microtubule drug carbetamide stops Nicotiana proteins in Lupinus angustifolius (lupin). Biochem. }. sylvestris pollen tube growth in the style. Protoplasma 232:673-679. 187:182-191. Johnson, K.A., and Sussex, I.M. 1990. Genomic amplification in the cotyledon parenchyma of common bean. Joppa, L.R., McNeal, F.H., and Welsh, J.R. 1966. Pollen Chromosoma 99:223-230. and anther development in cytoplasmic male sterile wheat (Triticum aestivum L.). Crop Sci. 6:296-297. Johnson, L.E.B., Wilcoxson, R.D., and Frosheiser, F.I. 1975. Transfer cells in tissues of the reproductive system of Jordaan, A., and Kruger, H. 1993. Pollen wall ontogeny of alfalfa. Can. ]. Bot. 53:952-956. Felicia muricata (Asteraceae: Astereae). Ann. Bot. 71:97-105. Johnson, S., Liu, C.-M., Hedley, C.L., and Wang, T.L. 1994. An analysis of seed development in Pisum sativum. Jordano, J., Almoguera, C, and Thomas, T.L. 1989. A sunXVIII. The isolation of mutants defective in embryo flower helianthinin gene upstream sequence ensemdevelopment. /. Expt. Bot. 45:1503-1511. ble contains an enhancer and sites of nuclear protein interaction. Plant Cell 1:855-866. Johnson-Flanagan, A.M., Huiwen, Z., Geng, X.-M, Brown, D.C.W., Nykiforuk, C.L., and Singh, J. 1992. Frost, Jose-Estanyol, M., Ruiz-Avila, L., and Puigdomenech, P. 1992. A maize embryo-specific gene encodes a proabscisic acid, and desiccation hasten embryo developline-rich and hydrophobic protein. Plant Cell ment in Brassica napus. Plant Physiol. 99:700-706. 4:413-423. Johnsson, H. 1944. Meiotic aberrations and sterility in Alopecurus myosuroides Huds. Hereditas 30:469-565. Josefsson, L.-G., Lenman, M., Ericson, M.L., and Rask, L. 1987. Structure of a gene encoding the 1.7 S storage Johnston, S.A., and Hanneman, R.E., Jr. 1982. protein, napin, from Brassica napus. ]. Biol. Chem. Manipulations of endosperm balance number over262:12196-12201. come crossing barriers between diploid Solanum species. Science 217:446-448. Juliano, J.B., and Aldama, M.J. 1937. Morphology of Oryza sativa Linnaeus. Philipp. Agr. 26:1-76. Johnston, S.A., den Nijs, T.P.M., Peloquin, S.J., and Hanneman, R.E., Jr. 1980. The significance of genie Juncosa, A.M. 1982. Developmental morphology of the embryo and seedling of Rhizophora mangle L. balance to endosperm development in interspecific (Rhizophoraceae). Am. ]. Bot. 69:1599-1611. crosses. Theor. Appl. Genet. 57:5-9. Johri, B.M., Ambegaokar, K.B., and Srivastava, P.S. 1992. Jiirgens, G. 1995. Axis formation in plant embryogenesis: cues and clues. Cell 81:467-470. Comparative Embryology of Angiosperms, Vols. 1 & 2. Berlin: Springer-Verlag. Jiirgens, G., Mayer, U., Torres Ruiz, R.A., Berleth, T, and Johri, B.M., and Vasil, I.K. 1961. Physiology of pollen. Bot. Misera, S. 1991. Genetic analysis of pattern formation Rev. 27:325-381. in the Arabidopsis embryo. Development Suppl. 1:27-38. Johri, M.M., and Coe, E.H., Jr. 1983. Clonal analysis of Jurgens, G., Torres Ruiz, R.A., and Berleth, T. 1994. corn plant development. I. The development of the Embryonic pattern formation in flowering plants. tassel and the ear shoot. Devel Biol. 97:154-172. Annu. Rev. Genet. 28:351-371. Jones, D.F. 1920. Heritable characters of maize. IV. A lethal factor-defective seeds. /. Hered. 11:160-167. Kadej, A.J., Wilms, H.J., and Willemse, M.T.M. 1985. Jones, H.A. 1927. Pollination and life history studies of Stigma and stigmatoid tissue of Lycopersicon esculentum Mil. Acta Bot. Neerl. 34:95-103. lettuce (Lactuca sativa L.). Hilgardia 2:425-479. Jones, P.A. 1977. Development of the quiescent center in Kadowaki, K.-L, Suzuki, T, and Kazama, S. 1990. A maturing embryonic radicles of pea (Pisum sativum chimeric gene containing 5' portion of atp6 is associL. cv. Alaska). Planta 135:233-240. ated with cytoplasmic male-sterility of rice. Mol. Jones, R.A. 1978. Effects oifloury-2 locus on zein accumuGen. Genet. 224:10-16. lation and RNA metabolism during maize Kahn, T.L., and DeMason, D.A. 1986. A quantitative and endosperm development. Biochem. Genet. 16:27-38. structural comparison of Citrus pollen tube developJones, R.A., Larkins, B.A., and Tsai, C.Y. 1977. Storage proment in cross-compatible and self-incompatible tein synthesis in maize. III. Developmental changes gynoecia. Can. ]. Bot. 64:2548-2555. in membrane-bound polyribosome composition and Kalinski, A., Loer, D.S., Weisemann, J.M., Matthews, B.F., in vitro protein synthesis of normal and opaque-2 and Herman, E.M. 1991. Isoforms of soybean seed maize. Plant Physiol. 59:733-737. oil body membrane protein 24 kDa oleosin are encoded by closely related cDNAs. Plant Mol. Biol. Jones, R.L. 1969. The fine structure of barely aleurone 17:1095-1098. cells. Planta 85:359-375. Jones, T.J., and Rost, T.L. 1989a. The developmental Kalla, R., Shimamoto, K., Potter, R., Nielsen, P.S., anatomy and ultrastructure of somatic embryos Linnestad, C, and Olsen, O.-A. 1994. The promoter of from rice (Oryza sativa L.) scutellum epithelial cells. the barley aleurone-specific gene encoding a putative Bot. Gaz. 150:41-49. 7 kDa lipid transfer protein confers aleurone cell-specific expression in transgenic rice. Plant J. 6:849-860. Jones, T.J., and Rost, T.L. 1989b. Histochemistry and ultrastructure of rice (Oryza sativa) zygotic embryogene- Kaltsikes, P.J. 1973. Early seed development in hexaploid sis. Am. ]. Bot. 76:504-520. triticale. Can. ]. Bot. 51:2291-2300. Jones-Villeneuve, E., Huang, B., Prudhomme, I., Bird, S., Kamada, H., and Harada, H. 1979a. Studies on the organogenesis in carrot tissue cultures. I. Effects of Kemble, R., Hattori, J., and Miki, B. 1995. growth regulators on somatic embryogenesis and Assessment of microinjection for introducing DNA root formation. Z. Pflanzenphysiol. 91:255-266. into uninuclear microspores of rapeseed. Plant Cell Tissue Organ Cult. 40:97-100. Kamada, H., and Harada, H. 1979b. Studies on the Joos, U., van Aken, J., and Kristen, U. 1994. Microtubules organogenesis in carrot tissue cultures. II. Effects of are involved in maintaining the cellular polarity in amino acids and inorganic nitrogenous compounds pollen tubes of Nicotiana sylvestris. Protoplasma on somatic embryogenesis. Z. Pflanzenphysiol. 179:5-15. 91:453-463. Joos, U., van Aken, J., and Kristen, U. 1995. The anti- Kamada, H., and Harada, H. 1981. Changes in the
588
References
endogenous level and effects of abscisic acid during Eichhornia crassipes (Mart.) Solms. Aquatic Bot. somatic embryogenesis of Daucus carota L. Plant Cell 16:41-47. Physiol. 22:1423-1429. Kandasamy, M.K., Dwyer, K.G., Paolillo, D.J., Doney, R.C., Nasrallah, J.B., and Nasrallah, M.E. 1990. Kamada, H., and Harada, H. 1984a. Studies on nitrogen metabolism during somatic embryogenesis in carrot. Brassica S-proteins accumulate in the intercellular I. Utilization of a-alanine as a nitrogen source. Plant matrix along the path of pollen tubes in transgenic Sci. Lett. 33:7-13. tobacco pistils. Plant Cell 2:39-49. Kamada, H., and Harada, H. 1984b. Changes in endoge- Kandasamy, M.K., Paolillo, D.J., Faraday, CD., Nasrallah, nous amino acid compositions during somatic J.B., and Nasrallah, M.E. 1989. The S-locus specific embryogenesis in Daucus carota L. Plant Cell Physiol. glycoproteins of Brassica accumulate in the cell wall 25:27-38. of developing stigma papillae. Devel Biol. 134:462-472. Kamada, H., Kobayashi, K., Kiyosue, T., and Harada, H. 1989. Stress induced somatic embryogenesis in car- Kandasamy, M.K., Thorsness, M.K., Rundle, S.J., rot and its application to synthetic seed production. Goldberg, M.L., Nasrallah, J.B., and Nasrallah, M.E. In Vitro Cell Devel Biol. 25:1163-1166. 1993. Ablation of papillar cell function in Brassica flowers results in the loss of stigma receptivity to Kamalay, J.C., and Goldberg, R.B. 1980. Regulation of pollination. Plant Cell 5:263-275. structural gene expression in tobacco. Cell 19:935-946. Kang, H.-G., Noh, Y.-S., Chung, Y.-Y., Costa, M.A., An, K., and An, G. 1995. Phenotypic alterations of petal and Kamalay, J.C., and Goldberg, R.B. 1984. Organ-specific sepal by ectopic expression of a rice MADS box gene nuclear RNAs in tobacco. Proc. Natl Acad. Sci. USA 81:2801-2805. in tobacco. Plant Mol. Biol. 29:1-10. Kamboj, R.K., and Jackson, J.F. 1986. Self-incompatibility Kanno, T., and Hinata, K. 1969. An electron microscopic alleles control a low molecular weight, basic protein study of the barrier against pollen-tube growth in in pistils of Petunia hybrida. Theor. Appl. Genet. self-incompatible Cruciferae. Plant Cell Physiol. 71:815-819. 10:213-216. Kamboj, R.K., Linskens, H.F., and Jackson, J.F. 1984. Kanta, K. 1960. Intra-ovarian pollination in Papaver rhoeas Energy-driven protein release from germinating L. Nature 188:683-684. pollen of Petunia hybrida. Ann. Bot. 54:647-652. Kanta, K., and Maheshwari, P. 1963a. Intraovarian pollination in some Papaveraceae.Phytomorphology Kameya, T., Hinata, K., and Mizushima, U. 1966. 13:215-230. Fertilization in vitro of excised ovules treated with calcium chloride in Brassica oleracea L. Proc. ]ap. Acad. Kanta, K., and Maheshwari, P. 1963b. Test-tube fertilization 42:165-167. in some angiosperms. Phytomorphology 13:230-237. Kameya, T., and Uchimiya, H. 1972. Embryoids derived Kanta, K., Ranga Swamy, N.S., and Maheshwari, P. 1962. from isolated protoplasts of carrot. Planta Test-tube fertilization in a flowering plant. Nature 103:356-360. 194:1214-1217. Kaminski, P.A., Buffard, D., and Strosberg, A.D. 1987. The Kapil, R.N., and Bhatnagar, A.K. 1975. A fresh look at the pea lectin gene family contains only one functional process of double fertilization in angiosperms. gene. Plant Mol. Biol. 9:497-507. Phytomorphology 25:334-368. Kandasamy, M.K., Kappler, R., and Kristen, U. 1988. Kapil, R.N., and Bhatnagar, A.K. 1981. Ultrastructure and Plasmatubules in the pollen tubes of Nicotiana biology of female gametophyte in flowering plants. sylvestris. Planta 173:35-41. Int. Rev. Cytol. 70:291-341. Kandasamy, M.K., and Kristen, U. 1987a. Developmental Kapil, R.N., and Tiwari, S.C. 1978a. Plant embryological aspects of ultrastructure, histochemistry and recepinvestigations and fluorescence microscopy: an tivity of the stigma of Nicotiana sylvestris. Ann. Bot. assessment of integration. Int. Rev. Cytol. 53:291-331. 60:427-^37. Kapil, R.N., and Tiwari, S.C. 1978b. The integumentary Kandasamy, M.K., and Kristen, U. 1987b. Pollen tube tapetum. Bot. Rev. 44:457-490. growth in the style of Nicotiana sylvestris is neither Kapil, R.N., and Vasil, I.K. 1963. Ovule. In Recent Advances in the Embryology of Angiosperms, ed. P. Maheshwari, influenced by the ovary nor directed by a gradient. /. Plant Physiol. 131:495-500. pp. 41-67. Delhi: International Society of Plant Morphologists. Kandasamy, M.K., and Kristen, U. 1989. Ultrastructural responses of tobacco pollen tubes to heat shock. Kaplan, D.R. 1969. Seed development in Downingia. Protoplasma 153:104-110. Phytomorphology 19:253-278. Kandasamy, M.K., and Kristen, U. 1990. Developmental Kapoor, M. 1959. Influence of growth substances on the aspects of ultrastructure and histochemistry of the ovules of Zephyranthes. Phytomorphology 9:313-315. stylar transmitting tissue of Nicotiana sylvestris. Bot. Kappler, R., Kristen, U., and Morre, D.J. 1986. Membrane Ada 103:384-391. flow in plants: fractionation of growing pollen tubes of tobacco by preparative free-flow electrophoresis Kandasamy, M.K., Nasrallah, J.B., and Nasrallah, M.E. and kinetics of labeling of endoplasmic reticulum 1994. Pollen-pistil interactions and developmental and Golgi apparatus with [3H]leucine. Protoplasma regulation of pollen tube growth in Arabidopsis. 132:38-50. Development 120:3405-3418. Kandasamy, M.K., Parthasarathy, M.V., and Nasrallah, Kapros, T., Bogre, L., Nemeth, K., Bak6, L., Gyorgyey, J., M.E. 1991. High pressure freezing and freeze substiWu, S.C, and Dudits, D. 1992. Differential exprestution improve immunolabeling of S-locus specific sion of histone H3 gene variants during cell cycle glycoproteins in the stigma papillae of Brassica. and somatic embryogenesis in alfalfa. Plant Physiol. Protoplasma 162:187-191. 98:621-625. Kandasamy, M.K., and Vivekanandan, M. 1983. Karas, I., and Cass, D.D. 1976. Ultrastructural aspects of Biochemical composition of stigmatic exudate of sperm cell formation in rye: evidence for cell
References
589
plate involvement in generative cell division. Kawagoe, Y., Campbell, B.R., and Murai, N. 1994. Phytomorphology 26:36-45. Synergism between CACGTG (G-box) and CACCTG cis-elements is required for activation of the Karim, M.A., Mehta, S.L., and Singh, M.P. 1984. Studies bean seed storage protein ji-phaseolin gene. Plant J. on esterase isoenzyme pattern in anthers and seeds 5:885-890. of male sterile wheats. Z. Pflanzenzuchtg 93:309-319. Karper, R.E., and Stephens, J.C. 1936. Floral abnormalities Kawagoe, Y., and Murai, N. 1992. Four distinct nuclear in sorghum. /. Hexed. 27:183-194. proteins recognize in vitro the proximal promoter of the bean seed storage protein P-phaseolin gene conKarssen, CM., Brinkhorst-van der Swan, D.L.C., ferring spatial and temporal control. Plant /. Breekland, A.E., and Koornneef, M. 1983. Induction 2:927-936. of dormancy during seed development by endogenous abscisic acid: studies on abscisic acid deficient Kawagoe, Y, and Murai, N. 1996. A novel basic genotypes of Arabidopsis thaliana (L.) Heynh. Planta region/helix-loop-helix protein binds to a G-box 157:158-165. motif CACGTG of the bean seed storage protein Pphaseolin gene. Plant Sci. 116:47-57. Kartha, K.K., Chibbar, R.N., Georges, F., Leung, N., Caswell, K., Kendall, E., and Qureshi, J. 1989. Kawahara, R., Sunabori, S., Fukuda, H., and Komamine, Transient expression of chloramphenicol acetylA. 1992. A gene expressed preferentially in the globtransferase (CAT) gene in barley cell cultures and ular stage of somatic embryogenesis encodes elongaimmature embryos through microprojectile bomtion-factor l a in carrot. Eur. ]. Biochem. 209:157-162. bardment. Plant Cell Rep. 8:429-432. Kawahara, R., Sunabori, S., Fukuda, H., and Komamine, A. 1994. Analysis by in situ hybridization of the Karunanandaa, B., Huang, S., and Kao, T.-H. 1994. expression of elongation factor l a in the carrot cells Carbohydrate moiety of the Petuniainflata S3 protein during somatic embryogenesis. /. Plant Res. is not required for self-incompatibility interactions 107:361-364. between pollen and pistil. Plant Cell 6:1933-1940. Karunanandaa, B., Singh, A., and Kao, T.-H. 1994. Kawashima, I., Kennedy, T.D., Chino, M., and Lane, B.G. 1992. Wheat Ec metallothionein genes. Like Characterization of a predominantly pistilmammalian Zn2+ metallothionein genes, wheat expressed gene encoding a y-thionin-like protein of Petunia inflata. Plant Mol. Biol. 26:459-464. Zn2+ metallothionein genes are conspicuously Kasarda, D.D., Okita, T.W., Bernardin, J.E., Baecker, P.A., expressed during embryogenesis. Eur. }. Biochem. Nimmo, C.C., Lew, E.J.-L., Dietler, M.D., and Greene, 209:971-976. EC. 1984. Nucleic acid (cDNA) and amino acid Kazimierska, E.M. 1978. Embryological studies of cross sequences of a-type gliadins from wheat (Triticum compatibility in the genus Trifolium L. II. aestivum). Proc. Natl Acad. Sci. USA 81:4712^716. Fertilization, development of embryo and endosperm in crossing T. repens L. with T. medium L. Kasembe, J.N.R. 1967. Phenotypic restoration of fertility Genet. Polon. 19:15-24. in a male-sterile mutant by treatment with gibberellic acid. Nature 215:668. Keddie, J.S., Edwards, E.-W., Gibbons, T, Shaw, C.H., and Murphy, D.J. 1992a. Sequence of an oleosin cDNA Kasha, K.J., and Kao, K.N. 1970. High frequency haploid from Brassica napus. Plant Mol. Biol. 19:1079-1083. production in barley (Hordeum vulgare L.). Nature 225:874-876. Keddie, J.S., Hubner, G., Slocombe, S.P., Jarvis, R.R, Kater, M.M., Koningstein, G.M., Nijkamp, H.J.J., and Cummins, I., Edwards, E.-W., Shaw, C.H., and Stuitje, A.R. 1991. cDNA cloning and expression of Murphy, D.J. 1992b. Cloning and characterisation of Brassica napus enoyl-acyl carrier protein reductase in an oleosin gene from Brassica napus. Plant Mol. Biol. Escherichia coli. Plant Mol. Biol. 17:895-909. 19:443-453. Kato, H. 1968. The serial observations of the adventive Keddie, J.S., Tsiantis, M., Piffanelli, P., Cella, R., embryogenesis in the microculture of carrot tissue. Hatzopoulos, P., and Murphy, D.J. 1994. A seed-speSci. Papers Coll. Gen. Edn Univ. Tokyo 18:191-197. cific Brassica napus oleosin promoter interacts with a Kato, H., and Takeuchi, M. 1963. Morphogenesis in vitro G-box-specific protein and may be bi-directional. starting from single cells of carrot root. Plant Cell Plant Mol. Biol. 24:327-340. Physiol. 4:243-245. Keeler, S.J., Sanders, P., Smith, J.K., and Mazur, B.J. 1993. Katti, R.Y., Giddanavar, H.S., Naik, S., Agadi, S.N., and Regulation of tobacco acetolactate synthase gene Hegde, R.R. 1994. Persistence of callose and tapetum expression. Plant Physiol. 102:1009-1018. in the microsporogenesis of genie male sterile Kehrel, B., and Wiermann, R. 1985. Immunochemical Cajanus cajan (L.) Millsp. with well formed endothelocalization of phenylalanineammonia-lyase and cium. Cytologia 59:65-72. chalcone synthase in anthers. Planta 163:183-190. Kaufmann, H., Salamini, E, and Thompson, R.D. 1991. Keijzer, C.J. 1987. The processes of anther dehiscence and Sequence variability and gene structure at the selfpollen dispersal. II. The formation and the transfer incompatibility locus of Solanum tuberosum. Mol. mechanism of pollenkitt, cell-wall development of Gen. Genet. 226:457-466. the loculus tissues and a function of orbicules in Kaul, C.L., and Singh, S.P. 1966. Studies in male-sterile pollen dispersal. New Phytol. 105:499-507. barley. II. Pollen abortion. Crop Sci. 6:539-541. Keijzer, C.J., and Cresti, M. 1987. A comparison of anther Kaul, M.L.H. 1988. Male Sterility in Higher Plants. Berlin: tissue development in male sterile Aloe vera and Springer-Verlag. male fertile Aloe ciliaris. Ann. Bot. 59:533-542. Kaul, M.L.H., and Nirmala, C. 1993. Male sterility in pea. Keijzer, C.J., and Willemse, M.T.M. 1988. Tissue interacII. Male sex specific dyssynapsis. Cytologia 58:67-76. tions in the developing locule of Gasteria verrucosa Kaul, V., Theunis, C.H., Palser, B.F., Knox, R.B., and during microgametogenesis. Ada Bot. Neerl. Williams, E.G. 1987. Association of the generative 37:475-492. cell and vegetative nucleus in pollen tubes of Keith, K., Kraml, M., Dengler, N.G., and McCourt, P. 1994. Rhododendron. Ann. Bot. 59:227-235. fusca3: a heterochronic mutation affecting late
590
References
Requirement of ethylene for growth of callus and embryo development in Arabidopsis. Plant Cell somatic embryogenesis in Medicago sativa L. /. Expt. 6:589-600. Bot. 43:1199-1202. Keller, W.A., and Armstrong, K.C. 1978. High frequency production of microspore-derived plants from Kerhoas, C, Gay, G., and Dumas, C. 1987. A multidisciBrassica napus anther cultures. Z. Pflanzenzuchtg plinary approach to the study of the plasma mem80:100-108. brane of Zea mays pollen during controlled dehydration. Planta 171:1-10. Keller, W.A., and Armstrong, K.C. 1979. Stimulation of embryogenesis and haploid production in Brassica Kerhoas, C, Knox, R.B., and Dumas, C. 1983. Specificity campestris anther cultures by elevated temperature of the callose response in stigmas of Brassica. Ann. treatments. Theor. Appl. Genet. 55:65-67. Bot. 52:597-602. Kemble, R.J., and Bedbrook, J.R. 1980. Low molecular Kermode, A.R., and Bewley, J.D. 1985. The role of matuweight circular and linear DNA in mitochondria ration drying in the transition from seed developfrom normal and male-sterile Zea mays cytoplasm. ment to germination. I. Acquisition of Nature 284:565-566. desiccation-tolerance and germinability during development of Ricinus communis L. seeds. /. Expt. Kemble, R.J., Flavell, R.B., and Brettell, R.I.S. 1982. Bot. 36:1906-1915. Mitochondrial DNA analyses of fertile and sterile maize plants derived from tissue culture with the Kermode, A.R., and Bewley, J.D. 1988. The role of matuTexas male sterile cytoplasm. Theor. Appl. Genet. ration drying in the transition from seed develop62:213-217. ment to germination. V. Responses of the immature castor bean embryo to isolation from the whole seed: Kemble, R.J., Gunn, R.E., and Flavell, R.B. 1980. a comparison with premature desiccation. /. Expt. Classification of normal and male-sterile cytoplasms Bot. 39:487-497. in maize. II. Electrophoretic analysis of DNA species in mitochondria. Genetics 95:451-458. Khan, M.R.I., Gatehouse, J.A., and Boulter, D. 1980. The seed proteins of cowpea (Vigna unguiculata L. Walp.). Kemble, R.J., and Mans, R.J. 1983. Examination of the /. Expt. Bot. 31:1599-1611. mitochondrial genome of revertant progeny from S cms maize with cloned S-l and S-2 hybridization Khavkin, E.E., Misharin, S.I., Ivanov, V.N., and Danovich, probes. /. Mol. Appl. Genet. 2:161-171. K.N. 1977. Embryonal antigens in maize caryopses: the temporal order of antigen accumulation during Kemp, C.L. 1964. The effects of inhibitors of RNA and embryogenesis. Planta 135:225-231. protein synthesis on cytological development during meiosis. Chromosoma 15:652-665. Khavkin, E.E., Misharin, S.I., Markov, Y.Y., and Peshkova, A.A. 1978. Identification of embryonal antigens of Kemp, C.L. 1966. Electron microscope autoradiographic maize: globulins as primary reserve proteins of the studies of RNA metabolism in Trillium erectum embryo. Planta 143:11-20. microspores. Chromosoma 19:137-148. Kempin, S.A., Mandel, M.A., and Yanofsky, MR 1993. Kheyr-Pour, A., Bintrim, S.B., Ioerger, T.R., Remy, R., Hammond, S.A., and Kao, T.-H. 1990. Sequence Conversion of perianth into reproductive organs by diversity of pistil S-proteins associated with gametoectopic expression of the tobacco floral homeotic phytic self-incompatibility in Nicotiana alata. Sex. gene NAG1. Plant Physiol. 103:1041-1046. Plant Reprod. 3:88-97. Kennell, J.C, and Horner, H.T. 1985. Megasporogenesis and megagametogenesis in soybean, Glycine max. Khoo, U., and Stinson, H.T., Jr. 1957. Free amino acid difAm. ]. Bot. 72:1553-1564. ferences between cytoplasmic male sterile and norKennell, J.C, and Pring, D.R. 1989. Initiation and processmal fertile anthers. Proc. Natl Acad. Sci. USA ing of atp6, T-urfl3 and ORF221 transcripts from 43:603-607. mitochondria of T cytoplasm maize. Mol. Gen. Genet. Khoo, U., and Wolf, M.J. 1970. Origin and development of 216:16-24. protein granules in maize endosperm. Am. ]. Bot. 57:1042-1050. Kennell, J.C, Wise, R.P., and Pring, D.R. 1987. Influence of nuclear background on transcription of a maize Kiang, A.-S., and Kavanagh, T.A. 1996. Cytoplasmic male sterility (CMS) in Lolium perenne L. 2. The mitochonmitochondrial region associated with Texas male drial genome of a CMS line is rearranged and consterile cytoplasm. Mol. Gen. Genet. 210:399-406. tains a chimaeric atp 9 gene. Theor. Appl. Genet. Kenrick, J., Kaul, V., and Williams, E.G. 1986. Self-incom92:308-315. patibility in Acacia retinodes: site of pollen-tube arrest Kiesselbach, T.A. 1949. The structure and reproduction is the nucellus. Planta 169:245-250. of corn. Univ. Nebraska Agr. Expt. Station Bull. Kenrick, J., and Knox, R.B. 1981a. Post-pollination exu161:3-96. date from stigmas of Acacia (Mimosaceae). Ann. Bot. Kikkert, J.R., Hebert-Soule, D., Wallace, P.G., Striem, M., 48:103-106. and Reisch, B.I. 1996. Transgenic plantlets of Kenrick, J., and Knox, R.B. 1981b. Structure and histo"Chancellor' grapevine (Vitis sp.) from biolistic chemistry of the stigma and style of some Australian transformation of embryogenic cell suspensions. species of Acacia. Aust. J. Bot. 29:733-745. Plant Cell Rep. 15:311-316. Kenrick, J., and Knox, R.B. 1985. Self-incompatibility in the nitrogen-fixing tree, Acacia retinodes: quantitative Kikuchi, A., Satoh, S., Nakamura, N., and Fujii, T. 1995. Differences in pectic polysaccharides between carrot cytology of pollen tube growth. Theor. Appl. Genet. embryogenic and non-embryogenic calli. Plant Cell 69:481-488. Rep. 14:279-284. Kent, N., and Brink, R.A. 1947. Growth in vitro of immaKim, M.Z., and Raghavan, V. 1988. Induction of pollen ture Hordeum embryos. Science 106:547-548. plantlets in rice by spikelet culture. Plant Cell Rep. Keown, A.C., Taiz, L., and Jones, R.L. 1977. The nuclear 7:560-563. DNA content of developing barley aleurone cells. Kim, S.-R., Kim, Y, and An, G. 1993. Molecular cloning Am.}. Bot. 64:1248-1253. and characterization of anther-preferential cDNA Kepczynski, J., McKersie, B.D., and Brown, D.C.W. 1992.
References
591
encoding a putative actin-depolymerizing factor. Kitada, K., Kurata, N., Satoh, H., and Omura, T. 1983. Plant Mol. Bib/. 21:39-45. Genetic control of meiosis in rice, Oryza sativa L. I. Classification of meiotic mutants induced by MNU Kim, S.-R., Finkel, D., Chung, Y.-Y, and An, G. 1994. and their cytogenetical characteristics. Jap. ]. Genet. Abundance patterns of lily pollen cDNAs: character58:231-240. ization of three pollen-preferential cDNA clones. Sex. Plant Reprod. 7:76-86. Kitano, H., Tamura, Y, Satoh, H., and Nagato, Y 1993. Hierarchical regulation of organ differentiation durKim, W.T., Li, X., and Okita, T.W. 1993. Expression of storing embryogenesis in rice. Plant ]. 3:607-610. age protein multigene families in developing rice endosperm. Plant Cell Physiol. 34:595-603. Kiyosue, T., and Shinozaki, K. 1995. Cloning of a carrot cDNA for a member of the family of ADP-ribosylaKim, W.T., and Okita, T.W. 1988a. Nucleotide and primary tion factors (ARFs) and characterization of the bindsequence of a major rice prolamine. FEBS Lett. ing of nucleotides by its product after expression in 231:308-310. £. coli. Plant Cell Physiol. 36:849-856. Kim, W.T., and Okita, T.W. 1988b. Structure, expression, and heterogeneity of the rice seed prolamines. Plant Kiyosue, T., Dong, J.G., Satoh, S., Kamada, H., and Physiol. 88:649-655. Harada, H. 1990a. Detection of an embryogenic cell antigen in carrot. Plant Cell Physiol. 31:947-950. Kim, W.T., Franceschi, V.R., Krishnan, H.B., and Okita, T.W. 1988. Formation of wheat protein bodies: Kiyosue, T., Nakajima, M., Yamaguchi, I., Satoh, S., Kamada, H., and Harada, H. 1992a. Endogenous levinvolvement of the Golgi apparatus in gliadin transels of abscisic acid in embryogenic cells, non-embryoport. Planta 176:173-182. genic cells and somatic embryos of carrot (Daucus Kim, Y., and An, G. 1992. Pollen-specific expression of the carota L.). Biochem. Physiol. Pflanz. 188:343-347. Arabidopsis thaliana aj-tubulin promoter assayed by P-glucuronidase, chloramphenicol acetyltransferase Kiyosue, T, Nakayama, J., Satoh, S., Isogai, A., Suzuki, and diphtheria toxin reporter genes. Transgen. Res. A., Kamada, H., and Harada, H. 1992b. Partial 1:188-194. amino-acid sequence of ECP31, a carrot embryoKim, Y.-H., and Janick, J. 1991. Abscisic acid and proline genic-cell protein, and enhancement of its accumuimprove desiccation tolerance and increase fatty lation by abscisic acid in somatic embryos. Planta acid content of celery somatic embryos. Plant Cell 186:337-342. Tissue Organ Cult. 24:83-89. Kiyosue, T., Satoh, S., Kamada, H., and Harada, H. 1991. Kindiger, B., Beckett, J.B., and Coe, E.H., Jr. 1991. Purification and immunohistochemical detection of Differential effects of specific chromosomal deficienan embryogenic cell protein in carrot. Plant Physiol. cies on the development of the maize pollen grain. 95:1077-1083. Genome 34:579-594. Kiyosue, T., Takano, K., Kamada, H., and Harada, H. King, R.W. 1976. Abscisic acid in developing wheat grains 1990b. Induction of somatic embryogenesis in carrot and its relationship to grain growth and maturation. by heavy metal ions. Can. ]. Bot. 68:2301-2303. Planta 132:43-51. Kiyosue, T., Yamaguchi-Shinozaki, K., Shinozaki, K., King, T.J., and Briggs, R. 1953. The transplantability of Higashi, K., Satoh, S., Kamada, H., and Harada, H. nuclei of arrested hybrid blastulae (R. pipiens 9 X R. 1992c. Isolation and characterization of a cDNA that encodes ECP31, an embryogenic-cell protein from catesbeiana a). ]. Expt. Zool. 123:61-78. Kini, A.V., Seetharam, A., and Joshi, S.S. 1994. Mechanism carrot. Plant Mol. Bib/. 19:239-249. of pollen abortion in cytoplasmic male sterile line of Kiyosue, T., Yamaguchi-Shinozaki, K., Shinozaki, K., sunflower. Cytologia 59:121-124. Kamada, H., and Harada, H. 1993. cDNA cloning of Kirby, E.G., and Vasil, I.K. 1979. Effect of pollen protein ECP40, an embryogenic-cell protein in carrot, and its diffusates on germination of eluted pollen samples expression during somatic and zygotic embryogeneof Petunia hybrida in vitro. Ann. Bot. 44:361-367. sis. Plant Mol. Bib/. 21:1053-1068. Kirch, H.-H., Li, Y.-Q., Seul, U., and Thompson, R.D. 1995. Klein, H.D. 1969. Male sterility in Pisum. Nucleus 12:167-172. The expression of a potato (Solanum tuberosum) SRNase gene in Nicotiana labacum pollen. Sex. PlantKlein, H.D., and Milutinovic, M. 1971. Genbedingte Storungen der Infloreszenz- und Bliitenbildung. Reprod. 8:77-84. Theor. Appl. Genet. 41:255-258. Kirch, H.H., Uhrig, H., Lottspeich, F., Salamini, F., and Thompson, R.D. 1989. Characterization of proteins Klein, T.M., Roth, B.A., and Fromm, M.E. 1989. associated with self-incompatibility in Solanum Regulation of anthocyanin biosynthetic genes introtuberosum. Theor. Appl. Genet. 78:581-588. duced into intact maize tissues by microprojectiles. Proc. Natl Acad. Sci. USA 86:6681-6685. Kirihara, J.A., Petri, J.B., and Messing, J. 1988. Isolation and sequence of a gene encoding a methionine-rich Klein, T.M., Gradziel, T, Fromm, M.E., and Sanford, J.C. 10-kDa zein protein from maize. Gene 71:359-370. 1988. Factors influencing gene delivery into Zea mays Kirihara, J.A., Hunsperger, J.P., Mahoney, W.C., and cells by high-velocity microprojectiles. Biotechnology Messing, J.W. 1988. Differential expression of a gene 6:559-563. for a methionine-rich storage protein in maize. Mol. Kleman-Mariac, C, Rougier, M., Cock, J.M., Gaude, T., and Dumas, C. 1995. S-locus glycoproteins are Gen. Genet. 211:477-484. Kirk, J.T.O., and Tilney-Bassett, R.A.E. 1978. The Plastids. expressed along the path of the pollen tubes in Their Chemistry, Structure, Growth and Inheritance. 2nd Brassica pistils. Planta 196:614-621. ed. Amsterdam: Elsevier/North-Holland Biomedical Klemsdal, S.S., Hughes, W., Lonneborg, A., Aalen, R.B., Press. and Olsen, O.-A. 1991. Primary structure of a novel barley gene differentially expressed in immature Kishitani, S., Yomoda, A., Konno, N., and Tanaka, Y. 1993. aleurone layers. Mol. Gen. Genet. 228:9-16. Involvement of phenylalanine ammonia-lyase in the development of pollen in broccoli (Brassica oleracea Klinge, B., and Werr, W. 1995. Transcription of the Zea mays homeobox (ZmHox) genes is activated early L.). Sex. Plant Reprod. 6:244-248.
592 in embryogenesis and restricted to meristems of the maize plant. Devel Genet. 16:349-357. Klosgen, R.B., Gierl, A., Schwarz-Sommer, Z., and Saedler, H. 1986. Molecular analysis of the waxy
References
cell determinants and control of fertilisation in plants. In Eukaryote Cell Recognition. Concepts and
Model Systems, ed. G.P. Chapman, C.C. Ainsworth and C.J. Chatham, pp. 175-193. Cambridge: Cambridge University Press. locus of Zea mays. Mol. Gen. Genet. 203:237-244. Kloti, A., Iglesias, V.A., Wunn, J., Burkhardt, P.K., Datta, Knox, R.B., Willing, R.R., and Ashford, A.E. 1972. Role of pollen-wall proteins as recognition substances in S.K., and Potrykus, I. 1993. Gene transfer by electrointerspecific incompatibility in poplars. Nature poration into intact scutellum cells of wheat 237:381-383. embryos. Plant Cell Rep. 12:671-675. Klucher, K.MV Chow, H., Reiser, L., and Fischer, R.L. 1996. Knox, R.B., Willing, R.R., and Pryor, L.D. 1972. The AINTEGUMENTA gene of Arabidopsis required Interspecific hybridization in poplars using recognifor ovule and female gametophyte development is tion pollen. Silvae Genet. 21:65-69. related to the floral homeotic gene APETALA2. Plant Knox, R.B., Clarke, A., Harrison, S., Smith, P., and Cell 8:137-153. Marchalonis, J.J. 1976. Cell recognition in plants: determinants of the stigma surface and their pollen Klyuchareva, M.V. 1960. On the changes in deoxyribonuinteractions. Proc. Natl Acad. Sci. USA 73:2788-2792. cleic acid of the egg cell during fertilization in barley, wheat, and rye. Doklady Bot. Sci. 134:193-196. Knudsen, S., and Miiller, M. 1991. Transformation of the Knox, R.B. 1971. Pollen-wall proteins: localization, developing barley endosperm by particle bombardenzymic and antigenic activity during development ment. Planta 185:330-336. in Gladiolus (Iridaceae). /. Cell Sci. 9:209-237. Knutzon, D.S., Thompson, G.A., Radke, S.E., Johnson, Knox, R.B. 1973. Pollen wall proteins: pollen—stigma W.B., Knauf, V.C., and Kridl, J.C. 1992. Modification interactions in ragweed and Cosmos (Compositae). /. of Brassica seed oil by antisense expression of a Cell Sci. 12-A21-U3. stearoyl-acyl carrier protein desaturase gene. Proc. Natl Acad. Sci. USA 89:2624-2628. Knox, R.B. 1984a. The pollen grain. In Embryology of Angiosperms, ed. B.M. Johri, pp. 197-271. Berlin: Kobayashi, T., Kobayashi, E., Sato, S., Hotta, Y., Miyajima, Springer-Verlag. N., Tanaka, A., and Tabata, S. 1994. Characterization of cDNAs induced in meiotic prophase in lily Knox, R.B. 1984b. Pollen-pistil interactions. In Encyclopedia microsporocytes. DNA Res. Tokyo 1:15-26. of Plant Physiology, Vol. 17, ed. H.F. Linskens and J. Heslop-Harrison, pp. 508-608. Berlin: Springer-Verlag. Kochba, J., Lavee, S., and Spiegel-Roy, P. 1977. Differences in peroxidase activity and isoenzymes in embryoKnox, R.B., Dickinson, H.G., and Heslop-Harrison, J. genie and non-embryogenic Shamouti' orange ovu1970. Cytochemical observations on changes in RNA lar callus lines. Plant Cell Physiol. 18:463-^67. content and acid phosphatase activity during the meiotic prophase in the anther of Cosmos bipinnatus Kochba, J., Spiegel-Roy, P., Neumann, H., and Saad, S. Cav. Ada Bot. Neerl. 19:1-6. 1978. Stimulation of embryogenesis in Citrus ovular callus by ABA, ethephon, CCC and Alar and its supKnox, R.B., and Heslop-Harrison, J. 1966. Control of pression by GA3. Z. Pflanzenphysiol. 89:427-432. pollen fertility through the agency of the light regime in the grass Dichanthium aristatum. Phyton Kodrzycki, R., Boston, R.S., and Larkins, B.A. 1989. The Austria 11:256-267. opaque-2 mutation of maize differentially reduces zein gene transcription. Plant Cell 1:105-114. Knox, R.B., and Heslop-Harrison, J. 1969. Cytochemical localization of enzymes in the wall of the pollen Koeppe, D.E., Cox, J.K., and Malone, C.P. 1978. grain. Nature 223:92-94. Mitochondrial heredity: a determinant in the toxic Knox, R.B., and Heslop-Harrison, J. 1970a. Pollen-wall response of maize to the insecticide methomyl. proteins: localization and enzymic activity. /. Cell Sci. Science 201:1227-1229. 6:1-27. Koes, R.E., van Blokland, R., Quattrocchio, F., van Tunen, Knox, R.B., and Heslop-Harrison, J. 1970b. Direct demonA.J., and Mol, J.N.M. 1990. Chalcone synthase promoters in Petunia are active in pigmented and unpigstration of the low permeability of the angiosperm meiotic tetrad using a fluorogenic ester. Z. mented cell types. Plant Cell 2:379-392. Pflanzenphysiol. 62:451-459. Kohler, F., and Wenzel, G. 1985. Regeneration of isolated barley microspores in conditioned media and trials Knox, R.B., and Heslop-Harrison, J. 1971a. Pollen-wall to characterize the responsible factor. /. Plant Physiol. proteins: localization of antigenic and allergenic pro121:181-191. teins in the pollen-grain walls of Ambrosia spp. (ragweeds). Cytobios 4:49-54. Kohler, R.H., Horn, R., Lossl, A., and Zetsche, K. 1991. Knox, R.B., and Heslop-Harrison, J. 1971b. Pollen-wall Cytoplasmic male sterility in sunflower is correlated proteins: electron-microscopic localization of acid with the co-transcription of a new open reading phosphatase in the intine of Crocus vernus. ]. Cell Sci. frame with the atpA gene. Mol. Gen. Genet. 8:727-733. 227:369-376. Knox, R.B., and Heslop-Harrison, J. 1971c. Pollen-wall Kohno, T., Chaen, S., and Shimmen, T. 1990. proteins: the fate of intine-held antigens on the Characterization of the translocator associated with stigma in compatible and incompatible pollinations pollen tube organelles. Protoplasma 154:179-183. of Phalaris tuberosa L. /. Cell Sci. 9:239-251. Kohno, T., and Shimmen, T. 1987. Ca2+-induced fragmentation of actin filaments in pollen tubes. Protoplasma Knox, R.B., Heslop-Harrison, J., and Reed, C. 1970. Localization of antigens associated with the pollen 141:177-179. grain wall by immunofluorescence. Nature Kohno, T., and Shimmen, T. 1988a. Mechanism of Ca2+ 225:1066-1068. inhibition of cytoplasmic streaming in lily pollen tubes. /. Cell Sci. 91:501-509. Knox, R.B., and Singh, M.B. 1987. New perspectives in pollen biology and fertilization. Ann. Bot. 60: Suppl. Kohno, T., and Shimmen, T. 1988b. Accelerated sliding of 4,15-37. pollen tube organelles along Characeae actin bundles regulated by Ca2+. /. Cell Bio/. 106:1539-1543. Knox, R.B., Southworth, D., and Singh, M.B. 1988. Sperm
References
593
Kohno, T., Ishikawa, Rv Nagata, T., Kohama, K., and tion in carrot somatic embryos in response to Shimmen, T. 1992. Partial purification of myosin abscisic acid. Plant Physiol. Biochem. 31:95-102. from lily pollen tubes by monitoring with in vitro Koornneef, M., Reuling, G., and Karssen, CM. 1984. The motility assay. Protoplasma 170:77-85. isolation and characterization of abscisic acid-insensitive mutants of Arabidopsis thaliana. Physiol. Plant. Kohno, T., Okagaki, T., Kohama, K., and Shimmen, T. 61:377-383. 1991. Pollen tube extract supports the movement of actin filaments in vitro. Protoplasma 161:75-77. Koornneef, M., Hanhart, C.J., Hilhorst, H.W.M., and Kohno-Murase, J., Murase, M., Ichikawa, H., and Karssen, CM. 1989. In vivo inhibition of seed develImamura, J. 1994. Effects of an antisense napin gene opment and reserve protein accumulation in recomon seed storage compounds in transgenic Brassica binants of abscisic acid biosynthesis and napus seeds. Plant Mol. Biol. 26:1115-1124. responsiveness mutants in Arabidopsis thaliana. Plant Physiol. 90:463-469. Kojima, K., Sasaki, S., and Yamamoto, N. 1994. Lightindependent and tissue-specific expression of a Korth, K.L., and Levings, C.E., III. 1993. Baculovirus reporter gene mediated by the pine cab-6 promoter expression of the maize mitochondrial protein in transgenic tobacco. Plant ]. 6:591-596. URF13 confers insecticidal activity in cell cultures Koltunow, A.M., 1994. Apomixis - other pathways for and larvae. Proc. Natl Acad. Sci. USA 90:3388-3392. reproductive development in angiosperms. In Korth, K.L., Kaspi, C.I., Siedow, J.N., and Levings, C.S., Genetic Control of Self-Incompatibility and Reproductive III. 1991. URF-13, a maize mitochondrial pore-formDevelopment in Flowering Plants, ed. E.G. Williams, ing protein, is oligomeric and has a mixed orientaA.E. Clarke and R.B. Knox, pp. 486-512. Dordrecht: tion in Escherichia coli plasma membranes. Proc. Natl Kluwer Academic Publishers. Acad. Sci. USA 88:10865-10869. Koltunow, A.M., Bicknell, R.A., and Chaudhury, A.M. Kortt, A.A., Caldwell, J.B., Lilley, G.G., and Higgins, T.J.V. 1995. Apomixis: molecular strategies for the genera1991. Amino acid and cDNA sequences of a methiotion of genetically identical seeds without fertilizanine-rich 2S protein from sunflower seed (Helianthus tion. Plant Physiol. 108:1345-1352. annuus L.). Eur. J. Biochem. 195:329-334. Koltunow, A.M., Truettner, J., Cox, K.H., Wallroth, M., Kost, B., Leduc, N., Sautter, C, Potrykus, I., and Neuhaus, and Goldberg, R.B. 1990. Different temporal and G. 1996. Transient marker-gene expression during spatial gene expression patterns occur during anther zygotic in-vitro embryogenesis of Brassica juncea development. Plant Cell 2:1201-1224. (Indian mustard) following particle bombardment. Planta 198:211-220. Komaki, M.S., Okada, K., Nishino, E., and Shimura, Y. 1988. Isolation and characterization of novel Kotilainen, M., Helariutta, Y, Elomaa, P., Paulin, L., and mutants of Arabidopsis thaliana defective in flower Teeri, T.H. 1994. A corolla- and carpel-abundant, development. Development 104:195-203. non-specific lipid transfer protein gene is expressed in the epidermis and parenchyma of Gerbera hybrida Konar, R.N., and Linskens, H.F. 1966a. The morphology var. Regina (Compositae). Plant Mol. Biol. and anatomy of the stigma of Petunia hybrida. Planta 26:971-978. 71:356-371. Konar, R.N., and Linskens, H.F. 1966b. Physiology and Kott, L.S., Polsoni, L., and Beversdorf, W.D. 1988. biochemistry of the stigmatic fluid of Petunia hybrida. Cytological aspects of isolated microspore culture of Planta 71:372-387. Brassica napus. Can. J. Bot. 66:1658-1664. Konar, R.N., and Nataraja, K. 1965. Experimental studies in Kott, L.S., Polsoni, L., Ellis, B., and Beversdorf, W.D. 1988. Ranunculus sceleratus L. Development of embryos Autotoxicity in isolated microspore cultures of from the stem epidermis. Phytomorphology 15:132—137. Brassica napus. Can. J. Bot. 66:1665-1670. Konar, R.N., and Stanley, R.G. 1969. Wall-softening Koul, A.K., and Karihaloo, J.L. 1977. In vivo embryoids from enzymes in the gynoecium and pollen of anthers of Narcissus biflorus Curt. Euphytica 26:97—102. Hemerocallis fulva. Planta 84:304-310. Kovacs, M., Barnabas, B., and Kranz, E. 1994. The isolaKonar, R.N., Thomas, E., and Street, H.E. 1972. Origin and tion of viable egg cells of wheat (Triticum aestivum structure of embryoids arising from epidermal cells of L.). Sex. Plant Reprod. 7:311-312. the stem of Ranunculus sceleratus L. J. Cell Sci. 11:77-93. Kovacs, M., Barnabas, B., and Kranz, E. 1995. ElectroKoning, A., Jones, A., Fillatti, J., Comai, L., and fused isolated wheat Triticum aestivum L.) gametes Lassner, M.W. 1992. Arrest of embryo developdevelop into multicellular structures. Plant Cell Rep. ment in Brassica napus mediated by modified 15:178-180. Pseudomonas aeruginosa exotoxin A. Plant Mol. Kovaleva, L.V., Milyaeva, E.L., and Chailakhyan, M.K. Biol. 18:247-258. 1978. Overcoming self-incompatibility by inhibitors of nucleic acid and protein metabolism. Kononowicz, A.J., and Janick, J. 1984. In vitro developPhytomorphology 28:445-449. ment of zygotic embryos of Theobroma cacao. ]. Am. Soc. Hort. Sci. 109:266-269. Kowles, R.V., and Phillips, R.L. 1985. DNA amplification Kononowicz, A.J., Nelson, D.E., Singh, N.K., Hasegawa, patterns in maize endosperm nuclei during kernel P.M., and Bressan, R.A. 1992. Regulation of the development. Proc. Natl Acad. Sci. USA 82:7010-7014. osmotin gene promoter. Plant Cell 4:513-524. Kowles, R.V., and Phillips, R.L. 1988. Endosperm develKononowicz, H., and Janick, J. 1984. Response of embryoopment in maize. Int. Rev. Cytol. 112:97-136. genie callus of Theobroma cacao L. to gibberellic acid Kowles, R.V., Srienc, F., and Phillips, R.L. 1990. and inhibitors of gibberellic acid synthesis. Z. Endoreduplication of nuclear DNA in the developPflanzenphysiol. 113:359-366. ing maize endosperm. Devel Genet. 11:125-132. Kononowicz, H., and Janick, J. 1988. Somatic embryogen- Kowles, R.V., McMullen, M.D., Yerk, G., Phillips, R.L., esis via callus of Theobroma cacao L. I. Cell cycle, DNA Kraemer, S., and Srienc, F. 1992. Endosperm mitotic content, RNA synthesis, and DNA template activity. activity and endoreduplication in maize affected by Ada Physiol. Plant. 10:93-105. defective kernel mutations. Genome 35:68-77. Koontz, D.A., and Choi, J.H. 1993. Protein phosphoryla- Kowyama, Y., Kakeda, K., Nakano, R., and Hattori, T.
594
References
Miflin, B.J. 1983. Molecular analysis of a mutation 1995. SLG/SRK-like genes are expressed in the conferring the high-lysine phenotype on the grain reproductive tissues of Ipomoea trifida. Sex. Plant of barley {Hordeum vulgare). Cell 34:161-167. Reprod. 8:333-338. Kowyama, Y., Kunz, C, Lewis, I., Newbigin, E., Clarke, Kress, W.J., and Stone, D.E. 1982. Nature of the sporoderm in monocotyledons, with special reference to the A.E., and Anderson, M.A. 1994. Self-compatibility in pollen grains of Canna and Heliconia. Grana a Lycopersicon penwianum variant (LA2157) is associated with a lack of style S-RNase activity. Theor. Appl. 21:129-148. Genet. 88:859-864. Kress, W.J., Stone, D.E., and Sellers, S.C. 1978. Kozar, F. 1974. Ultrastructure of pollen of Opnntia polyaUltrastructure of exine-less pollen: Heliconia cantha. Can. J. Bot. 52:313-315. (Heliconiaceae). Am. J. Bot. 65:1064-1076. Koziel, M.G., Beland, G.L., Bowman, C, Carozzi, N.B., Kreuger, M., and van Hoist, G.-J. 1993. Arabinogalactan Crenshaw, R., Crossland, L., Dawson, J., Desai, N., proteins are essential in somatic embryogenesis of Hill, M., Kadwell, S., Launis, K., Lewis, K., Daucus carota L. Planta 189:243-248. Maddox, D., McPherson, K., Meghji, M.R., Merlin, Kreuger, M., and van Hoist, G.-J. 1995. ArabinogalactanE., Rhodes, R., Warren, G.W., Wright, M., and protein epitopes in somatic embryogenesis of Evola, S.V. 1993. Field performance of elite transDaucus carota L. Planta 197:135-141. genic maize plants expressing an insecticidal pro- Kriete, G., Niehaus, K., Perlick, A.M., Piihler, A., and tein derived from Bacillus thuringiensis. Broer, I. 1996. Male sterility in transgenic tobacco Biotechnology 11:194-200. plants induced by tapetum-specific deacetylation of the externally applied non-toxic compound NKragh, K.M., Jacobsen, S., Mikkelsen, J.D., and Nielsen, acetyl-L-phosphinothricin. Plant J. 9:809-818. K.A. 1991. Purification and characterization of three chitinases and one P-l,3-glucanase accumulating in Krishnan, H.B., Franceschi, V.R., and Okita, T.W. 1986. the medium of cell suspension cultures of barley Immunochemical studies on the role of the Golgi (Hordenm vnlgare L.). Plant Sci. 76:65-77. complex in protein-body formation in rice seeds. Planta 169:471-480. Kranz, E., Bautor, J., and Lorz, H. 1991a. In vitro fertilization of single, isolated gametes of maize mediated Krishnan, H.B., and White, J.A. 1995. Morphometric analysis of rice seed protein bodies. Implication for a by electrofusion. Sex. Plant Reprod. 4:12-16. significant contribution of prolamine to the total proKranz, E., Bautor, J., and Lorz, H. 1991b. Elecrrofusiontein content of rice endosperm. Plant Physiol. mediated transmission of cytoplasmic organelles 109:1491-1495. through the in vitro fertilization process, fusion of sperm cells with synergids and central cells, and cell Krishnan, H.B., White, J.A., and Pueppke, S.G. 1988. reconstitution in maize. Sex. Plant Reprod. 4:17-21. Immunogold localization of prolamines in developing Haynaldia villosa endosperm. Protoplasma Kranz, E., and Lorz, H. 1990. Micromanipulation and in vitro fertilization with single pollen grains of maize. 144:25-33. Sex. Plant Reprod. 3:160-169. Krishnan, H.B., White, J.A., and Pueppke, S.G. 1989. Immunocytochemical analysis of protein body forKranz, E., and Lorz, H. 1993. In vitro fertilization with isomation in seeds of Sorghum bicolor. Can. ]. Bot. lated, single gametes results in zygotic embryogene67:2850-2856. sis and fertile maize plants. Plant Cell 5:739-746. Kranz, E., and Lorz, H. 1994. In vitro fertilisation of maize Krishnan, H.B., White, J.A., and Pueppke, S.G. 1990. Immunocytochemical evidence for the involvement by single egg and sperm cell protoplast fusion mediof the Golgi apparatus in the transport of the vacuoated by high calcium and high pH. Zygote 2:125-128. lar protein, y-secalin, in rye (Secale cereale) Kranz, E., von Wiegen, P., and Lorz, H. 1995. Early cytoendosperm. Cereal Chem. 67:360-366. logical events after induction of cell division in egg cells and zygote development following in vitro fer- Krishnaraj, S., and Vasil, I.K. 1995. Somatic embryogenetilization with angiosperm gametes. Plant ]. 8:9-23. sis in herbaceous monocots. In In Vitro Embryogenesis in Plants, ed. T. A. Thorpe, pp. Krasowski, M.J., and Owens, J.N. 1993. Ultrastructural 417-470. Dordrecht: Kluwer Academic Publishers. and histochemical postfertilization megagametophyte and zygotic embryo development of white Krishnasamy, S., Grant R.A., and Makaroff, C.A. 1994. Subunit 6 of the Fo-ATP synthase complex from cytospruce (Picea glauca) emphasizing the deposition of plasmic male-sterile radish: RNA editing and NH2seed storage products. Can. ]. Bot. 71:98-112. terminal protein sequencing. Plant Mol. Biol. Krebbers, E., Herdies, L., de Clercq, A., Seurinck, J., 24:129-141. Leemans, J., van Damme, J., Segura, M., Gheysen, G., van Montagu, M., and Vandekerckhove, J. 1988. Krishnasamy, S., and Makaroff, C.A. 1993. Characterization of the radish mitochondrial orfB Determination of the processing sites of an Arabidopsis 2S albumin and characterization of the locus: possible relationship with male sterility in complete gene family. Plant Physiol. 87:859-866. Ogura radish. Curr. Genet. 24:156-163. Kreis, M., Forde, B.G., Rahman, S., Miflin, B.J., and Krishnasamy, S., and Makaroff, C.A. 1994. Organ-specific reduction in the abundance of a mitochondrial proShewry, PR. 1985. Molecular evolution of the seed tein accompanies fertility restoration in cytoplasmic storage proteins of barley, rye and wheat. /. Mol. Biol. male-sterile radish. Plant Mol. Biol. 26:935-946. 183:499-502. Kreis, M., Shewry, PR., Forde, B.G., Rahman, S., Kristen, U. 1977. Granulocrine Ausscheidung von Narbensekret durch Vesikel des Endoplasmatischen Bahramian, M.B., and Miflin, B.J. 1984. Molecular Retikulums bei Aptenia cordifolia. Protoplasma analysis of the effects of the lys 3a gene on the 92:243-251. expression of Hor loci in developing endosperms of barley (Hordenm vulgare L.). Biochem. Genet. Kristen, U., Biedermann, M., Liebezeit, G., Dawson, R., and Bohm, L. 1979. The composition of stigmatic 22:231-255. exudate and the ultrastructure of the stigma papillae Kreis, M., Shewry, P.R., Forde, B.G., Rahman, S., and
References
in Aptenia cordifolia. Eur. J. Cell Biol. 19:281-287. Kriz, A.L. 1989. Characterization of embryo globulins encoded by the maize Gib genes. Biochem. Genet. 27:239-251. Kriz, A.L., Boston, R.S., and Larkins, B.A. 1987. Structural and transcriptional analysis of DNA sequences flanking genes that encode 19 kilodalton zeins. Mol. Gen. Genet. 207:90-98. Kriz, A.L., and Schwartz, D. 1986. Synthesis of globulins in maize embryos. Plant Physiol. 82:1069-1075. Kriz, A.L., and Wallace, N.H. 1991. Characterization of the maize Globulin-2 gene and analysis of two null alleles. Biochem. Genet. 29:241-254. Kriz, A.R., Wallace, M.S., and Paiva, R. 1989. Globulin gene expression in embryos of maize viviparous mutants. Evidence for regulation of the Glbl gene by ABA. Plant Physiol. 92:538-542. Krizek, B.A., and Meyerowitz, E.M. 1996a. The Arabidopsis homeotic genes APETALA3 and PISTILLATA are sufficient to provide the B class organ identity function. Development 122:11-22. Krizek, B.A., and Meyerowitz, E.M. 1996b. Mapping the protein regions responsible for the functional specificities of the Arabidopsis MADS domain organidentity proteins. Proc. Natl Acad. Sci. USA 93:4063-4070. Krochko, J.E., Pramanik, S.K., and Bewley, J.D. 1992. Contrasting storage protein synthesis and messenger RNA accumulation during development of zygotic and somatic embryos of alfalfa (Medicago sativa L.). Plant Physiol. 99:46-53. Krochko, J.E., Bantroch, D.J., Greenwood, J.S., and Bewley, J.D. 1994. Seed storage proteins in developing somatic embryos of alfalfa: defects in accumulation compared to zygotic embryos. /. Expt. Bot. 45:699-708. Kroh, M. 1966. Reaction of pollen after transfer from one stigma to another (Contribution to the character of the incompatibility mechanism in Cruciferae). Zuchter 36:185-189. Kroh, M. 1967a. Fine structure of Petunia pollen germinated in vivo. Rev. Palaeobot. Palynol. 3:197-203. Kroh, M. 1967b. Bildung und Transport des Narbensekrets von Petunia hybrida. Planta 77:250-260. Kroh, M., Gorissen, M.H., and Pfahler, P.L. 1979. Ultrastructural studies on styles and pollen tubes of Zea mays L. General survey on pollen tube growth in vivo. Ada Bot. Neerl. 28:513-518. Kroh, M., and Knuiman, B. 1982. Ultrastructure of cell wall and plugs of tobacco pollen tubes after chemical extraction of polysaccharides. Planta 154:241-250. Kroh, M., and Loewus, F. 1968. Biosynthesis of pectic substance in germinating pollen: labeling with myoinositol-2-14C. Science 160:1352-1354. Kroh, M., and Munting, A.J. 1967. Pollen-germination and pollen tube growth in Diplotaxis tenuifolia after crosspollination. Acta Bot. Neerl. 16:182-187. Kroh, M., and van Bakel, C.H.J. 1973. Incorporation of label into the intercellular substance of stylar transmitting tissue from Petunia pistils labeled with tritiated myo-inositol. An electron microscopic autoradiographic study. Acta Bot. Neerl. 22:106-111. Kroh, M., Miki-Hirosige, H., Rosen, W., and Loewus, F. 1970. Incorporation of label into pollen tube walls
595 from myoinositol-labeled Lilium longiflorwn pistils. Plant Physiol. 45:92-94. Krug, C.A., and Bacchi, O. 1943. Triploid varieties of Citrus. J.Hered. 34:277-283. Kubieri, E. 1968. Cytological processes during the development of antipodals in Ammophila arenaria Link. Acta Biol. Cracow Ser. Bot. 11:21-29. Kucera, V., and Poldk, J. 1975. The serological specificity of S alleles of homozygous incompatible lines of the marrow-stem kale (Brassica oleracea var. acephala DC). Bio/. Plant. 17:50-54. Kuehnle, A.R., and Earle, E.D. 1989. In vitro selection for methomyl resistance in CMS-T maize. Theor. Appl. Genet. 78:672-682. Kuhlmann, U., Foroughi-Wehr, B., Graner, A., and Wenzel, G. 1991. Improved culture system for microspores of barley to become a target for DNA uptake. Plant Breed. 107:165-168. Kuhtreiber, W.M., and Jaffe, L.F. 1990. Detection of extracellular calcium gradients with a calcium-specific vibrating electrode. /. Cell Biol. 110:1565-1573. Kulikauskas, R., Hou, A., Muschietti, J., and McCormick, S. 1995. Comparisons of diverse plant species reveal that only grasses show drastically reduced levels of ubiquitin monomer in mature pollen. Sex. Plant Reprod. 8:326-332. Kumamaru, T, Satoh, H., Iwata, N., Omura, T., Ogawa, M., and Tanaka, K. 1988. Mutants for rice storage proteins. 1. Screening of mutants for rice storage proteins of protein bodies in the starchy endosperm. Theor. Appl. Genet. 76:11-16. Kumar, V., and Trick, M. 1993. Sequence complexity of the S receptor kinase gene family in Brassica. Mol. Gen. Genet. 241:440-446. Kumar, V, and Trick, M. 1994. Expression of the S-locus receptor kinase multigene family in Brassica oleracea. Plant/. 6:807-813. Kunce, CM., and Trelease, R.N. 1986. Heterogeneity of catalase in maturing and germinated cotton seeds. Plant Physiol. 81:1134-1139. Kunst, L., Klenz, J.E., Martinez-Zapater, J., and Haughn, G.W. 1989. API gene determines the identity of perianth organs in flowers of Arabidopsis thaliana. Plant Cell 1:1195-1208. Kunz, C, Chang, A., Faure, J.-D., Clarke, A.E., Polya, G.M., and Anderson, M.A. 1996. Phosphorylation of style S-RNases by Ca2+-dependent protein kinases from pollen tubes. Sex. Plant Reprod. 9:25-34. Kuran, H. 1972. Callose localization in the walls of megasporocytes and megaspores in the course of development of monospore embryo sacs. Acta Soc. Bot. Polon. 41:519-534. Kurata, N., and Ito, M. 1978. Electron microscope autoradiography of 3H-thymidine incorporation during the zygotene stage in microsporocytes of lily. Cell Struct. Funct. 3:349-356. Kuroiwa, H. 1989. Ultrastructural examination of embryogenesis in Crepis capillaris (L.) Wallr.: 1. The synergid before and after pollination. Bot. Mag. Tokyo 102:9-24. Kuroiwa, H., and Kuroiwa, T. 1992. Giant mitochondria in the mature egg cell of Pelargonium zonale. Protoplasma 168:184-188. Kush, A., Brunelle, A., Shevell, D., and Chua, N.-H. 1993. The cDNA sequence of two MADS box proteins in Petunia. Plant Physiol. 102:1051-1052. Kvaale, A., and Olsen, O.A. 1986. Rates of cell division in
596 developing barley endosperms. Ann. Bot. 57:829-833. Kyle, D.J., and Styles, E.D. 1977. Development of aleurone and sub-aleurone layers in maize. Planta 137:185-193. Kyo, M., and Harada, H. 1985. Studies on conditions for cell division and embryogenesis in isolated pollen culture of Nicotiana rustica. Plant Physiol. 79:90-94. Kyo, M., and Harada, H. 1986. Control of the developmental pathway of tobacco pollen in vitro. Planta 168:427-432. Kyo, M., and Harada, H. 1990. Specific phosphoproteins in the initial period of tobacco pollen embryogenesis. Planta 182:58-63. Kyo, M., and Ohkawa, T. 1991. Investigation of subcellular localization of several phosphoproteins in embryogenic pollen grains of tobacco. /. Plant Physiol. 137:525-529. Kyozuka, J., Olive, M., Peacock, W.J., Dennis, E.S., and Shimamoto, K. 1994. Promoter elements required for developmental expression of the maize Adhl gene in transgenic rice. Plant Cell 6:799-810. Kysely, W., and Jacobsen, H.-J. 1990. Somatic embryogenesis from pea embryos and shoot apices. Plant Cell Tissue Organ Cult. 20:7-14.
References
and placenta on development of Capsella embryos in ovules cultivated in vitro. J. Plant Phx/siol. 118:127-137. Lai, F.-M., Senaratna, T., and McKersie, B.D. 1992. Glutamine enhances storage protein synthesis in Medicago sativa L. somatic embryos. Plant Sci. 87:69-77. Laibach, F. 1925. Das Taubwerden von Bastardsamen und kiinstliche Aufzucht friih absterbender Bastardembryonen. Z. Bot. 17:417-459. Lalonde, B.A., Nasrallah, M.E., Dwyer, K.G., Chen, C.-H., Barlow, B., and Nasrallah, J.B. 1989. A highly conserved Brassica gene with homology to the S-locusspecific glycoprotein structural gene. Plant Cell 1:249-258. Lam, E., and Chua, N.-H. 1991. Tetramer of a 21-base pair synthetic element confers seed expression and transcriptional enhancement in response to water stress and abscisic acid. /. Biol. Chem. 266:17131-17135. Lamb, C.J. 1994. Plant disease resistance genes in signal perception and transduction. Cell 76:419-422. Lambert, A.-M., and Bajer, A.S. 1972. Dynamics of spindle fibers and microtubules during anaphase and phragmoplast formation. Chromosoma 39:101-144. Lancelle, S.A., Cresti, M., and Hepler, P.K. 1987. Ultrastructure of the cytoskeleton in freeze-substituted pollen tubes of Nicotiana alata. Protoplasma 140:141-150. La Cour, L.F. 1949. Nuclear differentiation in the pollen grain. Heredity 3:319-337. Lancelle, S.A., and Hepler, P.K. 1991. Association of actin with cortical microtubules revealed by immunogold Labarca, C, Kroh, M., and Loewus, F. 1970. The composilocalization in Nicotiana pollen tubes. Protoplasma tion of stigmatic exudate from Lilium longiflorum. 165:167-172. Labeling studies with mt/o-inositol, D-glucose, and L-proline. Plant Physiol. 46:150-156. Lancelle, S.A., and Hepler, P.K. 1992. Ultrastructure of Labarca, C, and Loewus, F. 1972. The nutritional role of freeze-substituted pollen tubes of Lilium longiflorum. pistil exudate in pollen tube wall formation in Lilium Protoplasma 167:215-230. longiflorum. I. Utilization of injected stigmatic exu- Lang, J.D., Ray, S., and Ray, A. 1994. sinl, a mutation date. Plant Physiol. 50:7-14. affecting female fertility in Arabidopsis, interacts with modi, its recessive modifier. Genetics Labarca, C, and Loewus, F. 1973. The nutritional role of 137:1101-1110. pistil exudate in pollen tube wall formation in Lilium longiflorum. II. Production and utilization of Langdale, J.A., Irish, E.E., and Nelson, T.M. 1994. Action of the tunicate locus on maize floral development. exudate from stigma and stylar canal. Plant Physiol. 52:87-92. Devel Genet. 15:176-187. Lacadena, J.-R. 1974. Spontaneous and induced partheno- Langridge, P., and Feix, G. 1983. A zein gene of maize is transcribed from two widely separated promoter genesis and androgenesis. In Haploids in Higher regions. Cell 34:1015-1022. Plants. Advances and Potential, ed. K.J. Kasha, pp. 13-32. Guelph: University of Guelph. Langridge, P., Pintor-Toro, J.A., and Feix, G. 1982a. Transcriptional effects of the opaque-2 mutation of Lackie, S., and Yeung, E.C. 1996. Zygotic embryo develZea mays L. Planta 156:166-170. opment in Daucus carota. Can. J. Bot. 74:990-998. Ladin, B.F., Doyle, J.J., and Beachy, R.N. 1984. Molecular Langridge, P., Pintor-Toro, J.A., and Feix, G. 1982b. Zein precursor mRNAs from maize endosperm. Mol. Gen. characterization of a deletion mutation affecting the Genet. 187:432-438. a'-subunit of f5-conglycinin of soybean. /. Mol. Appl. Genet. 2:372-380. Langridge, P., Brettschneider, R., Lazzeri, P., and Lorz, H. 1992. Transformation of cereals via Agrobacterium Ladin, B.F., Tierney, M.L., Meinke, D.W., Hosangadi, P., and the pollen pathway: a critical assessment. Plant Veith, M., and Beachy, R.N. 1987. Developmental ]. 2:631-638. regulation of P-conglycinin in soybean axes and Langridge, P., Brown, J.W.S., Pintor-Toro, J.A., Feix, G., cotyledons. Plant Physiol. 84:35-41. Neuhaus, G., Neuhaus-Url, G., and Schweiger, H.-G. Lafleur, G.J., and Mascarenhas, J.P. 1978. The dependence 1985. Expression of zein genes in Acetabularia of generative cell division in Tradescantia pollen tubes mediterranea. Eur. J. Cell Biol. 39:257-264. on protein and RNA synthesis. Plant Sci. Lett. Langridge, P., Eibel, H., Brown, J.W.S., and Feix, G. 1984. 12:251-255. Transcription from maize storage protein gene proLaFountain, J.R., and LaFountain, K.L. 1973. Comparison moters in yeast. EMBO J. 3:2467-2471. of density of nuclear pores on vegetative and generative nuclei in pollen of Tradescantia. Expt. Cell Res.Larkins, B.A. 1981. Seed storage proteins: characterization 78:472-476. and biosynthesis. In The Biochemistry of Plants. A Comprehensive Treatise, Vol. 6, ed. A. Marcus, pp. LaFountain, K.L., and Mascarenhas, J.P. 1972. Isolation of 449-489. New York: Academic Press. vegetative and generative nuclei from pollen tubes. Expt. Cell Res. 73:233-236. Larkins, B.A., Bracker, C.E., and Tsai, C.Y. 1976. Storage protein synthesis in maize. Isolation of zein-syntheLagriffol, J., and Monnier, M. 1985. Effects of endosperm
References
597
regeneration of transgenic plants expressing the coat sizing polyribosomes. Plant Physiol. 57:740-745. protein of grapevine chrome mosaic nepovirus Larkins, B.A., and Dalby, A. 1975. In vitro synthesis of (GCMV). Plant Sci. 102:161-170. zein-like protein by maize polyribosomes. Biochem. Leah, R., Skriver, K., Knudsen, S., Ruud-Hansen, }., Biophys. Res. Commun. 66:1048-1054. Raikhel, N.V., and Mundy, J. 1994. Identification of Larkins, B.A., and Hurkman, WJ. 1978. Synthesis and an enhancer/silencer sequence directing the aleudeposition of zein in protein bodies of maize rone-specific expression of a barley chitinase gene. endosperm. Plant Physiol. 62:256-263. Plant J. 6:579-589. Larkins, B.A., Jones, R.A., and Tsai, C.Y. 1976. Isolation and in vitro translation of zein messenger ribonucleic Leal, I., and Misra, S. 1993a. Developmental gene expresacid. Biochemistry 15:5506-5511. sion in conifer embryogenesis and germination. III. Analysis of crystalloid protein mRNAs and desiccaLarkins, B.A., Pedersen, K., Handa, A.K., Hurkman, W.J., tion protein mRNAs in the developing embryo and and Smith, L.D. 1979. Synthesis and processing of megagametophyte of white spruce (Picea glauca maize storage proteins in Xenopus laevis oocytes. (Moench) Voss). Plant Sci. 88:25-37. Proc. Natl Acad. Sci. USA 76:6448-6452. Laroche-Raynal, M., and Delseny, M. 1986. Identification Leal, I., and Misra, S. 1993b. Molecular cloning and characterization of a legumin-like storage protein cDNA and characterization of the mRNA for major storof Douglas fir seeds. Plant Mol. Biol. 21:709-715. age proteins from radish. Eur. J. Biochem. Leal, I., Misra, S., Attree, S.M., and Fowke, L.C. 1995. 157:321-327. Effect of abscisic acid, osmoticum and desiccation on Laroche-Raynal, M., Aspart, L., Delseny, M., and Penon, P. US storage protein gene expression in somatic 1984. Characterization of radish mRNA at three embryos of white spruce. Plant Sci. 106:121-128. developmental stages. Plant Sci. Lett. 35:139-146. Larsen, K. 1977. Self-incompatibility in Beta vulgaris L. I. Leduc, N., Matthys-Rochon, E., and Dumas, C. 1995. Four gametophytic, complementary S-loci in sugar Deleterious effect of minimal enzymatic treatments beet. Hereditas 85:227-248. on the development of isolated maize embryo sacs in culture. Sex. Plant Reprod. 8:313-317. Larsen, P.B., Ashworth, E.N., Jones, M.L., and Woodson, W.R. 1995. Pollination-induced ethylene in carna- Leduc, N., Matthys-Rochon, E., Rougier, M., Mogensen, L., Holm, P., Magnard, J.-L., and Dumas, C. 1996. tion. Role of pollen tube growth and sexual compatIsolated maize zygotes mimic in vivo embryonic ibility. Plant Physiol. 108:1405-1412. development and express microinjected genes when Larson, D.A. 1963. Cytoplasmic dimorphism within cultured in vitro. Devel Biol. 177:190-203. pollen grains. Nature 200:911-912. Larson, D.A. 1965. Fine-structural changes in the cyto- Lee, C.H., and Power, J.B. 1988. Intraspecific gametosomatic hybridisation in Petunia hybrida. Plant Cell Rep. plasm of germinating pollen. Am. J. Bot. 52:139-154. 7:17-18. Larson, D.A., and Lewis, C.W., Jr. 1963. Pollen wall development in Parkinsonia aculeata. Grana Palynol. 3 Lee, D.W., and Dougall, D.K. 1973. Electrophoretic varia() tion in glutamate dehydrogenase and other Larson, D.A., Skvarla, J.J., and Lewis, C.W., Jr. 1962. An isozymes in wild carrot cells cultured in the presence electron microscope study of exine stratification and and absence of 2,4-dichlorophenoxyacetic acid. In fine structure. Pollen Spores 4:232-246. Vitro 8:347-352. Laser, K.D., and Lersten, N.R. 1972. Anatomy and cytol- Lee, H.-S., Huang, S., and Kao, T.-H. 1994. S proteins conogy of microsporogenesis of cytoplasmic male sterile trol rejection of incompatible pollen in Petunia angiosperms. Bot. Rev. 38:425-454. inflata. Nature 367:560-563. Laurain, D., TWmouillaux-Guiller, J., and Ch^nieux, J.-C. Lee, H.-S., Singh, A., and Kao, T.-H. 1992. RNase X2, a pis1993. Embryogenesis from microspores of Cinkgo til-specific ribonuclease from Petunia inflata, shares sequence similarity with solanaceous S proteins. biloba L., a medicinal woody species. Plant Cell Rep. 12:501-505. Plant Mol. Biol. 20:1131-1141. Laux, T., Mayer, K.F.X., Berger, J., and Jurgens, G. 1996. The Lee, H.S., Kim, S.W., Lee, K.-W., Eriksson, T., and Liu, J.R. WUSCHEL gene is required for shoot and floral meris1995. Agrobacterium-mediated transformation of gintem integrity in Arabidopsis. Development 122:87-96. seng (Panax ginseng) and mitotic stability of the inserted (3-glucuronidase gene in regenerants from Laver, H.K., Reynolds, S.J., Mon£ger, F., and Leaver, C.J. isolated protoplasts. Plant Cell Rep. 14:545-549. 1991. Mitochondrial genome organization and expression associated with cytoplasmic male steril- Lee, K., and Huang, A.H.C. 1991. Genomic nucleotide ity in sunflower (Helianthus annuus). Plant J. sequence of a Brassica napus 20-kilodalton oleosin 1:185-193. gene. Plant Physiol. 96:1395-1397. Lawrence, M.J. 1975. The genetics of self-incompatibility Lee, K., and Huang, A.H.C. 1994. Genes encoding in Papaver rhoeas. Proc. Roy. Soc. Lond. 188B:275-285. oleosins in maize kernel of inbreds Mol7 and B73. Lawrence, M.J., Marshall, D.F., Curtis, V.E., and Fearon, Plant Mol. Biol. 26:1981-1987. C.H. 1985. Gametophytic self-incompatibility re- Lee, K.H., Jones, R.A., Dalby, A., and Tsai, C.Y. 1976. examined: a reply. Heredity 54:131-138. Genetic regulation of storage protein content in Le Coq, C. 1972a. La megasporog£nese chez Vlris pseudamaize endosperm. Biochem. Genet. 14:641-650. corus L. I. Etude cytologique qualitative. Rev. Cytol. Lee, S.-H., Karunanandaa, B., McCubbin, A., Gilroy, S., Biol. Viget. 35:41-163. and Kao, T.-H. 1996. PRK1, a receptor-like kinase of Le Coq, C. 1972b. La megasporog^nese chez Vlris pseudaPetunia inflata, is essential for postmeiotic developcorus L. II. Etude cytologique quantitative. Rev. Cytol. ment of pollen. Plant J. 9:613-624. Biol. Veg&t. 35:303-329. Lee, S.-L.J., Earle, E.D., and Gracen, V.E. 1980. The cytolLe Gall, O., Torregrosa, L., Danglot, Y., Candresse, T, and ogy of pollen abortion in S cytoplasmic male-sterile Bouquet, A. 1994. /Igrobflcteriwm-mediated genetic corn anthers. Am. J. Bot. 67:237-245. transformation of grapevine somatic embryos and Lee, S.-L.J., Gracen, V.E., and Earle, E.D. 1979. The cytol-
598
References
ogy of pollen abortion in C-cytoplasmic male-sterile Levings, C.S., III. 1993. Thoughts on cytoplasmic male corn anthers. Am. J. Bot. 66:656-667. sterility in cms-T maize. Plant Cell 5:1285-1290. Lee, S.-L.J., and Warmke, H.E. 1979. Organelle size and Levings, C.S., III., Kim, B.D., Pring, D.R., Conde, M.R, number in fertile and T-cytoplasmic male-sterile Mans, R.J., Laughnan, J.R., and Gabay-Laughnan, corn. Am. J. Bot. 66:141-148. S.J. 1980. Cytoplasmic reversion of cms-S in maize: association with a transpositional event. Science Lee, W.S., Tzen, J.T.C., Kridl, J.C., Radke, S.E., and Huang, 209:1021-1023. A.H.C. 1991. Maize oleosin is correctly targeted to seed oil bodies in Brassica napus transformed with Levings, C.S., III., and Pring, D.R. 1976. Restriction the maize oleosin gene. Proc. Natl Acad. Sci. USA endonuclease analysis of mitochondrial DNA from 88:6181-6185. normal and Texas cytoplasmic male-sterile maize. Science 193:158-160. Leisy, D.J., Hnilo, J., Zhao, Y, and Okita, T.W. 1989. Expression of a rice glutelin promoter in transgenic Lewandowska, E., and Charzyriska, M. 1977. Tradescantia tobacco. Plant Mol. Biol. 14:41-50. bracteata pollen in vitro; pollen tube development Leite, A., Yunes, J.A., Turcinelli, S.R., and Arruda, P. 1992. and mitosis. Ada Soc. Bot. Polon. 46:587-598. Cloning and characterization of a cDNA encoding a Lewis, D. 1942. The physiology of incompatibility in sulfur-rich coixin. Plant Mol. Biol. 18:171-174. plants. I. The effect of temperature. Proc. Roy. Soc. Lond. 131B:13-26. Lelievre, J.-M., Oliveira, L.O., and Nielsen, N.C. 1992. 5'CATGCAT-3' elements modulate the expression of Lewis, D. 1943. The physiology of incompatibility in glycinin genes. Plant Physiol. 98:387-391. plants. II. Linum grandiflorum. Ann. Bot. 7:115-122. Lending, C.R., and Larkins, B.A. 1989. Changes in the Lewis, D. 1949. Structure of the incompatibility gene. II. Induced mutation rate. Heredity 3:339-355. zein composition of protein bodies during maize Lewis, D. 1952. Serological reactions of pollen incompatiendosperm development. Plant Cell 1:1011-1023. Lending, C.R., and Larkins, B.A. 1992. Effect of the bility substances. Proc. Roy. Soc. Lond. 140B:127-135. floury-2 locus on protein body formation during Lewis, D. 1954. Comparative incompatibility in maize endosperm development. Protoplasma angiosperms and fungi. Adv. Genet. 6:235-285. 171:123-133. Lewis, D. 1960. Genetic control of specificity and activity of the S antigen in plants. Proc. Roy. Soc. Lond. Lending, C.R., Chesnut, R.S., Shaw, K.L., and Larkins, 151B:468-477. B.A. 1989. Immunolocalization of avenin and globulin storage proteins in developing endosperm of Lewis, D. 1965. A protein dimer hypothesis on incompatAvena sativa L. Planta 178:315-324. ibility. In Genetics Today, Vol. 3, ed. S.J. Geerts, pp. 657-663. Oxford: Pergamon Press. Lending, C.R., Kriz, A.L., Larkins, B.A., and Bracker, C.E. 1988. Structure of maize protein bodies and Lewis, D., Burrage, S., and Walls, D. 1967. Immunological immunocytochemical localization of zeins. reactions of single pollen grains, electrophoresis and Protoplasma 143:51-62. enzymology of pollen protein exudates. /. Expt. Bot. Leon-Kloosterziel, K.M., Keijzer, C.J., and Koornneef, M. 18:371-378. 1994. A seed shape mutant of Arabidopsis that is Lewis, D., and Crowe, L.K. 1958. Unilateral interspecific affected in integument development. Plant Cell incompatibility in flowering plants. Heredity 6:385-392. 12:233-256. Lerner, D.R., and Raikhel, N.V. 1989. Cloning and charac- Lewis, D., and Jones, D.A. 1992. The genetics of hetterization of root-specific barley lectin. Plant Physiol. erostyly. In Evolution and Function of Heterostyly, ed. 91:124-129. S.C.H. Barrett, pp. 129-150. Berlin: Springer-Verlag. Lessard, P.A., Allen, R.D., Bernier, R, Crispino, J.D., Li, H.-J., and Sodmergen. 1995. Maternal cytoplasmic inheritance and pollen nucleolytic activities in some Fujiwara, T., and Beachy, R.N. 1991. Multiple nuclear factors interact with upstream sequences of differenPoaceae species. Cytologia 60:173-181. tially regulated (3-conglycinin genes. Plant Mol. Biol. Li, ]., and Liu, Y.-N. 1983. Chloroplast DNA and cytoplas16:397-413. mic male-sterility. Theor. Appl. Genet. 64:231-238. Leung, D.W.M. 1992. Involvement of plant chitinase in Li, L., Qu, R., de Kochko, A., Fauquet, C, and Beachy, sexual reproduction of higher plants. Phytochemistry R.N. 1993. An improved rice transformation system 31:1899-1900. using the biolistic method. Plant Cell Rep. 12:250-255. Leung, J., Bouvier-Durand, M., Morris, P.-C, Guerrier, D., Chefdor, R, and Giraudat, J. 1994. Arabidopsis ABA Li, M.-X. 1982. Pollen dimorphism and androgenesis of Paeonia in vivo. Acta Bot. Sinica 24:17-20. response gene ABU: features of a calcium-modulated protein phosphatase. Science 264:1448-1452. Li, W.-D., Xu, H., Cheng, X.-R, and Ma, R-S. 1991. The Levanony, H., Rubin, R., Altschuler, Y, and Galili, G. 1992. behaviour of pollen tubes on the stigma in the interEvidence for a novel route of wheat storage proteins sectional crosses in Populus and its relation to seedto vacuoles. /. Cell Biol. 119:1117-1128. setting. Chinese J. Bot. 3:102-109. Levieil, C. 1986. Evolution de l'association des cellules Li, Y.-Q., Croes, A.R, and Linskens, H.R 1983. Cell-wall proteins in pollen and roots of Lilium longiflorum: mSles dans le tube pollinique et dans le sac embryonnaire chez Cichorium intybus L. Compt. Rend. Acad. extraction and partial characterization. Planta Sci. Paris Ser. Ill 303:769-774. 158:422-427. Levin, J.Z., and Meyerowitz, E.M. 1995. UFO: an Li, Y.-Q., and Linskens, H.F. 1983a. Wall-bound proteins of pollen tubes after self- and cross-pollination in Arabidopsis gene involved in both floral meristem Lilium longiflorum. Theor. Appl. Genet. 67:11-16. and floral organ development. Plant Cell 7:529-548. Levings, C.S., III. 1983. Cytoplasmic male sterility. In Li, Y.-Q., and Linskens, H.F. 1983b. Neutral sugar composition of pollen tube walls of Lilium longiflorum. Acta Genetic Engineering of Plants. An Agricultural Bot. Need. 32:437-445. Perspective, ed. T. Kosuge, C.P. Meredith and A. Hollaender, pp. 81-92. New York: Plenum Press. Li, Y.-Q., and Tsao, T.H. 1985. Covalently bound wall pro-
References
599
tion on germination. Plant Physiol. 74:746-748. teins of pollen grains and pollen tubes grown in vitro and in styles after self- and cross-pollination in Lin, J.-J., Dickinson, D.B., and Ho, T.-H.D. 1987. Phytic Lilium longiflorum. Theor. Appl. Genet. 71:263-267. acid metabolism in lily {Lilium longiflorum Thunb.) pollen. Plant Physiol. 83:408-413. Li, Y.-Q., Tsao, T.H., and Linskens, H.F. 1986. Dependence of Lilium pollen germination and tube growth on Lin, Y., Wang, Y, Zhu, J.-K., and Yang, Z. 1996. Localization of a Rho GTPase implies a role in tip protein synthesis and glycosylation after inhibitor growth and movement of the generative cell in treatments. Proc. Koninkl. Nederl. Akad. Wetensch. pollen tubes. Plant Cell 8:293-303. 89G61-73. Li, Y.-Q., Bruun, L., Pierson, E.S., and Cresti, M. 1992. Lin, Y.-T., Chow, T.-Y, and Lin, C.-Y. 1971. Nitrogen Periodic deposition of arabinogalactan epitopes in metabolism associated with pollen grain germinathe cell wall of pollen tubes of Nicotiana tabacum. tion. Taiwania 16:67-84. Planta 188:532-538. Linck, A.J., and Blaydes, G.W. 1960. Demonstration of the Li, Y.-Q., Chen, R, Linskens, H.F., and Cresti, M. 1994a. chemotropism of pollen tubes in vitro in four plant Distribution of unesterified and esterified pectins in species. Ohio J. Sci. 60:274-278. cell walls of pollen tubes of flowering plants. Sex. Lind, J.L., Bacic, A., Clarke, A.E., and Anderson, M.A. Plant Reprod. 7:145-152. 1994. A style-specific hydroxyproline-rich glycoproLi, Y.-Q., Faleri, C, Geitmann, A., Zhang, H.-Q., and tein with properties of both extensins and arabinoCresti, M. 1995a. Immunogold localization of aragalactan proteins. Plant J. 6:491-502. binogalactan proteins, unesterified and esterified Lind, J.L., Bonig, I., Clarke, A.E., and Anderson, M.A. pectins in pollen grains and pollen tubes of Nicotiana 1996. A style-specific 120-kDa glycoprotein enters tabacum L. Protoplasma 189:26-36. pollen tubes of Nicotiana alata in vivo. Sex. Plant Li, Y.Q., Faleri, C, Thompson, R.D., Tiezzi, A., Eijlander, Reprod. 9:75-86. R., and Cresti, M. 1994b. Cytochemical immunolo- Lindsey, K., and Topping, J.F. 1993. Embryogenesis: a calization of the abundant pistil protein S^ in question of pattern. /. Expt. Bot. 44:359-374. potato (Solanum tuberosum). Sex. Plant Reprod. Lindstrom, J.T., Vodkin, L.O., Harding, R.W., and Goeken, 7:164-168. R.M. 1990. Expression of soybean lectin gene deletions in tobacco. Devel Genet. 11:160-167. Li, Y.Q., Southworth, D., Linskens, H.F., Mulcahy, D.L., and Cresti, M. 1995b. Localization of ubiquitin in Lindzen, E., and Choi, J.H. 1995. A carrot cDNA encoding anthers and pistils of Nicotiana. Sex. Plant Reprod. an atypical protein kinase homologous to plant cal8:123-128. cium-dependent protein kinases. Plant Mol. Biol. 28:785-797. Li, X., Franceschi, V.R., and Okita, T.W. 1993. Segregation of storage protein mRNAs on the rough endoplas- Linnestad, C, Lonneborg, A., Kalla, R., and Olsen, O.-A. mic reticulum membranes of rice endosperm cells. 1991. Promoter of a lipid-transfer protein gene Cell 72:869-879. expressed in barley aleurone cells contains similar Li, X., Nield, J., Hayman, D., and Langridge, P. 1994. myb and myc recognition sites as the maize Bz-McC Cloning a putative self-incompatibility gene from allele. Plant Physiol. 97:841-843. the pollen of the grass Phalaris coerulescens. Plant CellLinskens, H. 1955. Physiologische Untersuchungen der 6:1923-1932. Pollenschlauch-Hemmung selbststeriler Petunien. Li, X., Wu, Y, Zhang, D.-Z., Gillikin, J.W., Boston, R.S., Z. Bot. 43:1-44. Franceschi, V.R., and Okita, T.W. 1993. Rice pro- Linskens, H.F. 1958. Physiologische Untersuchungen zur lamine protein body biogenesis: a BiP-mediated Reifeteilung. II. Mitteilung. Uber die Anderung des process. Science 262:1054-1056. Nukleinsaurengehaltes wahrend der Pollenmeiose und Pollenentwicklung von Lilium henryi. Ada Bot. Li, X., and Okita, T.W. 1993. Accumulation of prolamines Neerl. 7:61-68. and glutelins during rice seed development: a quantitative evaluation. Plant Cell Physiol. Linskens, H.F. 1959. Zur Frage der Entstehung der 34:385-390. Abwehrkorper bei der Inkompatibilitatsreaktion von Petunia. Ber. Deut. Bot. Ges. 72:84-92. Liang, X., Dron, M., Schmid, J., Dixon, R.A., and Lamb, CJ. 1989. Developmental and environmental regula- Linskens, H.F. 1960. Zur Frage der Entstehung der tion of a phenylalanine ammonia-lyase-pi-gluAbwehr-Korper bei der Inkompatibilitatsreaktion curonidase gene fusion in transgenic tobacco plants. von Petunia. III. Mitteilung: serologische Teste mit Proc. Natl Acad. Sci. USA 86:9284-9288. Leitgewebs- und Pollen-Extrakten. Z. Bot. Lima-de-Faria, A. 1965. Labeling of the cytoplasm and the 48:126-135. 3 meiotic chromosomes of Agapanthus with H -thymi- Linskens, H.F. 1964. Pollen physiology. Annu. Rev. Plant dine. Hereditas 53:1-11. Physiol. 15:255-270. Lima-de-Faria, A., Pero, R., Avanzi, S., Durante, M., Linskens, H.F. 1966. Die Anderung des Protein- und Stahle, U., D'Amato, F., and Granstrom, H. 1975. Enzym-Musters wahrend der Pollenmeiose und Relation between ribosomal RNA genes and the Pollenentwicklung. Physiologische Untersuchungen DNA satellites of Phaseolus coccineus. Hereditas zur Reifeteilung. Planta 69:79-91. 79:5-19. Linskens, H.F. 1967. Isolation of ribosomes from pollen. Lin, B.-Y. 1984. Ploidy barrier to endosperm development Planta 73:194-200. in maize. Genetics 107:103-115. Linskens, H.F. 1969. Fertilization mechanisms in higher Lin, J., Uwate, W.J., and Stallman, V. 1977. Ultrastructural plants. In Fertilization, Vol. II, ed. C.B. Metz and A. localization of acid phosphatase in the pollen tube Monroy, pp. 189-253. New York: Academic Press. of Prunus avium L. (sweet cherry). Planta Linskens, H.F. 1974. Study of growth of Petunia styles. 135:183-190. Soviet Plant Physiol. 21:878-882. Lin, J.-J., and Dickinson, D.B. 1984. Ability of pollen to Linskens, H.F. 1975. Incompatibility in Petunia. Proc. Roy. germinate prior to anthesis and effect of desiccaSoc. Land. 188B:299-311.
600
References
Linskens, H.F., and Heinen, W. 1962. Cutinase-Nachweis Liu, C.-M., Xu, Z.-H., and Chua, N.-H. 1993b. Auxin polar in Pollen. Z. Bot. 50:338-347. transport is essential for the establishment of bilateral symmetry during early plant embryogenesis. Linskens, H.F., Kochuyt, A.S.L., and So, A. 1968. Plant Cell 5:621-630. Regulation der Nucleinsauren-Synthese durch Polyamine in keimendem Pollen von Petunia. Planta Liu, C.-N., and Rubenstein, I. 1993. Transcriptional char82:111-122. acterization of an -zein gene cluster in maize. Plant Mol. Bid. 22:323-336. Linskens, H.F., and Schrauwen, J. 1968a. Anderung des Ribosomen-Musters wahrend der Meiose. Natur- Liu, G.-Q., Cai, G., Del Casino, C, Tiezzi, A., and Cresti, wissenschaften 55:91. M. 1994. Kinesin-related polypeptide is associated with vesicles from Corylus avellana pollen. Cell Mot. Linskens, H.F., and Schrauwen, J. 1968b. Quantitative Cytoskel. 29:155-166. nucleic acid determinations in the microspore and tapetum fractions of lily anthers. Proc. Koninkl. Liu, Q., Golubovskaya, I., and Cande, W.Z. 1993. Nederl. Akad. Wetensch. 71G267-279. Abnormal cytoskeletal and chromosome distribution in po, ms4 and ms6; mutant alleles of polymitotic Linskens, H.F., and Schrauwen, J. 1969. The release of free that disrupt the cell cycle progression from meiosis amino acids from germinating pollen. Ada Bot. to mitosis in maize. /. Cell Sci. 106:1169-1178. Need. 18:605-614. Linskens, H.F., Schrauwen, J.A.M., and Konings, R.N.H. Liu, R., Olsen, O.-A., Kreis, M., and Halford, N.G. 1992. 1970. Cell-free protein synthesis with polysomes Molecular cloning of a novel barley seed protein from germinating Petunia pollen grains. Planta gene that is repressed by abscisic acid. Plant Mol. Biol. 18:1195-1198. 90:153-162. Linskens, H.F., and Tupy, J. 1966. The amino acids pool in Liu, W., Hildebrand, D.F., Grayburn, W.S., Phillips, G.C., and Collins, G.B. 1991. Effects of exogenous auxins the style of self-incompatible strains of Petunia after on expression of Iipoxygenases in cultured soybean self- and cross-pollination. Ziichter 36:151-158. embryos. Plant Physiol. 97:969-976. Linskens, H.F., van der Donk, J.A.W.M., and Schrauwen, J. 1971. RNA synthesis during pollen germination. Liu, X., and Meyerowitz, E.M. 1995. LEUNIG regulates AGAMOUS expression in Arabidopsis flowers. Planta 97:290-298. Development 121:975-991. Linskens, H.F., Havez, R., Linder, R., Salden, M., Randoux, A., Laniez, D., and Coustaut, D. 1969. Etude des gly- Liu, X.C., and Dickinson, H.G. 1989. Cellular energy levcanne-hydrolases au cours de la croissance du pollen els and their effect on male cell abortion in cytoplaschez Petunia hybrida auto-incompatible. Compt. Rend. mically male sterile lines of Petunia hybrida. Sex. Acad. Sci. Paris 269D: 1855-1857. Plant Reprod. 2:167-172. Lintilhac, P.M. 1974. Differentiation, organogenesis, and Liu, X.C., Jones, K., and Dickinson, H.G. 1987. DNA synthesis and cytoplasmic differentiation in tapetal cells the tectonics of cell wall orientation. II. Separation of of normal and cytoplasmically male sterile lines of stresses in a two-dimensional model. Am. ]. Bot. 61:135-140. Petunia hybrida. Theor. Appl. Genet. 74:846-851. Lippi, M.M., Cimoli, F., Maugini, E., and Tani, G. 1994. A Liu, X.C, Jones, K., and Dickinson, H.G. 1988. comparative study of the tapetal behaviour in male Cytoplasmic male sterility in Petunia hybrida: factors affecting mitochondrial ATP export in normal and fertile and male sterile Iris pallida Lam. during cytoplasmically male sterile plants. Theor. Appl. microsporogenesis. Caryologia 47:109-120. Genet. 76:305-310. Lippmann, B., and Lippmann, G. 1984. Induction of somatic embryos in cotyledonary tissue of soybean, Liu, X.-Y., Rocha-Sosa, M., Hummel, S., Willmitzer, L., Glycine max L. Merr. Plant Cell Rep. 3:215-218. and Frommer, W. B. 1991. A detailed study of the Litts, J.C., Colwell, G.W., Chakerian, R.L., and Quatrano, regulation and evolution of the two classes of R.S. 1987. The nucleotide sequence of a cDNA clone patatin genes in Solanum tuberosum L. Plant Mol. Biol. 17:1139-1154. encoding the wheat E protein. Nucl. Adds Res. 15:3607-3618. Lloyd, J.R., Wang, T.L., and Hedley, C.L. 1996. Analysis of seed development in Pisum sativum. XIX. Effect of Litts, J.C, Colwell, G.W., Chakerian, R.L., and Quatrano, mutant alleles at the r and rb loci on starch grain size R.S. 1991. Sequence analysis of a functional member and on the content and composition of starch in of the Em gene family from wheat. DNA Sequence developing pea seeds. /. Expt. Bot. 47:171-180. 1:263-274. Liu, B., and Palevitz, B.A. 1991. Kinetochore fiber forma- Lo Schiavo, F., Giuliano, G., and Sung, Z.R. 1988. Characterization of a temperature-sensitive carrot tion in dividing generative cells of Tradescantia. cell mutant impaired in somatic embryogenesis. Kinetochore reorientation associated with the transiPlant Sd. 54:157-164. tion between lateral microtubule interactions and end-on kinetochore fibers. /. Cell Sd. 98:475-482. Lo Schiavo, E, Quesada-Allue, L.A., and Sung, Z.R. 1986. Liu, B., and Palevitz, B.A. 1992. Anaphase chromosome sepTunicamycin affects somatic embryogenesis but not cell proliferation of carrot. Plant Sci. 44:65-71. aration in dividing generative cells of Tradescantia. Changes in microtubule organization and kinetochore Lo Schiavo, F., Filippini, F., Cozzani, F., Vallone, D., and distribution visualized by antitubulin and CREST Terzi, M. 1991. Modulation of auxin-binding proimmunocytochemistry. Protoplasma 166:122-133. teins in cell suspensions. I. Differential responses of carrot embryo cultures. Plant Physiol. 97:60-64. Liu, C.-M., Johnson, S., and Wang, T.L. 1995. cyd, a mutant of pea that alters embryo morphology is defective in Lo Schiavo, E, Giuliano, G., de Vries, S.C., Genga, A., Bollini, R., Pitto, L., Cozzani, E, Nuti-Ronchi, V., and cytokinesis. Devel Genet. 16:321-331. Terzi, M. 1990. A carrot cell variant temperature senLiu, C.-M., Xu, Z.-H., and Chua, N.-H. 1993a. Proembryo sitive for somatic embryogenesis reveals a defect in culture: in vitro development of early globular-stage the glycosylation of extracellular proteins. Mol. Gen. zygotic embryos from Brassica juncea. Plant ]. Genet. 223:385-393. 3:291-300.
References
Lo Schiavo, R, Pitto, L., Giuliano, G., Torti, G., NutiRonchi, V., Marazziti, D., Vergara, Rv Orselli, S., and Terzi, M. 1989. DNA methylation of embryogenic carrot cell cultures and its variations as caused by mutation, differentiation, hormones and hypomethylating drugs. Theor. Appl. Genet. 77:325-331. Loader, N.M., Woolner, E.M., Hellyer, A., Slabas, A.R., and Safford, R. 1993. Isolation and characterization of two Brassica napus embryo acyl-ACP thioesterase cDNA clones. Plant Mol. Biol. 23:769-778. Loeb, T.A., and Reynolds, T.L. 1994. Transient expression of the uidA gene in pollen embryoids of wheat following microprojectile bombardment. Plant Sci. 104:81-91. Loer, D.S., and Herman, E.M. 1993. Cotranslational integration of soybean {Glycine max) oil body membrane protein oleosin into microsomal membranes. Plant Physiol. 101:993-998. Lohmer, S., Maddaloni, M., Motto, M., di Fonzo, N., Hartings, H., Salamini, R, and Thompson, R.D. 1991. The maize regulatory locus opaque-2 encodes a DNA-binding protein which activates the transcription of the b-32 gene. EMBO ]. 10:617-624. Lolle, S.J., and Cheung, A.Y. 1993. Promiscuous germination and growth of wildtype pollen from Arabiopsis and related species on the shoot of the Arabidopsis mutant,fiddlehead.Devel Biol. 155:250-258. Lombardo, G., and Carraro, L. 1976a. Tapetal ultrastructural changes during pollen development. I. Studies on Antirrhinum majus. Caryologia 29:113-125. Lombardo, G., and Carraro, L. 1976b. Tapetal ultrastructural changes during pollen development. III. Studies on Gentiana acaulis. Caryologia 29:345-349. Lombardo, G., and Gerola, RM. 1968a. Cytoplasmic inheritance and ultrastructure of the male generative cell of higher plants. Planta 82:105-110. Lombardo, G., and Gerola, RM. 1968b. Ultrastructure of the pollen grain and taxonomy. Giorn. Bot. Hal. 102:353-380. Long, J.A., Moan, E.I., Medford, J.I., and Barton, M.K. 1996. A member of the KNOTTED class of homeodomain proteins encoded by the STM gene of Arabidopsis. Nature 379:66-69. Long, S.R., Dale, R.M.K., and Sussex, I.M. 1981. Maturation and germination of Phaseolus vulgaris embryonic axes in culture. Planta 153:405-415. Lonsdale, D., Onde, S., and Cuming, A. 1990. Transient expression of exogenous DNA in intact, viable wheat embryos following particle bombardment. /. Expt. Bot. 41:1161-1165. Lopes, M.A., Coleman, C.E., Kodrzycki, R., Lending, C.R., and Larkins, B.A. 1994. Synthesis of an unusual azein protein is correlated with the phenotypic effects of the floury! mutation in maize. Mol. Gen. Genet. 245:537-547. Lopes, M.A., and Larkins, B.A. 1991. Gamma-zein content is related to endosperm modification in quality protein maize. Crop Sci. 31:1655-1662. Lopes, M.A., and Larkins, B.A. 1993. Endosperm origin, development and function. Plant Cell 5:1383-1399. Lopez, I., Anthony, R.G., Maciver, S.K., Jiang, C.-J., Khan, S., Weeds, A.G., and Hussey, P.J. 1996. Pollen specific expression of maize genes encoding actin depolymerizing factor-like proteins. Proc. Natl Acad. Sci. USA 93:7415-7420. Lord, E.M., and Eckard, K.J. 1984. Incompatibility
601 between the dimorphic flowers of Collomia grandiflora, a cleistogamous species. Science 223:695-696. Lord, E.M., and Eckard, K.J. 1986. Ultrastructure of the dimorphic pollen and stigmas of the cleistogamous species, Collomia grandiflora (Polemoniaceae). Protoplasma 132:12-22. Lord, E.M., and Heslop-Harrison, Y. 1984. Pollen-stigma interaction in the Leguminosae: stigma organization and the breeding system in Vicia faba L. Ann. Bot. 54:827-836. Lord, E.M., and Kohorn, L.U. 1986. Gynoecial development, pollination, and the path of pollen tube growth in the tepary bean, Phaseolus acutifolius. Am. ]. Bot. 73:70-78. Lord, E.M., and Webster, B.D. 1979. The stigmatic exudate of Phaseolus vulgaris L. Bot. Gaz. 140:266-271. Lorenzi, R., Bennici, A., Cionini, P.G., Alpi, A., and D'Amato, R 1978. Embryo-suspensor relations in Phaseolus coccineus: cytokinins during seed development. Planta 143:59-62. Lotan, T., Ori, N., and Fluhr, R. 1989. Pathogenesis-related proteins are developmentally regulated in tobacco flowers. Plant Cell 1:881-887. Louis, J.-P, Augur, C, and Teller, G. 1990. Cytokinins and differentiation processes in Mercurialis annua. Genetic regulation, relations with auxins, indoleacetic acid oxidases, and sexual expression patterns. Plant Physiol. 94:1535-1541. Love, A. 1943. A Y-linked inheritance of asynapsis in Rumex acetosa. Nature 152:358-359. Lowe, J., and Nelson, O.E., Jr. 1946. Miniature seed - a study in the development of a defective caryopsis in maize. Genetics 31:525-533. Lu, Y, and Rutger, J.N. 1984. Cytological observations on induced genetic male sterile mutants in rice (Oryza sativa L.). Sci. Sinica 27B:322-331. Lu, Z.-X., Wu, M., Loh, C.-S., Yeong, C.-Y, and Goh, C.-J. 1993. Nucleotide sequence of a flower-specific MADS box cDNA clone from orchid. Plant Mol. Biol. 23:901-904. Luck, B.T., and Jordan, E.G. 1980. The mitochondria and plastids during microsporogenesis in Hyacinthoides non-scripta (L.) Chouard. Ann. Bot. 45:511-514. Ludevid, M.D., Freire, M.A., Gomez, J., Burd, C.G., Albericio, F., Giralt, E., Dreyfuss, G., and Pages, M. 1992. RNA binding characteristics of a 16 kDa glycine-rich protein from maize. Plant ]. 2:999-1003. Ludevid, M.D., Torrent, M., Martinez-Izquierdo, J.A., Puigdomenech, P., and Palau, J. 1984. Subcellular localization of glutelin-2 in maize (Zea mays L.) endosperm. Plant Mol. Biol. 3:227-234. Luegmayr, E. 1993. The generative cell and its close association with the endoplasmic reticulum of the vegetative cell in pollen of Cyrtandra pendula (Gesneriaceae). Protoplasma 177:73-81. Lund, H.A. 1956. Growth hormones in the styles and ovaries of tobacco responsible for fruit development. Am. J. Bot. 43:562-568. Lundqvist, A. 1956. Self-incompatibility in rye. I. Genetic control in the diploid. Hereditas 42:293-348. Lundqvist, A. 1975. Complex self-incompatibility systems in angiosperms. Proc. Roy. Soc. Land. 188B:235-245. Lundqvist, A. 1993. The self-incompatibility system in Lotus tennis (Fabaceae). Hereditas 119:59-66. Lukowitz, W., Mayer, U., and Jurgens, G. 1996. Cytokinesis in the Arabidopsis embryo involves the syntaxinrelated KNOLLE gene product. Cell 84:61-71.
602 Luo, Z.-X., and Wu, R. 1988. A simple method for the transformation of rice via the pollen-tube pathway. Plant Mol. Biol. Rep. 6:165-174. Lupotto, E. 1983. Propagation of an embryogenic culture of Medicago sativa L. Z. Pflanzenphysiol. 111:95-104. Lupotto, E. 1986. The use of single somatic embryo culture in propagating and regenerating lucerne (Medicago sativa L.). Ann. Bot. 57:19-24. Lur, H.-S., and Setter, T.L. 1993. Role of auxin in maize endosperm development. Timing of nuclear DNA endoreduplication, zein expression, and cytokinin. Plant Physiol. 103:273-280. Luthe, D.S. 1983. Storage protein accumulation in developing rice (Oryzfl sativa L.) seeds. Plant Sci. 32:147-158. Luthe, D.S. 1987. Storage protein synthesis during oat (Avena sativa L.) seed development. Plant Physiol. 84:337-340. Luthe, D.S., and Peterson, D.M. 1977. Cell-free synthesis of globulin by developing oat (Avena sativa L.) seeds. Plant Physiol. 59:836-841. Lutz, R.W., and Sjolund, R.D. 1973. Development of the generative cell wall in Monotropa uniflora L. pollen. Plant Physiol. 52:498-500. Lycett, G.W., Croy, R.R.D., Shirsat, A.H., and Boulter, D. 1984. The complete nucleotide sequence of a legumin gene from pea (Pisum sativum L.). Nucl. Acids Res. 12:4493^506. Lycett, G.W., Croy, R.R.D., Shirsat, A.H., Richards, D.M., and Boulter, D. 1985. The 5-flanking regions of three pea legumin genes: comparison of the DNA sequences. Nucl. Acids Res. 13:6733-6743. Lycett, G.W., Delauney, A.J., Gatehouse, J.A., Gilroy, J., Croy, R.R.D., and Boulter, D. 1983. The vicilin gene family of pea (Pisum sativum L.): a complete cDNA coding sequence for preprovicilin. Nucl. Acids Res. 11:2367-2380.
References
fertility in cytoplasmic male sterile Phaseolus vulgaris L. Proc. Natl Acad. Sci. USA 85:2714-2717. Maddaloni,, M., Donini, G., Balconi, C, Rizzi, E., Gallusci, P., Forlani, F., Lohmer, S., Thompson, R., Salamini, F., and Motto, M. 1996. The transcriptional activator opaque-2 controls the expression of a cytosolic form of pyruvate orthophosphate dikinase-1 in maize endosperm. Mol. Gen. Genet. 250:647-654. Maddock, S.E., Lancaster, V.A., Risiott, R., and Franklin, J. 1983. Plant regeneration from cultured immature embryos and inflorescences of 25 cultivars of wheat (Triticum aestivum). J. Expt. Bot. 34:915-926. Maeda, E., and Maeda, K. 1990. Ultrastructure of egg apparatus of rice (Oryza sativa) after anthesis. ]ap. }. Crop Sci. 59:179-197. Maeda, M., Yoshioka, M., and Ito, M. 1979. Studies on the behavior of meiotic protoplasts. IV. Protoplasts isolated from microsporocytes of liliaceous plants. Bot. Mag. Tokyo 92:111-121. Magoon, M.L., Jos, J.S., and Vasudevan, K.N. 1968. Male sterile cassava. Nucleus 11:1-6. Maherchandani, N.J., and Nay lor, J.M. 1971. Variability in DNA content and nuclear morphology of the aleurone cells of Avena fatua (wild oats). Can. J. Genet. Cytol. 13:578-584. Maheshwari, N. 1958. In vitro culture of excised ovules of Papaver somniferum. Science 127:342. Maheshwari, N., and Lai, M. 1961. In vitro culture of excised ovules of Papaver somniferum L. Phytomorphology 11:307-314. Maheshwari, P. 1950. An Introduction to the Embryology of Angiosperms. New York: McGraw-Hill Book Co. Maheshwari, S.C, and Prakash, R. 1965. Physiology of anther development in Agave americana. Physiol. Plant. 18:841-852. Maheswaran, G., and Williams, E.G. 1984. Direct somatic embryoid formation on immature embryos of Trifolium repens, T. pratense and Medicago sativa, and Ma, H. 1994. The unfolding drama of flower developrapid clonal propagation of T. repens. Ann. Bot. ment: recent results from genetic and molecular 54:201-211. analyses. Genes Devel. 8:745-756. Maheswaran, G., and Williams, E.G. 1985. Origin and Ma, H., McMullen, M.D., and Finer, J.J. 1994. development of somatic embryoids formed directly Identification of a homeobox-containing gene with on immature embryos of Trifolium repens in vitro. enhanced expression during soybean (Glycine max Ann. Bot. 56:619-630. L.) somatic embryo development. Plant Mol. Biol. Mahlberg, P.G. 1960. Embryogeny and histogenesis in 24:465-473. Nerium oleander L. I. Organization of primary meristematic tissues. Phytomorphology 10:118-131. Ma, H., Yanofsky, M.F., and Meyerowitz, E.M. 1991. AGL1-AGL6, an Arabidopsis gene family with simi- Maier, U.-G., Brown, J.W.S., Schmitz, L.M., Schwall, M., larity to floral homeotic and transcription factor Dietrich, G., and Feix, G. 1988. Mapping of tissuegenes. Genes Devel. 5:484-495. dependent and independent protein binding sites to the 5' upstream region of a zein gene. Mol. Gen. Mackenzie, A., Heslop-Harrison, ]., and Dickinson, H.G. Genet. 212:241-245. 1967. Elimination of ribosomes during meiotic prophase. Nature 215:997-999. Maier, U.-G., Brown, J.W.S., Toloczyki, C, and Feix, G. MacKenzie, C.J., Yoo, B.Y., and Seabrook, J.E.A. lf990. 1987. Binding of a nuclear factor to a consensus Stigma of Solanum tuberosum cv Shepody: morpholsequence in the 5 flanking region of zein genes from ogy, ultrastructure, and secretion. Am. ]. Bot. maize. EMBO }. 6:17-22. 77:1111-1124. Maiti, I.B., Kolattukudy, P.E., and Shaykh, M. 1979. Purification and characterization of a novel cutinase Mackenzie, S.A. 1991. Identification of a sterility-inducing from nasturtium (Tropaeolum majus) pollen. Arch. cytoplasm in a fertile accession line of Phaseolus vulgaris L. Genetics 127:411^416. Biochem. Biophys 196:412-423. Mackenzie, S.A., and Chase, CD. 1990. Fertility restora- Majewska, A., and Rodriguez-Garcia, M.I. 1996. rRNA tion is associated with a loss of a portion of the mitodistribution during microspore development in chondrial genome in cytoplasmic male-sterile anthers of Beta vulgaris L.: quantitative in situ common bean. Plant Cell 2:905-912. hybridization analysis. /. Cell Sci. 109:859-866. Mackenzie, S.A., Pring, D.R., Bassett, M.J., and Chase, CD. Majewska-Sawka, A., Rodriguez-Garcia, M.I., 1988. Mitochondrial DNA rearrangement associated Nakashima, H., and Jassen, B. 1993. Ultrastructural with fertility restoration and cytoplasmic reversion to expression of cytoplasmic male sterility in sugar
References
603
meristems and is negatively regulated by beet (Beta vulgaris L.). Sex. Plant Reprod. 6:22-32. APETALA1. Plant Cell 7:1763-1771. Makaroff, C.A., Apel, I.J., and Palmer, J.D. 1989. The atp6 coding region has been disrupted and a novel read- Mandel, M.A., Bowman, J.L., Kempin, S.A., Ma, H., Meyerowitz, E.M., and Yanofsky, M.F. 1992a. ing frame generated in the mitochondrial genome of Manipulation of flower structure in transgenic cytoplasmic male-sterile radish. J. Biol. Chem. tobacco. Cell 71:133-143. 264:11706-11713. Makaroff, C.A., and Palmer, J.D. 1988. Mitochondrial Mandel, M.A., Gustafson-Brown, C, Savidge, B., and Yanofsky, M.F. 1992b. Molecular characterization of DNA rearrangements and transcriptional alterations the Arabidopsis floral homeotic gene APETALA1. in the male-sterile cytoplasm of Ogura radish. Mol. Nature 360:273-277. Cell. Biol. 8:1474-1480. Makinen, Y, and Brewbaker, J.L. 1967. Isoenzyme poly- Manen, J.F., and Pusztai, A. 1982. Immunocytochemical localisation of lectins in cells of Phaseolus vulgaris L. morphism in flowering plants. I. Diffusion of seeds. Planta 155:328-334. enzymes out of intact pollen grains. Physiol. Plant. 20:477-482. Mangelsdorf, P.C. 1923. The inheritance of defective seeds in maize. /. Hered. 14:119-126. Makinen, Y.L.A., and Lewis, D. 1962. Immunological analysis of incompatibility (S) proteins and of cross- Manickam, A., van Damme, E.J.M., Kaliselvi, K., Verhaert, P., and Peumans, W.J. 1996. Isolation and cDNA reacting material in a self-compatible mutant of Oenothera organensis. Genet. Res. 3:352-363. cloning of an Em-like protein from mung bean (Vigna radiata) axes. Physiol. Plant. 97:524-530. Malho, R., and Pais, M.S.S. 1992. Kinetics and hydrodynamics of Agapanthus umbellatus pollen-tube growth: Mann, V., Mclntosh, L., Theurer, C, and Hirschberg, J. 1989. A new cytoplasmic male sterile genotype in the a structural and stereological study. Sex. Plant Reprod. 5:163-168. sugar beet Beta vulgaris L.: a molecular analysis. Theor. Appl. Genet. 78:293-297. Malh6, R., Read, N.D., Pais, M.S., and Trewavas, A.J. 1994. Role of cytosolic free calcium in the reorientation of Mansfield, M.A., and Raikhel, N.V. 1990. Abscisic acid enhances the transcription of wheat-germ agglutinin pollen rube growth. Plant}. 5:331-341. mRNA without altering its tissue-specific expresMalh6, R., Read, N.D., Trewavas, A.J., and Pais, M.S. 1995. sion. Planta 180:548-554. Calcium channel activity during pollen tube growth and reorientation. Plant Cell 7:1173-1184. Mansfield, S.G., and Briarty, L.G. 1991. Early embryogenMalhotra, K., and Maheshwari, S.C. 1977. Enhancement esis in Arabidopsis thaliana. II. The developing by cold treatment of pollen embryoid development embryo. Can. ]. Bot. 69:461-476. in Petunia hybrida. Z. Pflanzenphysiol. 85:177-180. Mansfield, S.G., and Briarty, L.G. 1992. Cotyledon cell Malik, C.P., and Gupta, S.C. 1976. Electrophoretic analysis development in Arabidopsis thaliana during reserve of protein from pollen and pollen tubes of Calotropis deposition. Can. ]. Bot. 70:151-164. procera R. Br. Indian J. Expt. Biol. 14:688-690. Mansfield, S.G., Briarty, L.G., and Erni, S. 1991. Early Malik, C.P., Singh, M., and Thapar, N. 1976. Physiology of embryogenesis in Arabidopsis thaliana. I. The mature sexual reproduction. IV. Histochemical characterisembryo sac. Can. J. Bot. 69:447-460. tics of suspensor of Brassica campestris. Manteuffel, R., and Panitz, R. 1993. In situ localization of Phytomorphology 26:384-389. faba bean and oat legumin-type proteins in transgenic tobacco seeds by a highly sensitive immunological Malik, C.P., and Singh, M.B. 1977. Dehydrogenase and tissue print technique. Plant Mol. Biol. 22:1129-1134. isocitrate lyase activity during pollen germination in Calotropis procera. Proc. Indian Acad. Sci. 86B:371-374. Manteuffel, R., Miintz, K., Piichel, M., and Scholz, G. Malik, C.P., Tewari, H.B., and Sood, P.P. 1969. On the func1976. Phase-dependent changes of DNA, RNA and tional significance of certain phosphatases in the protein accumulation during the ontogenesis of germinating pollen grains of Portulaca grandiflora. broad bean seeds (Viciafaba L., var. minor). Biochem. Portug. Ada Biol. HA:245-252. Physiol. Pflanz. 169:595-605. Malik, C.P., and Vermani, S. 1975. Physiology of sexual Manzocchi, L.A., Bianchi, M.W., and Viotti, A. 1989. reproduction. I. A histochemical study of the embryo Expression of zein in long term cultures of wildtype sac development in Zephyranthes rosea and Lagenaria and opaaue-2 maize endosperms. Plant Cell Rep. vulgaris. Ada Histochem. 53:244-280. 7:639-643. Malik, C.P., Vermani, S., and Bhatia, D.S. 1976. Physiology Manzocchi, L.A., Daminati, M.G., and Gentinetta, E. 1980. of sexual reproduction. III. Histochemical characterViable defective endosperm mutants in maize. II. istics of suspensor during embryo development in Kernel weight, nitrogen and zein accumulation durBrassica campestris Linn, var., Sarson. Ada Histochem. ing endosperm development. Maydica 25:199-210. 57:178-182. Manzocchi, L.A., Daminati, M.G., Gentinetta, E., and Malti, and Shivanna, K.R. 1984. Structure and cytochemSalamini, F. 1980. Viable defective endosperm istry of the pistil of Crotalaria retusa L. Proc. Indian mutants in maize. I. Kernel weight, protein fractions Natl Sci. Acad. 50B:92-102. and zein subunits in mature endosperms. Maydica Mandaokar, A.D., Koundal, K.R., Kansal, R., and Bansal, 25:105-116. H.C. 1993. Characterization of vicilin seed storage Mao, Z., Paiva, R., Kriz, A.L., and Juvik, J.A. 1995. protein of chickpea (Cicer arietinum L.). /. Plant Dehydrin gene expression in normal and viviparous Biochem. Biotech. 2:35-38. embryos of Zea mays during seed development and Mandaron, P., Niogret, M.F., Mache, R., and Mongger, F. germination. Plant Physiol. Biochem. 33:649-653. 1990. In vitro protein synthesis in isolated Maquoi, E., Hanke, D.E., and Deltour, R. 1993. The effects microspores of Zea mays at several stages of develof abscisic acid on the maturation of Brassica napus opment. Theor. Appl. Genet. 80:134-158. somatic embryos. An ultrastructural study. Mandel, M.A., and Yanofsky, M.F. 1995. The Arabidopsis Protoplasma 174:147-157. AGL8 MADS box gene is expressed in inflorescence Marciniak, K. 1993. DNA endoreplication level in
604
References
endosperm during seed development in three Marsh, G., and Beams, H.W. 1945. The orientation of monocotyledonous species. Acta Soc. Bot. Polon. pollen tubes of Vinca in the electric current. /. Cell. 62:143-147. Cotnp. Physiol. 25:195-204. Marco, Y.A., Thanh, V.H., Turner, N.E., Scallon, B.J., and Marsolier, M.-C, Carrayol, E., and Hirel, B. 1993. Multiple Nielsen, N.C. 1984. Cloning and structural analysis functions of promoter sequences involved in organof DNA encoding an A2Bla subunit of glycinin. /. Biol. specific expression and ammonia regulation of a Chem. 259:13436-13441. cytosolic soybean glutamine synthetase gene in Marcotte, W.R., Jr., Russell, S.H., and Quatrano, R.S. 1989. transgenic Lotus corniculatus. Plant ]. 3:405-414. Abscisic acid-responsive sequences from the Em Martin, F.W. 1969. Compounds from the stigmas of ten gene of wheat. Plant Cell 1:969-976. species. Am. J. Bot. 56:1023-1027. Mares, D., Jeffery, C, and Norstog, K. 1976. The differen- Martin, F.W. 1970a. The stigmatic exudate of Strelitzia. tiation of the modified aleurone of wheat Phyton Argentina 27:47-53. endosperm: nuclear membrane-mediated endoplas- Martin, F.W. 1970b. Compounds of the stigmatic surface mic reticulum development. Plant Sci. Lett. of Zea mays L. Ann. Bot. 34:835-842. 7:305-311. Martin, F.W. 1970c. The ultraviolet absorption profile of Mares, D.J., Norstog, K., and Stone, B.A. 1975. Early stigmatic extracts. New Phytol. 69:425-430. stages in the development of wheat endosperm. I. Martin, F.W, and Telek, L. 1971. The stigmatic secretion of The change from free nuclear to cellular endosperm. the sweetpotato. Am. ]. Bot. 58:317-322. Aust. J. Bot. 23:311-326. Martin, P.G. 1960. Differentiation of the nuclei of pollen Mares, D.J., Stone, B.A., Jeffery, C, and Norstog, K. 1977. grains. Heredity 14:125-132. Early stages in the development of wheat Martin-Tanguy, J., Perdrizet, E., Prevost, J., and Martin, C. endosperm. II. Ultrastructural observations on cell 1982. Hydroxycinnamic acid amides in fertile and wall formation. Aust. J. Bot. 25:599-613. cytoplasmic male sterile lines of maize. Phytochemistry 21:1939-1945. Mariani, C, de Beuckeleer, M., Truettner, J., Leemans, J., and Goldberg, R.B. 1990. Induction of male sterility Martinez-Palle, E., and Herrero, M. 1995. The ponticulus: in plants by a chimaeric ribonuclease gene. Nature a structure bridging pollen tube access to the ovules 347:737-741. of Pistacia vera. Sex. Plant Reprod. 8:217-222. Mariani, G, Gossele, V., de Beuckeleer, M., de Block, M., Martinussen, I., Junttila, O., and Twell, D. 1994. Goldberg, R.B., de Greef, W, and Leemans, J. 1992. A Optimization of transient gene expression in pollen chimaeric ribonuclease-inhibitor gene restores fertilof Norway spruce (Picea abies) by particle acceleraity to male sterile plants. Nature 357:384-387. tion. Physiol. Plant. 92:412-416. Marinos, N.G. 1970a. Embryogenesis of the pea (Pisum Martinussen, I., Bate, N., Weterings, K., Junttila, O., and sativum). I. The cytological environment of the develTwell, D. 1995. Analysis of gene regulation in growoping embryo. Protoplasma 70:261-279. ing pollen tubes of angiosperm and gymnosperm species using microprojectile bombardment. Physiol. Marinos, N.G. 1970b. Embryogenesis of the pea (Pisum Plant. 93:445-450. sativum). II. An unusual type of plastid in the suspensor cells. Protoplasma 71:227-233. Maruyama, K. 1968. Electron microscopic observation of Marks, M.D., and Larkins, B.A. 1982. Analysis of plastids and mitochondria during pollen developsequence microheterogeneiry among zein messenger ment in Tradescantia paludosa. Cytologia 33:482-497. RNAs. /. Biol. Chem. 257:9976-9983. Maruyama, K., Gay, H., and Kaufmann, B.P. 1965. The nature of the wall between generative and vegetaMarks, M.D., Lindell, J.S., and Larkins, B.A. 1985a. tive nuclei in the pollen grain of Tradescantia paluQuantitative analysis of the accumulation of zein dosa. Am. ]. Bot. 52:605-610. mRNA during maize endosperm development. /. Biol. Chem. 260:16445-16450. Masand, P., and Kapil, R.N. 1966. Nutrition of the embryo Marks, M.D., Lindell, J.S., and Larkins, B.A. 1985b. sac and embryo - a morphological approach. Nucleotide sequence analysis of zein mRNAs from Phytomorphology 16:158-175. maize endosperm. /. Biol. Chem. 260:16451-16459. Mascarenhas, J.P. 1966a. The distribution of ionic calcium Marocco, A., Santucci, A., Cerioli, S., Motto, M., di Fonzo, in the tissues of the gynoecium of Antirrhinum majus. N., Thompson, R., and Salamini, F. 1991. Three highProtoplasma 62:53-58. lysine mutations control the level of ATP-binding Mascarenhas, J.P. 1966b. Pollen tube growth and ribonuHSP70-like proteins in the maize endosperm. Plant cleic acid synthesis by vegetative and generative Cell 3:507-515. nuclei of Tradescantia. Am. J. Bot. 53:563-569. Marris, C, Gallois, P., Copley, J., and Kreis, M. 1988. The Mascarenhas, J.P. 1970. A new intermediate in plant cell 5' flanking region of a barley B hordein gene conwall synthesis. Biochem. Biophys. Res. Commun. trols tissue and developmental specific CAT 41:142-149. expression in tobacco plants. Plant Mol. Biol. Mascarenhas, J.P. 1975. The biochemistry of angiosperm 10:359-366. pollen development. Bot. Rev. 41:259-314. Marrs, K.A., Casey, E.S., Capitant, S.A., Bouchard, R.A., Mascarenhas, J.P. 1989. The male gametophyte of flowerDietrich, P.S., Mettler, I.J., and Sinibaldi, R.M. 1993. ing plants. Plant Cell 1:657-664. Characterization of two maize HSP90 heat shock Mascarenhas, J.P. 1990. Gene activity during pollen develprotein genes: expression during heat shock, opment. Annu. Rev. Plant Physiol. Plant Mol. Biol. embryogenesis, and pollen development. Devel 41:317-338. Genet. 14:27-^1. Mascarenhas, J.P. 1992. Pollen gene expression: molecular Marsden, M.P.F., and Meinke, D.W. 1985. Abnormal evidence. Int. Rev. Cytol. 140:3-18. development of the suspensor in an embryo- Mascarenhas, J.P. 1993. Molecular mechanisms of pollen lethal mutant of Arabidopsis thaliana. Am. }. Bot. tube growth and differentiation. Plant Cell 72:1801-1812. 5:1303-1314.
References
605
Mascarenhas, J.P., and Bell, E. 1969. Protein synthesis dur- Matsubara, S. 1964. Effect of nitrogen compounds on the ing germination of pollen. Studies on polyribosome growth of isolated young embryos of Datura. Bot. formation. Biochim. Biophys. Ada 179:199-203. Mag. Tokyo 77:253-259. Mascarenhas, J.P., and Bell, E. 1970. RNA synthesis during Matsubayashi, M., and Kuranuki, K. 1975. Embryogenic development of the male gametophyte of responses of the pollen to varied sucrose concentraTradescantia. Bevel Biol. 21:475-490. tions in anther culture. Sci. Rep. Vac. Agr. Kobe Univ. Mascarenhas, J.P., and Goralnick, R.D. 1971. Synthesis of 11:215-230. small molecular weight RNA in the pollen tube of Matsuda, N., Tsuchiya, T., Kishitani, S., Tanaka, Y., and Tradescantia paludosa. Biochim. Biophys. Ada 240:56-61. Toriyama, K. 1996. Partial male sterility in transgenic tobacco carrying antisense and sense PAL cDNA Mascarenhas, J.P., and Lafountain, J. 1972. Protoplasmic under the control of a tapetum-specific promoter. streaming, cytochalasin B, and growth of the pollen Plant Cell Physiol. 37:215-222. tube. Tissue Cell 4:11-14. Mascarenhas, J.P., and Machlis, L. 1962a. Chemotropic Matters, G.L., and Boyer, CD. 1982. Soluble starch synthases and starch branching enzymes from cotyleresponse of Antirrhinum majus pollen to calcium. dons of smooth- and wrinkled-seeded lines of Pisum Nature 196:292-293. sativum L. Biochem. Genet. 20:833-848. Mascarenhas, J.P., and Machlis, L. 1962b. The pollen-tube chemotropic factor from Antirrhinum majus: bioas- Matthews, B.F., Abdul-Baki, A.A., and Saunders, J.A. say, extraction, and partial purification. Am. ]. Bot. 1990. Expression of a foreign gene in electroporated 49:482-489. pollen grains of tobacco. Sex. Plant Reprod. 3:147-151. Mascarenhas, J.P., and Machlis, L. 1964. Chemotropic Matthews, J.A., Brown, J.W.S., and Hall, T.C. 1981. response of the pollen of Antirrhinum majus to calPhaseolin mRNA is translated to yield glycosylated cium. Plant Physiol. 39:70-77. polypeptides in Xenopus oocytes. Nature 294:175-176. Mascarenhas, J.P., and Mermelstein, J. 1981. Messenger RNAs: their utilization and degradation during Matthews, J.A., and Miflin, B.J. 1980. In vitro synthesis of pollen germination and tube growth. Ada Soc. Bot. barley storage proteins. Planta 149:262-268. Polon. 50:13-20. Matthys-Rochon, E., M61, R., Heizmann, P., and Dumas, C. 1994. Isolation and microinjection of active sperm Mascarenhas, N.T., Bashe, D., Eisenberg, A., Willing, R.P., nuclei into egg cells and central cells of isolated Xiao, C.-M., and Mascarenhas, J.P. 1984. Messenger maize embryo sacs. Zygote 2:29-35. RNAs in corn pollen and protein synthesis during germination and pollen tube growth. Theor. Appl. Matthys-Rochon, E., Vergne, P., Detchepare, S., and Genet. 68:323-326. Dumas, C. 1987. Male germ unit isolation from three tricellular pollen species: Brassica oleracea, Zea mays, Masuda, H., Oohashi, S.-I., Tokuji, Y., and Mizue, Y. 1995. and Triticum aestivum. Plant Physiol. 83:464—466. Direct embryo formation from epidermal cells of carrot hypocotyls. /. Plant Physiol. 145:531-534. Matton, D.P., Lau, S.-M., Okamoto, S., Clarke, A.E., and Newbigin, E. 1995. The S-locus of Nicotiana alata: Masuda, K., Kikuta, Y, and Okazawa, Y. 1984. Embryogenesis and ribosomal DNA in carrot cell genomic organization and sequence analysis of two suspensions cultured in vitro. Plant Sci. Lett. SRNase alleles. Plant Mol. Biol. 28:847-858. 33:23-29. Mattsson, J., Borkird, C, and Engstrom, P. 1992. Spatial and temporal expression patterns directed by the Masumori, M., Yamamoto, N., and Sasaki, S. 1992. Stored Agrobacterium tumefaciens T-DNA gene 5 promoter mRNA in pine seeds: prolonged preservation in dry during somatic embryogenesis in carrot. Plant Mol. seeds and disappearance during germination. Plant Biol. 18:629-637. Cell Physiol. 33:485^188. Masumura, T., Hibino, T., Kidzu, K., Mitsukawa, N., Mattsson, O., Knox, R.B., Heslop-Harrison, J., and Tanaka, K., and Fujii, S. 1990. Cloning and characterHeslop-Harrison, Y. 1974. Protein pellicle of stigization of a cDNA encoding a rice 13 kDa prolamin. matic papillae as a probable recognition site in Mol. Gen. Genet. 221:1-7. incompatibility reactions. Nature 247:298-300. Masumura, T., Kidzu, K., Sugiyama, Y., Mitsukawa, N., Matzke, A.J.M., Stoger, E.M., and Matzke, M.A. 1993. A Hibino, T., Tanaka, K., and Fujii, S. 1989a. Nucleotide zein gene promoter fragment drives GUS expression sequence of a cDNA encoding a major rice glutelin. in a cell layer that is interposed between the Plant Mol. Biol. 12:723-725. endosperm and the seed coat. Plant Mol. Biol. 22:553-554. Masumura, T., Shibata, D., Hibino, T., Kato, T., Kawabe, K., Takeba, G., Tanaka, K., and Fujii, S. 1989b. cDNA Matzke, A.J.M., Stoger, E.M., Schernthaner, J.P., and cloning of an mRNA encoding a sulfur-rich 10 kDa Matzke, M.A. 1990. Deletion analysis of a zein gene prolamin polypeptide in rice seeds. Plant Mol. Biol. promoter in transgenic tobacco plants. Plant Mol. 12:123-130. Biol. 14:323-332. Matousek, J., and Tupy, J. 1983. The release of nucleases Matzke, M.A., Susani, M., Binns, A.N., Lewis, E.D., from tobacco pollen. Plant Sci. Lett. 30:83-89. Rubenstein, I., and Matzke, A.J.M. 1984. Matousek, J., and Tupy, J. 1984. Purification and properTranscription of a zein gene introduced into sunties of extracellular nuclease from tobacco pollen. flower using a Ti plasmid vector. EMBO ]. Biol. Plant. 26:62-73. 3:1525-1531. Matousek, J., and Tupy, J. 1985. The release and some Mau, S.L., Raff, J., and Clarke, A.E. 1982. Isolation and partial characterization of components of Prunus properties of nuclease from various pollen species. /. avium L. styles, including an antigenic glycoprotein Plant Physiol. 119:169-178. associated with a self-incompatibility genotype. Matsubara, S. 1962. Studies on a growth promoting subPlanta 156:505-516. stance, "embryo factor", necessary for the culture of young embryos of Datura tatula in vitro. Bot. Mag. Mau, S.-L., Williams, E.G., Atkinson, A., Anderson, M.A., Tokyo 75:10-18. Cornish, E.C., Grego, B., Simpson, R.J., Kheyr-Pour,
606
References
A., and Clarke, A.E. 1986. Style proteins of a McConchie, C.A., Jobson, S., and Knox, R.B. 1985. wild tomato (Lycopersicon peruvianum) associated Computer-assisted reconstruction of the male germ with expression of self-incompatibility. Planta unit in pollen of Brassica campestris. Protoplasma 169:184-191. 127:57-63. Maurel, C, Brevet, J., Barbier-Brygoo, H., Guern, J., and McConchie, C.A., Knox, R.B., and Ducker, S.C. 1982. Temp6, J. 1990. Auxin regulates the promoter of the Pollen wall structure and cytochemistry in the searoot-inducing rolB gene of Agrobacterium rhizogenes grass Amphibolis griffithii (Cymodoceaceae). Ann. in transgenic tobacco. Mol. Gen. Genet. 223:58-64. Bot. 50:729-732. May, K.W., and Kasha, K.J. 1980. The cytological expres- McConchie, C.A., Russell, S.D., Dumas, G, Tuohy, M., sion and inheritance of desynapsis in a clone of and Knox, R.B. 1987. Quantitative cytology of the diploid timothy (Phleum nodosum L.). Euphytica sperm cells of Brassica campestris and B. oleracea. 29:233-240. Planta 170:446-452. Mayer, U., Buttner, G., and Jurgens, G. 1993. Apical-basal McConn, M., and Browse, J. 1996. The critical requirement pattern formation in the Arabidopsis embryo: studies for linolenic acid is pollen development, not photosynthesis, in an Arabidopsis mutant. Plant Cell on the role of the gnom gene. Development 8:403-416. 117:149-162. Mayer, U., Torres Ruiz, R.A., Berleth, T., Misera, S., and McCormick, S. 1991. Molecular analysis of male gametoJurgens, G. 1991. Mutations affecting body organizagenesis in plants. Trends Genet. 7:298-303. tion in the Arabidopsis embryo. Nature 353:402-407. McCoy, K., and Knox, R.B. 1988. The plasma membrane and generative cell organization in pollen of the Maze, J., and Bohm, L.R. 1974. Embryology of Agrostis mimosoid legume, Acacia retinodes. Protoplasma interrupta (Gramineae). Can. ]. Bot. 52:365-379. 143:85-92. Maze, J., and Bohm, L.R. 1977. Embryology of Festuca microstachys (Gramineae). Can. J. Bot. 55:1768-1782. McDaniel, C.N., and Poethig, R.S. 1988. Cell-lineage patMaze, J., and Lin, S.-C. 1975. A study of the mature terns in the shoot apical meristem of the germinating megagametophyte of Stipa elmeri. Can. J. Bot. maize embryo. Planta 175:13-22. 53:2958-2977. McDonnell, R.E., and Conger, B.V. 1984. Callus induction and plantlet formation from mature embryo McCabe, D.E., and Martinell, B.J. 1993. Transformation of explants of Kentucky bluegrass. Crop Sci. 24:573-578. elite cotton cultivars via particle bombardment of meristems. Biotechnology 11:596-598. McGee, J.D., Williams, E.G., Collins, G.B., and Hildebrand, D.F. 1989. Somatic embryogenesis in McCabe, D.E., Swain, W.F., Martinell, B.F., and Christou, P. Trifolium: protein profiles associated with high- and 1988. Stable transformation of soybean (Glycine max) low-frequency regeneration. /. Plant Physiol. by particle acceleration. Biotechnology 6:923-926. 135:306-312. McCarty, D.R., and Carson, C.B. 1991. The molecular genetics of seed maturation in maize. Physiol. Plant. McGranahan, G.H., Leslie, C.A., Uratsu, S.L., and Dandekar, A.M. 1990. Improved efficiency of the 81:267-272. walnut somatic embryo gene transfer system. Plant McCarty, D.R., Shaw, J.R., and Hannah, L.C. 1986. The Cell Rep. 8:512-516. cloning, genetic mapping, and expression of the constitutive sucrose synthase locus of maize. Proc. Natl McGranahan, G.H., Leslie, C.A., Uratsu, S.L., Martin, L.A., and Dandekar, A.M. 1988. Agrobacterium-mediAcad. Sci. USA 83:9099-9103. ated transformation of walnut somatic embryos and McCarty, D.R., Carson, C.B., Lazar, M., and Simonds, S.C. regeneration of transgenic plants. Biotechnology 1989a. Transposable element-induced mutations of 6:800-804. the viviparous-1 gene in maize. Devel Genet. 10:473-481. McGuire, D.C, and Rick, CM. 1954. Self-incompatibility in species of Lycopersicon sect. Eriopersicon and McCarty, D.R., Carson, C.B., Stinard, P.S., and Robertson, hybrids with L. esculentum. Hilgardia 23:101-124. D.S. 1989b. Molecular analysis of viviparous-1: an abscisic acid-insensitive mutant of maize. Plant Cell McHughen, A. 1980. The regulation of tobacco floral 1:523-532. organ initiation. Bot. Gaz. 141:389-395. McCarty, D.R., Hattori, T., Carson, C.B., Vasil, V., Lazar, McWilliam, A.A., Smith, S.M., and Street, H.E. 1974. The origin and development of embryoids in suspension M., and Vasil, I.K. 1991. The viviparous-1 developcultures of carrot (Daucus carota). Ann. Bot. mental gene of maize encodes a novel transcrip38:243-250. tional activator. Cell 66:895-905. McClure, B.A., Du, H., Liu, Y.-H., and Clarke, A.E. 1993. Meakin, P.J., and Gatehouse, J.A. 1991. Interaction of seed nuclear proteins with transcriptionally-enhancing S-locus products in Nicotiana alata pistils are subject regions of the pea (Pisum sativum L.) legA gene proto organ-specific post-transcriptional processing but moter. Planta 183:471-477. not post-translational processing. Plant Mol. Biol. Mecham, D.K., Fullington, J.G., and Greene, F.C. 1981. 22:177-181. Gliadin proteins in the developing wheat seed. /. Sci. McClure, B.A., Gray, J.E., Anderson, M.A., and Clarke, Food Agr. 32:773-780. A.E. 1990. Self-incompatibility in Nicotiana alata involves degradation of pollen rRNA. Nature Medford, J.I., and Sussex, I.M. 1989. Regulation of chlorophyll and Rubisco levels in embryonic cotyledons of 347:757-760. Phaseolus vulgaris. Planta 179:309-315. McClure, B.A., Haring, V, Ebert, P.E., Anderson, M.A., Simpson, R.J., Sukiyama, R, and Clarke, A.E. 1989. Medina, F.J., and Risueno, M.C. 1981. Nucleolar structure Style self-incompatibility gene products of Nicotiana and dynamics during meiotic prophase in pea alata are ribonucleases. Nature 342:955-957. ovules. Biol. Cell. 42:79-86. McConchie, C.A., Hough, T., and Knox, R.B. 1987. Medina, F.J., Risueno, M.C, and Rodriguez-Garcia, M.I. 1981. Evolution of the cytoplasmic organelles during Ultrastructural analysis of the sperm cells of female meiosis in Pisum sativum L. Planta mature pollen of maize, Zea mays. Protoplasma 151:215-225. 139:9-19.
References
607
Medina, F.J., Risuefio, M.C., Rodriguez-Garcia, M.I., and Mengoli, M., Bagni, N., Luccarini, G., Nuti Ronchi, V., and Serafini-Fracassini, D. 1989. Dancus carota cell Sanchez-Pina, M.A. 1983. The nucleolar organizer cultures: polyamines and effect of polyamine (NOR) and fibrillar centers during plant gametogenbiosynthesis inhibitors in the preembryogenic phase esis. /. Ultrastr. Res. 85:300-310. and different embryo stages. /. Plant Physiol. Meier, H., and Grant Reid, J.S. 1977. Morphological 134:389-394. aspects of the galactomannan formation in the endosperm of Trigonella foenum-graecum L. Menzel, G., Apel, K., and Melzer, S. 1995. Isolation and analysis of SaMADS C, the apetala 1 cDNA homolog (Leguminosae). Planta 133:243-248. from mustard. Plant Physiol. 108:853-854. Meijer, E.G.M., and Brown, D.C.W. 1987a. Role of exogenous reduced nitrogen and sucrose in rapid high fre- Mepham, R.H., and Lane, G.R. 1969. Formation and development of the tapetalperiplasmodium in quency somatic embryogenesis in Medicago sativa. Plant Cell Tissue Organ Cult, 10:11-19. Tradescantia bracteata. Protoplasma 68:175-192. Meijer, E.G.M., and Brown, D.C.W. 1987b. A novel system Mepham, R.H., and Lane, G.R. 1970. Observations on the fine structure of developing microspores of for rapid high frequency somatic embryogenesis in Tradescantia bracteata. Protoplasma 70:1-20. Medicago sativa. Physiol. Plant. 69:591-596. Meijer, E.G.M., and Simmonds, J. 1988. Polyamine levels Mericle, L.W., and Mericle, R.P. 1957. Irradiation of developing plant embryos. I. Effects of external irradiain relation to growth and somatic embryogenesis in tion (X rays) on barley embryogeny, germination tissue cultures of Medicago sativa L. /. Expt. Bot. and subsequent seedling development. Am. }. Bot. 39:787-794. 44:747-756. Meikle, P. J., Bonig, I., Hoogenraad, N.J., Clarke, A.E., and Stone, B.A. 1991. The location of (l-»3)-f3-glucans in Mericle, L.W., and Mericle, R.P. 1961. Radiosensitivity of the walls of pollen tubes of Nicotiana alata using a the developing plant embryo. In Fundamental Aspects of Radiosensitivity. Brookhaven Symp. Biol. 14:262-286. (l->3)-|3-glucan-specific monoclonal antibody. Planta Mericle, L.W., and Mericle, R.P. 1970. Nuclear DNA com185:1-8. plement in young proembryos of barley. Mutation Meinke, D.W. 1982. Embryo-lethal mutants of Arabidopsis Res. 10:515-518. thaliana: evidence for gametophytic expression of the Merkle, S.A., Parrott, W.A., and Flinn, B.S. 1995. mutant genes. Theor. Appl. Genet. 63:381-386. Morphogenetic aspects of somatic embryogenesis. Meinke, D.W. 1985. Embryo-lethal mutants of Arabidopsis In In Vitro Embryogenesis in Plants, ed. T.A. Thorpe, thaliana: analysis of mutants with a wide range of pp. 155-203. Dordrecht: Kluwer Academic lethal phases. Theor. Appl. Genet. 69:543-552. Publishers. Meinke, D.W. 1986. Embryo-lethal mutants and the study of plant embryo development. Oxford Surv. Plant Mertz, E.T., Bates, L.S., and Nelson, O.E. 1964. Mutant gene that changes protein composition and increases Mol. Cell Biol. 3:122-165. lysine content of maize endosperm. Science Meinke, D.W. 1991a. Embryonic mutants of Arabidopsis thaliana. Devel Genet. 12:382-392. 145:279-280. Meinke, D.W. 1991b. Genetic analysis of plant develop- Meurs, C, Basra, A.S., Karssen, CM., and van Loon, L.C. ment. In Plant Physiology. A Treatise, Vol. X, ed. F.C. 1992. Role of abscisic acid in the induction of desicSteward and R.G.S. Bidwell, pp. 437-490. San Diego: cation tolerance in developing seeds of Arabidopsis thaliana. Plant Physiol. 98:1484-1493. Academic Press. Meinke, D.W. 1991c. Perspectives on genetic analysis of Meyer, K., Leube, M.P., and Grill, E. 1994. A protein phosphatase 2C involved in ABA signal transduction in plant embryogenesis. Plant Cell 3:857-866. Arabidopsis thaliana. Science 264:1452-1455. Meinke, D.W. 1992. A homeotic mutant of Arabidopsis thaliana with leafy cotyledons. Science 258:1647-1650. Meyer, V.G. 1969. Some effects of genes, cytoplasm, and environment on male sterility of cotton (Gossypium). Meinke, D.W., Chen, J., and Beachy, R.N. 1981. Expression Crop Sci. 9:237-242. of storage-protein genes during soybean seed develMeyer, V.G., and Buffet, M. 1962. Cytoplasmic effects on opment. Planta 153:130-139. external-ovule production in cotton. /. Hered. Meinke, D.W., and Sussex, I.M. 1979a. Embryo-lethal 53:251-253. mutants of Arabidopsis thaliana. A model system for genetic analysis of plant embryo development. Devel Meyerowitz, E.M. 1994. The genetics of flower development. Sci. Am. 271(5):40-47. Biol. 72:50-61. Meinke, D.W., and Sussex, I.M. 1979b. Isolation and char- Meyerowitz, E.M., Bowman, J.L., Brockman, L.L., Drews, G.N., Jack, T., Sieburth, L., and Weigel, D. 1991. A acterization of six embryo-lethal mutants of genetic and molecular model for flower developArabidopsis thaliana. Devel Biol. 72:62-72. ment in Arabidopsis thaliana. Development Suppl. Meinke, D.W., Franzmann, L.H., Nickle, T.C., and Yeung, 1:157-167. E.C. 1994. Leafy cotyledon mutants of Arabidopsis. Mian, H.R., Kuspira, J., Walker, G.W.R., and Muntjewerff, Plant Cell 6:1049-1064. N. 1974. Histological and cytochemical studies on Melcher, U. 1979. In vitro synthesis of a precursor to the five genetic male-sterile lines of barley (Hordeum vitlmethionine-rich polypeptide of the zein fraction of gare). Can. ]. Genet. Cytol. 16:355-379. corn. Plant Physiol. 63:354-358. Melchers, G., Mohri, Y, Watanabe, K., Wakabayashi, S., Miao, S.-H., Kuo, C.-S., Kwei, Y.-L., Sun, A.-T., Ku, S.-Y, Lu, W.-L., Wang, Y.-Y, Chen, M.-L., Wu, M.-K., and and Harada, K. 1992. One-step generation of cytoplasmic male sterility by fusion of mitochondrHang, L. 1978. Induction of pollen plants of maize ial-inactivated tomato protoplasts with nuclear-inacand observations on their progeny. In Proceedings of tivated Solanum protoplasts. Proc. Natl Acad. Sci. Symposium on Plant Tissue Culture, pp. 23-33. Peking: USA 89:6832-6836. Science Press. Mena, M., Mandel, M.A., Lerner, D.R., Yanofsky, M.F., Michalczuk, L., Cooke, T.J., and Cohen, J.D. 1992. Auxin and Schmidt, R.J. 1995. A characterization of the levels at different stages of carrot somatic embryogeMADS-box gene family in maize. Plant ]. 8:845-854. nesis. Phytochemistry 31:1097-1103.
608
References
Michalczuk, L., Ribnicky, D.M., Cooke, T.J., and Cohen, deficient miniature-1 seed mutation is associated J.D. 1992. Regulation of indole-3-acetic acid biosynwith aberrant pedicel and endosperm development. thetic pathways in carrot cell cultures. Plant Physiol. Plant Cell 4:297-305. 100:1346-1353. Miller, R.J., and Koeppe, D.E. 1971. Southern corn leaf Michel, D., Salamini, R, Bartels, D., Dale, P., Baga, M., and blight: susceptible and resistant mitochondria. Szalay, A. 1993. Analysis of a desiccation and ABAScience 173:67-69. responsive promoter isolated from the resurrection Millerd, A. 1975. Biochemistry of legume seed proteins. plant Craterostigma planlagineum. Plant J. 4:29-40. Annu. Rev. Plant Physiol. 26:53-72. Miernyk, J.A., and Trelease, R.N. 1981. Control of enzyme Millerd, A., Simon, M., and Stern, H. 1971. Legumin synactivities in cotton cotyledons during maturation thesis in developing cotyledons of Viciafaba L. Plant and germination. IV. fi-Oxidation. Plant Physiol. Physiol. 48:419-425. 67:341-346. Millerd, A., and Spencer, D. 1974. Changes in RNA-synMiflin, B.J., Burgess, S.R., and Shewry, P.R. 1981. The thesizing activity and template activity in nuclei development of protein bodies in the storage tissues from cotyledons of developing pea seeds. Aust. ]. of seeds: subcellular separations of homogenates of Plant Physiol. 1:331-341. barley, maize, and wheat endosperms and of pea Millerd, A., Thomson, J.A., and Schroeder, H.E. 1978. cotyledons. /. Expt. hot. 32:199-219. Cotyledonary storage proteins in Pisum sativum. III. Mignouna, H., Virmani, S.S., and Briquet, M. 1987. Patterns of accumulation during development. Aust. J. Plant Physiol. 5:519-534. Mitochondrial DNA modifications associated with cytoplasmic male sterility in rice. Theor. Appl. Genet. Millerd, A., and Whitfeld, P.R. 1973. Deoxyribonucleic 74:666-669. acid and ribonucleic acid synthesis during the cell Mii, M. 1973. Studies on the haploid formation from expansion phase of cotyledon development in Vicia immature pollen grain. I. Relation between pollen faba L. Plant Physiol. 51:1005-1010. stage and the plantlet formation and the process of Millerd, A., Spencer, D., Dudman, W.F., and Stiller, M. embryoid formation in Nicotiana tabacum. ]ap. ]. 1975. Growth of immature pea cotyledons in culture. Breed. 23:27-34. Aust. J. Plant Physiol. 2:51-59. Mikami, T., Kishima, Y, Sugiura, M., and Kinoshita, T. Milligan, S.B., and Gasser, C.S. 1995. Nature and regula1985. Organelle genome diversity in sugar beet with tion of pistil-expressed genes in tomato. Plant Mol. normal and different sources of male sterile cytoBiol. 28:691-711. plasms. Theor. Appl. Genet. 71:166-171. Minocha, S.C, Papa, N.S., Khan, A.J., and Samuelsen, A.I. Mikami, T., Sugiura, M., and Kinoshita, T. 1984. Molecular 1991. Polyamines and somatic embryogenesis in carheterogeneity in mitochondrial and chloroplast rot. III. Effects of methylglyoxal bis(guanylhydraDNAs from normal and male sterile cytoplasms in zone). Plant Cell Physiol. 32:395-402. sugar beets. Curr. Genet. 8:319-322. Mis£ra, S., Miiller, A.J., Weiland-Heidecker, U., and Miki, H. 1964. A study of the tropism of pollen tubes to the Jurgens, G. 1994. The FUSCA genes of Arabidopsis: pistil. I. Tropism in Lilium. Bot. Mag. Tokyo 67:143-147. negative regulators of light responses. Mol. Gen. Miki-Hirosige, H., Hoek, I.H.S., and Nakamura, S. 1987. Genet. 244:242-252. Secretions from the pistil of Lilium longiflorum. Am. J. Mishkind, M., Raikhel, N.V., Palevitz, B.A., and Keegstra, Bot. 74:1709-1715. K. 1982. Immunocytochemical localization of wheat germ agglutinin in wheat. /. Cell Biol. 92:753-764. Miki-Hirosige, H., and Nakamura, S. 1982. Process of metabolism during pollen tube wall formation. /. Mishkind, M.L., Palevitz, B.A., Raikhel, N.V., and Electron Micros. 31:51-62. Keegstra, K. 1983. Localization of wheat germ aggluMike-Hirosige, H., Nakamura, S., and Tanaka, I. 1988. tinin-Iike lectins in various species of the Gramineae. Ultrastructural research on cell wall regeneration by Science 220:1290-1292. cultured pollen protoplasts of Lilium longiflorum. Sex. Misoo, S., Yoshida, K., and Mastubayashi, M. 1979. Plant Reprod. 1:36-45. Studies on the mechanisms of pollen embryogenesis. III. Mitotic responses of the pollen to varied sucrose Miki-Hirosige, H., Nakamura, S., Yasueda, H., Shida, T., concentrations and the process of embryoid formaand Takahashi, Y. 1994. Immunocytochemical localtion in tobacco anther culture. Sci. Rep. Fac. Agr. Kobe ization of the allergenic proteins in the pollen of Univ. 13:193-202. Cryptomeria japonica. Sex. Plant Reprod. 7:95-100. Mikulska, E., and Rodkiewicz, B. 1967a. infrastructure of Misra, S., and Bewley, J.D. 1985a. Reprogramming of prothe maturing embryo sac of Lilium regale. Ada Soc. tein synthesis from a developmental to a germinaBot Polon. 36:555-566. tive mode induced by desiccation of the axes of Phaseolus vulgaris. Plant Physiol. 78:876-882. Mikulska, E., and Rodkiewicz, B. 1967b. Fine structure of four-nucleate stages and the central cell of Lilium Misra, S., and Bewley, J.D. 1985b. The messenger RNA regale embryo sac. Flora 157:365-372. population in the embryonic axes of Phaseolus vidgaris during development and following germinaMiller, D.D., Scordilis, S.P., and Hepler, P.K. 1995. tion. /. Expt. Bot. 36:1644-1652. Identification and localization of three classes of myosins in pollen tubes of Lilium longiflorum and Misra, S., and Green, M.J. 1991. Developmental gene Nicotiana alata. J. Cell Sci. 108:2549-2563. expression in conifer embryogenesis and germination. II. Crystalloid protein synthesis in the developMiller, D.D., Callaham, D.A., Gross, D.J., and Hepler, P.K. 2+ ing embryo and megagametophyte of white spruce 1992. Free Ca gradient in growing pollen tubes of (Picea glauca [Moench] Voss.). Plant Sci. 78:61-71. Lilium. J. Cell Sci. 101:7-12. Miller, E.C. 1919. Development of the pistillate spikelet Misra, S., Attree, S.M., Leal, I., and Fowke, L.C. 1993. Effect of abscisic acid, osmoticum, and desiccation on synand fertilization in Zea mays L. /. Agr. Res. thesis of storage proteins during the development of 18:255-265. white spruce somatic embryos. Ann. Bot. 71:11-22. Miller, M.E., and Chourey, P.S. 1992. The maize invertase-
References
609
Misset, M.T., and Gourret, J.P. 1984. Accumulation of Moens, P.B. 1968. The structure and function of the synapsmooth cisternae in the tapetal cells of Ulex europaeits tonemal complex in Lilium longiflorum sporocytes. L. (Papilionoideae). /. Cell Sci. 72:65-74. Chromosoma 23:418-451. Mitchell, J.C., and Petolino, J.F. 1988. Heat stress effects on Moewus, F. 1950. Zur Physiologie und Biochemie der Selbststerilitat bei Forsythia. Biol. Zentral. 69:181-197. isolated reproductive organs of maize. /. Plant Moffatt, B., Pethe, C, and Laloue, M. 1991. Metabolism of Physiol. 133:625-628. benzyladenine is impaired in a mutant of Arabidopsis Mitten, D.H., Sato, S.J., and Skokut, T.A. 1984. In vitro thaliana lacking adenine phosphoribosyltransferase regenerative potential of alfalfa germplasm sources. activity. Plant Physiol. 95:900-908. Crop Sci. 24:943-945. Mittermann, I., Swoboda, I., Pierson, E., Eller, N., Kraft, Moffatt, B., and Somerville, C. 1988. Positive selection for D., Valenta, R., and Heberle-Bors, E. 1995. Molecular male-sterile mutants of Arabidopsis lacking adenine cloning and characterization of profilin from tobacco phosphoribosyl transferase activity. Plant Physiol. (Nicotiana tabacum): increased profilin expression 86:1150-1154. during pollen maturation. Plant Mol. Biol. Mogensen, H.L. 1972. Fine structure and composition of 27:137-146. the egg apparatus before and after fertilization in Mix, G., Wilson, H.M., and Foroughi-Wehr, B. 1978. The Quercus gambelii: the functional ovule. Am. ]. Bot. cytological status of plants of Hordeum vulgare L. 59:931-941. regenerated from microspore callus. Z. Pflanzen- Mogensen, H.L. 1973. Some histochemical, ultrastruczilchtg 80:89-99. tural, and nutritional aspects of the ovule of Quercus gambelii. Am. J. Bot. 60:48-54. Miyake, T., Kuroiwa, H., and Kuroiwa, T. 1995. Differential mechanisms of movement between a generative cell Mogensen, H.L. 1977. Ultrastructural analysis of female and a vegetative nucleus in pollen tubes of Nicotiana pachynema and the relationship between synaptonemal complex length and crossing-over in Zea tabacum as revealed by additions of colchicine and mays. Carlsberg Res. Commun. 42:475-497. nonanoic acid. Sex. Plant Reprod. 8:228-230. Miyamura, S., Kuroiwa, T., and Nagata, T. 1987. Mogensen, H.L. 1978a. Synergids of Proboscidea louisianica before fertilization. Phytomorphology 28:114-122. Disappearance of plastid and mitochondrial nucleoids during the formation of generative cells of Mogensen, H.L. 1978b. Pollen tube-synergid interactions in Proboscidea louisianica (Martineaceae). Am. ]. Bot. higher plants revealed by fluorescence microscopy. Protoplasma 141:149-159. 65:953-964. Miyoshi, H., Usami, T., and Tanaka, I. 1995. High level of Mogensen, H.L. 1981a. Ultrastructural localization of GUS gene expression driven by pollen-specific proadenosine triphosphatase in the ovules of Saintpaulia moters in electroporated lily pollen protoplasts. Sex. ionantha (Gesneriaceae) and its relation to synergid Plant Reprod. 8:205-209. function and embryo sac nutrition. Am. ]. Bot. 68:183-194. Mizelle, M.B., Sethi, R., Ashton, M.E., and Jensen, W.A. 1989. Development of the pollen grain and tapetum Mogensen, H.L. 1981b. Translocation of uranin within the of wheat {Triticwn aestivum) in untreated plants and living ovules of selected species. Am. ]. Bot. plants treated with chemical hybridizing agent 68:195-199. RH0007. Sex. Plant Reprod. 2:231-253. Mogensen, H.L. 1982. Double fertilization in barley and the cytological explanation for haploid embryo forMizukami, Y., and Ma, H. 1992. Ectopic expression of the mation, embryoless caryopses, and ovule abortion. floral homeotic gene AGAMOUS in transgenic Arabidopsis plants alters floral organ identity. Cell Carlsberg Res. Commun. 47:313-354. 71:119-131. Mogensen, H.L. 1984. Quantitative observations on the Mizukami, Y., and Ma, H. 1995. Separation of AG function pattern of synergid degeneration in barley. Am. ]. in floral meristem determinacy from that in reproBot. 71:1448-1451. ductive organ identity by expressing antisense AG Mogensen, H.L. 1985. Ultracytochemical localization of RNA. Plant Mol. Biol. 28:767-784. plasma membrane-associated phosphatase activity Mo, L.H., and von Arnold, S. 1991. Origin and development in developing tobacco seeds. Am. ]. Bot. 72:741-754. of embryogenic cultures from seedlings of Norway Mogensen, H.L. 1986. On the male germ unit in an spruce (Picea abies). ]. Plant Physiol. 138:223-230. angiosperm with bicellular pollen, Hippeastrum vitaMo, L.H., Egertsdotter, U., and von Arnold, S. 1996. turn. In Biotechnology and Ecology of Pollen, ed. D.L. Secretion of specific extracellular proteins by Mulcahy, G.B. Mulcahy and E. Ottaviano, pp. somatic embryos of Picea abies is dependent on 297-305. New York: Springer-Verlag. embryo morphology. Ann. Bot. 77:143-152. Mogensen, H.L. 1988. Exclusion of male mitochondria Mo, Y, Nagel, C, and Taylor, L.P. 1992. Biochemical comand plastids during syngamy in barley as a basis for plementation of chalcone synthase mutants defines a maternal inheritance. Proc. Natl Acad. Sci. USA role for flavonols in functional pollen. Proc. Natl 85:2594-2597. Acad. Sci. USA 89:7213-7217. Mogensen, H.L. 1992. The male germ unit: concept, comMo, Y.-S., and Yang, H.-Y. 1991. Mass isolation and preserposition, and significance. Int. Rev. Cytol. vation of viable sperm cells in Brassica campestris var. 140:129-147. purpurea. Ada Bot. Sinica 33:649-657. Mogensen, H.L. 1996. The hows and whys of cytoplasmic inheritance in seed plants. Am. ]. Bot. 83:383-404. Mo, Y.-S., and Yang, H.-Y. 1992. Isolation and fusion of sperm cells in several bicellular pollen species. Ada Mogensen, H.L., and Holm, P.B. 1995. Dynamics of Bot. Sinica 34:688-697. nuclear DNA quantities during zygote development Modrusan, Z., Reiser, L., Feldmann, K.A., Fischer, R.L., in barley. Plant Cell 7:487-494. and Haughn, G.W. 1994. Homeotic transformation Mogensen, H.L., and Pollak, P.E. 1983. Mg2+, ^-depenof ovules into carpel-like structures in Arabidopsis. dent phosphatase activity during seed development in Nicotiana tabacum. Am. J. Bot. 70:1105-1108. Plant Cell 6:333-349.
670
References
Mogensen, H.L., and Rusche, M.L. 1985. Quantitative ultrastructural analysis of barley sperm. I. Occurrence and mechanism of cytoplasm and organelle reduction and the question of sperm dimorphism. Protoplasma 128:1-13. Mogensen, H.L., and Suthar, H.K. 1979. Ultrastructure of the egg apparatus of Nicotiana tabacum (Solanaceae) before and after fertilization. Bot. Gaz. 140:168-179. Mogensen, H.L., and Wagner, V.T. 1987. Associations among components of the male germ unit following in vivo pollination in barley. Protoplasma 138:161-172. Mogensen, H.L., Wagner, V.T., and Dumas, C. 1990. Quantitative, three-dimensional ultrastructure of isolated corn (Zea mays) sperm cells. Protoplasma 153:136-140. Mogensen, H.L., Leduc, N., Matthys-Rochon, E., and Dumas, C. 1995. Nuclear DNA amounts in the egg and zygote of maize (Zea mays L.). Planta 197:641-645. Mohan Ram, H.Y., and Jaiswal, V.S. 1971. Feminization of male flowers of Cannabis sativa L. by a morphactin.
Monfort, S. 1985. Androgenesis of coconut: embryos from anther culture. Z. Pflanzenzilchtg 94:251-254. Monnier, M. 1975. Action d'un gel de polyacrylamide employe' comme support pour la culture de l'embryon immature de Capsella bursa-pastoris. Compt. Rend. Acad. Sci. Paris 280D:705-708.
Monnier, M. 1976a. Culture in vitro de l'embryon immature de Capsella bursa-pastoris Moench. Rev. Cytol. Biol. Veget. 39:1-120.
Monnier, M. 1976b. Action de la pression partielle d'oxygene sur le deVeloppement de l'embryon de Capsella bursa-pastoris cultiv£ in vitro. Compt. Rend. Acad. Sci. Paris 282D:1009-1012. Monnier, M. 1984. Survival of young immature Capsella
embryos cultured in vitro. ]. Plant Physiol. 115:105-113. Montague, M.J., Armstrong, T.A., and Jaworski, E.G. 1979. Polyamine metabolism in embryogenic cells of Daucus carota. II. Changes in arginine decarboxylase activity. Plant Physiol. 63:341-345. Montague, M.J., Koppenbrink, J.W., and Jaworski, E.G. Naturwissenschaften 58:149-150. 1978. Polyamine metabolism in embryogenic cells of Daucus carota. I. Changes in intracellular content and Mohapatra, S.S., and Knox, R.B., ed. 1996. Pollen rates of synthesis. Plant Physiol. 62:430-433. Biotechnology. Gene Expression and Allergen Characterization. New York: Chapman & Hall. Moore, C.W., and Creech, R.G. 1972. Genetic fine structure analysis of the amylose-extender locus in Zea mays Mohapatra, S.S., Hill, R., Astwood, }., Ekramoddoullah, L. Genetics 70:611-619. A.K.M., Olsen, E., Silvanovitch, A., Hatton, T., Kisil, F., and Sehon, A.H. 1990. Isolation and characteriza- Moore, H.M., and Nasrallah, J.B. 1990. A Brassica selftion of a cDNA clone encoding an IgE-binding proincompatibility gene is expressed in the stylar transtein from Kentucky bluegrass (Poa pratensis) pollen. mitting tissue of transgenic tobacco. Plant Cell Int. Arch. Allergy Appl. Immunol. 91:362-368. 2:29-38. Mohr, S., Schulte-Kappert, E., Oldenbach, W., Oettler, G., Moore, P.J., Moore, A.J., and Collins, G.B. 1994. Genotypic and Kiick, U. 1993. Mitochondrial DNA of cytoplasand developmental regulation of transient expresmic male-sterile Tritkum timopheevi: rearrangement sion of a reporter gene in soybean zygotic cotyleof upstream sequences of the atp6 and or/25 genes. dons. Plant Cell Rep. 13:556-560. Theor. Appl. Genet. 86:259-268. Moreira, M.A., Hermodson, M.A., Larkins, B.A., and Mok, D.W.S., Mok, M.C., and Rabakoarihanta, A. 1978. Nielsen, N.C. 1979. Partial characterization of the Interspecific hybridization of Phaseolus vulgaris with acidic and basic polypeptides of glycinin. /. Biol. P. lunatus and P. acutifolius. Theor. Appl. Genet. Chem. 254:9921-9926. 52:209-215. Moreira, M.A., Hermodson, M.A., Larkins, B.A., and M61, R. 1986. Isolation of protoplasts from female gameNielsen, N.C. 1981. Comparison of the primary tophytes of Torenia fournieri. Plant Cell Rep. structure of the acidic polypeptides of glycinin. Arch. Biochem. Biophys 210:633-642. 5:202-206. M61, R., Betka, A., and Wojciechowicz, M. 1995. Induction Moreno, J., and Chrispeels, M.J. 1989. A lectin gene of autonomous endosperm in Lupinus luteus, encodes the a-amylase inhibitor of the common Helleborus niger and Melandrium album by in vitro cul-
ture of unpollinated ovaries. Sex. Plant Reprod. 8:273-277. M61, R., Matthys-Rochon, E., and Dumas, C. 1993. In vitro culture of fertilized embryo sacs of maize: zygotes and two-celled proembryos can develop into plants. Planta 189:213-217. M61, R., Matthys-Rochon, E., and Dumas, C. 1994. The kinetics of cytological events during double fertilization in Zea mays L. Plant ]. 5:197-206. M61, R., Matthys-Rochon, E., and Dumas, C. 1995. Embryogenesis and plant regeneration from maize zygotes by in vitro culture of fertilized embryo sacs. Plant Cell Rep. 14:743-747.
Moneger, F., Smart, C.J., and Leaver, C.J. 1994. Nuclear restoration of cytoplasmic male sterility in sunflower is associated with the tissue-specific regulation of a novel mitochondrial gene. EMBO ]. 13:8-17. Moneger, E, Mandaron, P., Niogret, M.-F., Freyssinet, G., and Mache, R. 1992. Expression of chloroplast and mitochondrial genes during microsporogenesis in maize. Plant Physiol. 99:396-100.
bean. Proc. Natl Acad. Sci. USA 86:7885-7889.
Morgan, J.M. 1980. Possible role of abscisic acid in reducing seed set in water-stressed wheat plants. Nature 285:655-657. Morris, D.A. 1978. Germination inhibitors in developing seeds of Phaseolus vulgaris L. Z. Pflanzenphysiol. 86:433-441. Morris, PC, Kumar, A., Bowles, D.J., and Cuming, A.C. 1990. Osmotic stress and abscisic acid induce expression of the wheat Em genes. Eur. J. Biochem. 190:625-630. Morris, PC, Maddock, S.E., Jones, M.G.K., and Bowles, D.J. 1985. Changes in the levels of wheat- and barleygerm agglutinin during embryogenesis in vivo, in vitro and during germination. Planta 166:407-413. Morris, PC, Weiler, E.W., Maddock, S.E., Jones, M.G.K., Lenton, J.R., and Bowles, D.J. 1988. Determination of endogenous abscisic acid levels in immature cereal embryos during in vitro culture. Planta 173:110-116. Morrison, I.N., Kuo, J., and O'Brien, T.P. 1975. Histochemistry and fine structure of developing wheat aleurone cells. Planta 123:105-116.
References
Morrison, I.N., and O'Brien, T.P. 1976. Cytokinesis in the developing wheat grain; division with and without a phragmoplast. Planta 130:57-67. Morrison, I.N., O'Brien, T.P., and Kuo, J. 1978. Initial cellularization and differentiation of the aleurone cells in the ventral region of the developing wheat grain. Planta 140:19-30. Morrison, R.A., and Evans, D.A. 1987. Gametoclonal variation. Plant Breed. Rev. 5:359-391. Morrison, R.A., and Evans, D.A. 1988. Haploid plants from tissue culture: new plant varieties in a shortened time frame. Biotechnology 6:684-690. Morton, CM., Lawson, D.L., and Bedinger, P. 1989. Morphological study of the maize male sterile mutant ms7. Maydica 34:239-245. Morton, H., Evans, I.M., Gatehouse, J.A., and Boulter, D. 1983. Sequence complexity of messenger RNA in cotyledons of developing pea (Pisum sativum) seed. Phytochemistry 22:807-812. Morton, R.K., Palk, B.A., and Raison, J.K. 1964. Intracellular components associated with protein synthesis in developing wheat endosperm. Biochem. J. 91:522-528. Moscatelli, A., Del Casino, C, Lozzi, L., Cai, G., Scali, M., Tiezzi, A., and Cresti, M. 1995. High molecular weight polypeptides related to dynein heavy chains in Nicotiana tabacum pollen tubes. /. Cell Sci. 108:1117-1125. Moses, M.J., and Taylor, J.H. 1955. Desoxypentose nucleic acid synthesis during microsporogenesis in Tradescantia. Expt. Cell Res. 9:474-488. Moss, G.I., and Heslop-Harrison, J. 1967. A cytochemical study of DNA, RNA, and protein in the developing maize anther. II. Observations. Ann. Bot. 31:555-572. Moss, G.I., and Heslop-Harrison, J. 1968. Photoperiod and pollen sterility in maize. Ann. Bot. 32:833-846. Motto, M., Maddaloni, M., Ponziani, G., Brembilla, M., Marotta, R., di Fonzo, N., Soave, C, Thompson, R., and Salamini, F. 1988. Molecular cloning of the o2-m5 allele of Zea mays using transposon marking. Mol. Gen. Genet. 212:488^94. Mouritzen, P., and Holm, P.B. 1995. Isolation and culture of barley megasporocyte protoplasts. Sex. Plant Reprod. 8:321-325. Mu, J.-R, Lee, H.-S., and Kao, T.-H. 1994. Characterization of a pollen-expressed receptor-like kinase gene of Petunia inflata and the activity of its encoded kinase. Plant Cell 6:709-721. Mu, J.-H., Stains, J.P., and Kao, T.-H. 1994. Characterization of a pollen-expressed gene encoding a putative pectin esterase of Petunia inflata. Plant Mol. Biol. 25:539-544. Mulcahy, D.L., and Mulcahy, G.B. 1983. Gametophytic selfincompatibility reexamined. Science 220:1247-1251. Mulcahy, D.L., Mulcahy, G.B., and Robinson, R.W. 1979. Evidence for postmeiotic genetic activity in pollen of Cucurbita species. /. Hered. 70:365-368. Mulcahy, D.L., Robinson, R.W., Ihara, M., and Kesseli, R. 1981. Gametophytic transcription for acid phosphatases in pollen of Cucurbita species hybrids. /. Hered. 72:353-354. Mulcahy, G.B., and Mulcahy, D.L. 1985. Ovarian influence on pollen tube growth, as indicated by the semivivo technique. Am. ]. Bot. 72:1078-1080. Mulcahy, G.B., and Mulcahy, D.L. 1987. Induced pollen tube directionality. Am. J. Bot. 74:1458-1459. Miiller, A.J. 1963. Embryonentest zum Nachweis rezes-
611 siver Letalfaktoren bei Arabidopsis thaliana. Biol. Zentral. 82:133-163. Miiller-Rober, B., la Cognata, U., Sonnewald, U., and Willmitzer, L. 1994. A truncated version of an ADPglucose pyrophosphorylase promoter from potato specifies guard cell-selective expression in transgenic plants. Plant Cell 6:601-612. Mullins, M.G., Tang, F.C.A., and Facciotti, D. 1990. Agrobacterium-mediated genetic transformation of grapevines: transgenic plants of Vitis rupestris Scheele and buds of Vitis vinifera L. Biotechnology 8:1041-1045. Mundy, J., and Chua, N.-H. 1988. Abscisic acid and waterstress induce the expression of a novel rice gene. EMBO ]. 7:2279-2286. Munjal, S.V., and Narayan, R.K.J. 1995. Restriction analysis of the mitochondrial DNA of male-sterile and maintainer lines of pearl millet, Pennisetum americanum L. Plant Breed. 114:256-258. Munksgaard, D., Mattsson, O., and Okkels, F.T. 1995. Somatic embryo development in carrot is associated with an increase in levels of S-adenosylmethionine, S-adenosylhomocysteine and DNA methylation. Physiol. Plant. 93:5-10. Mufioz, C.A.P., Webster, B.D., and Jernstedt, J.A. 1995. Spatial congruence between exine pattern, microtubules and endomembranes in Vigna pollen. Sex. Plant Reprod. 8:147-151. Muntz, K. 1987. Developmental control of storage protein formation and its modulation by some internal and external factors during embryogenesis in plant seeds. Biochem. Physiol. Pflanz. 182:93-116. Murai, N., Sutton, D.W., Murray, M.G., Slightom, J.L., Merlo, D.J., Reichert, N.A., Sengupta-Gopalan, C, Stock, C.A., Barker, R.F., Kemp, J.D., and Hall, T.C. 1983. Phaseolin gene from bean is expressed after transfer to sunflower via tumor-inducing plasmid vectors. Science 222:476-482. Murfett, J., Bourque, J.E., and McClure, B.A. 1995. Antisense suppression of S-RNase expression in Nicotiana using RNA polymerase II- and Ill-transcribed gene constructs. Plant Mol. Biol. 29:201-212. Murfett, J., Atherton, T.L., Mou, B., Gasser, C.S., and McClure, B.A. 1994. S-RNase expressed in transgenic Nicotiana causes S-allele-specific pollen rejection. Nature 367:563-566. Murfett, J., Cornish, E.C., Ebert, P.R., Bonig, I., McClure, B.A., and Clarke, A.E. 1992. Expression of a selfincompatibility glycoprotein (S2-ribonuclease) from Nicotiana alata in transgenic Nicotiana tabncum. Plant Cell 4:1063-1074. Murfett, J., Ebert, PR., Haring, V, and Clarke, A.E. 1995. An S-RNase promoter from Nicotiana alata functions in transgenic N. alata plants but not in Nicotiana tabacum. Plant Mol. Biol. 28:957-963. Murfett, J., Strabala, T.J., Zurek, D.M., Mou, B., Beecher, B., and McClure, B.A. 1996. S RNase and interspecific pollen rejection in the genus Nicotiana: multiple pollen-rejection pathways contribute to unilateral incompatibility between self-incompatible and selfcompatible species. Plant Cell 8:943-958. Murgia, M., and Wilms, H.J. 1988. Three-dimensional image and mitochondrial distribution in sperm cells of Euphorbia dulcis. In Plant Sperms Cells as Tools for Biotechnology, ed. H.J. Wilms and C.J. Keijzer, pp. 75-79. Wageningen: Pudoc. Murgia, M., Charzyrlska, M., Rougier, M., and Cresti, M.
672
References
1991a. Secretory tapetum of Brassica oleracea L.: agriculturally important lupin species (Lupinus polarity and ultrastructural features. Sex. Plant angustifolius, L. albus, L. mutabilis). Plant Cell Tissue Reprod. 4:28-35. Organ Cult. 28:19-25. Murgia, M., Detchepare, S., van Went, J.L., and Cresti, M. Nagahashi, J., and Beevers, L. 1978. Subcellular localization of glycosyl transferases involved in glycopro1991b. Brassica napus pollen development during generative cell and sperm cell formation. Sex. Plant tein biosynthesis in the cotyledons of Pisum sativum L. Plant Physiol. 61:451-i59. Reprod. 4:176-181. Murgia, M., Huang, B.-Q., Tucker, S.C., and Musgrave, Nagata, R., Wada, M., Satoh, S., Kamada, H., and Harada, M.E. 1993. Embryo sac lacking antipodal cells in H. 1993. A cDNA differentially expressed between Arabidopsis thaliana (Brassicaceae). Am. J. Bot. embryogenic and nonembryogenic carrot (Daucus 80:824-838. carota L.) cells. Plant Physiol. 103:1021. Murgia, M., Wilms, H.J., Cresti, M., and Cesca, G. 1986. Nagato, Y. 1978. Analyses on the growth of embryo in Infrastructure of pollen development in Euphorbia some Gramineae. Jap. }. Breed. 28:97-105. dulcis L. 1. Diploid plants. Ada Bot. Need. 35:405-424. Nagato, Y, 1979. Incorporation of 3H-uridine and 3HMurphy, D.J., and Cummins, I. 1989a. Purification and leucine during early embryogenesis of rice and barimmunogold localisation of the major oil-body ley in caryopsis culture. Plant Cell Physiol. membrane protein of oilseed rape. Plant Sci. 20:765-773. 60:47-54. Nagato, Y., Kitano, Y, Kamijima, O., Kikuchi, S., and Murphy, D.J., and Cummins, I. 1989b. Biosynthesis of Satoh, H. 1989. Developmental mutants showing seed storage products during embryogenesis in abnormal organ differentiation in rice embryos. rapeseed, Brassica napus. J. Plant Physiol. 135:63-69. Theor. Appl. Genet. 78:11-15. Murphy, D.J., Cummins, I., and Kang, A.S. 1989. Synthesis Nagl, W. 1967. Die Riesenchromosomen von Phaseolus coccineus L.: Baueigentumlichkeiten, Strukturmodifikationen, of the major oil-body membrane protein in developzusatzliche Nukleolen und Vergleich mit den mitoing rapeseed (Brassica napus) embryos. Biochem. J. tischen Chromosomen. Osterr. Bot. Z. 114:171-182. 258:285-293. Murphy, D.J., Cummins, I., and Ryan, AJ. 1989. Nagl, W. 1969a. Banded polytene chromosomes in the Immunocytochemical and biochemical study of the legume Phaseolus vulgaris. Nature 221:70-71. biosynthesis and mobilisation of the major seed stor- Nagl, W. 1969b. Puffing of polytene chromosomes in a plant age proteins of Brassica napus. Plant Physiol. Biochem. (Phaseolus vulgaris).Naturwissenscliaften 56:221-222. 27:647-657. Nagl, W. 1974. The Phaseolus suspensor and its polytene chromosomes. Z. Pflanzenphysiol. 73:1-44. Murphy, J.J., and Dalby, A. 1971. Changes in the protein fractions of developing normal and opaque-2 maize Nagl, W. 1976a. The polytenic antipodal cells in Scilla bifoendosperm. Cereal Chem. 48:336-349. lia: DNA replication pattern and possibility of nucleMurray, B.G. 1990. Heterostyly and pollen-tube interacolar DNA amplification. Cytobiologie 14:165-170. tions in Luculia gratissima (Rubiaceae). Ann. Bot. Nagl, W. 1976b. Early embryogenesis in Tropaeolum majus 65:691-698. L.: ultrastructure of the embryo-suspensor. Biochem. Murray, M.G., and Kennard, W.C. 1984. Altered chroPhysiol. Pflanz. 170:253-260. matin conformation of the higher plant gene phase- Nagl, W. 1976c. Ultrastructural and developmental olin. Biochemistry 23:4225-4232. aspects of autolysis in embryo-suspensors. Ber. Deut. Bot. Ges. 89:301-311. Murty, N.S.R., and Lakshmi, N. 1979. Male sterile mutant Nagl, W. 1977. "Plastolysomes" - plastids involved in the in Capsicum annuum L. Curr. Sci. 48:312. Muschietti, J., Dircks, L., Vancanneyt, G., and McCormick, autolysis of the embryo-suspensor in Phaseolus. Z. Pflanzenphysiol. 85:45-51. S. 1994. LAT52 protein is essential for tomato pollen development: pollen expressing antisense LAT52 Nagl, W. 1978. Endopolyploidy and Polyteny in RNA hydrates and germinates abnormally and canDifferentiation and Evolution. Amsterdam: Northnot achieve fertilization. Plant J. 6:321-338. Holland Publishing Co. Musgrave, M.E., Antonovics, J., and Siedow, J.N. 1986. Is Nagl, W. 1990. Translocation of putrescine in the ovule, male-sterility in plants related to lack of cyanidesuspensor and embryo of Phaseolus coccineus. J. Plant resistant respiration in tissues? Plant Sci. 44:7-11. Physiol. 136:587-591. Mutschler, M.A., Bliss, F.A., and Hall, T.C. 1980. Variation Nagl, W. 1992. The polytenic endosperm haustorium of in the accumulation of seed storage protein among Rhinanthus minor (Scrophulariaceae): functional genotypes of Phaseolus vulgaris (L.). Plant Physiol. ultrastructure. Can. J. Bot. 70:1997-2004. 65:627-630. Nagl, W., and Kiihner, S. 1976. Early embryogenesis in Tropaeolum majus L.: diversification of plastids. Nacken, W.K.F., Huijser, P., Beltran, J.-P, Saedler, H., and Planta 133:15-19. Sommer, H. 1991a. Molecular characterization of Naito, S., Dube, PH., and Beachy, R.N. 1988. Differential two stamen-specific genes, tapl and fill, that are expression of conglycinin a' and |3 subunit genes in expressed in the wild type, but not in the deficiens transgenic plants. Plant Mol. Biol. 11:109-123. mutant of Antirrhinum majus. Mol. Gen. Genet. Naito, S., Hirai, M.Y., Chino, M., and Komeda, Y. 1994a. 229:129-136. Expression of a soybean (Glycine max [L.] Merr.) seed storage protein gene in transgenic Arabidopsis Nacken, W.K.F., Huijser, P., Saedler, H., and Sommer, H. 1991b. Molecular analysis of tap2, an anther-specific thaliana and its response to nutritional stress and to gene from Antirrhinum majus. FEBS Lett. 280:155-158. abscisic acid mutations. Plant Physiol. 104:497-503. Nadeau, J.A., Zhang, X.S., Li, J., and O'Neill. S.D. 1996. Naito, S., Inaba-Higano, K., Kumagai, T., Kanno, T., Ovule development: identification of stage-specific Nambara, E., Fujiwara, T., Chino, M., and Komeda, and tissue-specific cDNAs. Plant Cell 8:213-239. Y. 1994b. Maternal effects of mtol mutation, that causes overaccumulation of soluble methionine, on Nadolska-Orczyk, A. 1992. Somatic embryogenesis of
References
613
the expression of a soybean (3-conglycinin gene pro- Nasrallah, J.B., and Nasrallah, M.E. 1984. Electrophoretic heterogeneity exhibited by the S-allele specific glymoter-GUS fusion in transgenic Arabidopsis thaliana. coproteins of Brassica. Experientia 40:279-281. Plant Cell Physiol. 35:1057-1063. Nakagawa, H., Ohmiya, Kv and Hattori, T. 1996. A rice Nasrallah, J.B., and Nasrallah, M.E. 1989. The molecular genetics of self-incompatibility in Brassica. Annu. bZIP protein, designated OSBZ8, is rapidly induced Rev. Genet. 23:123-139. by abscisic acid. Plant J. 9:217-227. Nakajima, T. 1962. Physiological studies of seed develop- Nasrallah, J.B., and Nasrallah, M.E. 1993. Pollen-stigma signaling in the sporophytic self-incompatibility ment, especially embryonic growth and endosperm response. Plant Cell 5:1325-1335. development. Bull. Univ. Osaka Pref. Ser. B 13:13-48. Nakamura, N., Sado, M., and Arai, Y. 1980. Sucrose Nasrallah, J.B., Nishio, T., and Nasrallah, M.E. 1991. The self-incompatibility genes of Brassica: expression and metabolism during the growth of Camellia japonica use in genetic ablation of floral tissues. Annu. Rev. pollen. Phytochemistry 19:205-209. Plant Physiol. Plant Mol. Biol. 42:393^122. Nakamura, N., and Suzuki, H. 1981. Sugar composition of pollen grain and pollen tube cell walls. Nasrallah, J.B., Rundle, S.J., and Nasrallah, M.E. 1994. Genetic evidence for the requirement of the Brassica Phytochemistry 20:981-984. S-locus receptor kinase gene in the self-incompatiNakamura, N., Fukushima, A., Iwayama, H., and Suzuki, bility response. Plant ]. 5:373-384. H. 1991. Electrotropism of pollen tubes of camellia Nasrallah, J.B., Yu, S.-M., and Nasrallah, M.E. 1988. Selfand other plants. Sex. Plant Repwd. 4:138-143. incompatibility genes of Brassica oleracea: expression, Nakamura, S. 1979. Development of the pollen grain wall in Lilium longiflorum. ]. Electron Micros. 28:275-284. isolation and structure. Proc. Natl Acad. Sci. USA Nakamura, S., and Miki-Hirosige, H. 1982. Coated vesi85:5551-5555. cles and cell plate formation in the microspore Nasrallah, J.B., Kao, T.-H., Chen, C.-H., Goldberg, M.L., mother cell. /. Ultrastr. Res. 80:302-311. and Nasrallah, M.E. 1987. Amino-acid sequence of glycoproteins encoded by three alleles of the S locus Nakamura, S., and Miki-Hirosige, H. 1985. Fine-structural of Brassica oleracea. Nature 326:617-619. study on the formation of the generative cell wall and intine-3 layer in a growing pollen grain of Lilium Nasrallah, J.B., Kao, T.-H., Goldberg, M.L., and Nasrallah, longiflorum. Am. J. Bot. 72:365-375. M.E. 1985. A cDNA clone encoding an S-locus-specific glycoprotein from Brassica oleracea. Nature Nakamura, S., Ikehara, T., Uchida, H., Suzuki, T., and 318:263-267. Sodmergen. 1992. Fluorescence microscopy of plastid nucleoids and a survey of nuclease C in higher Nasrallah, J.B., Stein, J.C., Kandasamy, M.K., and plants with respect to mode of plastid inheritance. Nasrallah, M.E. 1994. Signaling the arrest of pollen Protoplasma 169:68-74. tube development in self-incompatible plants. Science 266:1505-1508. Nakanishi, T., Esashi, Y, and Hinata, K. 1969. Control of self-incompatibility by CO2 gas in Brassica. Plant Cell Nasrallah, M.E. 1974. Generic control of quantitative variation in self-incompatibility proteins detected by Physiol. 10:925-927. immunodiffusion. Genetics 76:45-50. Nakanishi, T., and Hinata, K. 1973. An effective time for CO2 gas treatment in overcoming self-incompatibil- Nasrallah, M.E. 1979. Self-incompatibility antigens and S ity in Brassica. Plant Cell Physiol. 14:873-879. gene expression in Brassica. Heredity 43:259-263. Nakanishi, T.M., Saeki, N., Maeno, M., Ozaki, T., Kawai, Nasrallah, M.E., Barber, J.T., and Wallace, D.H. 1970. Selfincompatibility proteins in plants: detection, genetY., and Ichii, T. 1991. Ultrastructural study on the ics, and possible mode of action. Heredity 25:23-27. stylar transmitting tissue in Japanese pear. Sex. Plant Reprod. 4:95-103. Nasrallah, M.E., Kandasamy, M.K., and Nasrallah, J.B. 1992. A genetically defined trans-acting locus reguNakano, M., Hoshino, Y, and Mii, M. 1994. Regeneration lates S-locus function in Brassica. Plant /. 2:497-506. of transgenic plants of grapevine (Vitis vinifera L.) via Agrobacterium r/iizogenes-mediated transformation of Nasrallah, M.E., and Nasrallah, J.B. 1986. Molecular biolembryogenic calli. /. Expt. Bot. 45:649-656. ogy of self-incompatibility in plants. Trends Genet. 2:239-244. Nambara, E., Naito, S., and McCourt, P. 1992. A mutant of Arabidopsis which is defective in seed development Nasrallah, M.E., and Wallace, D.H. 1967a. Immunoand storage protein accumulation is a new abi3 genetics of self-incompatibility in Brassica oleracea L. allele. Plant J. 2:435-441. Heredity 22:519-527. Nambara, E., Keith, K., McCourt, P., and Naito, S. 1995. A Nasrallah, M.E., and Wallace, D.H. 1967b. Immunoregulatory role for the AB13 gene in the establishchemical detection of antigens in self-incompatibilment of embryo maturation in Arabidopsis thaliana. ity genotypes of cabbage. Nature 213:700-701. Nasrallah, M.E., Wallace, D.H., and Savo, R.M. 1972. Development 121:629-636. Genotype, protein, phenotype relationships in selfNarayanaswamy, S., and George, L. 1972. Morphogenesis incompatibility of Brassica. Genet. Res. 20:151-160. of belladonna (Atropa belladonna L.) plantlets from pollen in culture. Indian J. Expt. Bid. 10:382-384. Natesh, S., and Rau, M.A. 1984. The embryo. In Nasatir, M., and Stern, H. 1959. Changes in the activities Embryology of Angiosperms, ed. B.M. Johri, pp. of aldolase and D-glyceraldehyde-3-phosphate 377-443. Berlin: Springer-Verlag. dehydrogenase during the mitotic cycle in Nath, J., and Watson, C.V. 1980. Acid phosphatase microspores of Lilium longiflorum.}. Biophys. Biochem. changes associated with development of male sterile Cytol. 6:189-192. and fertile maize (Zea mays L.). Biochem. Genet. Nasrallah, J.B., Doney, R.C., and Nasrallah, M.E. 1985. 18:377-387. Biosynthesis of glycoproteins involved in the Naumova, T.N. 1993. Apomixis in Angiosperms. Nucellar pollen-stigma interaction of incompatibility in and Integumentary Embryony. Boca Raton: CRC Press. developing flowers of Brassica oleracea L. Planta Naumova, T.N., and Willemse, M.T.M. 1995. Ultrastructural characterization of apospory in 165:100-107.
614
References
Panicum maximum. Sex. Plant Reprod. 8:107-204. A., Blagrove, R.J., March, J.F., Kortt, A.A., and Nave, E.B., and Sawhney, V.K. 1986. Enzymatic changes Higgins, T.J.V. 1990. Pea convicilin: structure and in post-meiotic anther development in Petunia primary sequence of the protein and expression of a hybrida. I. Anther ontogeny and isozyme analyses. /. gene in the seeds of transgenic tobacco. Planta Plant Physiol. 125:451^65. 180:461-170. Nawaschin, S.G. 1898. Resultate einer Revision der Newcomb, W. 1973a. The development of the embryo sac Befruchtungsvorgange bei Lilium martagon und of sunflower Helianthus annuus before fertilization. Fritillaria tenella. Bull. Acad. Imp. Sci. St. Petersburg Can. J. Bot. 51:863-878. 9:377-382. Newcomb, W. 1973b. The development of the embryo sac of sunflower Helianthus annuus after fertilization. Negruk, V.I., Cherny, D.I., Nififorova, I.D., Aleksandrov, A.A., and Butenko, R.G. 1982. Isolation and characCan. J. Bot. 51:879-890. terization of minicircular DNAs found in mitochon- Newcomb, W. 1978. The development of cells in the drial fraction of Viciafaba. FEBS Lett. 142:115-117. coenocytic endosperm of the African blood lily Haemanthus katherinae. Can. J. Bot. 56:483-501. Nehra, N.S., Chibbar, R.N., Leung, N., Caswell, K., Mallard, C, Steinhauer, L., Baga, M., and Kartha, Newcomb, W., and Fowke, L.C. 1973. The fine structure of K.K. 1994. Self-fertile transgenic wheat plants regenthe change from the free-nuclear to cellular condierated from isolated scutellar tissues following tion in the endosperm of chickweed Stellaria media. microprojectile bombardment with two distinct gene Bot. Gaz. 134:236-241. constructs. Plant ]. 5:285-297. Newcomb, W., and Fowke, L.C. 1974. Stellaria media Neill, J.D., Litts, J.C., Anderson, O.D., Greene, F.C., and embryogenesis: the development and ultrastructure Stiles, J.I. 1987. Expression of a wheat a-gliadin gene of the suspensor. Can. ]. Bot. 52:607-614. in Saccharomyces cerevisiae. Gene 55:303-317. Newcomb, W., and Steeves, T.A. 1971. Helianthus annuus embryogenesis: embryo sac wall projections before Neill, S.J., Horgan, R., and Parry, A.D. 1986. The and after fertilization. Bot. Gaz. 132:367-371. carotenoid and abscisic acid content of viviparous kernels and seedlings of Zea mays L. Planta 169:87-96. Newton, C.H., Flinn, B.S., and Sutton, B.C.S. 1992. VicilinNeill, S.J., Horgan, R., and Rees, A.F. 1987. Seed developlike seed storage proteins in the gymnosperm intement and vivipary in Zea mays L. Planta 171:358-364. rior spruce (Picea glauca/engelmanii). Plant Mol. Biol. Nelson, O.E., Chourey, P.S., and Chang, M.T. 1978. 20:315-322. Nucleoside diphosphate sugar-starch glucosyl trans- Ng, J.D., Ko, T.-Z., and McPherson, A. 1993. Cloning, ferase activity of wx starch granules. Plant Physiol. expression, and crystallization of jack bean 62:383-386. (Canavalia ensiformis) canavalin. Plant Physiol. 101:713-728. Nelson, O.E., Mertz, E.T., and Bates, L.S. 1965. Second mutant gene affecting the amino acid pattern of Nielsen, K.A., and Hansen, I.B. 1992. Appearance of extramaize endosperm proteins. Science 150:1469-1470. cellular proteins associated with somatic embryogeNelson, O.E., and Rines, H.W. 1962. The enzymatic definesis in suspension cultures of barley (Hordeum ciency in the waxy mutant of maize. Biochem. vulgare L.). /. Plant Physiol. 139:489-497. Biophys. Res. Commun. 9:297-300. Nielsen, N.C., Dickinson, CD., Cho, T.-J., Thanh, V.H., Nelson, O.E., and Tsai, C.Y. 1964. Glucose transfer from Scallon, B.J., Fischer, R.L., Sims, T.L., Drews, G.N., adenosine diphosphate-glucose to starch in preparaand Goldberg, R.B. 1989. Characterization of the tions of waxy seeds. Science 145:1194-1195. glycinin gene family in soybean. Plant Cell 1:313-328. Nelson, P.M., and Rossman, E.C. 1958. Chemical induc- Niimi, Y. 1970. In vitro fertilization in the self-incompatition of male sterility in inbred maize by use of gibble plant, Petunia hybrida. J. Jap. Soc. Hort. Sci. berellins. Science 127:1500-1501. 39:346-352. Nepi, M., Ciampolini, F., and Pacini, E. 1995. Niitsu, T., Hanaoka, A., and Uchiyama, K. 1972. Development of Cucurbita pepo pollen: ultrastructure Reinvestigations of the spindle body and related and histochemistry of the sporoderm. Can. J. Bot. problems. II. Effects of cycloheximide upon the 73:1046-1057. development of kinetochore fibers studied in the pollen mitosis of Ornithogalum virens in vivo. Neto, G.C., Yunes, J.A., da Silva, M.J., Vettore, A.L., Cytologia 37:143-154. Arruda, P., and Leite, A. 1995. The involvement of opaque 2 on pS-prolamin gene regulation in maize Niizeki, H., and Oono, K. 1968. Induction of haploid rice and Coix suggests a more general role for this tranplant from anther culture. Proc. ]ap. Acad. 44:554-557. scriptional activator. Plant Mol. Biol. 27:1015-1029. Ninnemann, H., and Epel, B. 1973. Inhibition of cell diviNeuffer, M.G., and Sheridan, W.F. 1980. Defective kernel sion by blue light. Expt. Cell Res. 79:318-326. mutants of maize. I. Genetic and lethality studies. Niogret, M.F., Culianez-Macia, Goday, A., Alba, M.M., Genetics 95:929-944. and Pages, M. 1996. Expression and cellular localizaNeuhaus, G., Spangenberg, G., Scheid, O.M., and tion of rab28 mRNA and RAB28 protein during Schweiger, H.-G. 1987. Transgenic rapeseed plants maize embryogenesis. Plant ]. 9:549-557. obtained by the microinjection of DNA into Niogret, M.-F., Dubald, M., Mandaron, P., and Mache, R. microspore-derived embryoids. Theor. Appl. Genet. 1991. Characterization of pollen polygalacturonase 75:30-36. encoded by several cDNA clones in maize. Plant Neumann, D., and Weber, E. 1978. Formation of protein Mol. Biol. 17:1155-1164. bodies in ripening seeds of Vicia faba L. Biochem. Nirmala, C, and Kaul, M.L.H. 1994. Male sterility in pea. Physiol. Pflanz. 173:167-180. V. Gene action during heterotypic and homotypic Newbigin, E., Anderson, M.A., and Clarke, A.E. 1993. divisions. Cytologia 59:43-50. Gametophytic self-incompatibility systems. Plant Nishihara, M., Ito, M., Tanaka, I., Kyo, M., Ono, K., Cell 5:1315-1324. Irifune, K., and Morikawa, H. 1993. Expression of the p-glucuronidase gene in pollen of lily (Lilium Newbigin, E.J., deLumen, B.O., Chandler, P.M., Gould,
References
615
longiflorum), tobacco (Nicotiana tabacum), Nicotiana megagametogenesis in two species of Oenothera. rustica, and peony (Paeonia lactiflora) by particle Ann. Bot. 46:473-477. bombardment. Plant Physiol. 102:357-361. Noher de Halac, I., Cismondi, I.A., and Harte, C. 1990. Pollen ontogenesis in Oenothera: a comparison of Nishio, T., and Hinata, K. 1977. Analysis of S-specific progenotypically normal anthers with the male-sterile teins in stigma of Brassica olemcea L. by isoelectric mutant sterilis. Sex. Plant Reprod. 3:41-53. focusing. Heredity 38:391-396. Nishio, T., and Hinata, K. 1982. Comparative studies on S- Noher de Halac, I., and Harte, C. 1977. Different patterns glycoproteins purified from different S-genotypes in of callose wall formation during megasporogenesis self-incompatible Brassica species. I. Purification and in two species of Oenothera (Onagraceae). Plant Syst. Evol. 127:23-38. chemical properties. Genetics 100:641-647. Nishio, T., Toriyama, K., Sato, T., Kandasamy, M.K., Noma, M., Huber, J., Ernst, D., and Pharis, R.P. 1982. Paolillo, D.J., Nasrallah, J.B., and Nasrallah, M.K. Quantitation of gibberellins and the metabolism of [3H]gibberellin A, during somatic embryogenesis 1992. Expression of S-locus glycoprotein genes from Brassica oleracea and B. campestris in transgenic plants in carrot and anise cell cultures. Planta of self-compatible B. napus cv Westar. Sex. Plant 155:369-376. Reprod. 5:101-109. Nomura, K., and Komamine, A. 1985. Identification and isolation of single cells that produce somatic Nishizawa, N.K., Mori, S., Watanabe, Y, and Hirano, H. embryos at a high frequency in a carrot suspension 1994. Ultrastructural localization of the basic 7S culture. Plant Physiol. 79:988-991. globulin in soybean (Glycine max) cotyledons. Plant Cell Physiol. 35:1079-1085. Nomura, K., and Komamine, A. 1986a. Polarized DNA Nissen, P. 1994. Stimulation of somatic embryogenesis in synthesis and cell division in cell clusters during carrot by ethylene: effects of modulators of ethylsomatic embryogenesis from single carrot cells. Neiu Phytol. 104:25-32. ene biosynthesis and action. Physiol. Plant. 92:397-403. Nomura, K., and Komamine, A. 1986b. In situ hybridization on tissue sections. A method for detection of Nitsch, C. 1974a. Pollen culture - a new technique for poly(A)+RNA. Plant Tissue Cult. Lett. Japan 3:92-94. mass production of haploid and homozygous plants. In Haploids in Higher Plants. Advances and Nomura, K., and Komamine, A. 1995. Physiological and biochemical aspects of somatic embryogenesis. In In Potential, ed. KJ. Kasha, pp. 123-135. Guelph: The Vitro Embryogenesis in Plants, ed. T.A. Thorpe, pp. University of Guelph. 249-265. Dordrecht: Kluwer Academic Publishers. Nitsch, C. 1974b. La culture de pollen isol£ sur milieu synth&ique. Compt. Rend. Acad. Set Paris 278D:1031-1034. Nong, V.H., Schlesier, B., Bassiiner, R., Repik, A., Horstmann, C, and Miintz, K. 1995. Narbonin, a Nitsch, C, and Norreel, B. 1973. Effet d'un choc thernovel 2S protein from Vicia narbonensis L. seeds: mique sur le pouvoir embryogene du pollen de Datura innoxia cultiv6 dans l'anthere ou isole de l'ancDNA, gene structure and developmentally reguthere. Compt. Rend. Acad. Sci. Paris 276D:303-306. lated formation. Plant Mol. Biol. 28:61-72. Nitsch, J.P. 1972. Haploid plants from pollen. Z. Norgaard, J.V., and Krogstrup, P. 1991. Cytokinin induced somatic embryogenesis from immature embryos of Pflanzenzuchtg 67:3-18. Abies nordmanniana Lk. Plant Cell Rep. 9:509-513. Nitsch, J.P., and Nitsch, C. 1969. Haploid plants from pollen grains. Science 163:85-87. Norreel, B. 1972. Etude comparative de la repartition des Nivison, H.T., and Hanson, M.R. 1989. Identification of a acides ribonucl&ques au cours de 1'embryogenese mitochondrial protein associated with cytoplasmic zygotique et de 1'embryogenese androge'ne'tique male sterility in Petunia. Plant Cell 1:1121-1130. chez le Nicotiana tabacum L. Compt. Rend. Acad. Sci. Nivison, H.T., Sutton, C.A., Wilson, R.K., and Hanson, Paris 275D:1219-1222. M.R. 1994. Sequencing, processing, and localization Norstog, K. 1961. The growth and differentiation of culof the petunia cms-associated mitochondrial protein. tured barley embryos. Am. }. Bot. 48:876-884. Plant ]. 5:613-623. Norstog, K. 1972a. Early development of the barley Nobiling, R., and Reiss, H.-D. 1987. Quantitative analysis embryo: fine structure. Am. J. Bot. 59:123-132. of calcium gradients and activity in growing pollen Norstog, K. 1972b. Factors relating to precocious germitubes of Lilium longiflorum. Protoplasma 139:20-24. nation in cultured barley embryos. Phytomorphology Nogler, G.A. 1984. Gametophytic apomixis. In Embryology 22:134-139. of Angiosperms, ed. B.M. Johri, pp. 475-518. Berlin: Norstog, K. 1974. Nucellus during early embryogeny in Springer-Verlag. barley: fine structure. Bot. Gaz. 135:97-103. Noguchi, T. 1990. Consumption of lipid granules and for- Norstog, K., and Klein, R.M. 1972. Development of culmation of vacuoles in the pollen tube of Tradescantia tured barley embryos. II. Precocious germination reflexa. Protoplasma 156:19-28. and dormancy. Can. }. Bot. 50:1887-1894. Noguchi, X, and Morr£, D.J. 1991. Membrane flow in Norstog, K., and Smith, J.E. 1963. Culture of small barley plants: preparation and kinetics of labelling of embryos on defined media. Science 142:1655-1656. plasma membranes from growing pollen tubes of Nozue, K., and Wada, M. 1993. Electrotropism of tobacco. Protoplasma 163:34-42. Nicotiana pollen tubes. Plant Cell Physiol. Noguchi, T., and Ueda, K. 1990. Structure of pollen grains 34:1291-1296. of Tradescantia reflexa with special reference to theNunberg, A.N., Li, Z., Bogue, M.A., Vivekananda, J., generative cell and the ER around it. Cell Struct. Reddy, A.S., and Thomas, T.L. 1994. Developmental Fund. 15:379-384. and hormonal regulation of sunflower helianthinin Noher de Halac, I. 1980a. Fine structure of nucellar cells genes: proximal promoter sequences confer regionduring development of the embryo sac in Oenothera alized seed expression. Plant Cell 6:473-486. biennis L. Ann. Bot. 45:515-521. Nuti Ronchi, V., Caligo, M.A., Nozzolini, M., and Noher de Halac, I. 1980b. Callose deposition during Luccarini, G. 1984. Stimulation of carrot somatic
676
References
embryogenesis by proline and serine. Plant Cell Rep. allows endosperm development without fertiliza3:210-214. tion. Proc. Natl Acad. Sci. USA 93:5319-5324. Nuti Ronchi, V., Giorgetti, L., Tonelli, M., and Martini, G. Ohl, S., Hedrick, S.A., Chory, J., and Lamb, C.J. 1990. 1992a. Ploidy reduction and genome segregation in Functional properties of a phenylalanine ammoniacultured carrot cell lines. I. Prophase chromosome lyase promoter from Arabidopsis. Plant Cell reduction. Plant Cell Tissue Organ Cult. 30:107-114. 2:837-848. Nuti Ronchi, V., Giorgetti, L., Tonelli, M. and Martini, G. Ohshika, K., and Ikeda, H. 1994. Isolation and preserva1992b. Ploidy reduction and genome segregation in tion of the living embryo sac of Crinum asiaticum L. cultured carrot cell lines. II. Somatic meiosis. Plant var. japonicum Baker. /. Plant Res. 107:17-21. Cell Tissue Organ Cult. 30:115-119. Ohta, Y. 1986. High-efficiency genetic transformation of maize by a mixture of pollen and exogenous DNA. Nygaard, P. 1977. Utilization of exogenous carbohydrates Proc. Natl Acad. Sci. USA 83:715-719. for tube growth and starch synthesis in pine pollen suspension cultures. Physiol. Plant. 39:206-210. Ohyama, T., Iwaikawa, Y, Kobayashi, T., Hotta, Y., and Tabata, S. 1992. Isolation of synaptonemal complexes Nyman, L.P., Webb, E.L., Gu, Z., and Arditti, J. 1986. from lily microsporocytes. Plant Sci. 86:115-124. Structure and in vitro growth of zygotic embryos of taro (Colocasia esculenta var. antiquorum). Ann. Bot.Oishi, M.Y., and Bewley, J.D. 1992. Premature drying, fluri57:623-630. done-treatment, and embryo isolation during development of maize kernels (Zea mays L.) induce Nyman, L.P., Webb, E.L., Gu, Z., and Arditti, J. 1987. germination, but the protein synthetic responses are Effects of growth regulators and glutamine on in different. Potential regulation of germination and provitro development of zygotic embryos of taro tein synthesis by abscisic acid. /. Expt. Bot. 43:759-767. (Colocasia esculenta var. antiquorum). Ann. Bot. 59:517-523. Okada, K., Ueda, J., Komaki, M.K., Bell, C.J., and Shimura, Y. 1991. Requirement of the auxin polar transport system in early stages of Arabidopsis floral O'Brien, S.P. 1994. Pistil structure and pollen tube pathways in Leptospermum myrsinoides and L. continentale bud formation. Plant Cell 3:677-684. (Myrtaceae). Ann. Bot. 73:225-230. Okamoto, H., Yano, A., Shiraishi, H., Okada, K., and O'Driscoll, D., Hann, C., Read, S.M., and Steer, M.W. 1993. Shimura, Y. 1994. Genetic complementation of a floEndocytotic uptake of fluorescent dextrans by pollen ral homeotic mutation, apelalai, with an Arabidopsis tubes grown in vitro. Protoplasma 175:126-130. thaliana gene homologous to DEFICIENS of Antirrhinum majus. Plant Mol. Biol. 26:465-472. O'Keefe, D.P., Tepperman, J.M., Dean, C., Leto, K.J., Erbes, D.L., and Odell, J.T. 1994. Plant expression of a bacter- Okamuro, J.K., den Boer, B.G.W., and Jofuku, K.D. 1993. Regulation of Arabidopsis flower development. Plant ial cytochrome P450 that catalyzes activation of a sulCell 5:1183-1193. fonylurea pro-herbicide. Plant Physiol. 105:473-482. O'Neill, P., Singh, M.B., and Knox, R.B. 1988. Cell biology Okamuro, J.K., and Goldberg, R.B. 1992. A mutant lectin gene is rescued from an insertion element that blocks of the stigma of Brassica campestris in relation to CO2 its expression. Plant Cell 4:1141-1146. effects on self-pollination. /. Cell Sci. 89:541-549. O'Neill, P.M., Singh, M.B., and Knox, R.B. 1989. Okamuro, J.K., Jofuku, K.D., and Goldberg, R.B. 1986. Biosynthesis of S-associated proteins following selfSoybean seed lectin gene and flanking nonseed proand cross-pollinations in Brassica campestris L. var. tein genes are developmentally regulated in transT.15'. Sex. Plant Reprod. 2:103-108. formed tobacco plants. Proc. Natl Acad. Sci. USA 83:8240-8244. O'Neill, S.D., Nadeau, J.A., Zhang, X.S., Bui, A.Q., and Halevy, A.H. 1993. Interorgan regulation of ethylene Okita, T.W. 1984. Identification and DNA sequence analybiosynthetic genes by pollination. Plant Cell sis of a y-type gliadin cDNA plasmid from winter 5:419-432. wheat. Plant Mol. Biol. 3:325-332. Obermeyer, G., and Blatt, M.R. 1995. Electrical properties Okita, T.W., Cheesbrough, V., and Reeves, CD. 1985. Evolution and heterogeneity of the a-/P-type and yof intact pollen grains of Lilium longiflorum: charactype gliadin DNA sequences./. Biol. Chem. 260:8203-8213. teristics of the non-germinating pollen grain. /. Expt. Bot. 46:803-813. Okita, T.W., and Greene, F.C. 1982. Wheat storage proteins. Isolation and characterization of the gliadin Obermeyer, G., and Weisenseel, M.H. 1991. Calcium messenger RNAs. Plant Physiol. 69:834-839. channel blocker and calmodulin antagonists affect the gradient of free calcium ions in lily pollen tubes. Okita, T.W., Hwang, Y.S., Hnilo, J., Kim, W.T., Aryan, A.P., Larson, R., and Krishnan, H.B. 1989. Structure and Eur. J. Cell Biol. 56:319-327. expression of the rice glutelin multigene family. /. Obermeyer, G., Liitzelschwab, M., Heumann, H.-G., and + Biol. Chem. 264:12573-12581. Weisenseel, M.H. 1992. Immunolocalization of H ATPases in the plasma membrane of pollen grains Okonkwo, S.N.C., and Raghavan, V. 1982. Studies on the and pollen tubes of Lilium longiflorum. Protoplasma germination of seeds of the root parasites, Alectra 171:55-63. vogelii and Striga gesnerioides. I. Anatomical changes in the embryos. Am. }. Bot. 69:1636-1645. Ockendon, D.J. 1968. Biosystematic studies in the Linum Oldknow, J., and Trick, M. 1995. Genomic sequence of an perenne group. New Phytol. 67:787-813. SRK-Iike gene linked to the S-locus of a self-incomOckendon, D.J., and Gates, P.J. 1975. Growth of cross- and patible Brassica oleracea line. Sex. Plant Reprod. self-pollen tubes in the styles of Brassica oleracea. Neiu 8:247-253. Phytol. 75:155-160. Ogur, M., Erickson, R.O., Rosen, G.U., Sax, K.B., and Oldknow, J., Franklin, T.M., Allard, S., Robert, L.S., and Trick, M. 1995. DNA sequences of the two homoeolHolden, C. 1951. Nucleic acids in relation to cell diviogous SLR1 genes of Brassica napus cv Westar. Sex. sion in Lilium longiflorum. Expt. Cell Res. 2:73-89. Plant Reprod. 8:254-255. Ohad, N., Margossian, L., Hsu, Y.-C, Williams, C, Repetti, P., and Fischer, R.L. 1996. A mutation that Olesen, P., and Bruun, L. 1990. A structural investigation
References
617
of the ovule in sugar beet, Beta vulgaris: integuments Ottaviano, E., and Mulcahy, D.L. 1989. Genetics of angiosperm pollen. Adv. Genet. 26:1-64. and micropyle. Nord. J. Bot. 9:499-506. Olff, H v Kuiper, D., van Damme, J.M.M., and Kuiper, Ottoboni, L.M.M., Leite, A., Yunes, J.A., Targon, M.L.P.N., P.J.C. 1989. Gynodioecy in Plantago lanceolata. VI. de Souza Filho, G.A., and Arruda, P. 1993. Sequence Functions of cytokinins in growth, development, analysis of 22 kDa-like a-coixin genes and their comand reproduction of two sex types. Can. ]. Bot. parison with homologous zein and kafirin genes 67:2765-2769. reveals highly conserved protein structure and regulatory elements. Plant Mol. Biol. 21:765-778. Olmedilla, A., Schrauwen, J.A.M., and Wullems, GJ. 1991. Visualization of starch-synthase expression by in situ Ouellet, T., Rutledge, R.G., and Miki, B.L. 1992. Members hybridization during pollen development. Planta of the acetohydroxyacid synthase multigene family 184:182-186. of Brassica napus have divergent patterns of expression. Plant J. 2:321-330. Olsen, O.-A., Jakobsen, K.S., and Schmelzer, E. 1990. Development of barley aleurone cells: temporal and Ouyang, T.-W., Hu, H., Chuang, C.-C., and Tseng, C.-C. spatial patterns of accumulation of cell-specific 1973. Induction of pollen plants from anthers of mRNAs. Planta 181:462-466. Triticum aestivum L. cultured in vitro. Sci. Sinica 16:79-95. Olsen, O.-A., Potter, R.H., and Kalla, R. 1992. Histo-differentiation and molecular biology of developing cereal Overman, M.A., and Warmke, H.E. 1972. Cytoplasmic male sterility in Sorghum. II. Tapetal behavior in ferendosperm. Seed Sci. Res. 2:117-131. Olson, A.R. 1980. Seed morphology of Monotropa uniflora tile and sterile anthers. /. Hered. 63:226-234. L. (Ericaceae). Am. J. Bot. 67:968-974. Owen, H.A., and Makaroff, C.A. 1995. Ultrastructure Olson, A.R. 1991. Postfertilization changes in ovules of of microsporogenesis and microgametogenesis Monotropa uniflora L. (Monotropaceae). Am. ]. Bot. in Arabidopsis thaliana (L.) Heynh. ecotype 78:99-107. Wassilewskija (Brassicaceae). Protoplasma 185:7-21. Olson, A.R. 1993. Patterns of embryo and endosperm for- Owens, SJ. 1981. Self-incompatibility in the Commelmation in Monotropa hypopitys (Monotropaceae) inaceae. Ann. Bot. 47:567-581. from North America and western Sweden. Am. J. Owens, S.J., and Dickinson, H.G. 1983. Pollen wall develBot. 80:839-846. opment in Gibasis (Commelinaceae). Ann. Bot. Olson, A.R. 1994. Pollen tube pathway through the 51:1-15. gynoecium of Monotropsis odorata (Monotropaceae). Owens, S.J., and Horsfield, NJ. 1982. Light and electron Am. J. Bot. 81:718-725. microscopic study of stigmas in Aneilema and Olson, A.R., and Cass, D.D. 1981. Changes in megagameCommelina species (Commelinaceae). Protoplasma tophyte structure in Papaver nudicaule L. 112:26-36. (Papaveraceae) following in vitro placental pollina- Owens, S.J., and Kimmins, F.M. 1981. Stigma morphology tion. Am. J. Bot. 68:1333-1341. in Commelinaceae. Ann. Bot. 47:771-783. Oommen, A., Dixon, R.A., and Paiva, N.L. 1994. The elic- Ozias-Akins, P., and Vasil, I.K. 1982. Plant regeneration itor-inducible alfalfa isoflavone reductase promoter from cultured immature embryos and inflorescences confers different patterns of developmental expresof Triticum aestivum L. (wheat): evidence for somatic sion in homologous and heterologous transgenic embryogenesis. Protoplasma 110:95-105. plants. Plant Cell 6:1789-1803. Ozias-Akins, P., and Vasil, I.K. 1983. Improved efficiency and normalization of somatic embryogenesis in Ooms, J.J.J., Leon-Kloosterziel, K.M., Bartels, D., Koornneef, M., and Karssen, CM. 1993. Acquisition Triticum aestivum (wheat). Protoplasma 117:40-44. of desiccation tolerance and longevity in seeds of Arabidopsis thaliana. A comparative study using Pacini, E., Bellani, L.M., and Lozzi, R. 1986. Pollen, tapeabscisic acid-insensitive abi3 mutants. Plant Physiol. turn and anther development in two cultivars of 102:1185-1191. sweet cherry (Prunus avium). Phytomorphology Oparka, K.J., and Harris, N. 1982. Rice protein-body for36:197-210. mation: all types are initiated by dilation of the Pacini, E., and Casadoro, G. 1981. Tapetum plastids of endoplasmic reticulum. Planta 154:184-188. Olea europaea L. Protoplasma 106:289-296. Or, E., Boyer, S.K., and Larkins, B.A. 1993. Opaque! modi- Pacini, E., and Cresti, M. 1976. Close association between fiers act post-transcriptionally and in a polar manner plastids and endoplasmic reticulum cisterns during on y-zein gene expression in maize endosperm. Plant pollen grain development in Lycopersicon peruCell 5:1599-1609. vianum. ]. Ultrastr. Res. 57:260-265. Ori, N., Sessa, G., Lotan, T., Himmelhoch, S., and Fluhr, R. Pacini, E., and Cresti, M. 1978. Ultrastructural characteris1990. A major stylar matrix polypeptide (sp41) is a tics of tapetum and microspore mother cells in member of the pathogenesis-related proteins superLycopersicon peruvianum during meiotic prophase. class. EMBO }. 9:3429-3436. Soc. Bot. Fr. Actual. Bot. 125:121-128. Ornduff, R. 1964. The breeding system of Oxalis suksdorfii. Pacini, E., Cresti, M., and Sarfatti, G. 1972. Incorporation Am. J. Bot. 51:307-314. of integumentary nuclei in Eranthis hiemalis Ornduff, R. 1966. The breeding system of Pontederia corendosperm and their disaggregation by the endoplasmic reticulum. /. Submicros. Cytol. 4:19-31. data L. Bull. Torrey Bot. Cl. 93:407-416. Osborn, T.C., Alexander, D.C., Sun, S.S.M., Cardona, C, Pacini, E., Franchi, G.G., and Hesse, M. 1985. The tapeand Bliss, F.A. 1988. Insecticidal activity and lectin tum: its form, function, and possible phylogeny in homology of arcelin seed protein. Science Embryophyta. Plant Syst. Evol. 149:155-185. 240:207-210. Pacini, E., and Juniper, B.E. 1979a. The ultrastructure of 0sterbye, U. 1975. Self-incompatibility in Ranunculus acris pollen-grain development in the olive (Olea L. I. Genetic interpretation and evolutionary aspects. europaea). 1. Proteins in the pore. Neiu Phytol. Hereditas 80:91-112. 83:157-163.
678
References
Pacini, Ev and Juniper, B.E. 1979b. The ultrastructure of Pandey, K.K. 1958. Time of the S allele action. Nature pollen grain development in the olive (Oka 181:1220-1221. europaea). 2. Secretion by the tapetal cells. New Phytol. Pandey, K.K. 1960. Incompatibility in Abutilon 'hybridum.' 83:165-174. Am. ]. Bot. 47:877-883. Pacini, E., and Juniper, B.E. 1983. The ultrastructure of Pandey, K.K. 1964. Elements of the S-gene complex. Genet. the formation and development of the amoeboid Res. 5:397-409. tapetum in Arum italicum Miller. Protoplasma Pandey, K.K. 1967. Origin of genetic variability: combina117:116-129. tions of peroxidase isozymes determine multiple allelism of the S gene. Nature 213:669-672. Pacini, E., and Keijzer, C.J. 1989. Ontogeny of intruding nonperiplasmodial tapetum in the wild chicory, Chichorium Pandey, K.K. 1970. Time and site of the S-gene action, intybus (Compositae). Plant Syst. Evol. 167:149-164. breeding systems and relationships in incompatibility. Euphytica 19:364-372. Pacini, E., Simoncioli, C, and Cresti, M. 1975. Ultrastructure of nucellus and endosperm of Pandey, K.K. 1973. Heat sensitivity of esterase isozymes Diplotaxis erucoides during embryogenesis. Caryologia in the styles of Lilium and Nicotiana. New Phytol. 28:525-538. 72:839-850. Pacini, E., Taylor, P.E., Singh, M.B., and Knox, R.B. 1992. Pandey, K.K. 1977. Mentor pollen: possible role of wallheld pollen growth promoting substances in overDevelopment of plastids in pollen and tapetum of coming intra- and interspecific incompatibility. rye-grass, Lolium perenne L. Ann. Bot. 70:179-188. Genetica 47:219-229. Pais, M.S., and Feijo, J.A. 1987. Microbody proliferation during the microsporogenesis of Ophrys lutea Cav. Pang, P.P., Pruitt, R.E., and Meyerowitz, E.M. 1988. (Orchidaceae). Protoplasma 138:149-155. Molecular cloning, genomic organization, expression and evolution of 12S seed storage protein genes Paiva, R., and Kriz, A.L. 1994. Effect of abscisic acid on of Arabidopsis thaliana. Plant Mol. Biol. 11:805-820. embryo-specific gene expression during normal and precocious germination in normal and viviparous Panitz, R., Borisjuk, L., Manteuffel, R., and Wobus, U. 1995. maize (Zea mays) embryos. Planta 192:332-339. Transient expression of storage-protein genes during early embryogenesis of Vicia faba: synthesis and Pakendorf, K.W. 1970. Male sterility in Lupinus mutabilis metabolization of vicilin and legumin in the embryo, Sweet. Z. Pflanzenzilchtg 63:227-236. suspensor and endosperm. Planta 196:765-774. Palevitz, B.A. 1990. Kinetochore behavior during generative cell division in Tradescantia virginiana. Protoplasma Pankow, H. 1957. Uber den Pollenkitt bei Galanthus nivalis 157:120-127. L. Flora 146:240-253. Palevitz, B.A. 1993. Organization of the mitotic apparatus Parchman, L.G., and Lin, K.-C. 1972. Nucleolar RNA synduring generative cell division in Nicotiana tabacum. thesis during meiosis of lily microsporocytes. Nature Protoplasma 174:25-35. New Biol. 239:235-237. Palevitz, B.A., and Cresti, M. 1988. Microtubule organiza- Parchman, L.G., and Roth, T.F. 1971. Pachytene synaption in the sperm of Tradescantia virginiana. tonemal complexes and meiotic achiasmatic chroProtoplasma 146:28-34. mosomes. Chromosoma 33:129-145. Palevitz, B.A., and Cresti, M. 1989. Cytoskeletal changes Parchman, L.G., and Stern, H. 1969. The inhibition of protein synthesis in meiotic cells and its effect on chroduring generative cell division and sperm formation mosome behavior. Chromosoma 26:298-311. in Tradescantia virginiana. Protoplasma 150:54—71. Palevitz, B.A., and Liu, B. 1992. Microfilaments (F-actin) Parcy, F., Valon, C, Raynal, M., Gaubier-Comella, P., in generative cells and sperm: an evaluation. Sex. Delseny, M., and Giraudat, J. 1994. Regulation of gene expression programs during Arabidopsis seed Plant Reprod 5:89-100. development: roles of the ABI3 locus and of endogePalevitz, B.A., and Tiezzi, A. 1992. Organization, componous abscisic acid. Plant Cell 6:1567-1582. sition, and function of the generative cell and sperm Pareddy, D.R., and Greyson, R.I. 1985. In vitro culture of cytoskeleton. Int. Rev. Cytol. 140:149-185. immature tassels of an inbred field variety of Zea Palmer, J.D., Shields, C.R., Cohen, D.B., and Orton, T.J. mays, cv. Oh43. Plant Cell Tissue Organ Cult. 1983. An unusual mitochondrial DNA plasmid in 5:119-128. the genus Brassica. Nature 301:725-728. Palmer, R.G. 1971. Cytological studies of ameiotic and Pareddy, D.R., and Petolino, J.F. 1992. Maturation of maize pollen in vitro. Plant Cell Rep. 11:535-539. normal maize with reference to premeiotic pairing. Paris, D., Rietsema, J., Satina, S., and Blakeslee, A.F. 1953. Chromosoma 35:233-246. Effect of amino acids, especially aspartic and gluPalmer, R.G., Johns, C.W., and Muir, PS. 1980. Genetics tamic acid and their amides, on the growth of Datura and cytology of the ms3 male-sterile soybean. /. stramonium embryos in vitro. Proc. Natl Acad. Sci. Hered. 71:343-348. USA 39:1205-1212. Palser, B.F., Rouse, J.L., and Williams, E.G. 1992. A scanning electron microscope study of the pollen tube Parker, M.L. 1982. Protein accumulation in developing endosperm of a high-protein line of Triticum dicocpathway in pistils of Rhododendron. Can. ]. Bot. coides. Plant Cell Environ. 5:37-43. 70:1039-1060. Pan, D., and Nelson, O.E. 1984. A debranching enzyme Parker, M.L., and Hawes, C.R. 1982. The Golgi apparatus in developing endosperm of wheat (Triticum aesdeficiency in endosperms of the sugary-1 mutants of tivum L.). Planta 154:277-283. maize. Plant Physiol. 74:324-238. Pan, J.-L., Gao, G.-H., and Ban, H. 1983. Initial patterns of Patton, D.A., Franzmann, L.A., and Meinke, D.W. 1991. Mapping genes essential for embryo development in androgenesis in wheat anther culture. Ada Bot. Arabidopsis thaliana. Mol. Gen. Genet. 227:337-347. Sinica 25:34-39. Panchaksharappa, M.G., and Syamasundar, J. 1975. A Patton, D.A., and Meinke, D.W. 1990. Ultrastructure of arrested embryos from lethal mutants of Arabidopsis cytochemical study of ovule development in Divcadi thaliana. Am. J. Bot. 77:653-661. montanum Dalz. Cytologia 40:141-149.
References
619
Patton, D.A., Volrath, S., and Ward, E.R. 1996. Pennell, R.I., Geltz, N.R., Koren, E., and Russell, S.D. 1987. Production and partial characterization of Complementation of an Arabidopsis thaliana biotin hybridoma antibodies elicited to the sperm of auxotroph with an Escherichia coli biotin biosynthetic Plumbago zeylanica. Bot. Gaz. 148:401-406. gene. Mol. Gen. Genet. 251:261-266. Paul, A.-L., and Ferl, RJ. 1994. In vivo footprinting identifies Pennell, R.I., Janniche, L., Kjellbom, P., Scofield, G.N., Peart, J.M., and Roberts, K. 1991. Developmental an activating element of the maize Adh2 promoter speregulation of a plasma membrane arabinogalactan cific for root and vascular tissues. Plant ]. 5:523-533. protein epitope in oilseed rape flowers. Plant Cell Paul, W., Hodge, R., Smartt, S., Draper, J., and Scott, R. 3:1317-1326. 1992. The isolation and characterisation of the tapetum-specific Arabidopsis thaliana A9 gene. Plant Mol.Pennell, R.I., Janniche, L., Scofield, G.N., Booij, H., de Biol. 19:611-622. Vries, S.C., and Roberts, K. 1992. Identification of a transitional cell state in the developmental pathway PavingerovS, D., Dostal, J., Biskov<S, R., and Benetka, V. to carrot somatic embryogenesis. /. Cell Biol. 1994. Somatic embryogenesis and Agrobacterium119:1371-1380. mediated transformation of chrysanthemum. Plant Pennell, R.I., Knox, J.P., Scofield, G.N., Selvendran, R.R., Sci. 97:95-101. and Roberts, K. 1989. A family of abundant plasma Pavlov, A.N., Konarev, V.G., Kolesnik, T.I., and membrane-associated glycoproteins related to the Shayakhmetov, I.F. 1975. Gliadins of the caryopsis of arabinogalactan proteins is unique to flowering wheat during the process of its development. Soviet plants. /. Cell Biol. 108:1967-1977. Plant Physiol. 22:63-67. Payne, G., Kono, Y, and Daly, J.M. 1980. A comparison of Pental, D., Mukhopadhyay, A., Grover, A., and Pradhan, A.K. 1988. A selection method for the synthesis of purified host specific toxin from Helminthosporium triploid hybrids by fusion of microspore protoplasts maydis, race T, and its acetate derivative on oxidation (n) with somatic cell protoplasts (2n). Theor. Appl. by mitochondria from susceptible and resistant Genet. 76:237-243. plants. Plant Physiol. 65:785-791. Pearson, O.H. 1981. Nature and mechanisms of cytoplas- Penza, R., Lurquin, P.F., and Filippone, E. 1991. Gene mic male sterility in plants. HortScience 16:482-487. transfer by cocultivation of mature embryos with Agrobacterium tumefaciens: application to cowpea Pechan, P.M. 1989. Successful cocultivation of Brassica napus microspores and proembryos with Agro(Vigna unguiculata Walp). /. Plant Physiol. 138:39-13. bacterium. Plant Cell Rep. 8:387-390. Perata, P., Picciarelli, P., and Alpi, A. 1990. Pattern of variPechan, P.M., and Keller, W.A. 1988. Identification of ations in abscisic acid content in suspensors, potentially embryogenic microspores in Brassica embryos, and integuments of developing Phaseolus coccineus seeds. Plant Physiol. 94:1776-1780. napus. Physiol. Plant. 74:377-384. Pechan, P.M., Bartels, D., Brown, D.C.W., and Schell, J. Perdue, T.D., Loukides, C.A., and Bedinger, PA. 1992. The 1991. Messenger-RNA and protein changes associformation of cytoplasmic channels between tapetal ated with induction of Brassica microspore embryocells in Zea mays. Protoplasma 171:75-79. genesis. Planta 184:161-165. Perdue, T.D., and Parthasarathy, M.V. 1985. In situ localization of F-actin in pollen tubes. Enr. J. Cell Biol. Peddada, L.B., and Mascarenhas, J.P. 1975. The synthesis 39:13-20. of 5S ribosomal RNA during pollen development. Devel. Growth Diffn 17:1-8. Pereira, A.S.R., and Linskens, H.F. 1963. The influence of Pedersen, K., Argos, P., Naravana, S.V.L., and Larkins, glutathione and glutathione antagonists on meiosis B.A. 1986. Sequence analysis and characterization of in excised anthers of Lilium henryi. Ada Bot. Neerl. a maize gene encoding a high-sulfur zein protein of 12:302-314. Mr 15,000. /. Biol. Chem. 261:6279-6284. Perez, M., Ishioka, G.Y., Walker, L.E., and Chesnut, R.W. 1990. cDNA cloning and immunological characteriPedersen, K., Bloom, K.S., Anderson, J.N., Glover, D.V., zation of the rye grass allergen Lol p I. /. Biol. Client. and Larkins, B.A. 1980. Analysis of the complexity 265:16210-16215. and frequency of zein genes in the maize genome. Biochemistry 19:1644-1650. Perez-Grau, L., and Goldberg, R.B. 1989. Soybean seed Pedersen, K., Devereux, J., Wilson, D.R., Sheldon, E., and protein genes are regulated spatially during Larkins, B.A. 1982. Cloning and sequence analysis embryogenesis. Plant Cell 1:1095-1109. reveal structural variation among related zein genes P6rez-Grau, L., Cortadas, J., Puigdomenech, P., and Palau, in maize. Cell 29:1015-1026. J. 1986. Accumulation and subcellular localization of Pedersen, S., and Andersen, S.B. 1993. Developmental glutelin-2 transcripts during maturation of maize expression of isozymes during embryogenesis in endosperm. FEBS Lett. 202:145-148. Perez-Munoz, C.A., Jernstedt, J.A., and Webster, B.D. barley anther culture. Plant Sci. 91:75-86. Pedersen, S., Simonsen, V., and Loeschcke, V. 1987. 1993a. Pollen wall development in Vigna vexillata. Overlap of gametophytic and sporophytic gene I. Characterization of wall layers. Am. ]. Bot. expression in barley. Theor. Appl. Genet. 75:200-206. 80:1183-1192. Peirson, B.N., Owen, H.A., Feldmann, K.A., and Perez-Munoz, C.A., Jernstedt, J.A., and Webster, B.D. Makaroff, C.A. 1996. Characterization of three male1993b. Pollen wall development in Vigna vexillata. II. sterile mutants of Arabidopsis thaliana exhibiting Ultrastructural studies. Am. j. Bot. 80:1193-1202. alterations in meiosis. Sex. Plant Reprod. 9:1-10. Periasamy, K., and Amalathas, J. 1991. Absence of callose Pelletier, G., and Ilami, M. 1972. Les facteurs de l'anand tetrad in the microsporogenesis of Pandanus drog^nese in vitro chez Nicotiana tabacum. Z. odoratissimus with well-formed pollen exine. Ann. Pflanzenphysiol. 68:97-114. Bot. 67:29-33. Pennell, R.I., and Roberts, K. 1990. Sexual development Pernollet, J.-C, and Vaillant, V. 1984. Characterization and complexity of wheat developing endosperm in the pea is presaged by altered expression of aramRNAs. Plant Physiol. 76:187-190. binogalactan protein. Nature 344:547-549.
620
References
fusion and tip extension in pollen tubes. Planta Pescitelli, S.M., and Sukhapinda, K. 1995. Stable transfor163:20-26. mation via electroporation into maize type II callus and regeneration of fertile transgenic plants. Plant Pierson, E.S. 1988. Rhodamine-phalloidin staining of FCell Rep. 14:712-716. actin in pollen after dimethylsulphoxide permeabilization. Sex. Plant Reprod. 1:83-87. Peterson, D.M., Saigo, R.H., and Holy, J. 1985. Development of oat aleurone cells and their protein Pierson, E.S., Derksen, J., and Traas, J.A. 1986. bodies. Cereal Cliem. 62:366-371. Organization of microfilaments and microtubules in pollen tubes grown in vitro or in vivo in various Peterson, P.A., Flavell, R.B., and Barratt, D.H.P. 1975. angiosperms. Eur. J. Cell Biol. 41:14-18. Altered mitochondrial membrane activities associated with cytoplasmically-inherited disease sensitiv- Pierson, E.S., Kengen, H.M.P., and Derksen, J. 1989. ity in maize. Theor. Appl. Genet. 45:309-314. Microtubules and actin filaments co-localize in pollen tubes of Nicotiana tabacum L. and Lilium longiPetolino, J.F., and Jones, A.M. 1986. Anther culture of elite florum Thunb. Protoplasma 150:75-77. genotypes of maize. Crop Sci. 26:1072-1074. Petolino, J.F., and Thompson, S.A. 1987. Genetic analysis Pierson, E.S., Lichtscheidl, I.K., and Derksen, J. 1990. Structure and behaviour of organelles in living of anther culture response in maize. Theor. Appl. pollen tubes of Lilium longiflorum. f. Expt. Bot. Genet. 74:284-286. 41:1461-1468. Pettitt, J.M. 1976. Pollen wall and stigma surface in the marine angiosperms Thalassia and Thalassodendron.Pierson, E.S., Li, Y.Q., Zhang, H.Q., Willemse, M.T.M., Linskens, H.F., and Cresti, M. 1995. Pulsatory Micron 7:21-32. growth of pollen tubes: investigation of a possible Pettitt, J.M. 1980. Reproduction in seagrasses: nature of relationship with the periodic distribution of cell the pollen and receptive surface of the stigma in the wall components. Ada Bot. Need. 44:121-128. Hydrocharitaceae. Ann. Bot. 45:257-271. Pettitt, J.M. 1981. Reproduction in seagrasses: pollen Pierson, E.S., Miller, D.D., Callaham, D.A., Shipley, A.M., development in Thalassia hemprichii, Halophila stipu- Rivers, B.A., Cresti, M., and Hepler, P.K. 1994. Pollen tube growth is coupled to the extracellular calcium lacea and Thalassodendron ciliatum. Ann. Bot. ion flux and the intracellular calcium gradient: effect 48:609-622. of BAPTA-type buffers and hypertonic media. Plant Phatak, S.C., Wittwer, S.H., Honma, S., and Bukovac, M.J. Cell 6:1815-1828. 1966. Gibberellin-induced anther and pollen development in a stamen-less tomato mutant. Nature Pierson, E.S., Willekens, P.G.M., Maessen, M., and 209:635-636. Helsper, J.P.F.G. 1986. The effect of lectins on germinating pollen of Lilium longiflorum. I. Effect on Piaggesi, A., Picciarelli, P., Lorenzi, R., and Alpi, A. 1989. pollen germination, pollen tube growth and organiGibberellins in embryo-suspensor of Phaseolus coczation of microfilaments. Acta Bot. Neerl. 35:249-256. cineus seeds at the heart stage of embryo developPipkin, J.L., Jr., and Larson, D.A. 1972. Characterization of ment. Plant Physiol. 91:362-366. the Very' lysine-rich histones of active and quiesPicciarelli, P., and Alpi, A. 1986. Gibberellins in suspencent anther tissues of Hippeastrum belladonna. Expt. sors of Phaseolus coccinens L. seeds. Plant Physiol. Cell Res. 71:249-260. 82:298-300. Picciarelli, P., and Alpi, A. 1987. Embryo-suspensor of Pipkin, J.L., Jr., and Larson, D.A. 1973. Changing patterns of nucleic acids, basic and acidic proteins in generaTropaeolum majus: identification of gibberellin A63. tive and vegetative nuclei during pollen germinaPhytochemistry 26:329-330. tion and pollen tube growth in Hippeastrum Picciarelli, P., Piaggesi, A., and Alpi, A. 1991. Gibberellins belladonna. Expt. Cell Res. 79:28-42. in suspensor, embryo and endosperm of developing seeds of Cytisus laburnum. Phytochemistry Pirrie, A., and Power, J.B. 1986. The production of fertile, triploid somatic hybrid plants (Nicotiana glutinosa (n) 30:1789-1792. + N. tabacum (2n)) via gameticsomatic protoplast Picciarelli, P., Alpi, A., Pistelli, L., and Scalet, M. 1984. fusion. Theor. Appl. Genet. 72:48-52. Gibberellin-like activity in suspensors of Tropaeolum majus L. and Cytisus laburnum L. Planta 162:566-568. Pitto, L., Lo Schiavo, F., and Terzi, M. 1985. a-amanitin resistance is developmentally regulated in carrot. Picton, J.M., and Steer, M.W. 1981. Determination of secreProc. Natl Acad. Sci. USA 82:2799-2803. tory vesicle production rates by dictyosomes in pollen tubes of Tradescantia using cytochalasin D. /. Pitto, L., Lo Schiavo, F., Giuliano, G., and Terzi, M. 1983. Analysis of the heat-shock protein pattern during Cell Sci. 49:261-272. somatic embryogenesis of carrot. Plant Mol. Biol. Picton, J.M., and Steer, M.W. 1982. A model for the mech2:231-237. anism of tip extension in pollen tubes. /. Theor. Biol. Pla, M., Goday, A., Vilardell, J., G6mez, J., and Pages, M. 98:15-20. 1989. Differential regulation of ABA-induced 23-25 Picton, J.M., and Steer, M.W. 1983a. Evidence for the role kDa proteins in embryo and vegetative tissues of the of Ca2+ ions in tip extension in pollen tubes. viviparous mutants of maize. Plant Mol. Biol. Protoplasma 115:11-17. 13:385-394. Picton, J.M., and Steer, M.W. 1983b. The effect of cycloheximide on dictyosome activity in Tradescantia Pla, M., Gomez, J., Goday, A., and Pages, M. 1991. Regulation of the abscisic acid-responsive gene rablS pollen tubes determined using cytochalasin D. Eur. J. in maize viviparous mutants. Mol. Gen. Genet. Cell Biol. 29:133-138. 230:394-400. Picton, J.M., and Steer, M.W. 1983c. Membrane recycling and the control of secretory activity in pollen tubes. Pla, M., Vilardell, J., Guiltinan, M.J., Marcotte, W.R., Niogret, M.F., Quatrano, R.S., and Pages, M. 1993. /. Cell Sci. 63:303-310. The Cfs-regulatory element CCACGTGG is involved Picton, J.M., and Steer, M.W. 1985. The effects of ruthein ABA and water-stress responses of the maize gene nium red, lanthanum, fluorescein isothiocyanate rab28. Plant Mol. Biol. 21:259-266. and trifluoperazine on vesicle transport, vesicle
References
621
induced male and female sterility in canola (Brassica Plant, A.L., van Rooijen, G.J.H., Anderson, C.P., and napus L.). Ann. Bot. 62:83-86. Moloney, M.M. 1994. Regulation of an Arabidopsis oleosin gene promoter in transgenic Brassica napus. Polowick, PL., and Sawhney, V.K. 1990. Microsporogenesis Plant Mol. Biol. 25:193-205. in a normal line and in the ogu cytoplasmic male-sterile line of Brassica napus. I. The influence of high temPlatt-Aloia, K.A., Lord, E.M., DeMason, D.A., and peratures. Sex. Plant Reprod. 3:263-276. Thomson, W.W. 1986. Freeze-fracture observations on membranes of dry and hydrated pollen from Polowick, P.L., and Sawhney, V.K. 1991. Microsporogenesis in a normal line and in the ogu cytoplasmic Collomia, Phoenix and Zea. Planta 168:291-298. male-sterile line of Brassica napus. II. The influence of Plaut, W.S. 1953. DNA synthesis in the microsporocytes of intermediate and low temperatures. Sex. Plant Lilium henryi. Hereditas 39:438-444. Reprod. 4:22-27. Pnueli, L., Abu-Abeid, M., Zamir, D., Nacken, W., SchwarzSommer, Z., and Lifschitz, E. 1991. The MADS box Polowick, P.L., and Sawhney, V.K. 1992. Ultrastructural changes in the cell wall, nucleus and cytoplasm of gene family in tomato: temporal expression during pollen mother cells during meiotic prophase I in floral development, conserved secondary structures Lycopersicon esculentum (Mill.). Protoplasma and homology with homeotic genes from Antirrhinum 169:139-147. and Arabidopsis. Plant J. 1:255-266. Pnueli, L., Hareven, D., Broday, L., Hurwitz, C, and Polowick, PL., and Sawhney, V.K. 1993a. Differentiation of the tapetum during microsporogenesis in tomato Lifschitz, E. 1994a. The TM5 MADS box gene medi(Lycopersicon esculentum Mill.), with special reference ates organ differentiation in the three inner whorls of to the tapetal cell wall. Ann. Bot. 72:595-605. tomato flowers. Plant Cell 6:175-186. Pnueli, L., Hareven, D., Rounsley, S.D., Yanofsky, M.F., Polowick, PL., and Sawhney, V.K. 1993b. An ultrastrucand Lifschitz, E. 1994b. Isolation of the tomato AGAtural study of pollen development in tomato MOUS gene TAG1 and analysis of its homeotic role (Lycopersicon esculentum). I. Tetrad to early binucleate in transgenic plants. Plant Cell 6:163-173. microspore stage. Can. J. Bot. 71:1039-1047. Poddubnaya-Arnoldi, V.A. 1967. Comparative embryol- Polowick, PL., and Sawhney, V.K. 1993c. An ultrastrucogy of the Orchidaceae. Phytomorphology 17:312-320. tural study of pollen development in tomato (Lycopersicon esculentum). II. Pollen maturation. Can. Poethig, R.S., Coe, E.H., Jr., and Johri, M.M. 1986. Cell linJ. Bot. 71:1048-1055. eage patterns in maize embryogenesis: a clonal analysis. Devel Biol. 117:392-404. Polowick, P.L., and Sawhney, V.K. 1995. Ultrastructure of Polito, V.S. 1983. Membrane-associated calcium during the tapetal cell wall in the stamenless-2 mutant of pollen grain germination: a microfluorometric tomato (Lycopersicon esculentum): correlation analysis. Protoplasma 117:226-232. between structure and male sterility. Protoplasma 189:249-255. Pollak, P.E. 1992. Cytological differences between a cytoplasmic male sterile tobacco cybrid and its fertile Polsoni, L., Kott, L.S., and Beversdorf, W.D. 1988. Largecounterpart during early anther development. Am. ]. scale microspore culture technique for mutationBot. 79:937-945. selection studies in Brassica napus. Can. f. Bot. 66:1681-1685. Pollak, P.E., Hansen, K., Astwood, J.D., and Taylor, L.P. 1995. Conditional male fertility in maize. Sex. Plant Polya, G.M., Micucci, V, Rae, A.L., Harris, P.J., and Reprod. 8:231-241. Clarke, A.E. 1986. Ca2+-dependent protein phosphoPollak, P.E., Vogt, T, Mo, Y., and Taylor, L.P. 1993. rylation in germinated pollen of Nicotiana alata, an Chalcone synthase and flavonol accumulation in ornamental tobacco. Physiol. Plant. 67:151-157. stigmas and anthers of Petunia hybrida. Plant Physiol.Pomeroy, M.K., Kramer, J.K.G., Hunt, D.J., and Keller, 102:925-932. W.A. 1991. Fatty acid changes during development of zygotic and microspore-derived embryos of Pollock, E.G., and Jensen, W.A. 1964. Cell development Brassica napus. Physiol. Plant. 81:447-454. during early embryogenesis in Capsella and Gossypium. Am. /. Bot. 51:915-921. Ponte, I., Guillen, P., Debon, R.M., Reina, M., Aragay, A., Pollock, E.G., and Jensen, W.A. 1967. Ontogeny and cytoEspel, E., di Fonzo, N., and Palau, J. 1994. Narrow chemistry of the chalazal proliferating cells of A/T-rich zones present at the distal 5-flanking Capsella bursa-pastoris (L.) Medic. New Phytol. sequences of the zein genes Zcl and Zc2 bind a 66:413-417. unique 30 kDa HMG-like protein. Plant Mol. Biol. 26:1893-1906. Polowick, PL., and Greyson, R.I. 1982. Anther development, meiosis and pollen formation in Zea tassels Ponz, F., Paz-Ares, J., Hernandez-Lucas, C, Carbonero, P., cultured in defined liquid medium. Plant Sci. Lett. and Garci'a-Olmedo, F. 1983. Synthesis and process26:139-145. ing of thionin precursors in developing endosperm Polowick, PL., and Greyson, R.I. 1984. The relative effifrom barley (Hordeum vulgare L.). EMBO /. ciency of cytokinins in the development of normal 2:1035-1040. spikelets on cultured tassels of Zea mays. Can. J. Bot.Ponzi, R., and Pizzolongo, P. 1973. Ultrastructure of plas62:830-834. tids in the suspensor cells of Ipomoea purpurea Roth. Polowick, PL., and Sawhney, V.K. 1985. Temperature /. Submicros. Cytol. 5:257-263. effects on male fertility and flower and fruit devel- Ponzi, R., and Pizzolongo, P. 1976. Cytinus hypocistis L. opment in Capsicum annuum L. Sci. Hort. embryogenesis: ultrastructural aspects of megas25:117-127. porogenesis and megagametogenesis. /. Submicros. Polowick, PL., and Sawhney, V.K. 1987. A scanning elecCytol. 8:327-336. tron microscopic study on the influence of tempera- Ponzi, R., and Pizzolongo, P. 1984. Ultrastructural study ture on the expression of cytoplasmic male sterility of change from free-nuclear to cellular endosperm in in Brassica napus. Can. ]. Bot. 65:807-814. Ipomoea purpurea Roth, and Cytinus hypocistis L. Polowick, P.L., and Sawhney, V.K. 1988. High temperature Giorn. Bot. Hal. 118:147-154.
622 Porath, D., and Galun, E. 1967. In vitro culture of hermaphrodite floral buds of Cucumis melo L.: Microsporogenesis and ovary formation. Ann. Bot. 31:283-290. Porter, E.K. 1981. Origins and genetic nonvariability of the proteins which diffuse from maize pollen. Environ. Health Pers. 37:53-59. Porter, E.K., Bird, J.M., and Dickinson, H.G. 1982. Nucleic acid synthesis in microsporocytes of Lilium cv. Cinnabar: events in the nucleus. /. Cell Sci. 57:229-246. Porter, E.K., Parry, D., and Dickinson, H.G. 1983. Changes in poly(A)+RNA during male meiosis in Lilium. J. Cell Sci. 62:177-186. Porter, E.K., Parry, D., Bird, J., and Dickinson, H.G. 1984. Nucleic acid metabolism in the nucleus and cytoplasm of angiosperm meiocytes. Symp. Soc. Expt. Biol. 38:363-379. Portnoi, L., and Horovitz, A. 1977. Sugars in natural and artificial pollen germination substrates. Ann. Bot. 41:21-27. Potrykus, 1.1990. Gene transfer to plants: assessment and perspectives. Physiol. Plant. 79:125-134. Potz, H., and Tatlioglu, T. 1993. Molecular analysis of cytoplasmic male sterility in chives (Allium schoenoprasum L.). Theor. Appl. Genet. 87:439-445. Poulson, R., and Beevers, L. 1973. RNA metabolism during the development of cotyledons of Pisum sativum L. Biochim. Biophys. Ada 308:381-389. Povvling, A. 1981. Species of small DNA molecules found in mitochondria from sugarbeet with normal and male sterile cytoplasms. Mol. Gen. Genet. 183:82-84. Powling, A. 1982. Restriction endonuclease analysis of mitochondrial DNA from sugarbeet with normal and male-sterile cytoplasms. Heredity 49:117-120. Powling, A., and Ellis, T.H.N. 1983. Studies on the organelle genomes of sugarbeet with male-fertile and male-sterile cytoplasms. Theor. Appl. Genet. 65:323-328. Prabhakar, K., and Vijayaraghavan, M.R. 1983. Histochemistry and ulrrastructure of suspensor cells in Alyssum maritimum. Cytologia 48:389-402. Prahl, A.-K., Springstubbe, H., Grumbach, K., and Wiermann, R. 1985. Studies on sporopollenin biosynthesis: the effect of inhibitors of carotenoid biosynthesis on sporopollenin accumulation. Z. Natnrfars. 40c:621-626. Pramanik, S.K., and Bewley, J.D. 1993. A ubiquitin carrier protein cDNA from developing alfalfa embryos. Plant Physiol. 102:1049-1050. Pramanik, S.K., Krochko, J.E., and Bewley, J.D. 1992. Distribution of cytosolic mRNAs between polysomal and ribonucleoprotein complex fractions in alfalfa embryos. Plant Physiol. 99:1590-1596. Prandl, R., Kloske, E., and Schoffl, F. 1995. Developmental regulation and tissue-specific differences of heat shock gene expression in transgenic tobacco and Arabidopsis plants. Plant Mol. Biol. 28:73-82. Prasad, K. 1977. Histochemistry of anther and ovule in Farsetia hamiltonii (Royle) and Ernca sativa Mill (Cruciferae). /. Indian Bot. Soc. 56:90-99. Prat, S., Perez-Grau, L., and Puigdomenech, P. 1987. Multiple variability in the sequence of a family of maize endosperm proteins. Gene 52:41-49. Prat, S., Cortadas, J., Puigdomenech, P., and Palau, J. 1985. Nucleic acid (cDNA) and amino acid sequences of the maize endosperm protein glutelin-2. Nucl. Acids Res. 13:1493-1504.
References
Preiss, J., Lammel, C, and Sabraw, A. 1971. A unique adenosine diphosphoglucose pyrophosphorylase associated with maize embryo tissue. Plant Physiol. 47:104-108. Pressey, R. 1991. Polygalacturonase in tree pollens. Phytochemistry 30:1753-1755. Pressey, R., and Reger, B.J. 1989. Polygalacturonase in pollen from corn and other grasses. Plant Sci. v 59:57-62. Pretov3, A. 1974. The influence of the osmotic potential of the cultivation medium on the development of excised flax embryos. Biol. Plant. 16:14-20. Pretov3, A. 1986. Growth of zygotic flax embryos in vitro and influence of kinetin. Plant Cell Rep. 3:210-211. Preuss, D., Rhee, S.Y., and Davis, R.W. 1994. Tetrad analysis possible in Arabidopsis with mutation of the QUARTET (QRT) genes. Science 264:1458-1460. Preuss, D., Lemieux, B., Yen, G., and Davis, R.W. 1993. A conditional sterile mutation eliminates surface components from Arabidopsis pollen and disrupts cell signaling during fertilization. Genes Devel. 7:974-985. Prevost, I., and Le Page-Degivry, M.T. 1985. Changes in abscisic acid content in axis and cotyledons of developing Phaseolus vulgaris embryos and their physiological consequences. /. Expt. Bot. 36:1900-1905. Price, S.D., and Barrett, S.C.H. 1982. Tristyly in Pontederia cordata (Pontederiaceae). Can. J. Bot. 60:897-905. Priestley, D.A., and de Kruijff, B. 1982. Phospholipid motional characteristics in a dry biological system. A 31 P-nuclear magnetic resonance study of hydrating Typha latifolia pollen. Plant Physiol. 70:1075-1078. Pring, D.R., Conde, M.R, and Levings, C.S., III. 1980. DNA heterogeneity within the C group of maize male-sterile cytoplasms. Crop Sci. 20:159-162. Pring, D.R., Conde, M.R, and Schertz, K.R 1982. Organelle genome diversity in Sorghum: male-sterile cytoplasms. Crop Sci. 22:414-421. Pring, D.R., and Levings, C.S., III. 1978. Heterogeneity of maize cytoplasmic genomes among male-sterile cytoplasms. Genetics 89:121-136. Pring, D.R., Conde, M.R, Schertz, K.R, and Levings, C.S., III. 1982. Plasmid-like DNAs associated with mitochondria of cytoplasmic male-sterile Sorghum. Mol. Gen. Genet. 186:180-184. Pring, D.R., Levings, C.S., III., Hu, W.W.L., and Timothy, D.H. 1977. Unique DNA associated with mitochondria in the "S"-type cytoplasm of male-sterile maize. Proc. Natl Acad. Sci. USA 74:2904-2908. Pritchard, A.J., and Hutton, E.M. 1972. Anther and pollen development in male-sterile Phaseolus atropurpureus. J. Hered. 63:280-282. Pritchard, H.N. 1964a. A cytochemical study of embryo sac development in Stellaria media. Am. J. Bot. 51:371-378. Pritchard, H.N. 1964b. A cytochemical study of embryo development in Stellaria media. Am. ]. Bot. 51:472-479. Pruitt, K.D., and Hanson, M.R. 1989. Cytochrome oxidase subunit II sequences in Petunia mitochondria: two intron-containing genes and an intron-less pseudogene associated with cytoplasmic male sterility. Curr. Genet. 16:281-291. Pruitt, K.D., and Hanson, M.R. 1991. Transcription of the Petunia mitochondrial CMS-associated Pcf locus in male sterile and fertility-restored lines. Mol. Gen. Genet. 227:348-355. Przybyllok, T., and Nagl, W. 1977. Auxin concentration in the embryo and suspensors of Tropaeolum majus, as
References
623
determined by mass fragmentation (single ion deteclated expression of a reintroduced napin gene. Theor. tion). Z. Pflanzenphysiol. 84:463-465. Appl. Genet. 75:685-694. Punnett, H.H. 1953. Cytological evidence of hexaploid Radlowski, M., Kalinowski, A., Krblikowski, Z., and Bartkowiak, S. 1994. Protease activity from maize cells in maize endosperm. /. Hered. 44:257-259. pollen. Phytochemistry 35:853-856. Puupponen-Pimia, R., Saloheimo, M., Vasara, T., Ra, R., Gaugecz,J., KurteYi, U., Knowles, J.K.C., Keranen, S., Radojevic, L., Zylberberg, L., and Kovoor, J. 1980. Etude ultrastructurale des embryons androgenetiques and Kauppinen, V. 1993. Characterization of a birch d'Aesculus hippocastannm L. Z. Pflanzenphysiol. (Betula pendula Roth.) embryogenic gene, BP8. Plant Mol. Biol. 23:423-428. 98:255-261. Pwee, K.-Hv and Gray, J.G. 1993. The pea plastocyanin Rae, A.L., Harris, P.J., Bade, A., and Clarke, A.E. 1985. promoter directs cell-specific but not full light-reguComposition of the cell walls of Nicotiana alata Link et Otto pollen tubes. Planta 166:128-133. lated expression in transgenic tobacco plants. Plant ]. 3:437-449. Raeber, J.G., and Bolton, A. 1955. A new form of male Pysh, L.D., Aukerman, M.J., and Schmidt, R.J. 1993. sterility in Nicotiana tabacum L. Nature 176:314-315. OHP1: a maize basic domain/ leucine zipper protein Rafalski, J.A. 1986. Structure of wheat gamma-gliadin genes. Gene 43:221-229. that interacts with opaque2. Plant Cell 5:227-236. Rafalski, J.A., Scheets, K., Metzler, M., Peterson, D.M., Hedgcoth, C, and Soil, D.G. 1984. Developmentally Qiu, X., and Erickson, L. 1996. A pollen-specific polyregulated plant genes: the nucleotide sequence of a galacturonase-like cDNA from alfalfa. Sex. Plant wheat gliadin genomic clone. EMBO ]. 3:1409-1415. Reprod. 9:123-124. Qu, R., and Huang, A.H.C. 1990. Oleosin KD 18 on the Raff, J.W., and Clarke, A.E. 1981. Tissue-specific antigens secreted by suspension-cultured callus cells of surface of oil bodies in maize. Genomic and cDNA Primus avium L. Planta 153:115-124. sequences and the deduced protein structure. /. Biol. Chem. 265:2238-2243. Raff, J.W., Knox, R.B., and Clarke, A.E. 1981. Style antiQu, R., Vance, V.B., and Huang, A.H.C. 1990. Expression gens of Primus avium L. Planta 153:125-129. of genes encoding oleosin isoforms in the embryos Raff, J.W., Pettitt, J.M., and Knox, R.B. 1981. of maturing maize kernels. Plant Sci. 72:223-232. Cytochemistry of pollen tube growth in stigma and style of Primus avium. Phytomorphology 31:214-231. Qu, R., Wang, S.-M., Lin, Y.-H., Vance, V.B., and Huang, A.H.C. 1986. Characteristics and biosynthesis of Rafnar, T., Griffith, I.J., Kuo, M.-C, Bond, J.F., Rogers, B.L., membrane proteins of lipid bodies in the scutella of and Klapper, D.G. 1991. Cloning of Amb a I (antigen maize (Zea mays L.). Biochem. ]. 235:57-65. E), the major allergen family of short ragweed Qu, R.-D., and Chen, Y. 1984. Pathways of androgenesis pollen. /. Biol. Chem. 266:1229-1236. and observations on cultured pollen grains in rice Raghavan, P., and Philip, V.]. 1982. Morphological and (Oryza sativa subsp. Keng). Ada Bot. Sinica histochemical changes in the egg and zygote of 26:580-587. Lagerstroemia speciosa. I. Cell size, vacuole and insoluble polysaccharides. Proc. Indian Acad. Sci. (Plant Quattrocchio, R, Tolk, M.A., Coraggio, I., Mol. J.N.M., Sci.) 91:465^172. Viotti, A., and Koes, R.E. 1990. The maize zein gene zE19 contains two distinct promoters which are Raghavan, V. 1976a. Role of the generative cell in androindependently activated in endosperm and anthers genesis in henbane. Science 191:388-389. of transgenic Petunia plants. Plant Mol. Biol. Raghavan, V. 1976b. Experimental Embryogenesis in 15:81-93. Vascular Plants. London: Academic Press. Quayle, T.J.A., Hetz, W., and Feix, G. 1991. Raghavan, V. 1977a. Patterns of DNA synthesis during Characterization of a maize endosperm culture pollen embryogenesis in henbane. /. Cell Biol. expressing zein genes and its use in transient trans73:521-526. formation assays. Plant Cell Rep. 9:544-548. Raghavan, V. 1977b. Applied aspects of embryo culture. Que, Q.-D., and Tang, X.-H. 1988. The changes of protein In Applied and Fundamental Aspects of Plant Cell, content and synthetic activity in squash pollen durTissue, and Organ Culture, ed. J. Reinert and Y.P.S. ing germination. Ada Bot. Sinica 30:501-507. Bajaj, pp. 375-397. Berlin: Springer-Verlag. Raghavan, V. 1978. Origin and development of pollen Rabakoarihanta, A., Mok, D.W.S., and Mok, M.C. 1979. embryoids and pollen calluses in cultured anther Fertilization and early embryo development in recisegments of Hyoscyamus niger (henbane). Am. ]. Bot. procal interspecific crosses of Phaseolus. Theor. Appl. 65:984-1002. Genet. 54:55-59. Raghavan, V. 1979a. Embryogenic determination and Racchi, M.L., Gavazzi, G., Monti, D., and Manitto, P. 1978. ribonucleic acid synthesis in pollen grains of An analysis of the nutritional requirements of the pro Hyoscyamus niger (henbane). Am. J. Bot. 66:36-39. mutant in Zea mays. Plant Sci. Lett. 13:357-364. Raghavan, V. 1979b. An autoradiographic study of RNA Racusen, R.H., and Schiavone, F.M. 1988. Detection of synthesis during pollen embryogenesis in spatially- and stage-specific proteins in extracts from Hyoscyamus niger (henbane). Am. ]. Bot. 66:784-795. single embryos of the domesticated carrot. Raghavan, V. 1981a. A transient accumulation of poly(A)Development 103:665-674. containing RNA in the tapetum of Hyoscyamus niger Radding, CM. 1982. Homologous pairing and strand during microsporogenesis. Devel Biol. 81:342-348. exchange in genetic recombination. Annu. Rev. Genet. Raghavan, V. 1981b. Distribution of poly(A)-containing 16:405-437. RNA during normal pollen development and during Radke, S.E., Andrews, B.M., Moloney, M.M., Crouch, induced pollen embryogenesis in Hyoscyamus niger. M.L., Kridl, J.C, and Knauf, V.C. 1988. }. Cell Biol. 89:593-606. Transformation of Brassica napus L. using Raghavan, V. 1984. Protein synthetic activity during norAgrobaderium tumefaciens: developmentally regumal pollen development and during induced pollen
624
References
embryogenesis in Hyoscyamus niger. Can. J. Bot. Raghavan, V., and Sharma, K.K. 1995. Zygotic embryoge62:2493-2513. nesis in gymnosperms and angiosperms. In In Vitro Raghavan, V. 1985. The applications of embryo rescue in Embryogenesis in Plants, ed. T.A. Thorpe, pp. 73-115. agriculture. In Biotechnology in International Dordrecht: Kluwer Academic Press. Agricultural Research, pp. 189-197. Manila: Raghavan, V., and Torrey, J.G. 1963. Growth and morphoInternational Rice Research Institute. genesis of globular and older embryos of Capsella in culture. Am. }. Bot. 50:540-551. Raghavan, V. 1986a. Pollen developmental biology in cultured anthers. In Cell Culture and Somatic Cell GeneticsRaghavan, V, and Torrey, J.G. 1964. Effects of certain of Plants, Vol. 3, ed. I.K. Vasil, pp. 275-304. Orlando: growth substances on the growth and morphogeneAcademic Press. sis of immature embryos of Capsella in culture. Plant Raghavan, V. 1986b. Variability through wide crosses and Physiol. 39:691-699. embryo rescue. In Cell Culture and Somatic Cell Rahman, S., Shewry, P.R., and Miflin, B.J. 1982. Genetics of Plants, Vol. 3, ed. I.K. Vasil, pp. 612-633. Differential protein accumulation during barley Orlando: Academic Press. grain development. /. Expt. Bot. 33:717-728. Raghavan, V. 1986c. Embryogenesis in Angiosperms. A Rahman, S., Kreis, M., Forde, B.G., Shewry, PR., and Developmental and Experimental Study. Cambridge: Miflin, B.J. 1984. Hordein-gene expression during Cambridge University Press. development of the barley (Hordeum vulgare) endosperm. Biochem. J. 223:315-322. Raghavan, V. 1988. Anther and pollen development in rice (Oryza sativa). Am. J. Bot. 75:183-196. Raikhel, N.V., Bednarek, S.Y., and Wilkins, T.A. 1988. Celltype-specific expression of a wheat-germ agglutinin Raghavan, V. 1989. mRNAs and a cloned histone gene are gene in embryos and young seedlings of Triticum aesdifferentially expressed during anther and pollen tivum. Planta 176:406-414. development in rice (Oryza sativa L.). /. Cell Sci. 92:217-229. Raikhel, N.V., and Lerner, D.R. 1991. Expression and regulation of lectin genes in cereals and rice. Devel Raghavan, V. 1990a. Origin of the quiescent center in the Genet. 12:255-260. root of Capsella bursa-pastoris (L.) Medik. Planta 181:62-70. Raikhel, N.V., and Quatrano, R.S. 1986. Localization of Raghavan, V. 1990b. Spatial distribution of mRNA during wheat-germ agglutinin in developing wheat embryos and those cultured in abscisic acid. Planta pre-fertilization ovule development in Capsella bursa-pastoris. Sex. Plant Reprod. 3:170-178. 168:433-440. Raghavan, V. 1990c. Hyoscyamus spp.: anther culture stud- Raikhel, N.V., and Wilkins, T.A. 1987. Isolation and ies. In Biotechnology in Agriculture and Forestry 12. characterization of a cDNA clone encoding wheat Haploids in Crop Improvement I, ed. Y.P.S. Bajaj, pp. germ agglutinin. Proc. Natl Acad. Sci. USA 290-305. Berlin: Springer-Verlag. 84:6745-6749. Raghavan, V. 1991. Some perspectives on the develop- Raina, A., and Datta, A. 1992. Molecular cloning of a gene ment of embryos in angiosperms. A synthesis of old encoding a seed-specific protein with nutritionally ideas and new concepts. Rev. Cytol. Biol. Veget.-Bot. balanced amino acid composition from Amaranthus. 14:163-174. Proc. Natl Acad. Sci. USA 89:11774-11778. Raghavan, V. 1993. Embryo culture: methods and appli- Raineri, D.M., Bottino, P., Gordon, M.P., and Nester, E.W. cations. In Plant Biotechnology. Commercial Prospects 1990. Agrobacterium-mediated transformation of rice and Problems, ed. J. Prakash and R.L.M. Pierik, pp. (Oryza sativa L.). Biotechnology 8:33-38. 143-168. New Delhi: Oxford & IBH Publishing Co. Rajam, M.V. 1989. Restriction of pollen germination and Raghavan, V. 1994. In vitro methods for the control of fertube growth in lily pollen by inhibitors of polyamine tilization and embryo development. In Plant Cell and metabolism. Plant Sci. 59:53-56. Tissue Culture, ed. I.K. Vasil and T.A. Thorpe, pp. Rajasekaran, K., Vine, J., and Mullins, M.G. 1982. 173-194. Dordrecht: Kluwer Academic Publishers. Dormancy in somatic embryos and seeds of Vitis: Raghavan, V. 1995. Manipulation of pollen grains for changes in endogenous abscisic acid during embryogametophytic and sporophytic types of growth. geny and germination. Planta 154:139-144. Meth. Cell Biol. 49:367-375. Rajasekaran, K., Hein, M.B., Davis, G.C., Carnes, M.G., Raghavan, V, and Goh, C.J. 1994. DNA synthesis and and Vasil, I.K. 1987. Endogenous growth regulators mRNA accumulation during germination of in leaves and tissue cultures of Pennisetum purpureum Schum. /. Plant Physiol. 130:13-25. embryos of the orchid Spathoglottis plicata. Protoplasma 183:137-147. Rajasekhar, E.W. 1973. Nuclear divisions in protoplasts isolated from pollen tetrads of Datura metel. Nature Raghavan, V., Jiang, C, and Bimal, R. 1992. Cell- and tis246:223-224. sue-specific expression of rice histone gene transcripts during anther and pollen development in Rajora, O.P., and Zsuffa, L. 1986. Sporophytic and gamehenbane (Hyoscyamus niger). Am. J. Bot. 77:778-783. tophytic gene expression in Populus deltoides Marsh., P. nigra L., and P. maximowiczii Henry. Can. ]. Genet. Raghavan, V., and Kamalay, J.C. 1993. Expression of two Cytol. 28:476^182. cloned mRNA sequences during development and germination of spores of the sensitive fern, Onoclea Ramachandran, C, and Raghavan, V. 1989. Changes in sensibilis L. Planta 189:1-9. nuclear DNA content of endosperm cells during grain development in rice (Oryza sativa). Ann. Bot. Raghavan, V, and Nagmani, R. 1989. Cytokinin effects on 64:459-468. pollen embryogenesis in cultured anthers of Ramachandran, C, and Raghavan, V. 1990. Intracellular Hyoscyamus niger. Can. J. Bot. 67:247-257. localization of glutelin mRNA during grain developRaghavan, V, and Olmedilla, A. 1989. Spatial patterns of ment in rice. /. Expt. Bot. 41:393-399. histone mRNA expression during grain development and germination in rice. Cell Diffn Devel. Ramachandran, C, and Raghavan, V. 1992a. Apomixis in 27:183-196. distant hybridization. In Distant Hybridization in
References
625
par culture d'antheres. Analyse cytophotom<§trique Crop Plants, ed. G. Kalloo and J.B. Chowdhury, pp. in situ et in vitro des microspores de Petunia et ble 106-121. Berlin: Springer-Verlag. tendre. Z. Pflanzenzuchtg 89:265-277. Ramachandran, C, and Raghavan, V. 1992b. Regulation of gene expression during rice grain development Rasch, E., and Woodard, J.W. 1959. Basic proteins of plant and germination. Trans. Malaysian Soc. Plant Physiol. nuclei during normal and pathological cell growth. 3:217-227. /. Biophys. Biochem. Cytol. 6:263-276. Ramaer, H. 1935. Cytology of Hevea. Genetica 17:193-236. Rashid, A., and Reinert, J. 1981. In vitro differentiation of embryogenic pollen, control by cold treatment and Raman, K., Walden, D.B., and Greyson, R.I. 1980. embryo formation in ab initio pollen cultures of Fertilization in Zea mays by cultured gametophytes. Nicotiana tabacum var. Badischer Burley. Protoplasma /. Hered. 71:311-314. 109:285-294. Ramanna, M.S. 1974. The origin and in vivo development of embryoids in the anthers of Solanum hybrids. Rashid, A., and Reinert, J. 1983. Factors affecting high-freEuphytica 23:623-632. quency embryo formation in ab initio pollen cultures of Nicotiana. Protoplasma 116:155-160. Ramanna, M.S., and Hermsen, J.G.T. 1974. Embryoid formation in the anthers of some interspecific hybrids Rashid, A., Siddiqui, A.W., and Reinert, J. 1981. Ultrastructure of embryogenic pollen of Nicotiana in Solarium. Euphytica 23:423^127. tabacum var. Badischer Burley. Protoplasma Ramaswamy, S.N., and Arekal, G.D. 1982. Embryology of 107:375-385. Eriocaulon xeranthemum Mart. (Eriocaulaceae). Ada Rashid, A., Siddiqui, A.W., and Reinert, J. 1982. Bot. Neerl. 31:41-54. Subcellular aspects of origin and structure of pollen Ramaswamy, S.N., Swamy, B.G.L., and Govindappa, D.A. embryos of Nicotiana. Protoplasma 113:202-208. 1981. From zygote to seedling in Eriocaulon robustobrownianum Ruhl. (Eriocaulaceae). Beitr. Biol. Pflanz.Rashid, A., and Street, H.E. 1973. The development of 55:179-188. haploid embryoids from anther cultures of Atropa belladonna L. Planla 113:263-270. Rambaud, C, and Rambour, S. 1989. Partial characterization of nitrate reductase in carrot cells: changes in Rashid, A., and Street, H.E. 1974. Segmentations in enzymatic activity during somatic embryogenesis. microspores of Nicotiana sylvestris and Nicotiana Plant Physiol. Biochem. 27:235-243. tabacum which lead to embryoid formation in anther cultures. Protoplasma 80:323-334. Randolph, L.F. 1936. Developmental morphology of the Rasmussen, J., and Hanson, M.R. 1989. A NADH dehycaryopsis in maize. /. Agr. Res. 53:881-916. drogenase subunit gene is co-transcribed with the Rangaswamy, N.S. 1967. Morphogenesis of seed germinaabnormal Petunia mitochondrial gene associated tion in angiosperms. Phytomorphology 17:477—487. with cytoplasmic male sterility. Mol. Gen. Genet. Rangaswamy, N.S., and Shivanna, K.R. 1967. Induction of 215:332-336. gamete compatibility and seed formation in axenic cultures of a diploid self-incompatible species of Rasmussen, N., and Green, P.B. 1993. Organogenesis in Petunia. Nature 216:937-939. flowers of the homeotic green pistillate mutant of tomato (Lycopersicon esculentum). Am. ]. Bot. Rangaswamy, N.S., and Shivanna, K.R. 1969. Test-tube fertilization in Dicranostigma franchetianum (Prain) 80:805-813. Rasmussen, S.K., and Brandt, A. 1986. Nucleotide Fedde. Curr. Sci. 38:257-259. Rangaswamy, N.S., and Shivanna, K.R. 1971a. sequences of cDNA clones for C-hordein polypeptides. Carlsberg Res. Commun. 51:371-379. Overcoming self-incompatibility in Petunia axillaris. II. Placenta! pollination in vitro. ]. Indian Bot. Soc. Rastogi, R., and Sawhney, V.K. 1988a. Flower culture of a male sterile stamenless-2 mutant of tomato 50A:286-296. (Lycopersicon esculentum). Am. ]. Bot. 75:513-518. Rangaswamy, N.S., and Shivanna, K.R. 1971b. Overcoming self-incompatibility in Petunia axillaris. Rastogi, R., and Sawhney, V.K. 1988b. Suppression of stamen development by CCC and ABA in tomato floral III. Two-site pollinations in vitro. Phytomorphology buds cultured in vitro. J. Plant Physiol. 133:620-624. 21:284-289. Rao, K.V. 1995. Transient gene expression in electropo- Rastogi, R., and Sawhney, V.K. 1990a. Polyamines and flower development in the male sterile stamenless-2 rated immature embryos of rice (Oryza sativa L.). /. mutant of tomato (Lycopersicon esculentum Mill.). I. Plant Physiol. 147:71-74. Level of polyamines and their biosynthesis in norRao, K.V., Suprasanna, P., and Reddy, G.M. 1990. mal and mutant flowers. Plant Physiol. 93:439-445. Biochemical changes in embryogenic and nonembryogenic calli of Zea mays L. Plant Sci. Rastogi, R., and Sawhney, V.K. 1990b. Polyamines and flower development in the male sterile stamenless-2 66:127-130. mutant of tomato (Lycopersicon esculentum Mill.). II. Rao, M.K., and Devi, K.U. 1983. Variation in expression of Effects of polyamines and their biosynthetic genie male sterility in pearl millet. /. Hered. 74:34-38. inhibitors on the development of normal and mutant Rao, M.K., and Koduru, P.R.K. 1978. Cytogenetics of a facfloral buds cultured in vitro. Plant Physiol. tor for syncyte formation and male sterility in 93:446-452. Pennisetum americanum. Theor. Appl. Genet. 53:1-7. Rao, P.S., and Rangaswamy, N.S. 1972. In vitro develop- Rathburn, H., Song, J., and Hedgcoth, C. 1993. Cytoplasmic male sterility and fertility restoration in ment of the pollinated pistils of Nicotiana rustica L. wheat are not associated with rearrangements of Bot. Gaz. 133:350-355. mitochondrial DNA in the gene regions for cob, coxll, Rao, P.V.L., and De, D.N. 1987. Haploid plants from in or coxl. Plant Mol. Biol. 21:195-201. vitro anther culture of the leguminous tree, Peltophorum pterocarpum (DC) K. Hayne (copper Rathore, K.S., Cork, R.J., and Robinson, K.R. 1991. A cytoplasmic gradient of Ca2+ is correlated with the pod). Plant Cell Tissue Organ Cult. 11:167-177. Raquin, C, Amssa, M., Henry, Y., de Buyser, J., and Essad, growth of lily pollen tubes. Devel Biol. 148:612-619. S. 1982. Origine des plantes polyploi'des obtenues Rathore, K.S., Hodges, T.K., and Robinson, K.R. 1988.
626
References
Ionic basis of currents in somatic embryos of Danais Nicotiana plumbaginifolia and Digitalis lanata. Plant carota. Planta 175:280-289. Sci. 82:47-58. Raudaskoski, M., Astrom, H., Pertilla, K., Virtanen, I., and Reinbothe, C, Tewes, A., Lehmann, J., Parthier, B., and Louhelainen, J. 1987. Role of the microtubule Reinbothe, S. 1994. Induction by methyl jasmonate cytoskeleton in pollen tubes: an immunocytochemiof embryogenesis-related proteins and mRNAs in cal and ultrastructural approach. Biol. Cell. Nicotiana plumbaginifolia. Plant Sci. 104:59-70. 61:177-188. Reinbothe, C, Tewes, A., Luckner, M., and Reinbothe, S. Ray, A., Robinson-Beers, K., Ray, S., Baker, S.C, Lang, J.D., 1992. Differential gene expression during somatic Preuss, D., Milligan, S.B., and Gasser, C.S. 1994. embryogenesis in Digitalis lanata analyzed by in vivo Arabidopsis floral homeotic gene BELL (BEL1) conand in vitro protein synthesis. Plant /. 2:917-926. trols ovule development through negative regula- Reinbothe, S., Machmudova, A., Wasternack, C, tion of AGAMOUS gene (AG). Proc. Natl Acad. Sci. Reinbothe, C, and Parthier, B. 1992. JasmonateUSA 91:5761-5765. induced proteins in cotton: immunological relationship to the respective barley proteins and homology Raynal, M, Depigny, D., Cooke, R., and Delseny, M. 1989. of transcripts to late embryogenesis abundant (Lea) Characterization of a radish nuclear gene expressed mRNAs. /. Plant Growth Regul. 11:7-14. during late seed maturation. Plant Physiol. 91:829-836. Reinert, J. 1959. Uber die Kontrolle der Morphogenese und die Induktion von Adventivembryonen an Raynal, M., Depigny, D., Grellet, R, and Delseny, M. 1991. Gewebekulturen aus Karotten. Planta 53:318-333. Characterization and evolution of napin-encoding genes in radish and related crucifers. Gene 99:77-86. Reinert, J., Tazawa, M., and Semenoff, S. 1967. Nitrogen compounds as factors of embryogenesis in vitro. Reddy, B.V.S., Green, J.M., and Bisen, S.S. 1978. Genetic Nature 216:1215-1216. male sterility in pigeon pea. Crop Sci. 18:362-364. Reddy, J.T., Dudareva, N., Evrard, J.-L., Krauter, R., Reinholz, E. 1959. Beeinflussung der Morphogenese Steinmetz, A., and Pillay, D.T.N. 1995. A pollen-speembryonaler Organe durch ionisierende cific gene from sunflower encodes a member of the Strahlungen. I. Keimlingsanomalien durch leucine-rich-repeat protein superfamily. Plant Sci. Rontgenbestrahlung von Arabidopsis thalianaEmbryonen in verschiedenen Entwicklungsstadien. 111:81-93. Strahlentherapie 109:537-553. Reddy, V.M., and Daynard, T.B. 1983. Endosperm characteristics associated with rate of grain filling and ker- Reinold, S., Hauffe, K.D., and Douglas, C.J. 1993. Tobacco and parsley 4-coumarate:coenzyme A ligase genes nel size in corn. Maydica 28:339-355. are temporally and spatially regulated in a cell typeReeves, CD., Krishnan, H.B., and Okita, T.W. 1986. Gene specific manner during tobacco flower developexpression in developing wheat endosperm. ment. Plant Physiol. 101:373-383. Accumulation of gliadin and ADPglucose pyrophosphorylase messenger RNAs. Plant Physiol. 82:34-40. Reiser, L., Modrusan, Z., Margossian, L., Samach, A., Ohad, N., Haughn, G.W., and Fischer, R.L. 1995. The Reeves, CD., and Okita, T.W. 1987. Analyses of a/p-type BELLI gene encodes a homeodomain protein gliadin genes from diploid and hexaploid wheats. involved in pattern formation in the Arabidopsis Gene 52:257-266. ovule primordium. Cell 83:735-742. Regan, S.M., and Moffatt, B.A. 1990. Cytochemical analysis of pollen development in wild-type Arabidopsis Reiss, H.-D., and Herth, W. 1978. Visualization of the Ca2+-gradient in growing pollen tubes of Lilium and a male-sterile mutant. Plant Cell 2:877-889. longiflorum with chlorotetracycline fluorescence. Reger, B.J., Chaubal, R., and Pressey, R. 1992. Protoplasma 97:373-377. Chemotropic responses by pearl millet pollen tubes. Sex. Plant Reprod. 5:47-56. Reiss, H.-D., and Herth, W. 1979a. Calcium ionophore A 23187 affects localized wall secretion in the tip Reger, B.J., Pressey, R., and Chaubal, R. 1992. In vitro region of pollen tubes of Lilium longiflorum. Planta chemotropism of pearl millet pollen tubes to stigma 145:225-232. tissue: a response to glucose produced in the medium by tissue-bound invertase. Sex. Plant Reiss, H.-D., and Herth, W. 1979b. Calcium gradients in Reprod. 5:201-205. tip growing plant cells visualized by chlorotetracycline fluorescence. Planta 146:615-621. Rehm, S. 1952. Male sterile plants by chemical treatment. Nature 170:38-39. Reiss, H.-D., and Herth, W. 1980. Broad-range effects of Reina, M., Guillen, P., Ponte, I., Boronat, A., and Palau, J. ionophore X-537A on pollen tubes of Lilium longiflo1990a. DNA sequence of the gene encoding the Zcl rum. Planta 147:295-301. protein from Zen mays W64 A. Nucl. Acids Res. 18:6425.Reiss, H.-D., and Herth, W. 1982. Disoriented growth of pollen tubes of Lilium longiflorum Thunb. induced by Reina, M., Ponte, I., Guillen, P., Boronat, A., and Palau, J. prolonged treatment with the calcium-chelating 1990b. Sequence analysis of a genomic clone encodantibiotic, chlorotetracycline. Planta 156:218-225. ing a Zc2 protein from Zea mays VV64 A. Nncl. Acids Reiss, H.-D., and Herth, W. 1985. Nifedipine-sensitive calRes. 18:6426. cium channels are involved in polar growth of lily Reinbothe, C, Diettrich, B., and Luckner, M. 1990. pollen tubes. /. Cell Sci. 76:247-254. Regeneration of plants from somatic embryos of Digitalis lanata. J. Plant Physiol. 137:224-228. Reiss, H.-D., Herth, W., and Nobiling, R. 1985. Development of membrane- and calcium-gradients Reinbothe, C, Tewes, A., and Reinbothe, S. 1992a. during pollen germination of Lilium longiflorum. Comparative molecular analysis of gene expression Planta 163:84-90. during plant embryogenesis: do evolutionarily conserved mechanisms control early plant develop- Reiss, H.-D., Herth, W., and Schnepf, E. 1985. Plasmament? AgBiotech Nezvs Inform. 4:381N-397N. membrane "rosettes" are present in the lily pollen tube. Naturwissenschaften 72:276. Reinbothe, C, Tewes, A., and Reinbothe, S. 1992b. Altered gene expression during somatic embryogenesis in Reiss, H.-D., and McConchie, C.A. 1988. Studies of Najas
References
627
pollen tubes. Fine structure and the dependence of anther cultures. Plant Physiol. 100:1744-1750. chlorotetracycline fluorescence on external free ions. Reynolds, T.L., and Raghavan, V. 1982. An autoradiProtoplasma 142:25-35. ographic study of RNA synthesis during maturation and germination of pollen grains of Hyoscyamus Reiss, H.-D., and Nobiling, R. 1986. Quin-2 fluorescence niger. Protoplasma 111:177-188. in lily pollen tubes. Distribution of free cytoplasmic calcium. Protoplasma 131:244-246. Reznickova, S.A., and Bogdanov, Y.F. 1972. Meiosis in Reiss, H.-D., Herth, W., Schnepf, E., and Nobilin, R. 1983. excised anthers of Lilium candidum. Biol. Zentral. The tip-to-base calcium gradient in pollen tubes of 91:409-428. Lilium longiflorum measured by proton-induced X- Reznickova, S.A., and Dickinson, H.G. 1982. Ultrastructural ray emission (PIXE). Protoplasma 115:153-159. aspects of storage lipid mobilization in the tapetum of Lilium hybrida var. Enchantment. Planta 155:400-408. Rembert, D.H., Jr. 1971. Phylogenetic significance of megaspore tetrad patterns in Leguminales. Reznickova, S.A., and Willemse, M.T.M. 1980. Formation Phytomorphology 21:1-9. of pollen in the anther of Lilium. II. The function of Rembur, J., Nougarede, A., Rondet, P., and Francis, D. the surrounding tissues in the formation of pollen 1992. Floral-specific polypeptides in Silene coeli-rosa. and pollen wall. Ada Bot. Neerl. 29:141-156. Can. J. Bot. 70:2326-2633. Richards, A.J., and Ibrahim, H.B. 1982. The breeding sysRemy, R., and Ambard-Bretteville, F. 1983. Two dimentem in Primula veris L. II. Pollen tube growth and sional analysis of chloroplast proteins from normal seed-set. New Phylol. 90:305-314. and cytoplasmic male sterile Brassica napus. Theor.Richards, A.J., and Mitchell, J. 1990. The control of incomAppl. Genet. 64:249-253. patibility in distylous Pulmonaria affinis Jordan (Boraginaceae). Bot. ]. Linn. Soc. 104:369-380. Renner, O. 1914. Befruchtung und Embryobildung bei Oenothera lamarckiana und einigen verwandten Richards, J.H., and Barrett, S.C.H. 1984. The developmenArten. Flora 107:115-150. tal basis of tristyly in Eichhornia paniculata (Pontederiaceae). Am. J. Bot. 71:1347-1363. Rerie, W.G., Whitecross, M., and Higgins, T.J.V. 1991. Developmental and environmental regulation of pea Richards, J.H., and Barrett, S.C.H. 1987. Development of legumin genes in transgenic tobacco. Mol. Gen. tristyly in Pontederia cordata (Pontederiaceae). I. Genet. 225:148-157. Mature floral structure and patterns of relative growth of reproductive organs. Am. ]. Bot. Rest, J.A., and Vaughan, J.G. 1972. The development of 74:1831-1841. protein and oil bodies in the seed of Sinapis alba L. Planta 105:245-262. Rick, CM. 1945. A survey of cytogenetic causes of unfruitReynolds, J.D., and Dashek, W.V. 1976. Cytochemical fulness in the tomato. Genetics 30:347-362. analysis of callose localization in Lilium longiflorum Rick, CM. 1948. Genetics and development of nine malepollen tubes. Ann. Bot. 40:409-416. sterile tomato mutants. Hilgardia 18:599-633. Reynolds, T.L. 1984a. Callus formation and organogenesis Rietsema, J., Satina, S., and Blakeslee, A.F. 1953. The effect in anther cultures of Solanum carolinense L. /. Plant of sucrose on the growth of Datura stramonium Physiol. 117:157-161. embryos in vitro. Am. ]. Bot. 40:538-545. Reynolds, T.L. 1984b. An ultrastructural and stereological Rifot, M. 1973. Evolution structurale du pole chalazien du analysis of pollen grains of Hyoscyamus niger during sac embryonnaire d'Aquilegia vulgaris en liaison avec normal ontogeny and induced embryogenic develson activite trophique. Compt. Rend. Acad. Sci. Paris opment. Am. J. Bot. 71:490-504. 277D:1313-1316. Reynolds, T.L. 1985. Ultrastructure of anomalous pollen Rigau, J., Capellades, M., Montoliu, L., Torres, M.A., development in embryogenic anther cultures of Romera, C, Martinez-Izquierdo, J.A., Tagu, D., and Hyoscyamus niger. Am. ). Bot. 72:44-51. Puigdomenech, P. 1993. Analysis of a maize a-tubulin gene promoter by transient expression and in Reynolds, T.L. 1986. Pollen embryogenesis in anther cultransgenic tobacco plants. Plant ]. 4:1043-1050. tures of Solanum carolinense L. Plant Cell Rep. 5:273-275. Riggs, CD., and Hasenkampf, C.A. 1991. Antibodies directed against a meiosis-specific, chromatin-assoReynolds, T.L. 1987. A possible role for ethylene during ciated protein identify conserved meiotic epitopes. IAA-induced pollen embryogenesis in anther culChromosoma 101:92-98. tures of S. carolinense L. Am. }. Bot. 74:967-969. Reynolds, T.L. 1989. Ethylene effects on pollen callus for- Riggs, CD., Voelker, T.A., and Chrispeels, M.J. 1989. mation and organogenesis in anther cultures of Cotyledon nuclear proteins bind to DNA fragments harboring regulatory elements of phytohemaggluSolanum carolinense L. Plant Sci. 61:131-136. tinin genes. Plant Cell 1:609-621. Reynolds, T.L. 1990. Interactions between calcium and auxin during pollen androgenesis in anther cultures Riggs, CD., Hunt, D.C, Lin, J., and Chrispeels, M.J. 1989. Utilization of luciferase fusion genes to monitor difof Solanum carolinense L. Plant Sci. 72:109-114. ferential regulation of phytohemagglutinin and Reynolds, T.L. 1993. A cytological analysis of microspores phaseolin promoters in transgenic tobacco. Plant Sci. of Triticum aestivum (Poaceae) during normal 63:47-57. ontogeny and induced embryogenic development. Am. ]. Bot. 80:569-576. Rihova, L., Capkova, V., and Tupy, J. 1996. Changes in Reynolds, T.L., and Crawford, R.L. 1996. Changes in glycoprotein patterns associated with male gametoabundance of an abscisic acid-responsive, early cysphyte development and with induction of pollen teine-labeled metallothionein transcript during embryogenesis in Nicotiana tabacum L. /. Plant pollen embryogenesis in bread wheat (Triticum aesPhysiol. 147:573-581. tivum). Plant Mol. Biol. 32:823-829. Rijven, A.H.G.C. 1952. In vitro studies on the embryo of Capsella bursa-pastoris. Ada Bot. Neerl. 1:157-200. Reynolds, T.L., and Kitto, S.L. 1992. Identification of embryoid-abundant genes that are temporally Rijven, A.H.G.C. 1956. Glutamine and asparagine as expressed during pollen embryogenesis in wheat nitrogen sources for the growth of plant embryos in
628
References
vitro: a comparative study of 12 species. Aust. ]. Bio/. Sri. 9:511-527. Riley, R., and Bennett, M.D. 1971. Meiotic DNA synthesis. Nature 230:182-185. Risuefio, M.C., Gimgnez-Marh'n, G., L6pez-Saez, J.F., and Rodrfguez-Garcia, M.I. 1969. Origin and development of sporopollenin bodies. Protoplasma 67:361-374. Ritala, A., Aikasalo, R., Aspegren, K., SalmenkallioMarttila, M., Akerman, Sv Mannonen, L., Kurt£n, U., Puupponen-Pimia, R., Teeri, T.H., and Kauppinen, V. 1995. Transgenic barley by particle bombardment. Inheritance of the transferred gene and characteristics of transgenic barley plants. Euphytica 85:81-88. Ritala, A., Aspegren, K., Kurten, U., and SalmenkallioMarttila, M. 1994. Fertile transgenic barley by particle bombardment of immature embryos. Plant Mol. Biol. 24:317-325. Rittscher, M., and Wiermann, R. 1983. Occurrence of phenylalanine ammonia-lyase (PAL) in isolated tapetum cells of Tulipa anthers. Protoplasma 118:219-224. Rivers, B.A., and Bernatzky, R. 1994. Protein expression of a self-compatible allele from Lycopersicon peruvianum: introgression and behavior in a self-incompatible background. Sex. Plant Reprod. 7:357-362. Rivers, B.A., Bernatzky, R., Robinson, S.J., and JahnenDechent, W. 1993. Molecular diversity at the selfincompatibility locus is a salient feature in natural populations of wild tomato {Lycopersicon peruvianum). Mol. Gen. Genet. 238:419^27. Rivin, C.J., and Grudt, T. 1991. Abscisic acid and the developmental regulation of embryo storage proteins in maize. Plant Physiol. 95:358-365. Roath, W.W., and Hockett, E.A. 1971. Genetic male sterility in barley. III. Pollen and anther characteristics. Crop Sri. 11:200-203. Robert, L.S., Thompson, R.D., and Flavell, R.B. 1989. Tissue-specific expression of a wheat high molecular weight glutenin gene in transgenic tobacco. Plant Cell 1:569-578. Robert, L.S., Allard, S., Franklin, T.M., and Trick, M. 1994a. Sequence and expression of endogenous Slocus glycoprotein genes in self-compatible Brassica
Ultrastructure and physiology of the stigmatic papillar cells. /. Cell Sci. 66:241-253. Roberts, I.N., Harrod, G., and Dickinson, H.G. 1984b. Pollen-stigma interactions in Brassica oleracea. II. The fate of stigma surface proteins following pollination and their role in the self-incompatibility response. /. Cell Sci. 66:255-264. Roberts, I.N., Stead, A.D., Ockendon, D.J., and Dickinson, H.G. 1979. A glycoprotein associated with the acquisition of the self-incompatibility system by maturing stigmas of Brassica oleracea. Planta 146:179-183.
Roberts, J.K., DeSimone, N.A., Lingle, W.L., and Dure, L., III. 1993. Cellular concentrations and uniformity of cell-type accumulation of two Lea proteins in cotton embryos. Plant Cell 5:769-780. Roberts, M.R., Boyes, E., and Scott, R.J. 1995. An investigation of the role of the anther tapetum during microspore development using genetic cell ablation. Sex. Plant Reprod. 8:299-307.
Roberts, M.R., Hodge, R., and Scott, R. 1995. Brassica napus pollen oleosins possess a characteristic C-terminal domain. Planta 195:469-470. Roberts, M.R., Foster, G.D., Blundell, R.P., Robinson, S.W., Kumar, A., Draper, J., and Scott, R. 1993a. Gametophytic and sporophytic expression of an anther-specific Arabidopsis thaliana gene. Plant J. 3:111-120. Roberts, M.R., Hodge, R., Ross, J.H.E., Sorensen, A., Murphy, D.J., Draper, J., and Scott, R. 1993b. Characterization of a new class of oleosins suggests a male gametophyte-specific lipid storage pathway. Plant J. 3:629-636. Roberts, M.R., Robson, F., Foster, G.D., Draper, J., and Scott, R.J. 1991. A Brassica napus mRNA expressed specifically in developing microspores. Plant Mol. Biol. 17:295-299. Robertson, D., Weissinger, A.K., Ackley, R., Glover, S., and Sederoff, R.R. 1992. Genetic transformation of Norway spruce (Picea abies (L.) Karst) using somatic embryo explants by microprojectile bombardment. Plant Mol. Biol. 19:925-935.
Robertson, D.S. 1952. The genotype of the endosperm and embryo as it influences vivipary in maize. Proc. Natl Acad. Sci. USA 38:580-583.
Robertson, D.S. 1955. The genetics of vivipary in maize. Genetics 40:745-760. Robert, L.S., Allard, S., Gerster, J.L., Cass, L., and Simmonds, J. 1993. Isolation and characterization of Robertson, M., Walker-Simmons, M., Munro, D., and Hill, R.D. 1989. Induction of oc-amylase inhibitor synthea polygalacturonase gene highly expressed in sis in barley embryos and young seedlings by Brassica napns pollen. Plant Mol. Biol. 23:1273-1278. abscisic acid and dehydration stress. Plant Physiol. Robert, L.S., Allard, S., Gerster, J.L., Cass, L., and 91:415^120. Simmonds, J. 1994b. Molecular analysis of two Brassica napus genes expressed in the stigma. Plant Robichaud, C, and Sussex, I.M. 1986. The response of viviparous-1 and wild type embryos of Zea mays to Mol. Biol. 26:1217-1222. culture in the presence of abscisic acid. /. Plant Robert, L.S., Gerster, J., Allard, S., Cass, L., and Simmonds, Physiol. 126:235-242. J. 1994c. Molecular characterization of two Brassica napus genes related to oleosins which are highly Robichaud, C, and Sussex, I.M. 1987. The uptake and metabolism of [2-14C]-ABA by excised wild type and expressed in the tapetum. Plant J. 6:927-933. viviparous-1 embryos of Zea mays L. /. Plant Physiol. Roberton, M., and Chandler, P.M. 1992. Pea dehydrins: 130:181-188. identification, characterisation and expression. Plant Robichaud, C.S., Wong, J., and Sussex, I.M. 1980. Control Mol. Biol. 19:1031-1044. of in vitro growth of viviparous embryo mutants of Roberts, D.R., Flinn, B.S., Webb, D.T., Webster, F.B., and maize by abscisic acid. Devel Genet. 1:325-330. Sutton, B.C.S. 1990. Abscisic acid and indole-3butyric acid regulation of maturation and accumula- Robie, C.A., and Minocha, S.C. 1989. Polyamines and tion of storage proteins in somatic embryos of somatic embryogenesis in carrot. I. The effects of interior spruce. Physiol. Plant. 78:355-360. difluoromethylornithine and difluoromethylarginine. Plant Sci. 65:45-54. Roberts, I.N., Harrod, G., and Dickinson, H.G. 1984a. Pollen-stigma interactions in Brassica oleracea I. Robinson-Beers, K., Pruitt, R.E., and Gasser, C.S. 1992. napus. Mol. Gen. Genet. 242:209-216.
References
629
Ovule development in wild-type Arabidopsis and observed with the electron microscope. Flora two female-sterile mutants. Plant Cell 4:1237-1249. 158:181-188. Rochange, R, Serrano, L., Marque, C, Teulieres, C, and Rodriguez-Garcia, M.I., and Fernandez, M.C. 1987. Boudet, A.-M. 1995. DNA delivery into Eucalyptus Cytoplasmic nucleoloids during microsporogenesis globulus zygotic embryos through biolistics: optiin Olea europaea L. Biol. Cell. 60:155-160. mization of the biological and physical parameters Rodn'guez-Garcia, M.I., Fernandez, M.C, and Alch6, J.D. of bombardment for two different particle guns. 1995. Immunocytochemical localization of allergenic Plant Cell Rep. 14:674-678. protein (Ole e I) in the endoplasmic reticulum of the Rode, A., Hartmann, C, de Buyser, J., and Henry, Y. 1988. developing pollen grain of olive (Olea europaea L.). Planta 196:558-563. Evidence for a direct relationship between mitochondrial genome organization and regeneration Rodriguez-Garcia, M.I., and Garcia, A. 1978. ability in hexaploid wheat somatic tissue cultures. Differentiation of the plastid population during Curr. Genet. 14:387-394. microsporogenesis and the development of the pollen grain in the Liliaceae. Biol. Cell. 33:63-70. Rodin, J., and Rask, L. 1990. Characterization of matteuccin, the 2.2S storage protein of the ostrich fern. Rodn'guez-Garcia, M.I., Fernandez, M.C, Alch£, J.D., and Evolutionary relationship to angiosperm seed storOlmedilla, A. 1995. Endoplasmic reticulum as a storage proteins. Eur. f. Biochem. 192:101-107. age site for allergenic proteins in pollen grains of several Oleaceae. Protoplasma 187:111-116. Rodin, ]., Ericson, M.L., Josefsson, L.-G., and Rask, L. 1990. Characterization of a cDNA clone encoding a Rodriguez-Sotres, R., and Black, M. 1994. Osmotic potenBrassica napus 12 S protein (cruciferin) subunit. tial and abscisic acid regulate triacylglycerol syntheRelationship between precursors and mature chains. sis in developing wheat embryos. Planta 192:9-15. /. Biol. Chem. 265:2720-2723. Roeckel, P., and Dumas, C. 1993. Survival at 20°C and cryRodin, S., Sjodahl, S., Josefsson, L.-G., and Rask, L. 1992. opreservation of isolated sperm cells from Zea mays Characterization of a Brassica napus gene encoding a pollen grains. Sex. Plant Reprod. 6:212-216. cruciferin subunit: estimation of sizes of cruciferin Roeckel, P., Heizmann, P., Dubois, M., and Dumas, C. gene families. Plant Mol. Biol. 20:559-563. 1988. Attempts to transform Zea mays via pollen Rodkiewicz, B. 1960. Measurements of desoxyribose grains. Effect of pollen and stigma nuclease activities. Sex. Plant Reprod. 1:156-163. nucleic acid by Feulgen-photometry in nuclei of pollen grains of Tradescantia bracteata. Ada Soc. Bot. Rogers, CM. 1979. Distyly and pollen dimorphism in Polon. 29:211-217. Linum suffruticosum (Linaceae). Plant Syst. Evol. 131:127-132. Rodkiewicz, B. 1970. Callose in cell walls during megasporogenesis in angiosperms. Planta 93:39-47. Rogers, CM., and Harris, B.D. 1969. Pollen exine deposition: a clue to its control. Am. j. Bot. 56:1209-1211. Rodkiewicz, B., and Bednara, J. 1974. Distribution of organelles and starch grains during megasporogen- Rogers, H.J., Greenland, A.J., and Hussey, P.J. 1993. Four esis in Epilobium. In Fertilization in Higher Plants, ed. members of the maize p-tubulin gene family are H.F. Linskens, pp. 89-95. Amsterdam: Northexpressed in the male gametophyte. Plant j . Holland Publishing Co. 4:875-882. Rodkiewicz, B., and Bednara, J. 1976. Cell wall ingrowths Rogers, H.J., Harvey, A., and Lonsdale, D.M. 1992. and callose distribution in megasporogenesis in Isolation and characterization of a tobacco gene with some Orchidaceae. Phyto?norphology 26:276-281. homology to pectate lyase which is specifically Rodkiewicz, B., Duda, E., and Bednara, J. 1989. Organelle expressed during microsporogenesis. Plant Mol. Biol. aggregations during microsporogenesis in 20:493-502. Nymphaea. Flora 183:397-404. Rogers, H.J., Allen, R.L., Hamilton, W.D.O., and Lonsdale, Rodkiewicz, B., Fyk, B., and Szczuka, E. 1994. Chlorophyll D.M. 1991. Pollen specific cDNA clones from Zea mays. Biochim. Biophys. Ada 1089:411-413. and cutin in early embryogenesis in Capsella, Arabidopsis, and Stellaria investigated by fluores- Roggen, H.P., and Stanley, R.G. 1971. Autoradiographic cence microscopy. Sex. Plant Reprod. 7:287-292. studies of pear pollen tube walls. Physiol. Plant. 24:80-84. Rodkiewicz, B., and G6rska-Brylass, A. 1967. Occurrence of callose in the walls of meiotically dividing cell in Roggen, H.P.J.R. 1967. Changes in enzyme activities durthe ovule of Orchis. Naturwissenschaften 54:499-500. ing the progame phase in Petunia hybrida. Ada Bot. Neerl. 16:1-31. Rodkiewicz, B., and G6rska-Brylass, A. 1968. Callose in the walls of the developing megasporocyte and Roggen, H.P.J.R. 1972. Scanning electron microscopical megaspores in the orchid ovule. Ada Soc. Bot. Polon. observations on compatible and incompatible 37:19-28. pollen-stigma interactions in Brassica. Euphytica Rodkiewicz, B., and Mikulska, E. 1963. Electron micro21:1-10. scope observations of cytoplasmic changes in devel- Roggen, H.P.J.R., and Stanley, R.G. 1969. Cell-walloping megasporocyte of Lilium candidum. Flora hydrolysing enzymes in wall formation as measured 154:383-387. by pollen-tube extension. Planta 84:295-303. Rodkiewicz, B., and Mikulska, E. 1965a. The development Roiz, L., Goren, R., and Shoseyov, O. 1995. Stigmatic of cytoplasmic structures in the embryo sac of Liliwn RNase in calamondin (Citrus reticulata var. austera x candidum, as observed with the electron microscope. Fortunella sp.). Physiol. Plant. 94:585-590. Planta 67:297-304. Roiz, L., and Shoseyov, O. 1995. Stigmatic RNase in selfRodkiewicz, B., and Mikulska, E. 1965b. Electron microcompatible peach (Prunus persica). Int. J. Plant Sci. scope observations of endoplasmic reticulum in 156:37-41. Lilium megasporocyte. Flora 155:341-346. Roman, H. 1948. Directed fertilization in maize. Proc. Natl Rodkiewicz, B., and Mikulska, E. 1967. The micropylar Acad. Sci. USA 34:36-42. and antipodal cells of the Lilium regale embryo sac Rondet, P. 1962. L'organogenese au cours de l'embryo-
630
References
genese chez VAlyssum marititnum Lamk. Compt. Rougier, M., Jnoud, N., and Dumas, C. 1988. Localization Rend. Acad. Sci. Paris 255:2278-2280. of adenylate cyclase activity in Populus: its relation to pollen-pistil recognition and incompatibility. Sex. Rose, R.E., Dejesus, C.E., Moylan, S.L., Ridge, N.P., Plant Reprod. 1:140-149. Scherer, D.E., and Knauf, V.C. 1987. The nucleotide sequence of a cDNA clone encoding acyl carrier pro- Rougier, M., Jnoud, N., Said, C, Russell, S., and Dumas, tein (ACP) from Brassica campestris seeds. Nucl. Acids C. 1991. Male gametophyte development and formaRes. 15:7197. tion of the male germ unit in Populus deltoides following compatible pollination. Protoplasma Rosen, W.G. 1961. Studies on pollen-tube chemotropism. 162:140-150. Am. J. Bot. 48:889-895. Rosen, W.G., and Gawlik, S.R. 1966. Fine structure of lily Rougier, M., Jnoud, N., Said, C, Russell, S., and Dumas, C. 1992. Interspecific incompatibility in Populus: pollen tubes following various fixation and staining inhibition of tube growth and behaviour of the male procedures. Protoplasma 61:181-191. germ unit in P. deltoides X P. alba cross. Protoplasma Rosen, W.G., and Thomas, H.R. 1970. Secretory cells of 168:107-112. lily pistils. I. Fine structure and function. Am. /. Bot. 57:1108-1114. Rounsley, S.D., Ditta, G.S., and Yanofsky, M.F. 1995. Diverse roles for MADS box genes in Arabidopsis Rosen, W.G., Gawlik, S.R., Dashek, W.V., and development. Plant Cell 7:1259-1269. Siegesmund, K.A. 1964. Fine structure and cytochemistry of Lilium pollen tubes. Am. ]. Bot. 51:61-71. Roussell, D.L., Boston, R.S., Goldsbrough, P.B., and Larkins, B.A. 1988. Deletion of DNA sequences Rosenberg, L.A., and Rinne, R.W. 1988. Protein synthesis flanking an Mr 19000 zein gene reduces its transcripduring natural and precocious soybean seed (Glycine max [L.] Merr.) maturation. Plant Physiol. 87:474-178. tional activity in heterologous plant tissues. Mol. Gen. Genet. 211:202-209. Rosenfield, C.-L., Fann, C, and Loewus, F.A. 1978. Metabolic studies on intermediates in the mi/o-inosi- Roustan, J.-P., Latche, A., and Fallot, J. 1989a. Stimulation of Daucus carota somatic embryogenesis by tol oxidation pathway in Lilium longiflorum pollen. I. inhibitors of ethylene synthesis: cobalt and nickel. Conversion to hexoses. Plant Physiol. 61:89-95. Plant Cell Rep. 8:182-185. Rosenfield, C.-L., and Matile, P. 1979. Glycosidases in pear pollen tube development. Plant Cell Physiol. Roustan, J.-P, Latche, A., and Fallot, J. 1989b. Effet de 20:605-613. l'acide salicylique et de l'acide acetylsalicylique sur la production d'£thylene et l'embryogenese somaRoss, A.H., Manners, J.M., and Birch, R.G. 1995. tique de suspensions cellulaires de carotte (Daucus Embryogenic callus production, plant regeneration carota L.). Compt. Rend. Acad. Sci. Paris Sen III and transient gene expression following particle 308:395-399. bombardment in the pasture grass, Cenchrus ciliaris (Gramineae). Atist. ]. Bot. 43:193-199. Roustan, J.-P., Latche, A., and Fallot, J. 1990. Control of carrot somatic embryogenesis by AgNO3, an Ross, J.H.E., and Murphy, D.J. 1992. Biosynthesis and inhibitor of ethylene action: effect on arginine decarlocalisation of storage proteins, oleosins and lipids boxylase activity. Plant Sci. 67:89-95. during seed development in Coriandrum sativnm and other Umbelliferae. Plant Sci. 86:59-70. Roustan, J.-P, Latche, A., and Fallot, J. 1992. Influence of ethylene on the incorporation of 3,4-[14C]methionine Ross, J.H.E., and Murphy, D.J. 1993. Differential accumuinto polyamines in Daucus carota cells during lation of storage products in developing seeds and somatic embryogenesis. Plant Physiol. Biochem. somatic cell cultures of Daiicus carota L. Plant Sci. 30:201-205. 88:1-11. Ross, J.H.E., and Murphy, D.J. 1996. Characterization of Rouwendal, G.J.A., van Damme, J.M.M., and Wessels, anther-expressed genes encoding a major class of J.G.H. 1987. Cytoplasmic male sterility in Plantago lanceolata L.: differences between male-sterile cytoextracellular oleosin-like proteins in the pollen coat plasms at the DNA- and RNA-level. Theor. Appl. of Brassicaceae. Plant}. 9:625-637. Genet. 75:59-65. Rost, T.L., and Lersten, N.R. 1970. Transfer aleurone cells in Setaria lutescens (Gramineae). Protoplasma 71:403-408.Rowland, L.J., and Chourey, P.S. 1990. In situ hybridization analysis of sucrose synthase expression in Roth, T.F., and Ito, M. 1967. DNA-dependent formation of developing kernels of maize. Maydica 35:373-382. the synaptinemal complex at meiotic prophase. /. Cell Biol. 35:247-255. Rowley, J.R. 1963. Ubisch body development in Poa anmia. Grana Palynol. 4:25-36. Rothenberg, M., and Hanson, M.R. 1987. Recombination between parental mitochondrial DNA following Rowley, J.R. 1964. Formation of the pore in pollen of Poa annua. In Pollen Physiology and Fertilization, ed. H.F. protoplast fusion can occur in a region which norLinskens, pp. 59-69. Amsterdam: North-Holland mally does not undergo intragenomic recombination Publishing Co. in parental plants. Curr. Genet. 12:235-240. Rothenberg, M., and Hanson, M.R. 1988. A functional Rowley, J.R., and Dunbar, A. 1967. Sources of membranes for exine formation. Svensk. Bot. Tidskr. 61:49-64. mitochondrial ATP synthase proteolipid gene produced by recombination of parental genes in a Rowley, J.R., and Erdtman, G. 1967. Sporoderm in Populus and Salix. Grana Palynol. 7:517-567. Petunia somatic hybrid. Genetics 118:155-161. Rothenberg, M., Boeshore, M.L., Hanson, M.R., and Izhar, Rowley, J.R., Muhlethaler, K., and Frey-Wyssling, A. 1959. A route for the transfer of materials through the S. 1985. Intergenomic recombination of mitochondrpollen grain wall. /. Biophys. Biochem. Cytol. ial genomes in a somatic hybrid plant. Curr. Genet. 6:537-538. 9:615-618. Rottmann, W.H., Brears, T., Hodge, T.P., and Lonsdale, Rowley, J.R., and Skvarla, J.J. 1975. The glycocalyx and initiation of exine spinules on microspores of Canna. D.M. 1987. A mitochondrial gene is lost via homoloAm. ]. Bot. 62:479-485. gous recombination during reversion of CMS T maize to fertility. EMBO J. 6:1541-1546. Rowley, J.R., and Southworth, D. 1967. Deposition of
631
References
sporopollenin on lamellae of unit membrane dimen- Russell, S.D. 1979. Fine structure of megagametophyte sions. Nature 213:703-704. development in Zea mays. Can. ]. Bot. 57:1093-1110. Royo, J., Kunz, C, Kowyama, Y., Anderson, M., Clarke, Russell, S.D. 1980. Participation of male cytoplasm during gamete fusion in an angiosperm, Plumbago zeylanica. A.E., and Newbigin, E. 1994. Loss of a histidine Science 210:200-201. residue at the active site of S-locus ribonuclease is associated with self-compatibility in Lycopersicon Russell, S.D. 1982. Fertilization in Plumbago zeylanica: peruviamim. Proc. Natl Acad. Sci. USA 91:6511-6514. entry and discharge of the pollen tube in the embryo Rozycka, M., Khan, S., Lopez, I., Greenland, A.J., and sac. Can. ]. Bot. 60:2219-2230. Hussey, P.J. 1995. A Zea mays pollen cDNA encoding Russell, S.D. 1983. Fertilization in Plumbago zeylanica: a putative actin depolymerizing factor. Plant Physiol. gamete fusion and the fate of the male cytoplasm. 107:1011-1012. Am.}. Bot. 70:416-434. Rubin, R., Levanony, H., and Galili, G. 1992. Evidence for Russell, S.D. 1984. Ultrastructure of the sperm of the presence of two different types of protein bodies Plumbago zeylanica. II. Quantitative cytology and in wheat endosperm. Plant Physiol. 99:718-724. three-dimensional organization. Planta 162:385-391. Rubinstein, A.L., Prata, R.T.N., and Bedinger, P.A. 1995. Russell, S.D. 1985. Preferential fertilization in Plumbago: Developmental accumulation of hydroxyproline ultrastructural evidence for gamete-level recogniand hydroxyproline-containing proteins in Zea mays tion in an angiosperm. Proc. Natl Acad. Sci. USA 82:6129-6132. pollen. Sex. Plant Reprod. 8:27-32. Rubinstein, A.L., Broadwater, A.H., Lowrey, K.B., and Russell, S.D. 1986. Isolation of sperm cells from the pollen Bedinger, P.A. 1995a. Pexl, a pollen-specific gene of Plumbago zeylanica. Plant Physiol. 81:317-319. with an extensin-like domain. Proc. Natl Acad. Sci. Russell, S.D. 1987. Quantitative cytology of the egg and USA 92:3086-3090. central cell of Plumbago zeylanica and its impact on cytoplasmic inheritance patterns. Theor. Appl. Genet. Rubinstein, A.L., Mdrquez, J., Suarez-Cervera, M., and 74:693-699. Bedinger, P.A. 1995b. Extensin-like glycoproteins in the maize pollen tube wall. Plant Cell 7:2211-2225. Russell, S.D. 1992. Double fertilization. Int. Rev. Cytol. Rudd, J.J., Franklin, F.C.H., Lord, J.M., and Franklin-Tong, 140:357-388. V.E. 1996. Increased phosphorylation of a 26-kD Russell, S.D., and Cass, D.D. 1981. Ultrastructure of the pollen protein is induced by the self-incompaibility sperms of Plumbago zeylanica. 1. Cytology and assoresponse in Papaver rhoeas. Plant Cell 8:713—724. ciation with the vegetative nucleus. Protoplasma Rueda, J., and Vazquez, A.M. 1985. Effect of auxins and 107:85-107. cytokinins upon the start of meiosis in cultured Russell, S.D., and Cass, D.D. 1988. Fertilization in anthers of rye (Secale cereale). Can. J. Genet. Cytol.
27:759-765. Ruiz-Avila, L., Ludevid, M.D., and Puigdomenech, P. 1991. Differential expression of a hydroxyprolinerich cell-wall protein gene in embryonic tissues of Zea mays L. Planta 184:130-136. Ruiz-Avila, L., Burgess, S.R., Stiefel, V., Ludevid, M.D., and Puigdomenech, P. 1992. Accumulation of cell wall hydroxyproline-rich glycoprotein mRNA is an early event in maize embryo cell differentiation.
Plumbagella micrantha. Am. J. Bot. 75:778-781.
Russell, S.D., Cresti, M., and Dumas, C. 1990. Recent progress on sperm characterization in flowering plants. Physiol. Plant. 80:669-676. Russell, S.D., and Mao, L.-J. 1990. Patterns of embryo-sac organization, synergid degeneration and cotyledon orientation in Linum usitatissimum L. Planta 182:52-57. Russell, S.D., Rougier, M., and Dumas, C. 1990. Organization of the early post-fertilization megagaProc. Natl Acad. Sci. USA 89:2414-2418. metophyte of Populus deltoides. Ultrastructure and implications for male cytoplasmic transmission. Rundle, S.J., and Nasrallah, J.B. 1992. Molecular characProtoplasma 155:153-165. terization of type 1 serine/threonine phosphatases from Brassica oleracea. Plant Mol. Biol. 20:367-375. Rutten, T.L.M., and Derksen, J. 1990. Organization of actin Rundle, S.J., Nasrallah, M.E., and Nasrallah, J.B. 1993. filaments in regenerating and outgrowing subprotoEffects of inhibitors of protein serine/threonine plasts from pollen tubes of Nicotiana tabacum L. phosphatases on pollination in Brassica. Plant Planta 180:471^79. Physiol. 103:1165-1171. Rutten, T.L.M., and Derksen, J. 1992. Microtubules in Rusche, M.L., and Mogensen, H.L. 1988. The male germ pollen tube subprotoplasts: organization during prounit of Zea mays: quantitative ultrastructure and toplast formation and protoplast outgrowth. three-dimensional analysis. In Sexual Reproduction in Protoplasma 167:231-237. Higher Plants, ed. M. Cresti, P. Gori and E. Pacini, pp. Ryan, A.J., Royal, C.L., Hutchinson, J., and Shaw, C.H. 221-226. Berlin: Springer-Verlag. 1989. Genomic sequence of a 12S seed storage proRusche, M.L., Mogensen, H.L., Zhu, T., and Smith, S.E. tein from oilseed rape (Brassica napus c.v. jet neuf). 1995. The zygote and proembryo of alfalfa: quantitaNucl. Acids Res. 17:3584. tive, three-dimensional analysis and implications for Ryczkowski, M. 1960. Changes of the osmotic value durbiparental plastid inheritance. Protoplasma ing the development of the ovule. Planta 55:343-356. 189:88-100. Ryczkowski, M. 1964. Physico-chemical properties of the Russell, D.A., and Sachs, M.M. 1991. The maize cytosolic central vacuolar sap in developing ovules. In Pollen glyceraldehyde-3-phosphate dehydrogenase gene Physiology and Fertilization, ed. H.F. Linskens, pp. family: organ-specific expression and genetic analy17-25. Amsterdam: North-Holland Publishing Co. sis. Mol. Gen. Genet. 229:219-228. Russell, D.R., Wallace, K.M., Bathe, J.H., Martinell, B.J., Saalbach, G., Christov, V., Jung, R., Saalbach, I., Manteuffel, R., Kunze, G., Brambarov, K., and and McCabe, D.E. 1993. Stable transformation of Muntz, K. 1995. Stable expression of vicilin from Phaseolus vulgaris via electric-discharge mediated Vicia faba with eight additional single methionine particle acceleration. Plant Cell Rep. 12:165-169.
632 residues but failure of accumulation of legumin with an attached peptide segment in tobacco seeds. Mol. Breed. 1:245-258. Saalbach, I., Pickardt, T., Machemehl, R, Saalbach, G., Schieder, O., and Miintz, K. 1994. A chimeric gene encoding the methionine-rich 2S albumin of the Brazil nut (Bertholletia excelsa H.B.K.) is stably expressed and inherited in transgenic grain legumes. Mol. Gen. Genet. 242:226-236. Saba-El-Leil, M.K., Rivard, S., Morse, D., and Cappadocia, M. 1994. The Sll and S13 self incompatibility alleles in Solannm chacoense Bitt. are remarkably similar. Plant Mol. Biol. 24:571-583. Safford, R., Windust, J.H.C., Lucas, C, de Silva, J., James, CM., Hellyer, A., Smith, C.G., Slabas, A.R., and Hughes, S.G. 1988. Plastid-localised seed acyl-carrier protein of Brassica napus is encoded by a distinct, nuclear multigene family. Eur. ]. Biochem. 174:287-295. Sage, T.L., and Williams, E.G. 1995. Structure, ultrastructure, and histochemistry of the pollen tube pathway in the milkweed Asclepias exaltata L. Sex. Plant Reprod. 8:257-265. Sagi, L., Panis, B., Remy, S., Schoofs, H., de Smet, K., Swennen, R., and Cammue, B.P.A. 1995a. Genetic transformation of banana and plantain {Musa spp.) via particle bombardment. Biotechnology 13:481-485. Sagi, L., Remy, S., Verelst, B., Panis, B., Cammue, B.P.A., Volckaert, G., and Swennen, R. 1995b. Transient gene expression in transformed banana {Musa cv. Bluggoe) protoplasts and embryogenic cell suspensions. Euphytica 85:89-95. Saigo, R.H., Peterson, D.M., and Holy, J. 1983. Development of protein bodies in oat starchy endosperm. Can. J. Bot. 61:1206-1215. Saini, H.S., and Aspinall, D. 1981. Effect of water deficit on sporogenesis in wheat (Triticum aestivnm L.). Ann. Bot. 48:623-633. Saini, H.S., Sedgley, M., and Aspinall, D. 1984. Developmental anatomy in wheat of male sterility induced by heat stress, water deficit or abscisic acid. Anst. ]. Plant Physiol. 11:243-253. Saito, G.Y., Chang, Y.C., Walling, L.L., and Thomson, W.W. 1989. A correlation in plastid development and cytoplasmic ultrastructure with nuclear gene expression during seed ripening in soybean. New Phytol. 113:459-469. Sakaguchi, K., Kurata, N., Takegami, M.H., and Ito, M. 1980. Inhibition of DNA and RNA syntheses and suppression of meiotic development in lily microsporocytes. Cell Struct. Fund. 5:367-377. Sakai, H., Medrano, L.J., and Meyerowitz, E.M. 1995. Role of SUPERMAN in maintaining Arabidopsis floral whorl boundaries. Nature 378:199-203. Salamini, R, di Ponzo, N., Pornasari, E., Gentinetta, E., Reggiani, R., and Soave, C. 1983. Mucronate, Me, a dominant gene of maize which interacts with opaque2 to suppress zein synthesis. Theor. Appl. Genet. 65:123-128. Salinas, J., Oeda, K., and Chua, N.-H. 1992. Two G-boxrelated sequences confer different expression patterns in transgenic tobacco. Plant Cell 4:1485-1493. San, L.H., and Gelebart, P. 1986. Production of gynogenetic haploids. In Cell Culture and Somatic Cell Genetics of Plants, Vol. 3 , ed. I.K. Vasil, pp. 305-322. Orlando: Academic Press. San Noeum, L.H. 1976. Haploides A'Hordeum vulgare L.
References
par culture in vitro d'ovaires non feconde's. Ann. Amelior. Plantes 26:751-754. Sanchez-Martinez, D., Puigdomenech, P., and Pages, M. 1986. Regulation of gene expression in developing Zea mays embryos. Protein synthesis during embryogenesis and early germination of maize. Plant Physiol. 82:543-549. Sanchez-Martinez, D., G6mez, J., Ludevid, M.D., Torrent, M., Puigdomenech, P., and Pages, M. 1987. Absence of storage protein synthesis in the embryo of Zea mays. Plant Sci. 53:215-221. Sanchez-Pina, M.A., Kieft, H., and Schel, J.H.N. 1989. Immunocytochemical detection of non-histone nuclear antigens in cryosections of developing somatic embryos from Daucus carota L. /. Cell Sci. 93:615-622. Sand, S.A., and Christoff, G.T. 1973. Cytoplasmic-chromosomal interactions and altered differentiation in tobacco. /. Hered. 64:24-30. Sanders, L.C., and Lord, E.M. 1989. Directed movement of latex particles in the gynoecia of three species of flowering plants. Science 243:1606-1608. Sanders, L.C., Wang, C.-S., Walling, L.S., and Lord, E.M. 1991. A homolog of the substrate adhesion molecule vitronectin occurs in four species of flowering plants. Plant Cell 3:629-635. Sanders, M.E., and Burkholder, PR. 1948. Influence of amino acids on growth of Datura embryos in culture. Proc. Natl Acad. Sci. USA 34:516-526. Sanford, J.C, Skubik, K.A., and Reisch, B.I. 1985. Attempted pollen-mediated plant transformation employing genomic donor DNA. Theor. Appl. Genet. 69:571-574. Sangduen, N., Kreitner, G.L., and Sorensen, E.L. 1983a. Light and electron microscopy of embryo development in perennial and annual Medicago species. Can. }. Bot. 61:837-849. Sangduen, N., Kreitner, G.L., and Sorensen, E.L. 1983b. Light and electron microscopy of embryo development in an annual X perennial Medicago species cross. Can. J. Bot. 61:1241-1257. Sanger, J.M., and Jackson, W.T. 1971a. Fine structure study of pollen development in Haemanthus katherinae Baker. I. Formation of vegetative and generative cells. /. Cell Sci. 8:289-301. Sanger, J.M., and Jackson, W.T. 1971b. Fine structure study of pollen development in Haemanthus katherinae Baker. II. Microtubules and elongation of the generative cells. /. Cell Sci. 8:303-315. Sanger, J.M., and Jackson, W.T. 1971c. Fine structure study of pollen development in Haemanthus katherinae Baker. III. Changes in organelles during development of the vegetative cell. /. Cell Sci. 8:317-329. Sangwan, R.S. 1978. Change in the amino-acid content during male gametophyte formation of Datura metel in situ. Theor. Appl. Genet. 52:221-225. Sangwan, R.S., Bourgeois, Y, and Sangwan-Norreel, B.S. 1991. Genetic transformation of Arabidopsis thaliana zygotic embryos and identification of critical parameters influencing transformation efficiency. Mol. Gen. Genet. 230:475-485. Sangwan, R.S., and Camefort, H. 1978. Action d'un choc thermique sur le contenu en acides amines des antheres et des grains de pollen embryogenes du Datura metel L. et du Nicotiana tabacum L. Compt. Rend. Acad. Sci. Paris 287D:471^74. Sangwan, R.S., and Camefort, H. 1982. Ribosomal bodies
References
633
specific to both pollen and zygotic embryogenesis in patibility-related RNases in styles of Japanese pear Datura. Experientia 38:395-397. (Pyrus serotina Rehd.). Plant Cell Physiol. 33:811-814. Sangwan, R.S., and Camefort, H. 1983. The tonoplast, a Sassa, H., Hirano, H., and Ikehashi, H. 1993. Identification specific marker of embryogenic microspores of and characterization of stylar glycoproteins associDatura cultured in vitro. Histochemistry 78:473-480. ated with self-incompatibility genes of Japanese pear, Pyrus serotina Rehd. Mol. Gen. Genet. 241:17-25. Sangwan, R.S., and Camefort, H. 1984. Cold-treatment related structural modifications in the embryogenic Sassa, R , Mase, N., Hirano, H., and Ikehashi, H. 1994. anthers of Datura. Cytologia 49:473-487. Identification of self-incompatibility-related glycoproteins in styles of apple (Mains X domestica). Theor. Sangwan, R.S., Ducrocq, C, and Sangwan-Norreel, B. Appl. Genet. 89:201-205. 1993. Agrobacterium-mediated transformation of pollen embryos in Datura innoxia and Nicotiana Sassa, H., Nishio, T., Kowyama, Y., Hirano, H., Koba, T., tabacum: production of transgenic haploid and fertile and Ikehashi, H. 1996. Self-incompatibility (S) alleles homozygous dihaploid plants. Plant Sci. 95:99-115. of the Rosaceae encode members of a distinct class of the T2/S ribonuclease superfamily. Mol. Gen. Genet. Sangwan, R.S., and Sangwan-Norreel, B.S. 1987a. 250:547-557. Ultrastructural cytology of plastids in pollen grains of certain androgenic and nonandrogenic plants. Sassen, M.M.A. 1964. Fine structure of Petunia pollen grain and pollen tube. Acta Bot. Neerl. 13:175-181. Protoplasma 138:11-22. Sangwan, R.S., and Sangwan-Norreel, B.S. 1987b. Sassen, M.M.A. 1974. The stylar transmitting tissue. Acta Bot. Neerl. 23:99-108. Biochemical cytology of pollen embryogenesis. Int. Rev. Cytol. 107:221-272. Sastri, D.C., and Shivanna, K.R. 1976. Attempts to overSangwan, R.S., Gauthier, D.A., Turpin, D.H., Pomeroy, come interspecific incompatibility in Sesamum by M.K., and Plaxton, W.C. 1992. Pyruvate-kinase using recognition pollen. Ann. Bot. 40:891-893. isoenzymes from zygotic and microspore-derived Sastri, D.C., and Shivanna, K.R. 1979. Role of pollen-wall embryos of Brassica napus. Planta 187:198-202. proteins in intraspecific incompatibility in Saccharum Sangwan-Norreel, B.S. 1978. Cytochemical and ultrabengalense. Phytomorphology 29:324-330. structural peculiarities of embryogenic pollen grains Satina, S. 1944. Periclinal chimeras in Datura in relation to and of young androgenic embryos of Datura innoxia. development and structure (a) of the style and Can. J. Bot. 56:805-817. stigma (b) of calyx and corolla. Am. J. Bot. 31:493-502. Sangwan-Norreel, B.S. 1979. Evolution du contenu en DNA nucle'aire dans les gametophytes males du Satina, S., and Rietsema, J. 1959. Seed development. In Datura innoxia au cours de la pe>iode favorable a Blakeslee: The Genus Datura, ed. A.G. Avery, S. Satina l'androgenese. Can. J. Bot. 57:450-457. and J. Rietsema, pp. 181-195. New York: Ronald Sangwan-Norreel, B.S. 1981. Evolution in vitro du contenu Press Co. en ADN nucl£aire et de la ploi'die des embryons Sato, S., Willson, C, and Dickinson, H.G. 1989. The RNA polliniques du Datura innoxia. Can. ]. Bot. 59:508-517. content of the nucleolus and nucleolus-like inclusions Sangwan-Norreel, B.S. 1983. Male gametophyte nuclear in the anther of Lilium estimated by an improved DNA content evolution during androgenic inducRNase-gold labelling method. /. Cell Sci. 94:675-683. tion in Datura innoxia Mill. Z. Pflanzenphysiol. Sato, S., Jones, K., de Los Dios Alche, J., and Dickinson, 111:47-54. H.G. 1991. Cytoplasmic nucleoloids of Lilium male Sano, Y. 1984. Differential regulation of waxy gene expresreproductive cells contain rDNA transcripts and sion in rice endosperm. Theor. Appl. Genet. share features of development with nucleoli. /. Cell 68:467-473. Sci. 100:109-118. Santanen, A., and Simola, L.K. 1992. Changes in Sato, S., Newell, C, Kolacz, K., Tredo, L., Finer, J., and polyamine metabolism during somatic embryogeneHinchee, M. 1993. Stable transformation via particle sis in Picea abies. J. Plant Physiol. 140:475-480. bombardment in two different soybean regeneration systems. Plant Cell Rep. 12:408-413. Sari Gorla, M., Frova, C, Binelli, G., and Ottaviano, E. 1986. The extent of gametophytic-sporophytic gene Sato, S., Toya, T., Kawahara, R., Whittier, R.F., Fukuda, H., expression in maize. Theor. Appl. Genet. 72:42-47. and Komamine, A. 1995. Isolation of a carrot gene Sarker, R.H., Elleman, C.J., and Dickinson, H.G. 1988. expressed specifically during early-stage somatic Control of pollen hydration in Brassica requires conembryogenesis. Plant Mol. Biol. 28:39-46. tinued protein synthesis, and glycosylation is neces- Sato, X, Thorsness, M.K., Kandasamy, M.K., Nishio, T., sary for intraspecific incompatibility. Proc. Natl Acad. Hirai, M., Nasrallah, J.B., and Nasrallah, M.E. 1991. Sci. USA 85:4340-4344. Activity of an S-locus gene promoter in pistils and Sasaki, K., Shimomura, K., Kamada, H., and Harada, H. anthers of transgenic Brassica. Plant Cell 3:867-876. 1994. IAA metabolism in embryogenic and non- Sato, Y, Hong, S.-K., Tagiri, A., Kitano, H., Yamamoto, N., embryogenic carrot cells. Plant Cell Physiol. Nagato, Y, and Matsuoka, M. 1996. A rice homeobox 35:1159-1164. gene, OSH1, is expressed before organ differentiaSasaki, Y., and Harada, H. 1991. Binding form of pollen tion in a specific region during early embryogenesis. mother cell protein in the nucleosomes of lily. Plant Proc. Natl Acad. Sci. USA 93:8117-8122. Physiol. 96:1161-1166. Satoh, H., and Omura, T. 1981. New endosperm mutaSasaki, Y, Yasuda, H., Ohba, Y, and Harada, H. 1990. tions induced by chemical mutagens in rice, Oryza Isolation and characterization of a novel nuclear prosativa L. Jap. J. Breed. 31:316-326. tein from pollen mother cells of lily. Plant Physiol. Satoh, S., Kamada, H., Harada, H., and Fujii, T. 1986. 94:1467-1471. Auxin-controlled glycoprotein release into the Sass, J.E., and Sprague, G.F. 1950. The embryology of medium of embryogenic carrot cells. Plant Physiol. "germless" maize. Iowa State Coll. ]. Sci. 24:209-218. 81:931-933. Sassa, H., Hirano, H., and Ikehashi, H. 1992. Self-incom- Saunders, J.W., and Bingham, E.T. 1972. Production of
634
References
alfalfa plants from callus tissue. Crop Sci. 12:804-808. Scalla, R., Due, G., Rigaud, J., Lefebvre, A., and Meignoz, Sauter, J.J. 1968. Histoautoradiographische Untersuchungen R. 1981. RNA containing intracellular particles in cytoplasmic male sterile faba bean (Vicia faba L.). zur Ribonudeinsaure-Synthese wahrend der Meiosis bei Paeonia teniiifolia L. Naturwissenschaften 55:236.
Sauter, J.J. 1969a. Autoradiographische Untersuchungen zur RNS- und Protein-synthese in Pollenmutterzellen, jungen Pollen und Tapetumzellen wahrend der
Plant Sci. Lett. 22:269-277.
Scallon, B., Thanh, V.H., Floener, L.A., and Nielsen, N.C. 1985. Identification and characterization of DNA clones encoding group-II glycinin subunits. Theor. Mikrosporogenese von Paeonia teniiifolia. Z. Appl. Genet. 70: 510-519. Pflatizenphysiol. 61:1-19. Scallon, B.J., Dickinson, CD., and Nielsen, N.C. 1987. Sauter, J.J. 1969b. Cytochemische Untersuchung der Characterization of a null-allele for the Gyt glycinin Histone in Zellen mit unterschiedlicher RNS- und gene from soybean. Mol. Gen. Genet. 208:107-113. Protein-Synthese. Z. Pflanzenphysiol. 60:434-449. Scanlon, M.J., Stinard, P.S., James, M.G., Myers, A.M., and Robertson, D.S. 1994. Genetic analysis of 63 mutaSauter, J.J., and Marquardt, H. 1967a. Nucleohistone und tions affecting maize kernel development isolated Ribonukleinsaure-Synthese wahrend der Pollenfrom mutator stocks. Genetics 136:281-294. entwicklung. Naturwissenschaften 54:546. Sauter, J.J., and Marquardt, H. 1967b. Die Rolle des Schaffner, M. 1906. The embryology of the shepherd's Nukleohistons bei der RNS- und Proteinsynthese purse. Ohio Naturl. 7:1-8. wahrend der Mikrosporogenese von Paeonia teniiifo- Schardl, C.L., Pring, D.R., and Lonsdale, D.M. 1985. lia L. Z. Pflanzenphysiol. 58:126-137. Mitochondrial DNA rearrangements associated with fertile revertants of S-type male-sterile maize. Cell Sautter, C, Leduc, N., Bilang, R., Iglesias, V.A., Gisel, A., 43:361-368. Wen, X., and Potrykus, I. 1995. Shoot apical meristems as a target for gene transfer by microballistics. Schardl, C.L., Lonsdale, D.M., Pring, D.R., and Rose, K.R. Euphytica 85:45-51. 1984. Linearization of maize mitochondrial chromosomes by recombination with linear episomes. Savidge, B., Rounsley, S.D., and Yanofsky, M.F. 1995. Nature 310:292-296. Temporal relationship between the transcription of two Arabidopsis MADS box genes and the floral Scharpe, A., and van Parijs, R. 1973. The formation of organ identity genes. Plant Cell 7:721-733. polyploid cells in ripening cotyledons of Pisum sativum L. in relation to ribosome and protein synSawhney, V.K. 1974. Morphogenesis of the stamenless-2 thesis. /. Expt. Bot. 24:216-222. mutant in tomato. III. Relative levels of gibberellins in the normal and mutant plants. /. Expt. Bot. Scheer, U., and Franke, W.W. 1972. Annulate lamellae in 25:1004-1009. plant cells: formation during microsporogenesis and pollen development in Canna generalis Bailey. Planta Sawhney, V.K. 1981. Abnormalities in pepper {Capsicum 107:145-159. annuum) flowers induced by gibberellic acid. Can. ]. Scheets, K., and Hedgcoth, C. 1988. Nucleotide sequence Bot. 59:8-16. of a ygliadin gene: comparisons with other y gliadin Sawhney, V.K. 1983. Temperature control of male sterility sequences show the structure of y gliadin genes and in a tomato mutant. /. Hered. 74:51-54. the general primary structure of y gliadins. Plant Sci. Sawhney, V.K., and Bhadula, S.K. 1987. Characterization 57:141-150. and temperature regulation of soluble proteins of a male sterile tomato mutant. Biochem. Genet. Scheike, R., Gerold, E., Brennicke, A., Mehring-Lemper, M., and Wricke, G. 1992. Unique patterns of mito25:717-728. chondrial genes, transcripts and proteins in different Sawhney, V.K., and Bhadula, S.K. 1988. Micromale-sterile cytoplasms of Daucus carota. Theor. Appl. sporogenesis in the normal and male-sterile staGenet. 83:419-427. menless-2 mutant of tomato (Lycopersicon esculenSchel, J.H.N., and Kieft, H. 1986. An ultrastructural study tum). Can. J. Bot. 66:2013-2021. of embryo and endosperm development during in Sawhney, V.K., Chen, K., and Sussex, I.M. 1985. Soluble vitro culture of maize ovaries (Zea mays). Can. ]. Bot. proteins of the mature floral organs of tomato 64:2227-2238. (Lycopersicon esculentum Mill.). /. Plant Physiol. 121:265-271. Schel, J.H.N., Kieft, H., and van Lammeren, A.A.M. 1984. Interactions between embryo and endosperm durSawhney, V.K., and Greyson, R.I. 1973a. Morphogenesis ing early developmental stages of maize caryopses of the stamenless-2 mutant in tomato. I. (Zea mays). Can. J. Bot. 62:2842-2853. Comparative description of the flowers and ontogeny of stamens in the normal and mutant Scherer, D., Sato, A., McCarter, D.W., Radke, S.E., Kridl, J.C., and Knauf, V.C. 1992. Non-essential repeats in plants. Am. ]. Bot. 60:514-523. the promoter region of a Brassica rapa acyl carrier Sawhney, V.K., and Greyson, R.I. 1973b. Morphogenesis of protein gene expressed in developing embryos. the stamenless-2 mutant in tomato. II. Modifications Plant Mol. Biol. 18:591-594. of sex organs in the mutant and normal flowers by Scheres, B., Wolkenfelt, H., Willemsen, V., Terlouw, M., plant hormones. Can. ]. Bot. 51:2473-2479. Lawson, E., Dean, C, and Weisbeek, P. 1994. Sawhney, V.K., and Greyson, R.I. 1979. Interpretations of Embryonic origin of the Arabidopsis primary root determination and canalisation of stamen developand root meristem initials. Development ment in a tomato mutant. Can. J. Bot. 57:2471-2477. 120:2475-2487. Sawhney, V.K., and Polowick, PL. 1986. Temperatureinduced modifications in the surface features of sta- Schernthaner, J.P., Matzke, M.A., and Matzke, A.J.M. 1988. Endosperm-specific activity of a zein gene promens of a tomato mutant: an SEM study. Protoplasma moter in transgenic tobacco plants. EMBO ]. 131:75-81. 7:1249-1255. Sawhney, V.K., and Shukla, A. 1994. Male sterility in flowering plants: are plant growth substances involved? Schiavone, F.M. 1988. Microamputation of somatic embryos of the domestic carrot reveals apical control Am. }. Bot. 81:1640-1647.
References
635
male sterility in interspecific hybrids of Epilobium. 2. of axis elongation and root regeneration. Development 103:657-664. Expression of mitochondrial genes and structure of the mitochondrial DNA. Theor. Appl. Genet. Schiavone, F.M., and Cooke, T.J. 1987. Unusual patterns of 76:565-569. somatic embryogenesis in the domesticated carrot: developmental effects of exogenous auxins and Schmulling, T., Schell, J., and Spena, A. 1988. Single genes auxin transport inhibitors. Cell Diffn 21:53-62. from Agrobacterium rhizogenes influence plant develSchiavone, F.M., and Racusen, R.H. 1990. Microsurgery opment. EMBO /. 7:2621-2629. reveals regional capabilities for pattern reestablish- Schmulling, T., Rohrig, H., Pilz, S., Walden, R., and Schell, ment in somatic carrot embryos. Devel Bio/. J. 1993. Restoration of fertility by antisense RNA in 141:211-219. genetically engineered male sterile tobacco plants. Mol. Gen. Genet. 237:385-394. Schiavone, F.M., and Racusen, R.H. 1991. Regeneration of the root pole in surgically transected carrot embryos Schnall, J.A., Cooke, T.J., and Cress, D.E. 1988. Genetic analysis of somatic embryogenesis in carrot cell culoccurs by position-dependent, proximodistal ture: initial characterization of six classes of temperreplacement of missing tissues. Development 113:1305-1313. ature-sensitive variants. Devel Genet. 9:49-67. Schick, R., and Stubbe, H. 1932. Die Gene von Antirrhinum Schnall, J.A., Hwang, C.H., Cooke, T.J., and Zimmerman, J.L. 1991. An evaluation of gene expression during majus. II. Z. Indukt. Abstamm. Vererbungs. 62:249-290. somatic embryogenesis of two temperature-sensiSchickler, H., Benner, M.S., and Messing, J. 1993. tive carrot variants unable to complete embryo Repression of the high-methionine zein gene in the development. Physiol. Plant. 82:498-504. maize inbred line Mol7. Plant J. 3:221-229. Schiefelbein, J., Galway, M., Masucci, J., and Ford, S. 1993. Schneider, T., Dinkins, R., Robinson, K., Shellhammer, J., Pollen tube and root-hair tip growth is disrupted in and Meinke, D.W. 1989. An embryo-lethal mutant of Arabidopsis thaliana is a biotin auxotroph. Devel Biol. a mutant of Arabidopsis thaliana. Plant Physiol. 131:161-167. 103:979-985. Schlag, M., and Hesse, M. 1992. The formation of the gen- Schneitz, K., Hiilskamp, M., and Pruitt, R.E. 1995. Wildtype ovule development in Arabidopsis thaliana: a erative cell in Polystachia pubescens (Orchidaceae). Sex. Plant Reprod. 5:131-137. light microscope study of cleared whole-mount tissue. Plant J. 7:731-749. Schlappi, M., and Hohn, B. 1992. Competence of immature maize embryos for Agrobacterium-mediated Schnepf, E., and Nagl, W. 1970. Uber einige gene transfer. Plant Cell 4:7-16. Strukturbesonderheiten der Suspensorzellen von Schlosser, K. 1961. Cytologische und cytochemische Phaseolus vulgaris. Protoplasma 69:133-143. Untersuchungen uber das Pollenschlauch wachstum Schopke, C, Taylor, N., Cdrcamo, R., Konan, N.K., selbststeriler Petunien. Z. Bot. 49:266-288. Marmey, P., Henshaw, G.G., Beachy, R.N., and Fauquet, C. 1996. Regeneration of transgenic cassava Schmidt, H., and Schmidt, V. 1981. Untersuchungen an plants (Manihot esculenta Crantz) from microbompollensterilen, stamenless-ahnlichen Mutanten von barded embryogenic suspension cultures. Nature Lycopersicon esculentum Mill. II. Normalisierung von Biotech. 14:731-735. ms-15 und ms-33 mit Gibberellinsaure (GA3). Biol. Zentral. 100:691-696. Schou, 0.1984. The dry and wet stigmas of Primula obconica: ultrastructural and cytochemical dimorphisms. Schmidt, R.J., Burr, F.A., and Burr, B. 1987. Transposon Protoplasma 121:99-113. tagging and molecular analysis of the maize regulaSchou, O., and Mattsson, 0.1985. Differential localization tory locus opaque-2. Science 238:960-963. of enzymes in the stigmatic exudates of Primula Schmidt, R.J., Burr, F.A., Aukerman, M.J., and Burr, B. 1990. Maize regulatory gene opaque-2 encodes a obconica. Protoplasma 125:65-74. protein with a "leucine-zipper" motif that binds to Schrauwen, J., and Linskens, H.F. 1972. Ribonuclease in styles. Planta 102:277-285. zein DNA. Proc. Natl Acad. Sci. USA 87:46-50. Schmidt, R.J., Kerudat, M., Aukerman, M.J., and Hoschek, Schrauwen, J.A.M., de Groot, P.F.M., van Herpen, M.M.A., van der Lee, T., Reynen, W.H., Weterings, G. 1992. Opaque-2 is a transcriptional activator that K.A.P., and Wullems, G.J. 1990. Stage-related expresrecognizes a specific target site in 22-kD zein genes. Plant Cell 4:689-700. sion of mRNAs during pollen development in lily and tobacco. Planta 182:298-304. Schmidt, R.J., Veit, B., Mandel, M.A., Mena, M., Hake, S., and Yanofsky, M.F. 1993. Identification and molecu- Schrauwen, J.A.M., Mettenmeyer, T., Croes, A.F., and lar characterization of ZAG1, the maize homolog of Wullems, G.J. 1996. Tapetum-specific genes: what the Arabidopsis floral homeotic gene AGAMOUS. role do they play in male gametophyte developPlant Cell 5:729-737. ment? Acta Bot. Neerl. 45:1-15. Schmit, A.-C, and Lambert, A.-M. 1987. Characterization Schrauwen, J.A.M., Reijnen, W.H., de Leeuw, H.C.G.M., and van Herpen, M.M.A. 1986. Response of pollen to and dynamics of cytoplasmic F-actin in higher plant heat stress. Acta Bot. Neerl. 35:321-327. endosperm cells during interphase, mitosis, and cytokinesis. /. Cell Biol. 105:2157-2166. Schroder, M.-B. 1984. Ultrastructural studies on plastids Schmit, A.-C, and Lambert, A.-M. 1990. Microinjected of generative and vegetative cells in the family fluorescent phalloidin in vivo reveals the F-actin Liliaceae. 1. Lilium martagon L. Biol. Zentral. dynamics and assembly in higher plant mitotic cells. 103:547-555. Schroder, M.-B. 1985a. Ultrastructural studies on plastids Plant Cell 2:129-138. of generative and vegetative cells in the family Schmitz, U.K., and Kowallik, K.V. 1987. Why are plastids Liliaceae. 2. Pritillaria imperialis and F. meleagris. Biol. maternally inherited in Epilobium? Ultrastructural observations during microgametogenesis. Plant Sci. Zentral. 104:21-27. Schroder, M.-B. 1985b. Ultrastructural studies on plastids 53:139-145. of generative and vegetative cells in Liliaceae. 3. Schmitz, U.K., and Michaelis, G. 1988. Dwarfism and
636
References
Plastid distribution during the pollen development the central cell. /. Cell Sci. 12:741-763. in Gasteria verrucosa (Mill.) Duval. Protoplasma Schulz, P., and Jensen, W.A. 1974. Capsella embryogenesis: 124:123-129. the development of the free nuclear endosperm. Schroder, M.-B. 1986a. Ultrastructural studies on plastids Protoplasma 80:183-205. of generative and vegetative cells in Liliaceae. 4. Schulz, P., and Jensen, W.A. 1977. Cotton embryogenesis: Plastid degeneration during generative cell maturathe early development of the free nuclear tion in Convallaria majalis L. Biol. Zentral. endosperm. Am. }. Bot. 64:384-394. 105:427^33. Schulz, P., and Jensen, W.A. 1981. Pre-fertilization ovule development in Capsella: ultrastructure and ultracySchroder, M.-B. 1986b. Ultrastructural studies on plastids of generative and vegetative cells in Liliaceae. 5. The tochemical localization of acid phosphatase in the behaviour of plastids during pollen development in meiocyte. Protoplasma 107:27-45. Chlorophytum comosum (Thunb.) Jacques. Theor. Appl.Schulz, P., and Jensen, W.A. 1986. Prefertilization ovule Genet. 72:840-844. development in Capsella: the dyad, tetrad, developSchroder, M.-B., and Hagemann, R. 1986. Ultrastructural ing megaspore, and two-nucleate gametophyte. Can. studies on plastids of generative and vegetative cells J. Bot. 64:875-884. in Liliaceae. 6. Patterns of plastid distribution during Schulz, P.J., Cross, J.W., and Almeida, E. 1993. Chemical generative cell formation in Aloe secundiflora and A. agents that inhibit pollen development: effects of the jucunda. Ada Bot. Neerl. 35:243-248. phenylcinnoline carboxylates SC-1058 and SC-1271 on the ultrastructure of developing wheat anthers Schroder, M.-B., and Oldenburg, H. 1990. Ultrastructural (Triticum aestivum L. var. Yecora rojb). Sex. Plant studies on plastids of generative and vegetative cells Reprod. 6:108-121. in Liliaceae. 7. Plastid distribution during generative cell development in Tulbaghia violacea Harv. FloraSchulz, R., and Jensen, W.A. 1968a. Capsella embryogene184:131-136. sis: the early embryo. /. Ultrastr. Res. 22:376-392. Schroeder, H.E. 1982. Quantitative studies on the cotyle- Schulz, R., and Jensen, W.A. 1968b. Capsella embryogenesis: the synergids before and after fertilization. Am. J. donary proteins in the genus Pisum. ]. Sci. Food Agr. 33:623-633. Bot. 55:541-552. Schroeder, H.E. 1984. Effects of applied growth regulators Schulz, R., and Jensen, W.A. 1968c. Capsella embryogenesis: the egg, zygote, and young embryo. Am. ]. Bot. on pod growth and seed protein composition in Pisum sativum L. /. Expt. Bot. 35:813-821. 55:807-819. Schroeder, H.E., Schotz, A.H., Wardley-Richardson, X, Schumann, CM., and Hancock, J.F. 1989. Paternal inheriSpencer, D., and Higgins, T.J.V. 1993. Transformation tance of plastids in Medicago sativa. Theor. Appl. Genet. 78:863-866. and regeneration of two cultivars of pea (Pisum sativum L.). Plant Physiol. 101:751-757. Schuster, W. 1961. Untersuchungen iiber kiinstlich induzierte Pollensterilitat bei Sonnenblumen Schubert, R., Baumlein, H., Czihal, A., and Wobus, U. (Helianthus annuus L.). Z. Pflanzenzuchtg 46:389-404. 1990. Genomic sequence of a 12S seed storage protein gene from oat (Avena sativa L. cv. ~Solidor'). Schwab, C.A. 1971. Callose in megasporogenesis of Nucl. Acids Res. 18:377. Diarrhena (Gramineae). Can. }. Bot. 49:1523-1524. Schubert, R., Panitz, R., Manteuffel, R., Nagy, I., Wobus, Schwall, M., and Feix, G. 1988. Zein promoter activity in transiently transformed protoplasts from maize. U., and Baumlein, H. 1994. Tissue-specific expression of an oat 12S seed globulin gene in developing Plant Sci. 56:161-166. tobacco seeds.: differential mRNA and protein accu- Schwartz, B.W., Yeung, E.C., and Meinke, D.W. 1994. mulation. Plant Mol. Biol. 26:203-210. Disruption of morphogenesis and transformation of Schuler, M.A., Schmitt, E.S., and Beachy, R.N. 1982. the suspensor in abnormal suspensor mutants of Closely related families of genes code for the a and Arabidopsis. Development 120:3235-3245. a' subunits of the soybean 7S storage protein com- Schwarz-Sommer, Z., Hue, I., Huijser, P., Flor, P.J., plex. Nucl. Acids Res. 10:8225-8244. Hansen, R., Tetens, F., Lonnig, W.-E., Saedler, F., and Sommer, H. 1992. Characterization of the Schuler, M.A., Ladin, B.R, Pollacco, J.C., Freyer, G., and Antirrhinum floral homeotic MADS-box gene defiBeachy, R.N. 1982. Structural sequences are conciens: evidence for DNA binding and autoregulation served in the genes coding for the a, a' and (5-subof its persistent expression throughout flower develunits of the soybean 7S seed storage protein. Nucl. opment. EMBOJ. 11:251-263. Acids Res. 10:8245-8261. Schultz, E.A., and Haughn, G.W. 1991. LEAFY, a homeotic Schwarz-Sommer, Z., Huijser, P., Nacken, W., Saedler, H., and Sommer, H. 1990. Genetic control of flower gene that regulates inflorescence development in development by homeotic genes in Antirrhinum Arabidopsis. Plant Cell 3:771-781. majus. Science 250:931-936. Schultz, E.A., and Haughn, G.W. 1993. Genetic analysis of the floral initiation process (FLIP) in Arabidopsis. Scofield, S.R., and Crouch, M.L. 1987. Nucleotide Development 119:745-765. sequence of a member of the napin storage proSchultz, E.A., Pickett, F.B., and Haughn, G.W. 1991. The tein family from Brassies napus. ]. Biol. Chem. FLOW gene product regulates the expression 262:12202-12208. domain of homeotic genes AP3 and PI in Arabidopsis Scoles, G.J., and Evans, L.E. 1979. Pollen development in male-fertile and cytoplasmic male-sterile rye. Can. J. flowers. Plant Cell 3:1221-1237. Bot. 57:2782-2790. Schulz, P., and Jensen, W.A. 1969. Capsella embryogenesis: the suspensor and the basal cell. Protoplasma Scorza, R., Cordts, J.M., Ramming, D.W., and Emershad, R.L. 1995. Transformation of grape (Vitis vinifera L.) 67:139-163. zygotic-derived somatic embryos and regeneration Schulz, P., and Jensen, W.A. 1971. Capsella embryogenesis: of transgenic plants. Plant Cell Rep. 14:589-592. the chalazal proliferating tissue. /. Cell Sci. 8:201-227. Schulz, P., and Jensen, W.A. 1973. Capsella embryogenesis: Scott, P., Lyne, R.L., and ap Rees, T. 1995. Metabolism of
References
637
maltose and sucrose by microspores isolated from Sedgley, M., Blesing, M.A., Bonig, I., Anderson, M.A., and Clarke, A.E. 1985. Arabinogalactan-proteins are barley (Hordeum vulgare L.). Planta 197:435-441. localized extracellularly in the transmitting tissue of Scott, R., Dagless, E., Hodge, R., Paul, W., Soufleri, I., and Nicotiana alata Link and Otto, an ornamental Draper, J. 1991. Patterns of gene expression in develtobacco. Micron Micros. Ada 16:247-254. oping anthers of Brassica napits. Plant Mol. Biol. 17:195-207. Seffens, W.S., Almoguera, C, Wilde, H.D., Vonder Haar, R.A., and Thomas, T.L. 1990. Molecular analysis of a Scribailo, R.W., and Barrett, S.C.H. 1991a. Pollen-pistil phylogenetically conserved carrot gene: developinteractions in tristylous Pontederia sagittata mental and environmental regulation. Devel Genet. (Pontederiaceae). I. Floral heteromorphism and 11:65-76. structural features of the pollen tube pathway. Am. ]. Bot. 78:1643-1661. Sehgal, C.B., and Gifford, E.M., Jr. 1979. Developmental and histochemical studies of the ovules of Nicotiana Scribailo, R.W., and Barrett, S.C.H. 1991b. Pollen-pistil mstica L. Bot. Gaz. 140:180-188. interactions in tristylous Pontederia sagittata (Pontederiaceae). II. Patterns of pollen tube growth. Sen, S.K. 1969. Chromatin-organisation during and after synapsis in cultured microsporocytes of Liliutn in Am. ]. Bot. 78:1662-1682. presence of mitomycin C and cycloheximide. Expt. Scutt, C.P., and Croy, R.R.D. 1992. An S5 self-incompatiCell Res. 55:123-127. bility allele-specific cDNA sequence from Brassica oleracea shows high homology to the SLR2 gene. Mol. Sen, S.K., Kundu, S.C., and Gaddipati, J.P. 1977. High Gen. Genet. 232:240-246. speed scintillation autoradiography of DNA fibres undergoing DNA synthesis at zygotene and Scutt, C.P., Fordham-Skelton, A.P., and Croy, R.R.D. 1993. pachytene in the lily. Expt. Cell Res. 108:471-i73. Okadaic acid causes breakdown of self-incompatibility in Brassica oleracea: evidence for the involve-Senda, M., Mikami, T., and Kinoshita, T. 1993. The sugar ment of protein phosphatases in the incompatible beet mitochondrial gene for the ATPase alpha-subunit: response. Sex. Plant Reprod. 6:282-285. sequence, transcription and rearrangements in cytoplasmic male-sterile plants. Curr. Genet. 24:164-170. Scutt, C.P., Gates, P.J., Gatehouse, J.A., Boulter, D., and Croy, R.R.D. 1990. A cDNA encoding an S-locus spe- Senda, M., Harada, T., Mikami, T., Sugiura, M., and cific glycoprotein from Brassica oleracea plants conKinoshita, T. 1991. Genomic organization and taining the S5 self-incompatibility allele. Mol. Gen. sequence analysis of the cytochrome oxidase subunit II gene from normal and male-sterile mitochondria Genet. 220:409-113. in sugar beet. Curr. Genet. 19:175-181. Seavey, S.R., and Bawa, K.S. 1986. Late-acting self-incomSengupta, C, and Raghavan, V. 1980a. Somatic embryopatibility in angiosperms. Bot. Rev. 52:195-219. Secor, D.L., and Russell, S.D. 1988. Megagametophyte genesis in carrot cell suspension. I. Pattern of protein organization in a polyembryonic line of Linum usiand nucleic acid synthesis. /. Expt. Bot. 31:247-258. tatissimum. Am. J. Bot. 75:114-122. Sengupta, C, and Raghavan, V. 1980b. Somatic embryogenesis in carrot cell suspension. II. Synthesis of Sedgley, M. 1974. The concentration of S-protein in stigribosomal RNA and poly (A)+ RNA. /. Expt. Bot. mas of Brassica oleracea plants homozygous and heterozygous for a given S-allele. Heredity 33:412-416. 31:259-268. Sedgley, M. 1975. Flavanoids in pollen and stigma of Sengupta, G, Deluca, V., Bailey, D.S., and Verma, D.P.S. Brassica oleracea and their effects on pollen germina1981. Post-translational processing of 7S and US tion in vitro. Ann. Bot. 39:1091-1095. components of soybean storage proteins. Plant Mol. Biol. 1:19-33. Sedgley, M. 1979. Structural changes in the pollinated and unpollinated avocado stigma and style. /. Cell Sci. Sengupta-Gopalan, C, Reichert, N.A., Barker, R.F., Hall, 38:49-60. T.C., and Kemp, J.D. 1985. Developmentally regulated expression of the bean fl-phaseolin gene in Sedgley, M. 1981a. Ultrastructure and histochemistry of tobacco seed. Proc. Nail Acad. Sci. USA 82:3320-3324. the watermelon stigma. /. Cell Sci. 48:137-146. Sedgley, M. 1981b. Anatomical aspects of compatible Serrano, L., Rochange, F., Semblat, J.P., Marque, C, pollen-stigma interaction. Phytomorphology Teulieres, C, and Boudet, A.-M. 1996. Genetic transformation of Eucalyptus globulus through biolistics: 31:158-165. Sedgley, M. 1981c. Early development of the Macadamia complementary development of procedures for ovary. Aust. ]. Bot. 29:185-193. organogenesis from zygotic embryos and stable Sedgley, M. 1982. Anatomy of the unpollinated and pollitransformation of corresponding proliferating tisnated watermelon stigma. /. Cell Sci. 54:341-355. sue. /. Expt. Bot. 47:285-290. Sedgley, M., and Blesing, M.A. 1982. Foreign pollination Sessa, G., and Fluhr, R. 1995. The expression of an abunof the stigma of watermelon (Citrullus lanatus dant transmitting tract-specific endoglucanase [Thunb.] Matsum and Nakai). Bot. Gaz. 143:210-215. (Sp41) is promoter-dependent and not essential for Sedgley, M., and Blesing, M.A. 1983. Developmental the reproductive physiology of tobacco. Plant Mol. anatomy of the avocado stigma papilla cells and Biol. 29:969-982. their secretion. Bot. Gaz. 144:185-190. Sessions, R.A., and Zambryski, P.C. 1995. Arabidopsis gynoecium structure in the wild type and in ettin Sedgley, M., and Buttrose, M.S. 1978. Structure of the mutants. Development 121:1519-1532. stigma and style of the avocado. Aust. J. Bot. 26:663-682. Sethi, R.S., and Jensen, W.A. 1981. Ultrastructural eviSedgley, M., and Clarke, A.E. 1986. Immuno-gold localidence for secretory phases in the life history of stigsation of arabinogalactan protein in the developing matic hairs of cotton: a dry type stigma. Am. }. Bot. style of Nicotiana alata. Nord. J. Bot. 6:591-598. 68:666-674. Sedgley, M., and Scholefield, P.B. 1980. Stigma secretion in Seurinck, J., Truettner, J., and Goldberg, R.B. 1990. The the watermelon before and after pollination. Bot. nucleotide sequence of an anther-specific gene. Nucl. Gaz. 141:428-434. Acids Res. 18:3403.
638
References
Shah, C.K., and Pandey, S.N. 1978. Histochemical studies mutants of maize. II. Morphological and embryo during embryogenesis in Limnophyton obtusifolium. culture studies. Genetics 95:945-960. Phytomorphology 28:31-42. Sheridan, W.F., and Neuffer, M.G. 1982. Maize developShannon, L.M., and Chrispeels, M.J. 1986. Molecular mental mutants. Embryos unable to form leaf priBiology of Seed Storage Proteins and Lectins. Rockville: mordia. /. Hered. 73:318-329. American Society of Plant Physiologists. Sheridan, W.F., and Stern, H. 1967. Histones of meiosis. Shannon, T.M., Picton, J.M., and Steer, M.W. 1984. The Expt. Cell Res. 45:323-335. inhibition of dictyosome vesicle formation in higher Sheridan, W.F., and Thorstenson, Y.R. 1986. plant cells by cytochalasin D. Eur. ]. Cell Bid. Developmental profiles of three embryo-lethal 33:144-147. maize mutants lacking leaf primordia: ptd*-1130, cp*Sharma, N., Bajaj, M., and Shivanna, K.R. 1985. 1418, and bno*-747B. Devel Genet. 7:35-19. Overcoming self-incompatibility through the use of Sheridan, W.F., Avalkina, N.A., Shamrov, I.I., Batygina, lectins and sugars in Petunia and Eruca. Ann. Bot. T.B., and Golubovskaya, I.N. 1996. The macl gene: 55:139-141. Controlling the commitment to the meiotic pathway Sharma, N., and Shivanna, K.R. 1982. Effects of pistil in maize. Genetics 142:1009-1020. extracts on in vitro responses of compatible and Sherwood, R.T. 1995. Nuclear DNA amount during incompatible pollen in Petunia hybrida Vilm. Indian }. sporogenesis and gametogenesis in sexual and Expt. Biol. 20:255-256. aposporous buffelgrass. Sex. Plant Reprod. 8:85-90. Sharma, N., and Shivanna, K.R. 1983. Lectin-like compo- Shevell, D.E., Leu, W.-M., Gillmor, C.S., Xia, G., nents of pollen and complementary saccharide moiFeldmann, K.A., and Chua, N.-H. 1994. EMB30 is ety of the pistil are involved in self-incompatibility essential for normal cell division, cell expansion, and recognition. Curr. Sci. 52:913-916. cell adhesion in Arabidopsis and encodes a protein that has similarity to Sec7. Cell 77:1051-1062. Sharma, N., and Shivanna, K.R. 1986. Treatment of the stigma with an extract of a compatible pistil over- Shewry, PR. 1995. Plant storage proteins. Biol. Rev. comes self-incompatibility in Petunia. New Phytol. 70:375-426. 102:443-447. Shewry, PR., Kreis, M., Burgess, S.R., Parmar, S., and Sharma, S., Singh, M.B., and Malik, C.P. 1981. Dark CO2 Miflin, B.J. 1983. The synthesis and deposition of the fixation during germination of Amaryllis vittata prolamin storage proteins (secalins) of rye. Planta pollen in suspension cultures. Indian ]. Expt. Biol. 159:439^55. 19:710-714. Shewry, PR., Pratt, H.M., Leggatt, M.M., and Miflin, B.J. 1979. Protein metabolism in developing endosperms Shayk, M., Kolattukudy, P.E., and Davis, R. 1977. of high-lysine and normal barley. Cereal Chem. Production of a novel extracellular cutinase by the 56:110-117. pollen and the chemical composition and ultrastructure of the stigma cuticle of nasturtium (Tropaeolum Shi, L., Mogensen, H.L., and Zhu, T. 1991. Dynamics of majus). Plant Physiol. 60:907-915. nuclear pore density and distribution patterns Sheffield, E., Cawood, A.H., Bell, P.R., and Dickinson, within developing pollen: implications for a funcH.G. 1979. The development of nuclear vacuoles tional relationship between the vegetative nucleus during meiosis in plants. Planta 146:597-601. and the generative cell. /. Cell Sci. 99:115-120. Sheldon, J.M., and Dickinson, H.G. 1983. Determination Shi, L., Zhu, T., Mogensen, H.L., and Keim, P. 1996. Sperm identification in maize by fluorescence in situ of patterning in the pollen wall of Lilium henryi. ]. hybridization. Plant Cell 8:815-821. Cell Sci. 63:191-208. Sheldon, J.M., and Dickinson, H.G. 1986. Pollen wall for- Shi, L., Zhu, T., Mogensen, H.L., and Smith, S.E. 1991. mation in Lilium: the effect of chaotropic agents, and Paternal plastid inheritance in alfalfa: plastid the organisation of the microtubular cytoskeleton nucleoid number within generative cells correlates during pattern development. Planta 168:11-23. poorly with plastid number and male plastid transmission strength. Curr. Genet. 19:399-401. Sheldon, J.M., and Hawes, C. 1988. The actin cytoskeleton during male meiosis in Lilium. Cell Biol. Int. Rep. Shiba, H., Hinata, K., Suzuki, A., and Isogai, A. 1995. 12:471^176. Breakdown of selfincompatibility in Brassica by the Shellhammer, ]., and Meinke, D. 1990. Arrested embryos antisense RNA of the SLG gene. Proc. Jap. Acad. 71B:81-83. from the biol auxotroph of Arabidopsis thaliana contain reduced levels of biotin. Plant Physiol. Shibata, D., Steczko, }., Dixon, J.E., Andrews, P C , Hermodson, M., and Axelrod, B. 1988. Primary 93:1162-1167. structure of soybean lipoxygenase L-2. /. Biol. Chem. Shen, J.B., and Hsu, F.C. 1992. Brassica anther-specific 263:6816-6821. genes: characterization and in situ localization of expression. Mol. Gen. Genet. 234:379-389. Shifriss, O. 1945. Male sterilities and albino seedlings in Shepard, ]., Boothroyd, E.R., and Stern, H. 1974. The effect cucurbits. A study in inbreeding. /. Hered. 36:47-52. of colchicine on synapsis and chiasma formation in Shii, C.T., Rabakoarihanta, A., Mok, M.C., and Mok, microsporocytes of Lilium. Chromosoma 44:423-437. D.W.S. 1982. Embryo development in reciprocal crosses of Phaseolus vulgaris L. and P. coccineus Lam. Sheridan, W.F. 1973. Nonaqueous isolation of nuclei from Theor. Appl. Genet. 62:59-64. lily pollen and an examination of their histones. Z. Pflanzenphysiol. 68:450-459. Shimamoto, K., Ackermann, M., and Dierks-Ventling, C. 1983. Expression of zein in long term endosperm Sheridan, W.F., and Clark, J.K. 1987. Maize embryogeny: a cultures of maize. Plant Physiol. 73:915-920. promising experimental system. Trend. Genet. 3:3-6. Sheridan, W.F., and Clark, J.K. 1993. Mutational analysis Shirsat, A., Wilford, N., Cray, R., and Boulter, D. 1989. Sequences responsible for the tissue specific proof morphogenesis of the maize embryo. Plant ]. moter activity of a pea legumin gene in tobacco. Mol. 3:347-358. Gen. Genet. 215:326-331. Sheridan, W.F., and Neuffer, M.G. 1980. Defective kernel
References
639
Shirsat, A.H., Meakin, P.J., and Gatehouse, J.A. 1990. Shotwell, M.A., Boyer, S.K., Chesnut, R.S., and Larkins, Sequences 5' to the conserved 28 bp leg box eleB.A. 1990. Analysis of seed storage protein genes of ment regulate the expression of pea seed storage oats. /. Biol. Chem. 265:9652-9658. protein gene legA. Plant Mol. Biol. 15:685-693. Shoup, J.R., Overton, J., and Ruddat, M. 1980. infrastructure and development of the sexine in the Shivanna, K.R. 1965. In vitro fertilization and seed formapollen wall of Silene alba (Caryophyllaceae). Bot. Gaz. tion in Petunia violacea Lindl. Phytomorphology 141:379-388. 15:183-185. Shivanna, K.R. 1982. Pollen-pistil interaction and control Shoup, J.R., Overton, J., and Ruddat, M. 1981. of fertilization. In Experimental Embryology of Vascular infrastructure and development of the nexine and Plants, ed. B.M. Johri, pp. 131-174. Berlin: Springerintine in the pollen wall of Silene alba Verlag. (Caryophyllaceae). Am. ]. Bot. 68:1090-1095. Shivanna, K.R., and Cresti, M. 1989. Effects of high Shufflebottom, D., Edwards, K., Schuch, W., and Bevan, M. 1993. Transcription of two members of a gene humidity and temperature stress on pollen memfamily encoding phenylalanine ammonia-Iyase brane integrity and pollen vigour in Nicotiana leads to remarkably different cell specificities and tabacum. Sex. Plant Reprod. 2:137-141. induction patterns. Plant J. 3:835-845. Shivanna, K.R., and Heslop-Harrison, J. 1981. Membrane state and pollen viability. Ann. Bot. 47:759-770. Shukla, A., and Sawhney, V.K. 1992. Cytokinins in a genie Shivanna, K.R., Heslop-Harrison, J., and Heslopmale sterile line of Brassica napus. Physiol. Plant. Harrison, Y. 1981. Heterostyly in Primula. 2. Sites of 85:23-29. pollen inhibition, and effects of pistil constituents on Shukla, A., and Sawhney, V.K. 1994. Abscisic acid: one of compatible and incompatible pollen-tube growth. the factors affecting male sterility in Brassica napus. Protoplasma 107:319-337. Physiol. Plant. 91:522-528. Shivanna, K.R., Heslop-Harrison, J., and Heslop- Shure, M., Wessler, S., and Fedoroff, N. 1983. Molecular Harrison, Y. 1983. Heterostyly in Primula. 3. Pollen identification and isolation of the waxy locus in water economy: a factor in the intramorph-incommaize. Cell 35:225-233. patibility response. Protoplasma 117:175-184. Shuttuck-Eidens, D.M., and Beachy, R.M. 1985. Shivanna, K.R., Heslop-Harrison, Y, and Heslop-Harrison, Degradation of p-conglycinin in early stages of soyJ. 1978. The pollen-stigma interaction: bud pollination bean embryogenesis. Plant Physiol. 78:895-898. in the Cruciferae. Ada Bot. Afeer//27:107-119. Siculella, L., and Palmer, J.D. 1988. Physical and gene Shivanna, K.R., Heslop-Harrison, Y, and Hesloporganization of mitochondrial DNA in fertile and Harrison, J. 1982. The pollen-stigma interaction in male sterile sunflower. CMS-associated alterations the grasses. 3. Features of the self-incompatibility in structure and transcription of the atpA gene. Nucl. response. Ada Bot. Neerl. 31:307-319. Acids Res. 16:3787-3799. Shivanna, K.R., Jaiswal, V.S., and Mohan Ram, H.Y 1974a. Sidorova, N.V. 1985. Characteristics of the ultrastructure Inhibition of gamete formation by cycloheximide in of isolated embryo sacs of tobacco. Doklady Akad. pollen tubes of Impatiens balsamina. Planta Nauk Ukr. SSR Ser. B 12:63-66. 117:173-177. Siebertz, B., Logemann, J., Willmitzer, L., and Schell, J. Shivanna, K.R., Jaiswal, V.S., and Mohan Ram, H.Y. 1974b. 1989. cis-Analysis of the wound-inducible promoter Effect of cycloheximide on cultured pollen grains of wunl in transgenic tobacco plants and histochemical Trigonella foenum-graecum. Plant Sci. Lett. 3:335-339. localization of its expression. Plant Cell 1:961-968. Shivanna, K.R., and Johri, B.M. 1985. The Angiosperm Silcock, D.J., Francis, D., Bryant, J.A., and Hughes, S.G. Pollen. Strudure and Function. New Delhi: Wiley 1990. Changes in nuclear DNA content, cell and Eastern Ltd. nuclear size, and frequency of cell division in the Shivanna, K.R., and Rangaswamy, N.S. 1969. Overcoming cotyledons of Brassica napus L. during embryogeneself-incompatibility in Petunia axillaris. I. Delayed sis. /. Expt. Bot. 41:401^07. pollination, pollination with stored pollen, and bud Silvanovich, A., Astwood, J., Zhang, L., Olsen, E., Kisil, F., pollination. Phytomorphology 19:372-380. Sehon, A., Mohapatra, S., and Hill, R. 1991. Nucleotide sequence analysis of three cDNAs codShivanna, K.R., and Rangaswamy, N.S. 1992. Pollen Biology. A Laboratory Manual. Berlin: Springer-Verlag. ing for Poa p IX isoallergens of Kentucky bluegrass Shivanna, K.R., and Sastri, D.C. 1981. Stigma-surface pollen. /. Biol. Chem. 266:1204-1210. esterase activity and stigma receptivity in some taxa Simon, A.E., Tenbarge, K.M., Scofield, S.R., Finkelstein, characterized by wet stigmas. Ann. Bot. 47:53-64. R.R., and Crouch, M.L. 1985. Nucleotide sequence of Shivanna, K.R., Xu, H., Taylor, P., and Knox, R.B. 1988. a cDNA clone of Brassica napus 12S storage protein Isolation of sperms from the pollen tubes of flowering shows homology with Iegumin from Pisum sativum. plants during fertilization. Plant Physiol. 87:647-650. Plant Mol. Biol. 5:191-201. Shoemaker, R.C., Christofferson, S.E., and Galbraith, D.W. Simon, R., Altschuler, Y, Rubin, R., and Galili, G. 1990. 1987. Storage protein accumulation patterns in Two closely related wheat storage proteins follow a somatic embryos of cotton (Gossypium hirsutum L.). markedly different subcellular route in Xenopus laePlant Cell Rep. 6:12-15. vis oocytes. Plant Cell 2:941-950. Shotwell, M.A., and Larkins, B.A. 1989. The biochemistry Simoncioli, C. 1974. Ultrastructural characteristics of and molecular biology of seed storage proteins. In "Diplotaxis erucoides (L.) DC." suspensor. Giorn. Bot. The Biochemistry of Plants. A Comprehensive Treatise, Hal. 108:175-189. Vol. 15, ed. A. Marcus, pp. 297-345. San Diego: Simpson, M.G., and Levin, G.A. 1994. Pollen ultrastrucAcademic Press. ture of the biovulate Euphorbiaceae. Int. J. Plant Sci. Shotwell, M.A., Afonso, C, Davies, E., Chesnut, R.S., and 155:313-341. Larkins, B.A. 1988. Molecular characterization of oat Sims, T.L. 1993. Genetic regulation of self-incompatibility. seed globulins. Plant Physiol. 87:698-704. Crit. Rev. Plant Sci. 12:129-167.
640
References
Singh, A., Ai, Y, and Kao, T.-H. 1991. Characterization of Temperature effects on endogenous indole-3-acetic ribonuclease activity of three S-allele-associated proacid levels in leaves and stamens of the normal and teins of Petunia inflata. Plant Physiol. 96:61-68. male sterile "stamenless-2' mutant of tomato (Lycopersicon esculentum Mill.). Plant Cell Environ. Singh, A., Evensen, K.B., and Kao, T.-H. 1992. Ethylene 15:373-377. synthesis and floral senescence following compatible and incompatible pollinations in Petunia inflata. Singh, S., and Walles, B. 1992. The ovarian transmitting Plant Physiol. 99:38-45. tissue in Lilium regale. Int. }. Plant Sci. 153:205-211. Singh, A., and Kao, T.-H. 1992. Gametophytic self-incom- Singh, S., and Walles, B. 1995. Ultrastructural differentiapatibility: biochemical, molecular genetic, and evotion of the ovarian transmitting tissue in Lilium lutionary aspects. Int. Rev. Cytol. 140:449-483. regale. Ann. Bot. 75:455-462. Singh, A., and Paolillo, D.J., Jr. 1990. Role of calcium in the Singh, S.P., and Hadley, H.H. 1961. Pollen abortion callose response of self-pollinated Brassica stigmas. in cytoplasmic male sterile Sorghum. Crop Sci. Am. J. Bot. 77:128-133. 1:430-432. Singh, A., Perdue, T.D., and Paolillo, D.J., Jr. 1989. Pollen- Singh, S.P., and Rhodes, A.M. 1961. A morphological and pistil interactions in Brassica oleracea: cell calcium in cytological study of male sterility in Cucurbita maxself and cross pollen grains. Protoplasma 151:57-61. ima. Proc. Am. Soc. Hort. Sci. 78:375-378. Singh, A.P., and Mogensen, H.L. 1975. Fine structure of Singleton, W.R., and Mangelsdorf, PC. 1940. Gametic the zygote and early embryo in Quercus gambelii. Am. lethals on the fourth chromosome of maize. Genetics J. Bot. 62:105-115. 25:366-390. Singh, A.P., and Mogensen, H.L. 1976. Fine structure of Sinha, S., Jha, K.K., and Roy, R.P. 1978. Segmentation patearly endosperm in Quercus gambelii. Cytologia tern of pollen in anther culture of Solanum surattense, 41:345-361. Luffa cylindrica and Luffa echinata. Phytomorphology Singh, M., Bhalla, P.L., and Malik, C.P. 1979. Peroxidase 28:43-49. localization during embryogenesis in Tropaeolum Sivaramakrishna, D. 1978. Size relationships of apical cell majiis. Phytomorphology 29:306-309. and basal cell in two-celled embryos in angiosperms. Can. ]. Bot. 56:1434-1438. Singh, M., and Brown, G.G. 1991. Suppression of cytoplasmic male sterility by nuclear genes alters expres- Sjodahl, S., Rodin, J., and Rask, L. 1991. Characterization sion of a novel mitochondrial gene region. Plant Cell of the 12S globulin complex of Brassica napus. 3:1349-1362. Evolutionary relationship to other 11-12S storage globulins. Eur. J. Biochem. 196:617-621. Singh, M.B., Bhalla, P.L., and Malik, C.P. 1980. Activity of some hydrolytic enzymes in autolysis of the embryo Sjodahl, S., Gustavsson, H.-O., Rodin, J., Lenman, M., suspensor in Tropaeolum majus L. Ann. Bot. Hoglund, A.-S., and Rask, L. 1993. Cruciferin gene 45:523-527. families are expressed coordinately but with tissueSingh, M.B., and Knox, R.B. 1984. Invertases of Liliwn specific differences during Brassica napus seed development. Plant Mol. Biol. 23:1165-1176. pollen. Characterization and activity during in vitro germination. Plant Physiol. 74:510-515. Sjodahl, S., Gustavsson, H.-O., Rodin, J., and Rask, L. Singh, M.B., and Knox, R.B. 1985. Gene controlling P1995. Deletion analysis of the Brassica napus crugalactosidase deficiency in pollen of oilseed rape. /. ciferin gene cru 1 promoter in transformed tobacco: Hered. 76:199-201. promoter activity during early and late stages of embryogenesis is influenced by as-acting elements Singh, M.B., and Malik, C.P. 1976. Histochemical studies in partially separate regions. Planta 197:264-271. on reserve substances and enzymes in female gametophyte of Zea mays. Ada Biol. Acad. Sci. Hung. Skokut, T.A., Manchester, J., and Schaefer, J. 1985. 27:231-244. Regeneration in alfalfa tissue culture. Stimulation of Singh, M.B., O'Neill, P.M., and Knox, R.B. 1985. Initiation somatic embryo production by amino acids and Nof postmeiotic p-galactosidase synthesis during 15 NMR determination of nitrogen utilization. Plant Physiol. 79:579-583. microsporogenesis in oilseed rape. Plant Physiol. 77:225-228. Skorupska, H.T., Desamero, N.V., and Palmer, R.G. 1993. Possible homeosis in flower development in cultiSingh, M.B., Hough, T., Theerakulpisut, P., Avjioglu, A., vated soybean [Glycine max (L.) Merr.]: morDavies, S., Smith, P.M., Taylor, P., Simpson, R.J., phoanatomy and genetics. /. Hered. 84:97-104. Ward, L.D., McCluskey, J., Puy, R., and Knox, R.B. 1991. Isolation of cDNA encoding a newly identified Skriver, K., Leah, R., Miiller-Uri, E, Olsen, F.-L., and Mundy, major allergenic protein of rye-grass pollen: intracelJ. 1992. Structure and expression of the barley lipid lular targeting to the amyloplast. Proc. Natl Acad. Sci. transfer protein gene Ltpl. Plant Mol. Biol. 18:585-589. USA 88:1384-1388. Skvarla, J.J., and Larson, D.A. 1966. Fine structural studies of Zea mays pollen I: cell membranes and exine Singh, N.K., and Shepherd, K.W. 1987. Solubility behaviour, synthesis, degradation and subcellular localizaontogeny. Am. }. Bot. 53:1112-1125. tion of a new class of disulfide-linked proteins in Skvarla, J.J., and Rowley, J.R. 1970. The pollen wall of Canna and its similarity to the germinal apertures of wheat endosperm. Aust. }. Plant Physiol. 14:245-252. other pollen. Am. }. Bot. 57:519-529. Singh, N.K., Donovan, G.R., Carpenter, H.C., Skerritt, J.H., and Langridge, P. 1993. Isolation and character- Sladky, Z., and Havel, L. 1976. The study of the conditions ization of wheat triticin cDNA revealing a unique for the fertilization in vitro in maize. Biol. Plant. lysine-rich repetitive domain. Plant Mol. Biol. 18:469-472. 22:227-237. Slay, R.M., Grimes, H.D., and Hodges, T.K. 1989. Plasma membrane proteins associated with undifferentiated Singh, S., and Sawhney, V.K. 1992. Cytokinins in a normal and embryonic Daucus carota tissue. Protoplasma line and the ogura (ogu) cytoplasmic male-sterile line 150:139-149. of rapeseed (Brassica napus). Plant Sci. 86:147-154. Singh, S., Sawhney, V.K., and Pearce, D.W. 1992. Slightom, J.L., Sun, S.M., and Hall, T.C. 1983. Complete
References
nucleotide sequence of a French bean storage protein gene: phaseolin. Proc. Nail Acad. Sci. USA 80:1987-1901. Slightom, J.L., Drong, R.F., Klassy, R.C., and Hoffman, L.M. 1985. Nucleotide sequences from phaseolin cDNA clones: the major storage proteins from Phaseolus vulgaris are encoded by two unique gene families. Nucl. Acids Res. 13:6483-6498. Slocombe, S.P., Cummins, I., Jarvis, R.P., and Murphy, D.J. 1992. Nucleotide sequence and temporal regulation of a seed-specific Brassica napus cDNA encoding a stearoyl-acyl carrier protein (ACP). Plant Mol. Biol. 20:151-155. Slocombe, S.P., Piffanelli, P., Fairbairn, D., Bowra, S., Hatzopoulos, P., Tsiantis, M., and Murphy, D.J. 1994. Temporal and tissue-specific regulation of a Brassica napus stearoyl-acyl carrier protein desaturase gene. Plant Physiol. 104:1167-1176. Smart, C.J., Moneger, F., and Leaver, CJ. 1994. Cell-specific regulation of gene expression in mitochondria during anther development in sunflower. Plant Cell 6:811-825. Smart, M.G., and O'Brien, T.P. 1983. The development of the wheat embryo in relation to the neighbouring tissues. Protoplasma 114:1-13. Smith, A.G., Gasser, C.G., Budelier, K.A., and Fraley, R.T. 1990. Identification and characterization of stamenand tapetum-specific genes from tomato. Mol. Gen. Genet. 222:9-16. Smith, A.M. 1988. Major differences in isoforms of starchbranching enzyme between developing embryos of round- and wrinkled-seeded peas (Pisum sativum L.). Planta 175:270-279. Smith, C.G. 1974. The ultrastructural development of spherosomes and oil bodies in the developing
641
tion of a novel gene in barley embryos. Plant Mol. Biol. 20:255-266. Smith, R.L., and Chowdhury, M.K.U. 1991. Characterization of pearl millet mitochondrial DNA fragments rearranged by reversion from cytoplasmic male sterility to fertility. Theor. Appl. Genet. 81:793-799. Smith, R.L., Chowdhury, M.K.U., and Pring, D.R. 1987. Mitochondrial DNA rearrangements in Pennisetum associated with reversion from cytoplasmic male sterility to fertility. Plant Mol. Biol. 9:277-286. Smith, S.M., and Street, H.E. 1974. The decline of embryogenic potential as callus and suspension cultures of carrot (Daucus carota L.) are serially subcultured. Ann. Bot. 38:223-241. Smyth, D.R. 1982. Proplastid DNA synthesis at pachytene in pollen mother cells of Lilium henryi. ]. Cell Sci. 56:293-302. Smyth, D.R., and Shaw, T. 1979. Cytoplasmic DNA synthesis at meiotic prophase in Lilium henryi. Aust. ]. Bot. 27:273-284. Smyth, D.R., and Stern, H. 1973. Repeated DNA synthesized during pachytene in Lilium henryi. Nature New Biol. 245:94-96. So, J.-S., and Larkins, B.A. 1991. Binding of an endosperm-specific nuclear protein to a maize betazein gene correlates with zein transcriptional activity. Plant Mol. Biol. 17:309-319. Soave, C, and Salamini, F. 1984. The role of structural and regulatory genes in the development of maize endosperm. Devel Genet. 5:1-25. Soave, C, Tardini, L., di Fonzo, N., and Salamini, F. 1981. Zein level in maize endosperm depends on a protein under control of the opaque-2 and opaque-6 loci. Cell 27:403-410. embryo of Crambe abyssinka. Planta 119:125-142. Sodmergen, Chen, G.H., Hu, Z.M., Guo, F.L., and Guan, Smith, C.R., Saunders, J.A., van Wert, S., Cheng, }., and X.L. 1995. Male gametophyte development in Matthews, B.F. 1994. Expression of GUS and CAT Plumbago zeylanica: cytoplasm localization and cell activities using electrotransformed pollen. Plant Sci. determination in the early generative cell. 104:49-58. Protoplasma 186:79-86. Smith, D.L. 1973. Nucleic acid, protein, and starch syn- Sodmergen, Luo, Y.Y., Kuroiwa, T., and Hu, S.Y. 1994. thesis in developing cotyledons of Pisum arvense L. Cytoplasmic DNA apportionment and plastid differAnn. Bot. 37:795-804. entiation during male gametophyte development in Smith, D.L., and Krikorian, A.D. 1988. Production of Pelargonium zonale. Sex. Plant Reprod. 7:51-56. somatic embryos from carrot tissues in hormone-free Sodmergen, Suzuki, T., Kawano, S., Nakamura, S., Tano, medium. Plant Sci. 58:103-110. S., and Kuroiwa, T. 1992. Behavior of organelle nuclei (nucleoids) in generative and vegetative cells Smith, D.L., and Krikorian, A.D. 1989. Release of somatic during maturation of pollen in Lilium longiflorum embryogenic potential from excised zygotic embryos of carrot and maintenance of proembryonic and Pelargonium zonale. Protoplasma 168:73-81. cultures in hormone-free medium. Am. ]. Bot. Somers, D.A., Rines, H.W., Gu, W., Kaeppler, H.F., and 76:1832-1843. Bushnell, W.R. 1992. Fertile, transgenic oat plants. Biotechnology 10:1589-1594. Smith, D.L., and Krikorian, A.D. 1990. Somatic embryogenesis of carrot in hormone-free medium: Somers, D.E., and Quail, P.H. 1995. Temporal and spatial external pH control over morphogenesis. Am. ]. Bot. expression patterns of PHYA and PHYB gene in 77:1634-1647. Arabidopsis. Plant J. 7:413-427. Smith, J.A., Krauss, M.R., Borkird, C, and Sung, Z.R. Sommer, H., Beltrdn, J.-P., Huijser, P., Pape, H., Lonnig, 1988. A nuclear protein associated with cell divisions W.-E., Saedler, H., and Schwarz-Sommer, Z. 1990. in plants. Planta 174:462-472. Deficiens, a homeotic gene involved in the control of Smith, J.G. 1973. Embryo development in Phaseolus vulflower morphogenesis in Antirrhinum majus: the progaris. II. Analysis of selected inorganic ions, ammotein shows homology to transcription factors. EMBO nia, organic acids, amino acids, and sugars in the /. 9:605-613. endosperm liquid. Plant Physiol. 51:454-458. Sommer, H., Nacken, W., Beltran, P., Huijser, P., Pape, H., Smith, L.G., Jackson, D., and Hake, S. 1995. Expression of Hansen, R., Flor, P., Saedler, H., and Schwarzknottedl marks shoot meristem formation during Sommer, Z. 1991. Properties of deficiens, a homeotic maize embryogenesis. Devel Genet. 16:344-348. gene involved in the control of flower morphogenesis Smith, L.M., Handley, }., Li, Y, Martin, H., Donovan, L., in Antirrhinum majus. Development Suppl. 1:169-175. and Bowles, D.J. 1992. Temporal and spatial regula- Sondheimer, E., and Linskens, H.F. 1974. Control of in
642
References
vitro germination and tube extension of Petunia Southworth, D., and Dickinson, D.B. 1975. |3-l,3-glucan hybrida pollen. Proc. Koninkl. Nederl. Akad. Wetensch. synthase from Lilium longiflorum pollen. Plant 77G116-124. Physiol. 56:83-87. Song,J., and Hedgcoth, C. 1994. A chimeric gene (orft.56) Southworth, D., and Dickinson, D.B. 1981. Ultrastructural is expressed as protein only in cytoplasmic malechanges in germinating lily pollen. Grana 20:29-35. sterile lines of wheat. Plant Mol. Biol. 26:535-539. Southworth, D., and Jernstedt, J.A. 1995. Pollen exine development precedes microtubule rearrangement Sood, P.P., Malik, C.P., and Tewari, H.B. 1968. A histoin Vigna unguiculata (Fabaceae): a model for pollen chemical study of the localization of succinicdehywall patterning. Protoplasma 187:79-87. drogenase in the germinating pollen grains of Portulaca grandiflora. Z. Biol. 116:215-220. Southworth, D., and Knox, R.B. 1989. Flowering plant sperm cells: isolation from pollen of Gerbera jamesonii Sood, R., Prabha, K., and Gupta, S.C. 1982. Is the Rejection (Asteraceae). Plant Sci. 60:273-277. reaction' inducing ability in sporophytic self-incompatibility systems restricted only to pollen and tape- Southworth, D., and Morningstar, P.A. 1992. Isolation of turn? Theor. Appl. Genet. 63:27-32. generative cells from pollen of Phoenix dactylifera. Sex. Plant Reprod. 5:270-274. Sood, R., Prabha, K., and Gupta, S.C. 1983. Overcoming self-incompatibility in Brassica campestris cv. Pusa Southworth, D., Platt-Aloia, K.A., and Thomson, W.W. Kalyani by bud and delayed pollinations. Beitr. Biol. 1988. Freeze fracture of sperm and vegetative cells in Pftanz. 58:59-65. Zea mays pollen. /. Ultrastr. Mol. Str. Res. 101:165-172. Sopory, S.K., and Maheshwari, S.C. 1976. Development of Southworth, D., Salvatici, P., and Cresti, M. 1994. Freeze fracture of membranes at the interface between vegpollen embryoids in anther cultures of Datura etative and generative cells in Amaryllis pollen. Int. J. innoxia. II. Effects of growth hormones. /. Expt. Bot. Plant Sci. 155:538-544. 27:58-68. Sorensen, M.B. 1992. Methylation of B-hordein genes in Southworth, D., Platt-Aloia, K.A., DeMason, D.A., and Thomson, W.W. 1989. Freeze-fracture of the generabarley endosperm is inversely correlated with gene tive cell of Phoenix dactylifera (Arecaceae). Sex. Plant activity and affected by the regulatory gene Lys3. Reprod. 2:270-276. Proc. Natl Acad. Sci. USA 89:4119-4123. Sorensen, M.B., Cameron-Mills, V, and Brandt, A. 1989. Southworth, D., Singh, M.B., Hough, T., Smart, I.J., Taylor, P., and Knox, R.B. 1988. Antibodies to pollen exines. Transcriptional and post-transcriptional regulation Planta 176:482-487. of gene expression in developing barley endosperm. Mol. Gen. Genet. 217:195-201. Souvr6, A., and Albertini, L. 1982. Etude des modifications de l'ultrastructure du tapis plasmodial du Sosnikhina, S.P., Fedotova, Y.S., Smirnov, V.G., Rhoeo discolor Hance au cours du developpement de Mikhailova, E.I., Kolomiets, O.L., and Bogdanov, Y.F. l'anthere, en relation avec les donnSes cytochim1992. Meiotic mutants of rye Secale cereale L. I. iques et autoradiographiques. Rev. Cytol. Biol. Veget.Synaptic mutant sy-1. Theor. Appl. Genet. 84:979-985. Bot. 5:151-169. Sossountzov, L., Ruiz-Avila, L., Vignols, F., Jolliot, A., Arondel, V., Tchang, F., Grosbois, M., Guerbette, F., Soyfer, V.N. 1980. Hereditary variability of plants under the action of exogenous DNA. Theor. Appl. Genet. Miginiac, E., Delseny, M., Puigdomenech, P., and 58:225-235. Kader, J.-C. 1991. Spatial and temporal expression of a maize lipid transfer protein gene. Plant Cell Spangenberg, G., Wang, Z.Y., Wu, X.L., Nagel, J., Iglesias, 3:923-933. V.A., and Potrykus, I. 1995a. Transgenic tall fescue (Festuca arundinacea) and red fescue (F. rubra) plants Soueges, R. 1919. Les premieres divisions de l'ceuf et les from microprojectile bombardment of embryogenic differenciations du suspenseur chez le Capsella bursasuspension cells. /. Plant Physiol. 145:693-701. pastoris Moench (1). Ann. Sci. Nat. Bot. Ser. X 1:1-28. Soueges, R. 1924. Developpement de l'embryon chez le Spangenberg, G., Wang, Z.-Y, Wu, X., Nagel, J., and Potrykus, I. 1995b. Transgenic perennial ryegrass Sagina procumbens L. Bull. Soc. Bot. Fr. 71:590-614. (Lolium perenne) plants from microprojectile bomSoueges, R. 1936. Embryog^nie des Saxifragac^es. bardment of embryogenic suspension cells. Plant Sci. Developpement de l'embryon chez le Saxifraga gran108:209-217. ulata L. Compt. Rend. Acad. Sci. Paris 202:240-242. Souer, E., van Houwelingen, A., Kloos, D., Mol, J., and Spanjers, A.W. 1978. Voltage variation in Lilium longifloKoes, R. 1996. The no apical meristem gene of Petunia rum pistils induced by pollination. Experientia is required for pattern formation in embryos and 34:36-37. flowers and is expressed at meristem and primordia Sparrow, A.H., Moses, M.J., and Steele, R. 1952. A cytoboundaries. Cell 85:159-170. logical and cytochemical approach to an understanding of radiation damage in dividing cells. Brit. Southworth, D. 1971. Incorporation of radioactive precurJ. Radiol. 25:182-188. sors into developing pollen walls. In Pollen: Development and Physiology, ed. J. Heslop-Harrison, Sparrow, A.H., Pond, V., and Kojan, S. 1955. pp. 115-120. London: Butterworth & Co. Microsporogenesis in excised anthers of Trillium erectum grown on sterile media. Am. J. Bot. 42:384-394. Southworth, D. 1975. Lectins stimulate pollen germinaSpassova, M., Moneger, F., Leaver, C.J., Petrov, P., tion. Nature 258:600-602. Atanassov, A., Nijkamp, H.J.J., and Hille, J. 1994. Southworth, D. 1983. Exine development in Gerbera Characterisation and expression of the mitochondrjamesonii (Asteraceae: Mutisieae). Am. J. Bot. ial genome of a new type of cytoplasmic male-sterile 70:1038-1047. sunflower. Plant Mol. Biol. 26:1819-1831. Southworth, D. 1988. Isolation of exines from gymSpena, A., and Schell, J. 1987. The expression of a heatnosperm pollen. Am. J. Bot. 75:15-21. inducible chimeric gene in transgenic tobacco plants. Southworth, D. 1990. Membranes of sperm and vegetaMol. Gen. Genet. 206:436^40. tive cells in pollen of Gerbera jamesonii. ]. Str. Biol. 103:97-103. Spena, A., Viotti, A., and Pirrotta, V. 1982. A homologous
References
643
repetitive block structure underlies the heterogene- Staiger, C.J., and Cande, W.Z. 1991. Microfilament distribution in maize meiotic mutants correlates with ity of heavy and light chain zein genes. EMBO /. microtubule organization. Plant Cell 3:637-644. 1:1589-1594. Spena, A., Estruch, J.J., Prinsen, Ev Nacken, W., van Staiger, C.J., Goodbody, K.C., Hussey, P.J., Valenta, R., Drabak, B.K., and Lloyd, C.W. 1993. The profilin Onckelen, H v and Sommer, H. 1992. Anther-specific multigene family of maize: differential expression of expression of the rolB gene of Agrobaderium rhizothree isoforms. Plant ]. 4:631-641. genes increases IAA content in anthers and alters anther development and whole flower growth. Staiger, D., and Apel, K. 1993. Molecular characterization Theor. Appl. Genet. 84:520-527. of two cDNAs from Sinapis alba L. expressed specifically at an early stage of tapetum development. Plant Spencer, ME., and Hodge, R. 1991. Cloning and sequencJ. 4:697-703. ing of the cDNA encoding the major albumin of Theobroma cacao. Identification of the protein as a Staiger, D., Kappeler, S., Muller, M., and Apel, K. 1994. member of the Kunitz protease inhibitor family. The proteins encoded by two tapetum-specific tranPlanta 183:528-535. scripts, Satap35 and S«tap44, from Sinapis alba L. are localized in the exine cell wall layer of developing Spencer, M.E., and Hodge, R. 1992. Cloning and sequencmicrospores. Planta 192:221-231. ing of a cDNA encoding the major storage proteins of Theobroma cacao. Identification of the proteins as Stalberg, K., Ellerstrom, M., Josefsson, L.-G., and Rask, L. 1993. Deletion analysis of a 2S seed storage protein members of the vicilin class of storage proteins. Planta 186:567-576. promoter of Brassica napus in transgenic tobacco. Plant Mol. Biol. 23:671-683. Speranza, A., and Calzoni, G.L. 1988. In vitro test of selfincompatibility in Mains domestica. Sex. Plant Reprod.Stanley, R.G., and Linskens, H.F. 1964. Enzyme activation 1:223-227. in germinating Petunia pollen. Nature 203:542-544. Speranza, A., and Calzoni, G.L. 1990. In vitro expression Stanley, R.G., and Linskens, H.F. 1965. Protein diffusion from germinating pollen. Physiol. Plant. 18:47-53. of self-incompatibility in Malus domestica mediated by stylar glycoproteins. Plant Physiol. Biochem. Stanley, R.G., and Linskens, H.F. 1967. Oxygen tension as a control mechanism in pollen tube rupture. Science 28:747-754. 157:833-834. Speranza, A., and Calzoni, G.L. 1992. Inhibition of pollentube growth and protein secretion by the monova- Stanley, R.G., and Linskens, H.F. 1974. Pollen: Biology, lent ionophore monensin. Sex. Plant Reprod. Biochemistry, Management. Berlin: Springer-Verlag. 5:232-238. Stapleton, A.E., and Bedinger, P.A. 1992. Immature maize Springer, B., Werr, W., Starlinger, P., Bennett, D.C., spikelets develop and produce pollen in culture. Zakolica, M., and Freeling, M. 1986. The shrunken Plant Cell Rep. 11:248-252. gene on chromosome 9 of Zea mays L. is expressed in Staswick, P., and Chrispeels, M.J. 1984. Expression of various plant tissues and encodes an anaerobic prolectin genes during seed development in normal and tein. Mol. Gen. Genet. 205:461^468. phytohemagglutinin-deficient cultivars of Phaseolus vulgaris. J. Mol. Appl. Genet. 2:525-535. Springer, P.S., McCombie, W.R., Sundaresan, V., and Martienssen, R.A. 1995. Gene trap tagging of PRO- Staswick, P.E., Hermodson, M.A., and Nielsen, N.C. 1984. LIFERA, an essential MCM2-3-5-like gene in The amino acid sequence of the A2Bla subunit of Arabidopsis. Science 268:877-880. glycinin. /. Biol. Chem. 259:13424-13430. Sree Ramulu, K., Bredemeijer, G.M.M., Dijkhuis, P., de Stauffer, C, Benito Moreno, R.M., and Heberle-Bors, E. Nettancourt, D., and Schibilla, H. 1979. Mentor 1991. Seed set after pollination with in-vitropollen effects on gametophytic incompatibility in matured, isolated pollen of Triticum aestivum. Theor. Nicotiana, Oenothera and Lycopersicum. Theor. Appl. Appl. Genet. 81:576-580. Genet. 54:215-218. Stax£n, I., Klimaszewska, K., and Bornman, C.H. 1994. Microtubular organization in protoplasts and cells of Sree Ramulu, K., Devreux, M., Ancora, G., and Laneri, U. somatic embryo-regenerating and non-regenerating 1976. Chimerism in Lycopersicum peruvianum plants cultures of Larix. Physiol. Plant. 91:680-686. regenerated from in vitro cultures of anthers and Stead, A.D., Roberts, I.N., and Dickinson, H.G. 1979. stem internodes. Z. Pflanzenziichtg 76:299-319. Pollen-pistil interaction in Brassica oleracea. Events Srivastava, H.K., Sarkissian, I.V., and Shands, H.L. 1969. prior to pollen germination. Planta 146:211-216. Mitochondrial complementation and cytoplasmic Stead, A.D., Roberts, I.N., and Dickinson, H.G. 1980. male sterility in wheat. Genetics 63:611-618. Pollen-stigma interaction in Brassica oleracea: the role Stacey, N.J., Roberts, K., and Knox, J.P. 1990. Patterns of of stigmatic proteins in pollen grain adhesion. /. Cell expression of the JIM4 arabinogalactan-protein epiSci. 42:417-423. tope in cell cultures and during somatic embryogenSteer, M.W. 1977a. Differentiation of the tapetum in Avena. esis in Daucus carota L. Planta 180:285-292. I. The cell surface. /. Cell Sci. 25:125-138. Staff, I.A., Taylor, P., Kenrick, J., and Knox, R.B. 1989. Ultrastructural analysis of plastids in angiosperm Steer, M.W. 1977b. Differentiation of the tapetum in Avena. II. The endoplasmic reticulum and Golgi pollen tubes. Sex. Plant Reprod. 2:70-76. apparatus. /. Cell Sci. 28:71-86. Staff, I.A., Taylor, P.E., Smith, P., Singh, M.B., and Knox, R.B. 1990. Cellular localization of water soluble, Steer, M.W., and Steer, J.M. 1989. Pollen tube tip growth. allergenic proteins in rye-grass (Lolium perenne) New Phytol. 111:323-358. pollen using monoclonal and specific IgE antibodies Stefaniak, B. 1984. Development of rye embryo. I. with immunogold probes. Histochem. J. 22:276-290. Preliminary investigations on ultrastructure of 7day-old proembryo. Ada Soc. Bot. Polon. 53:11-16. Staiger, C.J., and Cande, W.Z. 1990. Microtubule distribution in dv, a maize meiotic mutant defective in the Stefaniak, B. 1987. The in vitro development of isolated prophase to metaphase transition. Devel Biol. proembryos. Ada Soc. Bot. Polon. 56:37-^2. 138:231-242. Steffen, K. 1963. Male gametophyte. In Recent Advances in
644
References
the Embryology of Angiosperms, ed. P. Maheshwari, Stevens, V.A.M., and Murray, B.G. 1982. Studies on hetpp. 15-40. Delhi: International Society of Plant eromorphic self-incompatibility systems: physiologMorphologists. ical aspects of the incompatibility system of Primula obconica. Theor. Appl. Genet. 61:245-256. Steffensen, D.M. 1966. Synthesis of ribosomal RNA during growth and division in Lilium. Expt. Cell Res. Steward, EC. 1963. The control of growth in plant cells. 44:1-12. Sci. Am. 209 (4):104-113. Stein, J.C., and Nasrallah, J.B. 1993. A plant receptor-like Steward, EC, Mapes, M.O., and Mears, K. 1958. Growth gene, the S-Iocus receptor kinase of Brassica oleracea and organized development of cultured cells. II. L, encodes a functional serine/threonine kinase. Organization in cultures grown from freely susPlant Physiol. 101:1103-1106. pended cells. Am. J. Bot. 45:705-708. Stein, J.C., Dixit, R., Nasrallah, M.E., and Nasrallah, J.B. Steward, EC, Mapes, M.O., and Smith, J. 1958. Growth 1996. SRK, the stigma-specific S locus receptor and organized development of cultured cells. I. kinase of Brassica, is targeted to the plasma memGrowth and division of freely suspended cells. Am. brane in transgenic tobacco. Plant Cell 8:429-445. J. Bot. 45:693-703. Stein, J.C., Howlett, B., Boyes, D.C., Nasrallah, M.E., and Steward, EC, Mapes, M.O., Kent, A.E., and Holsten, R.D. 1964. Growth and development of cultured plant Nasrallah, J.B. 1991. Molecular cloning of a putative cells. Science 143:20-27. receptor protein kinase gene encoded at the selfincompatibility locus of Brassica oleracea. Proc. NatlStewart, J.M., and Hsu, CL. 1977. In ovulo embryo culture Acad. Sci. USA 88:8816-8820. and seedling development of cotton (Gossypium hirStejskal, J., and Griga, M. 1995. Comparative analysis of sutum L.). Planta 137:113-117. some isozymes and proteins in somatic and zygotic Stickland, R.G., and Wilson, K.E. 1983. Sugars and starch embryos of soybean (Glycine max [L.] Merr.). /. Plant in developing round and wrinkled pea seeds. Ann. Physiol. 146:497-502. Bot. 52:919-921. Stelly, D.M., and Palmer, R.G. 1982. Variable development Stieglitz, H. 1977. Role of P-l,3-glucanase in post-meiotic in anthers of partially male-sterile soybeans. /. Hered. microspore release. Devel Biol. 57:87-97. 73:101-108. Stieglitz, H., and Stern, H. 1973. Regulation of P-l,3-gluStenram, U., Heneen, W.K., and Skerritt, J.H. 1991. canase activity in developing anthers of Lilium. Devel Immunocytochemical localization of wheat storage Biol. 34:169-173. proteins in endosperm cells 30 days after anthesis. /. Stinissen, H.M., Peumans, W.J., and Carlier, A.R. 1983. Expt. Bot. 42:1347-1355. Occurrence and immunological relationships of lectins in gramineous species. Planta 159:105-111. Sterk, P., Booij, H., Schellekens, G.A., van Kammen, A., and de Vries, S.C. 1991. Cell-specific expression of Stinissen, H.M., Peumans, W.J., and Chrispeels, M.J. 1984. the carrot EP2 lipid transfer protein gene. Plant Cell Subcellular site of lectin synthesis in developing rice 3:907-921. embryos. EMBO J. 3:1979-1985. Sterling, C. 1955. Embryogeny in the lima bean. Bull. Stinissen, H.M., Peumans, W.J., and de Langhe, E. 1984. Torrey Bot. Cl. 82:325-338. Abscisic acid promotes lectin biosynthesis in developing and germinating rice embryos. Plant Cell Rep. Stern H. 1956. Sulfhydryl groups and cell division. Science 124:1292-1293. 3:55-59. Stern, H. 1958. Variations in sulfhydryl concentration dur- Stinson, J., and Mascarenhas, J.P. 1985. Onset of alcohol dehydrogenase synthesis during microsporogenesis ing microsporocyte meiosis in anthers of Lilium and in maize. Plant Physiol. 77:222-224. Trillium. J. Biophys. Biochem. Cytol. 4:157-161. Stern, H. 1961. Periodic induction of deoxyribonuclease Stinson, J.R., Eisenberg, A.J., Willing, P.R., Pe, M.E., Hanson, D.D., and Mascarenhas, J.P. 1987. Genes activity in relation to the mitotic cycle. /. Biophys. expressed in the male gametophyte of flowering Biochem. Cytol. 9:271-277. plants and their isolation. Plant Physiol. 83:442-447. Stern, H., and Hotta, Y. 1977. Biochemistry of meiosis. Phil. Trans. Roy. Soc. Lond. 277B:277-294. Stirn, S., and Jacobsen, H.-J. 1987. Marker proteins for embryogenic differentiation patterns in pea callus. Stern, H., and Hotta, Y. 1984. Chromosome organization in the regulation of meiotic prophase. Symp. Soc. Plant Cell Rep. 6:50-54. Expt. Biol. 38:161-175. Stoger, E., Benito Moreno, R.M., Ylstra, B., Vicente, O., Stern, H., and Kirk, PL. 1948. The oxygen consumption of and Heberle-Bors, E. 1992. Comparison of different the microspores of Trillium in relation to the mitotic techniques for gene transfer into mature and immature tobacco pollen. Transgen. Res. 1:71-78. cycle. /. Gen. Physiol. 31:243-248. Stern, H., and Timonen, S. 1954. The position of the cell Stoger, E., Fink, C, Pfosser, M., and Heberle-Bors, E. 1995. Plant transformation by particle bombardment of nucleus in the pathways of hydrogen transfer: embryogenic pollen. Plant Cell Rep. 14:273-278. cytochrome C, flavoproteins, glutathione, and ascorStorchova, H., Capkova V, and Tupy, J. 1994. A Nicotiana bic acid. /. Gen. Physiol. 38:41-52. tabacum mRNA encoding a 69-kDa glycoprotein Stern H., Westergaard, M., and von Wettstein, D. 1975. occurring abundantly in pollen tubes is transcribed Presynaptic events in meiocytes of Lilium longiflorum but not translated during pollen development in the and their relation to crossing-over: a preselection anthers. Planta 192:441^45. hypothesis. Proc. Natl Acad. Sci. USA 72:961-965. Stettler, R.F., Koster, R., and Steenackers, V. 1980. Stout, A.B. 1931. Pollen-tube behavior in Brassica pekinensis with reference to self-incompatibility in fertilizaInterspecific crossability studies in poplars. Theor. tion. Am. ]. Bot. 18:686-695. Appl. Genet. 58:273-282. Stevens, V.A.M., and Murray, B.G. 1981. Studies on het- Stout, A.B., and Chandler, C. 1933. Pollen-tube behavior in Hemerocallis with special reference to incompatieromorphic self-incompatibility systems: the cytobilities. Bull. Torrey Bot. Cl. 60:397-416. chemistry and ultrastructure of the tapetum of Stout, A.B., and Chandler, C. 1941. Change from selfPrimula obconica. J. Cell Sci. 50:419-431.
References
645
sion and differentiation of pollen grains in Triticale incompatibility to self-compatibility accompanying anthers cultured in vitro. Sci. Sinica 17:47-54. change from diploidy to tetraploidy. Science 94:118. Strasburger, E. 1884. Neue Untersuchungen tiber den Sun, M., and Ganders, F.R. 1987. Microsporogenesis in Befruchtungsvorgang bei den Phanerogamen als Grnndlagemale-sterile and hermaphroditic plants of nine gynodioecious taxa of Hawaiian Bidens (Asteraceae). fur eine Theorie der Zeugung. Jena: Gustav Fischer. Am. J. Bot. 74:209-217. Street, H.E., and Withers, L.A. 1974. The anatomy of embryogenesis. In Tissue Culture and Plant Science Sun, S.M., Slightom, J.L., and Hall, T.C. 1981. Intervening sequences in a plant gene - comparison of the partial 1974, ed. H.E. Street, pp. 71-100. London: Academic sequence of cDNA and genomic DNA of French Press. bean phaseolin. Nature 289:37-41. Strokov, A.A., Bogdanov, Y.F., and Reznickova, S.A. 1973. A quantitative study of histones of meiocytes. II. Sun, S.M., Mutschler, M.A., Bliss, F.A., and Hall, T.C. 1978. Protein synthesis and accumulation in bean cotylePolyacrylamide gel electrophoresis of isolated hisdons during growth. Plant Physiol. 61:918-923. tones from Lilium microsporocytes. Chromosoma 43:247-260. Sunderland, N. 1971. Anther culture: a progress report. Stuart, D.A., and Strickland, S.G. 1984a. Somatic embryoSci. Prog. Oxford 59:527-549. genesis from cell cultures of Medicago saliva L. I. The Sunderland, N., Collins, G.B., and Dunwell, J.M. 1974. role of amino acid additions to the regeneration The role of nuclear fusion in pollen embryogenesis medium. Plant Sci. Lett. 34:165-174. of Datura innoxia Mill. Planta 117:227-241. Stuart, D.A., and Strickland, S.G. 1984b. Somatic embryo- Sunderland, N., and Dunwell, J.M. 1977. Anther and genesis from cell cultures of Medicago sativa L. II. The pollen culture. In Plant Tissue and Cell Culture, 2nd interaction of amino acids with ammonium. Plant ed., ed. H.E. Street, pp. 223-265. Berkeley: University Sci. Lett. 34:175-181. of California Press. Sturaro, M, Vernieri, P., Castiglioni, P., Binelli, G., and Sunderland, N., and Evans, L.J. 1980. Multicellular pollen Gavazzi, G. 1996. The rea (red embryonic axis) pheformation in cultured barley anthers. II. The A, B, and C pathways. /. Expt. Bot. 31:501-514. notype describes a new mutation affecting the response of maize embryos to abscisic acid and Sunderland, N., and Roberts, M. 1977. New approach to osmotic stress. /. Expt. Bot. 47:717-728. pollen culture. Nature 270:236-238. Sturm, A., Voelker, T.A., Herman, E.M., and Chrispeels, Sunderland, N., and Roberts, M. 1979. Cold-pretreatment MJ. 1988. Correct glycosylation, Golgi-processing, of excised flower buds in float culture of tobacco and targeting to protein bodies of the vacuolar proanthers. Ann. Bot. 43:405-414. tein phytohemagglutinin in transgenic tobacco. Sunderland, N., and Wicks, F.M. 1971. Embryoid formaPlanta 175:170-183. tion in pollen grains of Nicotiana tabacum. ]. Expt. Bot. 22:213-226. Sugimoto, T, Momma, M., Hashizume, K., and Saio, K. 1987. Components of storage protein in hypocotyl- Sunderland, N., and Wildon, D.C. 1979. A note on the preradicle axis of soybean (Glycine max cv. Enrei) seeds. treatment of excised flower buds in float culture of Agr. Biol. Chem. 51:1231-1238. Hyoscyamus anthers. Plant Sci. Lett. 15:169-175. Sugiyama, T., Rafalski, A., and Soil, D. 1986. The Sunderland, N., Roberts, M., Evans, L.J., and Wildon, D.C. nucleotide sequence of a wheat y-gliadin genomic 1979. Multicellular pollen formation in cultured barclone. Plant Sci. 44:205-209. ley anthers. I. Independent division of the generative and vegetative cells. /. Expt. Bot. 30:1133-1144. Sugiyama, T, Rafalski, A., Peterson, D., and Soil, D. 1985. A wheat HMW glutenin subunit gene reveals a Sung, Z.R. 1979. Relationship of indole-3-acetic acid and tryptophan concentrations in normal and 5-methylhighly repeated structure. Nucl. Acids Res. tryptophan-resistant cell lines of wild carrots. Planta 13:8729-8737. 145:339-345. Sullivan, T.D., and Kaneko, Y. 1995. The maize brittlel gene encodes amyloplast membrane polypeptides. Sung, Z.R., Lazar, G.B., and Dudits, D. 1981. CycloPlanta 196:477-484. heximide resistance in carrot culture: a differentiated function. Plant Physiol. 68:261-264. Sullivan, T.D., Strelow, L.I., Illingworth, C.A., Phillips, R.L., and Nelson, O.E., Jr. 1991. Analysis of maize Sung, Z.R., and Okimoto, R. 1981. Embryonic proteins in somatic embryos of carrot. Proc. Natl Acad. Sci. USA brittle-1 alleles and a defective suppressor-mutatorinduced mutable allele. Plant Cell 3:1337-1348. 78:3683-3687. Sumner, M.J. 1992. Embryology of Brassica campestris: the Sung, Z.R., and Okimoto, R. 1983. Coordinate gene entrance and discharge of the pollen tube in the synexpression during somatic embryogenesis in carrots. Proc. Natl Acad. Sci. USA 80:2661-2665. ergid and the formation of the zygote. Can. J. Bot. 70:1577-1590. Sung, Z.R., Smith, R., and Horowitz, J. 1979. Quantitative Sumner, M.J., and van Caeseele, L. 1989. The ultrastrucstudies of embryogenesis in normal and 5-methylture and cytochemistry of the egg apparatus of tryptophan-resistant cell lines of wild carrot. Planta Brassica campestris. Can. J. Bot. 67:177-190. 147:236-240. Sumner, M.J., and van Caeseele, L. 1990. The develop- Sung, Z.R., Belachew, A., Shunong, B., and Bertrandment of the central cell of Brassica campestris prior to Garcia, R. 1992. EMF, an Arabidopsis gene required fertilization. Can. ]. Bot. 68:2553-2563. for vegetative shoot development. Science Sumner-Smith, M., Rafalski, J.A., Sugiyama, T., Stoll, M., 258:1645-1647. and Soil, D. 1985. Conservation and variability of Suss, J., and Tupy, J. 1976. On the nature of RNA synthesized in pollen tubes of Nicotiana alata. Biol. Plant. wheat a/ P-gliadin genes. Nucl. Acids Res. 13:3905-3916. Sun, C.-S. 1978. Androgenesis of cereal crops. In 18:140-146. Proceedings of Symposium on Plant Tissue Culture, pp. Suss, J., and Tupy, J. 1978. tRNA synthesis in germinating pollen. Biol. Plant. 20:70-72. 117-123. Peking: Science Press. Sun, C.-S., Wang, C.-C, and Chu, C.-C. 1974. Cell divi- Suss, J., and Tupy, J. 1979. Poly(A)+RNA synthesis in ger-
646
References
minating pollen of Nicotiana tabacum L. Biol. Plant. O., Heberle-Bors, E., Ahorn, H., and Breitenbach, M. 21:365-371. 1995b. Isoforms of Bet v 1, the major birch pollen allergen, analyzed by liquid chromatography, mass Suss, J., and Tupy, J. 1982. Kinetics of uridine uptake and spectrometry, and cDNA cloning. /. Biol. Chem. incorporation into RNA in tobacco pollen culture. Biol. Plant. 24:72-79. 270:2607-2613. Sussex, I. 1975. Growth and metabolism of the embryo Syamasundar, J., and Panchaksharappa, M.G. 1975. A and attached seedling of the viviparous mangrove, cytochemical study of the hypertrophied synergid in Rhizophora mangle. Am. J. Bot. 62:948-953. Allium cepa L. Cytologia 40:371-376. Sussex, I., Clutter, M., Walbot, V., and Brady, T. 1973. Syamasundar, J., and Panchaksharappa, M.G. 1976. A hisBiosynthetic activity of the suspensor of Phaseolus tochemical study of some post-fertilization developcoccineus. Caryologia 25: Suppl. 261-272. mental stages in Dipcadi montanum Dalz. Cytologia 41:123-130. Sussmuth, J., Dressier, K., and Hess, D. 1991. Agrobacterium-mediated transfer of the GUS gene Szakacs, E., and Barnabas, B. 1988. Cytological aspects of into pollen of Petunia. Bot. Ada 104:72-76. in vitro androgenesis in wheat (Triticum aestivum L.) using fluorescent microscopy. Sex. Plant Reprod. Suzuki, G., Watanabe, M., Toriyama, K., Isogai, A., and 1:217-222. Hinata, K. 1995. Molecular cloning of members of the S-multigene family in self-incompatible Brasska Szakacs, E., and Barnabas, B. 1995. The effect of colchicine campestris L. Plant Cell Physiol. 36:1273-1280. treatment on microspore division and microsporederived embryo differentiation in wheat (Triticum Suzuki, M., Koide, Y., Hattori, T., Nakamura, K., and aestivum L.) anther culture. Euphytica 83:209-213. Asahi, T. 1995. Different sets of cis-elements contribute to the expression of a catalase gene from cas- Szymkowiak, E.J., and Sussex, I.M. 1992. The internal tor bean during seed formation and postembryonic meristem layer (L3) determines floral meristem size development in transgenic tobacco. Plant Cell and carpel number in tomato periclinal chimeras. Physiol. 36:1067-1074. Plant Cell 4:1089-1100. Suzuki, T., Nakamura, C., Mori, N., and Kaneda, C. 1995. Overexpression of mitochondrial genes in alloplas- Tabata, S., Sato, S., Watanabe, Y., Yamamoto, M., and mic common wheat with a cytoplasm of wheatgrass Hotta, Y. 1993. Evidence of meiosis-specific regula(Agropyron trichophorum) showing depressed vigor tion of gene expression in lily microsporocytes. Plant and male sterility. Plant Mol. Biol. 27:553-565. Sci. 89:31-41. Swain, E., Li, C.P., and Poulton, J.E. 1992a. Development Tagliasacchi, A.M., Forino, L.M.C., Cionini, P.G., of the potential for cyanogenesis in maturing black Cavallini, A., Durante, M., Cremonini, R., and cherry (Primus serotina Ehrh.) fruits. Plant Physiol. Avanzi, S. 1984. Different structure of polytene chro98:1423-1428. mosome of Phaseolus coccineus suspensors during early embryogenesis. 3. Chromosome pair VI. Swain, E., Li, C.P., and Poulton, J.E. 1992b. Tissue and subProtoplasma 122:98-107. cellular localization of enzymes catabolizing (R)amygdalin in mature Prunus serotina seeds. Plant Takahashi, M. 1987. Development of omniaperturate Physiol. 100:291-300. pollen in Trillium kamtschaticum (Liliaceae). Am. ]. Swaminathan, M.S., Chopra, V.L., and Bhaskaran, S. 1962. Bot. 74:1842-1852. Cytological aberrations observed in barley embryos Takahashi, M. 1989a. Development of the echinate pollen cultured in irradiated potato mash. Radiation Res. wall in Farfugium japonicum (Compositae: 16:182-188. Senecioneae). Bot. Mag. Tokyo 102:219-234. Swamy, B.G.L. 1979. Embryogenesis in Cheirostylis flabel- Takahashi, M. 1989b. Pattern determination of the exine in Caesalpinia japonica (Leguminosae: Caesallata. Phytomorphology 29:199-203. pinioideae). Am. J. Bot. 76:1615-1626. Swamy, B.G.L. 1980. Embryogenesis in Sagittaria sagittaeTakahashi, M. 1993. Exine initiation and substructure in folia. Phytomorphology 30:204-212. pollen of Caesalpinia japonica (Leguminosae: Swamy, B.G.L., and Ganapathy, P.M. 1957. A new type of Caesalpinioideae). Am. J. Bot. 80:192-197. endosperm haustorium in Nothapodytes foetida. Phytomorphology 7:331-336. Takahashi, M. 1994. Pollen development in a submerged plant, Ottelia alismoides (L.) Pers. (Hydrocharitaceae). Swamy, B.G.L., and Lakshmanan, K.K. 1962a. The origin /. Plant Res. 107:161-164. of epicotylary meristem and cotyledon in Halophila ovata Gaudich. Ann. Bot. 26:243-249. Takahashi, M. 1995. Three-dimensional aspects of exine initiation and development in Lilium longiflorum Swamy, B.G.L., and Lakshmanan, K.K. 1962b. (Liliaceae). Am. J. Bot. 82:847-854. Contributions to the embryology of the Najadaceae. /. Indian Bot. Soc. 41:247-267. Takahashi, M., and Kouchi, J. 1988. Ontogenetic developSwegle, M., Kramer, K.J., and Muthukrishnan, S. 1992. ment of spinous exine in Hibiscus syriacus Properties of barley seed chitinases and release of (Malvaceae). Am. }. Bot. 75:1549-1558. embryo-associated isoforms during early stages of Takahashi, M., and Skvarla, J.J. 1991. Exine pattern forimbibition. Plant Physiol. 99:1009-1014. mation by plasma membrane in Bougainvillea spectabilis Willd. (Nyctaginaceae). Am. J. Bot. Swift, H. 1950. The constancy of desoxyribose nucleic acid 78:1063-1069. in plant nuclei. Proc. Natl Acad. Sci. USA 36:643-654. Swoboda, I., Dang, T.C.H., Heberle-Bors, E., and Vicente, Takahashi, T, Naito, S., and Komeda, Y. 1992. The O. 1995a. Expression of Bet c 1, the major birch Arabidopsis HSP18.2 promoter/GUS gene fusion in pollen allergen, during anther development. transgenic Arabidopsis plants: a powerful tool for the isolation of regulatory mutants of the heat-shock Protoplasma 187:103-110. response. Plant J. 2:751-761. Swoboda, I., Jilek, A., Ferreira, R, Engel, E., HoffmannSommergruber, K., Scheiner, O., Kraft, D., Takaiwa, F., Kikuchi, S., and Oono, K. 1986. The strucBreiteneder, H., Pittenauer, E., Schmid, E., Vicente, ture of rice storage protein glutelin precursor
References
647
Tanaka, 1.1988. Isolation of generative cells and their prodeduced from cDNA. FEBS Lett. 206:33-35. toplasts from pollen of Lilium longiflorum. Takaiwa, R, Kikuchi, S., and Oono, K. 1987. A rice glutelin Protoplasma 142:68-73. gene family - a major type of glutelin mRNAs can be divided into two classes. Mol. Gen. Genet. 208:15-22. Tanaka, 1.1991. Microtubule-determined plastid distribution during microsporogenesis in Lilium longiflorum. Takaiwa, R, Kikuchi, S., and Oono, K. 1989. The comJ. Cell Sci. 99:21-31. plete nucleotide sequence of new type cDNA coding for rice storage protein glutelin. Nucl. Acids Tanaka, I. 1993. Development of male gametes in flowerRes. 17:3289. ing plants. /. Plant Res. 106:55-63. Takaiwa, R, and Oono, K. 1991. Genomic DNA sequences Tanaka, I., and Ito, M. 1980. Induction of typical cell division in isolated microspores of Lilium longiflorum of two new genes for new storage protein glutelin in and Tulipa gesneriana. Plant Sci. Lett. 17:279-285. rice. Jap. ]. Genet. 66:161-171. Takaiwa, P., Oono, K., and Kato, A. 1991. Analysis of the Tanaka, I., and Ito, M. 1981a. Studies on microspore development in liliaceous plants. III. Pollen tube develop5' flanking region responsible for the endospermment in lily pollens cultured from the uninucleate specific expression of a rice glutelin chimeric gene in microspore stage. Plant Cell Physiol. 22:149-153. transgenic tobacco. Plant Mol. Biol. 16:49-58. Takaiwa, R, Ebinuma, H., Kikuchi, S., and Oono, K. 1987. Tanaka, I., and Ito, M. 1981b. Control of division patterns Nucleotide sequence of a rice glutelin gene. FEBS in explanted microspores of Tulipa gesneriana. Lett. 221:43-47. Protoplasma 108:329-340. Takaiwa, R, Oono, K., Wing, D., and Kato, A. 1991. Tanaka, I., Kitazume, C, and Ito, M. 1987. The isolation and culture of lily pollen protoplasts. Plant Sci. Sequence of three members and expression of a new 50:205-211. major subfamily of glutelin genes from rice. Plant Mol. Biol. 17:875-885. Tanaka, I., Nakamura, S., and Miki-Hirosige, H. 1989. Structural features of isolated generative cells and Takats, S.T. 1959. Chromatin extrusion and DNA transfer their protoplasts from pollen of some liliaceous during microsporogenesis. Chromosoma 10:430-453. plants. Gamete Res. 24:361-374. Takats, S.T. 1962. An attempt to detect utilization of DNA breakdown products from the tapetum for DNA Tanaka, I., Taguchi, T, and Ito, M. 1979. Studies on microspore development in liliaceous plants. I. The synthesis in the microspores of Lilium longiflorum. duration of the cell cycle and developmental aspects Am. ]. Bot. 49:748-758. in lily microspores. Bot. Mag. Tokyo 92:291-298. Takats, S.T. 1965. Non-random nuclear orientation during DNA synthesis in Tradescantia pollen grains. /. Cell Tanaka, I., Taguchi, T., and Ito, M. 1980. Studies on Biol. 25:151-157. microspore development in liliaceous plants. II. The behavior of explanted microspores of the lily, Takats, S.T, and Wever, G.H. 1971. DNA polymerase and Lilium longiflorum. Plant Cell Physiol. 21:667-676. DNA nuclease activities in S-competent and Sincompetent nuclei from Tradescantia pollen grains. Tanaka, I., and Wakabayashi, T. 1992. Organization of the Expt. Cell Res. 69:25-28. actin and microtubule cytoskeleton preceding pollen germination. An analysis using cultured Takayama, S., Isogai, A., Tsukamoto, C, Ueda, Y., Hinata, pollen protoplasts of Lilium longiflorum. Planta K., Okazaki, K., and Suzuki, A. 1987. Sequences of S186:473-482. glycoproteins, products of the Brassica campestris self-incompatibility locus. Nature 326:102-105. Tanaka, T., Nishihara, M., Seki, M., Sakamoto, A., Tanaka, K., Irifune, K., and Morikawa, H. 1995. Successful Takegami, M.H., and Ito, M. 1975. Studies on the behavior expression in pollen of various plant species of in of meiotic protoplasts. III. Peatures of nuclear and vitro synthesized mRNA introduced by particle cell division for lily protoplasts in the cell-wall-free bombardment. Plant Mol. Biol. 28:337-341. state. Bot. Mag. Tokyo 88:187-196. Takegami, M.H., Yoshioka, M., Tanaka, I., and Ito, M. Tang, W., Diaz, P., Wilson, K.A., and Tan-Wilson, A.L. 1993. Spatial distribution of protein accumulation in 1981. Characteristics of isolated microsporocytes soybean seed. Phytochemistry 33:1289-1295. from liliaceous plants for studies of the meiotic cell Tang, X., Hepler, P.K., and Scordilis, S.P. 1989. cycle in vitro. Plant Cell Physiol. 22:1-10. Immunochemical and immunocytochemical identiTakei, Y., Yamauchi, D., and Minamikawa, T. 1989. fication of a myosin heavy chain polypeptide in Nucleotide sequence of the canavalin gene from Nicotiana pollen tubes. /. Cell Sci. 92:569-574. Canavalia gladiata seeds. Nucl. Acids Res. 17:4381. Takeno, K., Koshioka, M., Pharis, R.P., Rajasekaran, K., Tang, X., Lancelle, S.A., and Hepler, P.K. 1989. and Mullins, M.G. 1983. Endogenous gibberellin-like Fluorescence microscopic localization of actin in substances in somatic embryos of grape (Vitis pollen tubes: comparison of actin antibody and phalvinifera x Vitis rupestris) in relation to embryogenesis loidin staining. Cell Mot. Cytoskel. 12:216-224. and the chilling requirement for subsequent devel- Tanksley, S.D., Zamir, D., and Rick, CM. 1981. Evidence opment of mature embryos. Plant Physiol. for extensive overlap of sporophytic and gameto73:803-808. phytic gene expression in Lycopersicon esculentum. Science 213:453-455. Talbot, D.R., Adang, M.J., Slightom, J.L., and Hall, T.C. 1984. Size and organization of a multigene family Tano, S., and Takahashi, H. 1964. Nucleic acid synthesis in encoding phaseolin, the major seed storage protein growing pollen tubes. /. Biochem. 56:578-580. of Phaseolus vulgaris L. Mol. Gen. Genet. 198:42^9. Tantikanjana, T., Nasrallah, M.E., and Nasrallah, J.B. 1996. Tamura, Y, Kitano, H., Satoh, H., and Nagato, Y. 1992. A The Brassica S gene family: molecular characterizagene profoundly affecting shoot organization in the tion of the SLR2 gene. Sex. Plant Reprod. 9:107-116. early phase of rice development. Plant Sci. 82:91-99. Tantikanjana, T, Nasrallah, M.E., Stein, J.G, Chen, C.-H., and Nasrallah, J.B. 1993. An alternative transcript of Tan, L.W., and Jackson, J.R 1988. Stigma proteins of the two loci self-incompatible grass Phalaris coerulescens. the S locus glycoprotein gene in a class II pollenrecessive self-incompatibility haplotype of Brassica Sex. Plant Reprod. 1:25-27.
648
References
oleracea encodes a membrane-anchored protein. Rhododendron: quantitative cytology, three-dimenPlant Cell 5:657-666. sional reconstruction, isolation and detection using fluorescent probes. Sex. Plant Reprod. 2:254-264. Tara, C.P., and Namboodiri, A.N. 1974. Aberrant microsporogenesis and sterility in Impatiens sultani Taylor, P.E., Kenrick, J., Blomstedt, C.K., and Knox, R.B. (Balsaminaceae). Am. ]. Bot. 61:585-591. 1991. Sperm cells of the pollen tubes of Brassica: Tarasenko, L.V., and Bannikova, V.P. 1991. Ultrastructure ultrastructure and isolation. Sex. Plant Reprod. of pollen grain development in some angiosperm 4:226-234. species. Phytomorphology 41:11-20. Tebbutt, S.J., and Lonsdale, D.M. 1995. Deletion analysis Tautorus, T.E., Wang, H., Fowke, L.C., and Dunstan, D.I. of a tobacco pollen-specific polygalacturonase. Sex. 1992. Microtubule pattern and the occurrence of prePlant Reprod. 8:242-246. prophase bands in embryogenic cultures of black Tebbutt, S.J., Rogers, H.J., and Lonsdale, D.M. 1994. spruce (Picea mariana Mill.) and non-embryogenic Characterization of a tobacco gene encoding a cultures of jack pine (Pinus banksiana Lamb.). Plant pollen-specific polygalacturonase. Plant Mol. Bib/. Cell Rep. 11:419-423. 25:283-297. Taylor, D.C., Barton, D.L., Rioux, K.P., MacKenzie, S.L., Telmer, C.A., Newcomb, W., and Simmonds, D.H. 1993. Reed, D.W., Underhill, E.W., Pomeroy, M.K., and Microspore development in Brassica napus and the Weber, N. 1992. Biosynthesis of acyl lipids containeffect of high temperature on division in vivo and in ing very-long chain fatty acids in microsporevitro. Protoplasma 172:154-165. derived and zygotic embryos of Brassica napus L. cv. Telmer, C.A., Newcomb, W., and Simmonds, D.H. 1995. Reston. Plant Physiol. 99:1609-1618. Cellular changes during heat shock induction and embryo development of cultured microspores of Taylor, D.C., Ferrie, A.M.R., Keller, W.A., Giblin, E.M., Pass, E.W., and MacKenzie, S.L. 1993. Bioassembly Brassica napus cv. Topas. Protoplasma 185:106-112. of acyl lipids in microspore-derived embryos of Temple, M., Makaroff, C.A., Mutschler, M.A., and Earle, Brassica campeslris L. Plant Cell Rep. 12:375-384. E.D. 1992. Novel mitochondrial genomes in Brassica Taylor, D.C., Weber, N., Barton, D.L., Underhill, E.W., napus somatic hybrids. Curr. Genet. 22:243-249. Hogge, L.R., Weselake, R.J., and Pomeroy, M.K. 1991. Templeman, T.S., DeMaggio, A.E., and Stetler, D.A. 1987. Triglycerol bioassembly in microspore-derived Biochemistry of fern spore germination: globulin embryos of Brassica napus L. cv. Reston. Plant Physiol. storage proteins in Matteuccia struthiopteris L. Plant 97:65-79. Physiol. 85:343-349. Taylor, D.C., Weber, N., Underhill, E.W., Pomeroy, M.K., Templeman, T.S., Stein, D.B., and DeMaggio, A.E. 1988. A Keller, W.A., Scowcroft, W.R., Wilen, R.W., Moloney, fern spore storage protein is genetically similar to M.M., and Holbrook, L.A. 1990. Storage-protein regthe 1.7 S seed storage protein of Brassica napus. ulation and lipid accumulation in microspore Biochem. Genet. 26:595-603. embryos of Brassica napus L. Planta 181:18-26. Tepfer, S.S. 1953. Floral anatomy and ontogeny in Taylor, J.H. 1950. The duration of differentiation in Aquilegia formosa var. truncata and Ranunculus repens. excised anthers. Am. J. Bot. 37:137-143. Univ. Calif. Publ. Bot. 25:513-647. Taylor, J.H. 1953. Autoradiographic detection of incorpo- Tepfer, S.S., Greyson, R.I., Craig, W.R., and Hindman, J.L. ration of P32 into chromosomes during meiosis and 1963. In vitro culture of floral buds of Aquilegia. Am. mitosis. Expt. Cell Res. 4:164-173. J. Bot. 50:1035-1045. Taylor, J.H. 1958. Incorporation of phosphorus-32 into Terada, R., Nakayama, T., Iwabuchi, M., and Shimamoto, nucleic acids and proteins during microgametogenK. 1993. A wheat histone H3 promoter confers cell esis in Tulbaghia. Am. }. Bot. 45:123-131. division-dependent and -independent expression of the gus A gene in transgenic rice plants. Plant ]. Taylor, J.H. 1959. Autoradiographic studies of nucleic 3:241-252. acids and proteins during meiosis in Lilium longiflorum. Am. J. Bot. 46:477-484. Terada, R., Nakayama, T, Iwabuchi, M., and Shimamoto, K. 1995. A type I element composed of the hexamer Taylor, J.H., and McMaster, R.D. 1954. Autoradiographic (ACGTCA) and octamer (CGCGGATC) motifs plays and microphotometric studies of desoxyribose a role(s) in meristematic expression of a wheat hisnucleic acid during microgametogenesis in Lilium longiflorum. Chromosoma 6:489-521. tone H3 gene in transgenic rice plants. Plant Mol. Biol. 27:17-26. Taylor, J.R.N., Schiissler, L., and Liebenberg, N.v.d.W. 1985. Protein body formation in the starchy Terada, S. 1928. Embryological studies in Oryza sativa L. /. endosperm of developing Sorghum bicolor (L.) Coll. Agr. Hokkaido Imp. Univ. 19:245-260. Moench seeds. S. Afr. }. Bot. 51:35-40. Terasaka, O., and Niitsu, T. 1989. Peculiar spindle configuration in the pollen tube revealed by the anti-tubuTaylor, L.P., and Jorgensen, R. 1992. Conditional male ferlin immunofluorescence method. Bot. Mag. Tokyo tility in chalcone synthase-deficient Petunia. ]. Hered. 102:143-147. 83:11-17. Taylor, M.G., and Vasil, I.K. 1991. Histology of, and phys- Terasaka, O., and Niitsu, T. 1990. Unequal cell division and chromatin differentiation in pollen grain cells. II. ical factors affecting, transient GUS expression in Microtubule dynamics associated with the unequal pearl millet (Pennisetum glaucum (L.) R. Br.) embryos following microprojectile bombardment. Plant Cell cell division. Bot. Mag. Tokyo 103:133-142. Rep. 10:120-125. Terasaka, O., and Niitsu, T. 1994. Differential roles of microTaylor, M.G., Vasil, V., and Vasil, I.K. 1993. Enhanced GUS tubule and actin-myosin cytoskeleton in the growth gene expression in cereal /grass cell suspensions and of Pinus pollen tubes. Sex. Plant Reprod. 7:264-272. immature embryos using the maize ubiquitin-based Terasaka, O., and Niitsu, T. 1995. The mitotic apparatus during unequal microspore division observed by a plasmid pAHC25. Plant Cell Rep. 12:491-495. confocal laser scanning microscope. Protoplasma Taylor, P., Kenrick, J., Li, Y., Kaul, V., Gunning, B.E.S., and Knox, R.B. 1989. The male germ unit of 189:187-193.
References
649
1992. Accumulation of group 3 late embryogenesis Terasawa, M., Shinohara, A., Hotta, Y., Ogawa, H., and abundant proteins in Zea mays embryos. Role of Ogawa, T. 1995. Localization of RecA-like recombiabscisic acid and the viviparous-1 gene product. Plant nation proteins on chromosomes of the lily at variPhysiol. 99:607-614. ous meiotic stages. Genes Devel. 9:925-934. Terc£-Laforgue, T., and Pernollet, J.-C. 1982. fitude quan- Thomas, E., Konar, R.N., and Street, H.E. 1972. The fine structure of the embryogenic callus of Ranunculus titative et qualitative de 1'accumulation des gliadines sceleratus L. /. Cell Sci. 11:95-109. au cours du deVeloppement du caryopse de bl£ (Triticum aestivtim L.). Compt. Rend. Acad. Sci. Paris Thomas, M.K., and Dnyansagar, V.R. 1975. Carbohydrate Ser. Ill 294:529-534. metabolism in pollen of Petunia nydaginiflora Juss. during germination and tube growth. Indian J. Expt. Testillano, P.S., Gonzaiez-Melendi, P., Ahmadian, P., Biol. 13:268-271. Fad6n, Bv and Risueno, M.C. 1995. The immunolocalization of nuclear antigens during the pollen Thomas, M.S., and Flavell, R.B. 1990. Identification of an developmental program and the induction of pollen enhancer element for the endosperm-specific embryogenesis. Expt. Cell Res. 221:41-54. expression of high molecular weight glutenin. Plant Thangavelu, Mv Belostotsky, D., Bevan, M.W., Flavell, Cell 2:1171-1180. R.B., Rogers, H.J., and Lonsdale, D.M. 1993. Partial Thompson, A.J., Bown, D., Yaish, S., and Gatehouse, J.A. 1991. Differential expression of seed storage protein characterization of the Nicotiana tabacum actin gene family: evidence for pollen-specific expression of genes in the pea leg] subfamily; sequence of gene one of the gene family members. Mol. Gen. Genet. legK. Biochem. Physiol. Pflanz. 187:1-12. 240:290-295. Thompson, A.J., Evans, I.M., Boulter, D., Croy, R.R.D., and Gatehouse, J.A. 1989. Transcriptional and post-tranThangstad, O.P., Iversen, T.-H., Slupphaug, G., and Bones, scriptional regulation of seed storage-protein gene A. 1990. Immunocytochemical localization of expression in pea (Pisum sativum L.). Planta myrosinase in Brassica napus L. Planta 180:245-248. 179:279-287. Thanh, V.H., and Shibasaki, K. 1977. Beta-conglycinin from soybean proteins. Isolation and immunological Thompson, G.A., Boston, R.S., Lyznik, L.A., Hodges, and physicochemical properties of the monomeric T.K., and Larkins, B.A. 1990. Analysis of promoter forms. Biochim. Biophys. Ada 490:370-384. activity from an a-zein gene 5' flanking sequence in transient expression assays. Plant Mol. Biol. Theerakulpisut, P., Xu, H., Singh, M.B., Pettitt, J.M., and 15:755-764. Knox, R.B. 1991. Isolation and developmental expression of Bcpl, an anther-specific cDNA clone in Thompson, G.A., Scherer, D.E., Foxall-van Aken, S., Kenny, J.W., Young, H.L., Shintani, D.K., Kridl, J.C, Brassica campestris. Plant Cell 3:1073-1084. and Knauf, V.C. 1991. Primary structures of the preTheunis, C.H. 1990. Ultrastructural analysis of Spinacia cursor and mature forms of stearoyl-acyl carrier prooleracea sperm cells isolated from mature pollen tein desaturase from safflower embryos and grains. Protoplasma 158:176-181. requirement of ferredoxin for enzyme activity. Proc. Theunis, C.H., Cresti, M., and Milanesi, C. 1991. Studies Natl Acad. Sci. USA 88:2578-2582. of the mature pollen of Spinacia oleracea after freeze substitution and observed with confocal laser scan- Thompson, K.F., and Taylor, J.P. 1971. Self-compatibility in kale. Heredity 27:459-471. ning fluorescence microscopy. Bot. Ada 104:324-329. Theunis, C.H., McConchie, C.A., and Knox, R.B. 1985. Thompson, R.D., and Bartels, D. 1983. Hordein messenger RNA levels in wild type and mutant barley Three-dimensional reconstruction of the generative endosperm. Plant Sci. Lett. 29:295-304. cell and its wall connection in mature bicellular pollen of Rhododendron. Micron Micros. Ada Thompson, R.D., Bartels, D., and Harberd, N.P. 1985. 16:225-231. Nucleotide sequence of a gene from chromosome ID of wheat encoding HMW-glutenin subunit. Nucl. Theunis, C.H., Pierson, E.S., and Cresti, M. 1991. Isolation Acids Res. 13:6833-6846. of male and female gametes in higher plants. Sex. Thompson, R.D., Bartels, D., Harberd, N.P., and Flavell, Plant Reprod. 4:145-154. R.B. 1983. Characterization of the multigene family Theunis, C.H., Pierson, E.S., and Cresti, M. 1992. The coding for HMW glutenin subunits in wheat using microtubule cytoskeleton and the rounding of isocDNA clones. Theor. Appl. Genet. 67:87-96. lated generative cells of Nicotiana tabacum. Sex. Plant Thompson, R.D., Kemble, R.J., and Flavell, R.B. 1980. Reprod. 5:64-71. Variations in mitochondrial DNA organisation Theunis, C.H., and van Went, J.L. 1989. Isolation of sperm between normal and male-sterile cytoplasms of cells from mature pollen grains of Spinacia oleracea. maize. Nucl. Acids Res. 8:1999-2008. Sex. Plant Reprod. 2:97-102. Thie-baud, C.H., and Ruch, F. 1978. Cytophotometric Thompson, R.D., and Kirch, H.-H. 1992. The S locus of flowering plants: when self-rejection is self-interest. study of nuclear differentiation during pollen develTrend. Genet. 8:381-387. opment in Tradescantia paludosa. Histochemistry Thompson-Coffe, C, Findlay, T.A.S., Wagner, B.A. and 57:119-128. Orr, A.R. 1992. Changes in protein complement This, P., Goffner, D., Raynal, M., Chartier, Y., and Delseny, accompany early organogenesis in the maize ear. Int. M. 1988. Characterization of major storage proteins ]. Plant Sci. 153:31-39. of sunflower and their accumulation. Plant Physiol. Biochem. 26:125-132. Thomson, J.A., and Schroeder, H.E. 1978. Cotyledonary Thoma, S., Hecht, U., Kippers, A., Botella, ]., de Vries, S., storage proteins in Pisum sativum. II. Hereditary and Somerville, C. 1994. Tissue-specific expression variation in components of the legumin and vicilin of a gene encoding a cell wall-localized lipid transfractions. Aust. ]. Plant Physiol. 5:281-294. fer protein from Arabidopsis. Plant Physiol. Thorsness, M.K., Kandasamy, M.K., Nasrallah, M.E., and Nasrallah, J.B. 1991. A Brassica S-locus gene pro105:35-45. moter targets toxic gene expression and cell death to Thomann, E.B., Sollinger, }., White, C, and Rivin, CJ.
650
References
the pistil and pollen of transgenic Nicotiana. Devel carrot (Daucus carota L.). Protoplasma 153:24-29. Biol. 143:173-184. Timmers, A.C.J., Kieft, H., and Schel, J.H.N. 1995. An immunofluorescence study on calmodulin distribuThorsness, M.K., Kandasamy, M.K., Nasrallah, M.E., and tion during somatic and zygotic embryogenesis of Nasrallah, J.B. 1993. Genetic ablation of floral cells in Arabidopsis. Plant Cell 5:253-261. carrot (Daucus carota L.). Acta Bot. Neerl. 44:19-32. Thoyts, P.J.E., Millichip, M.I., Stobart, A.K., Griffiths, W.T., Timmers, A.C.J., Reiss, H.-D., Bohsung, J., Traxel, K., and Shewry, P.R., and Napier, J.A. 1995. Expression and Schel, J.H.N. 1996. Localization of calcium during in vitro targeting of a sunflower oleosin. Plant Mol. somatic embryogenesis of carrot (Daucus carota L.). Biol. 29:403^10. Protoplasma 190:107-118. Tian, H.-Q., and Yang, H.-Y. 1991. Ultrastructural obser- Tirlapur, U.K., and Cresti, M. 1992. Computer-assisted vations on parthenogenesis and antipodal apogamy video image analysis of spatial variations in memof Allium tuberosum Roxb. Acta Bot. Sinica brane-associated Ca2+ and calmodulin during pollen 33:819-824. hydration, germination and tip growth in Nicotiana tabacum L. Ann. Bot. 69:503-508. Tierney, M.L., Bray, E.A., Allen, R.D., Ma, Y, Drong, R.F., Slightom, J., and Beachy, R.N. 1987. Isolation and Tirlapur, U.K., Kranz, E., and Cresti, M. 1995. characterization of a genomic clone encoding the PCharacterisation of isolated egg cells, in vitro fusion subunit of (3-conglycinin. Planta 172:356-363. products and zygotes of Zea mays L. using the technique of image analysis and confocal laser scanning Tiezzi, A., Moscatelli, A., Cai, G., Bartalesi, A., and Cresti, microscopy. Zygote 3:57-64. M. 1992. An immunoreactive homolog of mammalian kinesin in Nicotiana tabacum pollen tubes. Cell Tirlapur, U.K., and Shiggaon, S.V. 1988. Distribution of Mot. Cytoskel. 21:132-137. Ca2+ and calmodulin in the papillate cells of the stigma surface, visualized by chlorotetracycline and Tiezzi, A., Moscatelli, A., Milanesi, C, Ciampolini, C, and fluorescing calmodulin-binding phenothiazines. Cresti, M. 1987. Taxol-induced structures derived Ann. Biol. India 4:49-53. from cytoskeletal elements of the Nicotiana pollen tube. /. Cell Sci. 88:657-661. Tirlapur, U.K., van Went, J.L., and Cresti, M. 1993. Tilton, V.R. 1980. The nucellar epidermis and micropyle of Visualization of membrane calcium and calmodulin in Ornithogalum caudatum (Liliaceae) with a review of embryo sacs in situ and isolated from Petunia hybrida L. these structures in other taxa. Can. }. Bot. and Nicotiana tabacum L. Ann. Bot. 71:161-167. 58:1872-1884. Tirlapur, U.K., and Willemse, M.T.M. 1992. Changes in Tilton, V.R. 1981a. Ovule development in Ornithogalum calcium and calmodulin levels during microsporocaudatum (Liliaceae) with a review of selected papers genesis, pollen development and germination in on angiosperm reproduction. II. Megasporogenesis. Gasteria verrucosa (Mill.) H. Duval. Sex. Plant Reprod. New Phytol. 88:459-476. 5:214-223. Tilton, V.R. 1981b. Ovule development in Ornithogalum Tirlapur, U.K., Cai, G., Faleri, C, Moscatelli, A., Scali, M., caudatum (Liliaceae) with a review of selected papers Del Casino, C, Tiezzi, A., and Cresti, M. 1995. on angiosperm reproduction. IV. Egg apparatus Confocal imaging and immunogold electron structure and function. New Phytol. 88:505-531. microscopy of changes in distribution of myosin Tilton, V.R. 1984. Stigma, style, and pollen recognition in during pollen hydration, germination and pollen Galinsoga quadriradiata Ruiz & Pav. (Compositae). tube growth in Nicotiana tabacum L. Eur. J. Cell Biol. Caryologia 37:423-433. 67:209-217. Tilton, V.R., and Homer, H.T., Jr. 1980. Stigma, style, and Tirlapur, U.K., Scali, M., Moscatelli, A., Del Casino, C, obturator of Ornithogalum caudatum (Liliaceae) and Cai, G., Tiezzi, A., and Cresti, M. 1994. Confocal their function in the reproductive process. Am. J. Bot. image analysis of spatial variations in immunocyto67:1113-1131. chemically identified calmodulin during pollen hydration, germination and pollen tube tip growth Tilton, V.R., and Lersten, N.R. 1981. Ovule development in Nicotiana tabacum L. Zygote 2:63-68. in Ornithogalum caudatum (Liliaceae) with a review of selected papers on angiosperm reproduction. III. Titz, W. 1965. Vergleichende Untersuchungen iiber den Nucellus and megagametophyte. New Phytol. Grad der somatischen Polyploidie an nahe ver88:477-504. wandten diploiden und polyploiden Sippen einschliefilich der Cytologie von Antipoden. Osterr. Tilton, V.R., and Mogensen, H.L. 1979. Ultrastructural Bot. Z. 112:101-172. aspects of the ovule of Agave parryi before fertilization. Phytomorphology 29:338-350. Tiwari, S.C. 1982. Callose in the walls of mature embryo Tilton, V.R., and Russell, S.H. 1984. Applications of in vitro sac of Torenia fournieri. Protoplasma 110:1-4. pollination/fertilization technology. BioScience Tiwari, S.C. 1983. The hypostase in Torenia fournieri Lind.: 34:239-242. a histochemical study of the cell walls. Ann. Bot. 51:17-26. Tilton, V.R., Wilcox, L.W., and Palmer, R.G. 1984. Postfertilization wandlabrinthe formation and func- Tiwari, S.C. 1989. Cytoskeleton during pollen develoption in the central cell of soybean, Glycine max (L.) ment in Tradescantia virginiana: a study employing Merr. (Leguminosae). Bot. Gaz. 145:334-339. chemical fixation, freeze-substitution, immunofluorescence, and colchicine administration. Can. }. Bot. Tilton, V.R., Wilcox, L.W., Palmer, R.G., and Albertsen, 67:1244-1253. M.C. 1984. Stigma, style, and obturator of soybean, Glycine max (L.) Merr. (Leguminosae) and their func- Tiwari, S.C. 1994. An intermediate-voltage electron tion in the reproductive process. Am. ]. Bot. microscopic study of freeze-substituted generative 71:676-686. cell in pear (Pyrus communis L.): features with releTimmers, A.C.J., de Vries, S.C, and Schel, J.H.N. 1989. vance to cell-cell communication between the two Distribution of membrane-bound calcium and acticells of a germinating pollen. Sex. Plant Reprod. vated calmodulin during somatic embryogenesis of 7:177-186.
References
651
Tiwari, S.C, and Gunning, B.E.S. 1986a. Cytoskeleton, cell Toriyama, K., Stein, J.C, Nasrallah, M.E., and Nasrallah, J.B. 1991a. Transformation of Brassica oleracea with an surface and the development of invasive plasmodial tapetum in Tradescantia virginiana L. Protoplasma S-locus gene from B. campestris changes the self133:89-99. incompatibility phenotype. Theor. Appl. Genet. 81:769-776. Tiwari, S.C, and Gunning, B.E.S. 1986b. An ultrastructural, cytochemical and immunofluorescence study Toriyama, K., Thorsness, M.K., Nasrallah, J.B., and of postmeiotic development of plasmodial tapetum Nasrallah, M.E. 1991b. A Brassica S locus gene promoter directs sporophytic expression in the anther in Tradescantia virginiana L. and its relevance to the pathway of sporopollenin secretion. Protoplasma tapetum of transgenic Arabidopsis. Devel Biol. 133:100-114. 143:427-431. Tiwari, S.C., and Gunning, B.E.S. 1986c. Colchicine Torrent, M., Poca, E., Campos, E., Ludevid, M.D., and Palau, J. 1986. In maize, glutelin-2 and low molecuinhibits plasmodium formation and disrupts pathlar weight zeins are synthesized by membraneways of sporopollenin secretion in the anther tapebound polyribosomes and translocated into tum of Tradescantia virginiana L. Protoplasma microsomal membranes. Plant Mol. Biol. 7:393-403. 133:115-128. Tiwari, S.C., and Gunning, B.E.S. 1986d. Development of Torres, M.-A., Rigau, J., Puigdomenech, P., and Stiefel, V. 1995. Specific distribution of mRNAs in maize growtapetum and microspores in Canna L.: an example of an invasive but non-syncytial tapetum. Ann. Bot. 57: ing pollen tubes observed by whole-mount in situ hybridization with non-radioactive probes. Plant J. 557-563. 8:317-321. Tiwari, S.C, and Polito, V.S. 1988a. Spatial and temporal organization of actin during hydration, activation, Torres-Ruiz, R.A., and Jiirgens, G. 1994. Mutations in the and germination of pollen in Pyrus communis L.: a PASS gene uncouple pattern formation and morphopopulation study. Protoplasma 147:5-15. genesis in Arabidopsis development. Development 120:2967-2978. Tiwari, S.C, and Polito, V.S. 1988b. Organization of the cytoskeleton in pollen tubes of Pyrus communis: a Touraev, A., Fink, C.S., Stoger, E., and Heberle-Bors, E. study employing conventional and freeze-substitu1995a. Pollen selection: a transgenic reconstruction electron microscopy, immunofluorescence, and tion approach. Proc. Natl Acad. Sci. USA rhodamine-phalloidin. Protoplasma 147:100-112. 92:12165-12169. Tiwari, S.C, and Polito, V.S. 1990a. An analysis of the role Touraev, A., Lezin, R, Heberle-Bors, E., and Vicente, O. 1995b. Maintenance of gametophytic development of actin during pollen activation leading to germinaafter symmetrical division in tobacco microspore tion in pear (Pyrus communis L.): treatment with culture. Sex. Plant Reprod. 8:70-76. cytochalasin D. Sex. Plant Reprod. 3:121-129. Tiwari, S.C, and Polito, V.S. 1990b. The initiation and orga- Traas, J., Bellini, C, Nacry, P., Kronenberger, J., Bouchez, D., and Caboche, M. 1995. Normal differentiation nization of microtubules in germinating pear (Pyrus communis L.) pollen. Eitr.}. Cell Biol. 53:384-389. patterns in plants lacking microtubular preprophase Tiwari, S.C, Polito, VS., and Webster, B.D. 1990. In dry bands. Nature 375:676-677. pear (Pyrus communis L.) pollen, membranes assume Traas, J.A., Burgain, S., and de Vaulx, R.D. 1989. The orgaa tightly packed multilamellate aspect that disapnization of the cytoskeleton during meiosis in eggpears rapidly upon hydration. Protoplasma plant [Solanum melongena (L.)]: microtubules and F-actin are both necessary for coordinated meiotic 153:157-168. division. /. Cell Sci. 92:541-550. Tobias, CM., Howlett, B., and Nasrallah, J.B. 1992. An Arabidopsis thaliana gene with sequence similarity to Trick, M. 1990. Genomic sequence of a Brassica S locusthe S-locus receptor kinase of Brassica oleracea. Plant related gene. Plant Mol. Biol. 15:203-205. Physiol. 99:284-290. Trick, M., and Flavell, R.B. 1989. A homozygous S genotype of Brassica oleracea expresses two S-like genes. Tohda, H. 1974. Development of the embryo of Pogonia Mol. Gen. Genet. 218:112-117. (Orchidaceae). Sci. Rep. Tohoku Univ. Ser. IV (Biol.) 37:89-93. Trick, M., and Heizmann, P. 1992. Sporophytic selfTong, Z., Wang, T, and Xu, Y. 1990. Evidence for involveincompatibility systems: Brassica S gene family. Int. Rev. Cytol. 140:485-524. ment of phytochrome regulation in male sterility of a mutant of Oryza sativa L. Photochem. Pliotobiol. Triplett, B.A., and Quatrano, R.S. 1982. Timing, localiza52:161-164. tion, and control of wheat germ agglutinin synthesis Toonen, M.A.J., Hendriks, T, Schmidt, E.D.L., Verhoeven, in developing wheat embryos. Devel Biol. H.A., van Kammen, A., and de Vries, S.C. 1994. 91:491^96. Description of somatic-embryo-forming single cells Trobner, W., Ramirez, L., Motte, P., Hue, I., Huijser, P., in carrot suspension cultures employing video cell Lonnig, W.-E., Saedler, H., Sommer, H., and tracking. Planta 194:565-572. Schwarz-Sommer, Z. 1992. GLOBOSA: a homeotic Topfer, R., Gronenborn, B., Schell, J., and Steinbiss, H.-H. gene which interacts with DEFIC1ENS in the control of Antirrhinum floral organogenesis. EMBO ]. 1989. Uptake and transient expression of chimeric 11:4693-4704. genes in seed-derived embryos. Plant Cell 1:133-139. Topfer, R., and Steinbiss, H.-H. 1985. Plant regeneration Trognitz, B.R., and Schmiediche, P.E. 1993. A new look at from cultured fertilized barley ovules. Plant Sci. incompatibility relationships in higher plants. Sex. Plant Reprod. 6:183-190. 41:49-54. Toriyama, K., Okada, T, Watanabe, M., Ide, T, Ashida, T, Trull, M.C, Holaway, B.L., Friedman, W.E., and Xu, H., and Singh, M.B. 1995. A cDNA clone encodMalmberg, R.L. 1991. Developmentally regulated ing an IgE-binding protein from Brassica anther has antigen associated with calcium crystals in tobacco 2+ significant sequence similarity to Ca -binding proanthers. Planta 186:13-16. teins. Plant Mol. Biol. 29:1157-1165. Tsaftaris, A.S., and Scandalios, J.G. 1986. Spatial pattern of
652 catalase (Cat!) gene activation in scutella during postgerminative development in maize. Proc. Natl Acad. Sci. USA 83:5549-5553. Tsai, C.-Y., and Nelson, O.E. 1966. Starch-deficient maize mutant lacking adenosine diphosphate glucose pyrophosphorylase activity. Science 151:341-343. Tsai, D.-S., Lee, H.-S., Post, L.C., Kreiling, K.M., and Kao, T.-H. 1992. Sequence of an S-protein of Lycopersicon peruvianwn and comparison with other solanaceous S-proteins. Sex. Plant Reprod. 5:256-263. Tsao, T.-H. 1949. A study of chemotropism of pollen tubes
References
self- and non-pollinated pistils of Nicotiana alata. Biol. Plant. 3:47-64.
Tupy, J. 1966. Synthesis of protein and RNA in pollen tubes stimulated with 2-thiouracil. Biol. Plant. 8:398-410. Tupy, J. 1977. RNA synthesis and polysome formation in pollen tubes. Biol. Plant. 19:300-308. Tupy, J. 1982. Alterations in polyadenylated RNA during pollen maturation and germination. Biol. Plant. 24:331-340. Tupy, J., Hrabetova\ E., and Balatkova, V. 1977. Evidence in vitro. Plant Physiol. 24:494-504. for ribosomal RNA synthesis in pollen tubes in culture. Biol. Plant. 19:226-230. Tsay, H.S., Yeh, C.C., and Hsu, J.Y. 1990. Embryogenesis and plant regeneration from anther culture of bam- Tupy, J., Hrabetova, E., and Capkova-Balatkovd, V. 1980. Interaction of microbial DNA with germinating boo (Simocalamus latiflora (Munro) McClure). Plant pollen of Nicotiana tabacum L. in culture. Physiol. Cell Rep. 9:349-351. Veget. 18:677-687. Tschermak-Woess, E. 1956. Notizen iiber die Riesenkerne und "Riesenchromosomen" in dem Antipoden von Tupy, J., and Rangaswamy, N.S. 1973. The investigation of Aconitum. Chromosoma 8:114-134. the effect of pollination on ribosomal RNA, transfer Tschermak-Woess, E. 1957. Uber Kernstrukturen in den RNA and DNA contents in styles of Nicotiana alata. endopolyploiden Antipoden von Clivia miniata. Biol. Plant. 15:95-101. Chromosoma 8:637-649. Tupy, J., and Rihovd, L. 1984. Changes and growth effect Tschermak-Woess, E. 1959. Endopolyploidie der of pH in pollen tube culture. /. Plant Physiol. 115:1-10. Narbenpapillen bei Spironema fragrans. Osterr. Bot. Z.Tupy, J., Ri'hov^, L., and Zarsky, V. 1991. Production of fer106:74-80. tile tobacco pollen from microspores in suspension culture and its storage for in situ pollination. Sex. Tschermak-Woess, E., and Enzenberg-Kunz, U. 1965. Die Plant Reprod. 4:284-287. Struktur der hoch endopolyploiden Kerne im Endosperm von Zea mays, das auffallende Verhalten Tupy, J., Stanley, R.G., and Linskens, H.F. 1965. ihrer Nucleolen und ihr Endopolyploidiegrad. Stimulation of pollen tube growth in vitro by Planta 64:149-169. thiouracil and other antimetabolites of nucleic acid bases. Ada Bot. Neerl. 14:148-154. Tsinger, N.V., and Petrovskaya-Baranova, T.P. 1961. The pollen grain wall - a living, physiologically active Tupy, J., Suss, J., and Ri'hovd, L. 1986. RNA synthesis and structure. Doklady Bot. Sci.. 138:104-107. ribosome status in pollen tube growth of Nicotiana Tsuchimoto, S., van der Krol, A., and Chua, N.-H. 1993. tabacum L.; effects of external pH. /. Plant Physiol. 123:467-476. Ectopic expression of pMADS3 in transgenic Petunia phenocopies the Petunia blind mutant. Plant Cell Tupy, J., Suss, J., Hrabetova\ E., and Rihova L. 1983. 5:843-853. Developmental changes in gene expression during Tsuchiya, T., Toriyama, K., Ejiri, S.-L, and Hinata, K. 1994. pollen differentiation and maturation in Nicotiana Molecular characterization of rice genes specifically tabacum L. Biol. Plant. 25:231-237. expressed in the anther tapetum. Plant Mol. Biol. Turala, K. 1958. Endomitosis in the tapetal cells of 26:1737-1746. Cucurbita pepo. Acta Biol. Cracow Ser. Bot. 1:25-34. Tsuchiya, T., Toriyama, K., Nasrallah, M.E., and Ejiri, S.-I. Turala, K. 1966. Strukturen endopolyploider Kerne im 1992. Isolation of genes abundantly expressed in rice Bereich der Samenanlage einiger Monocotylen. anthers at the microspore stage. Plant Mol. Biol. Osterr. Bot. Z. 113:529-541. 20:1189-1193. Turbin, N.V., Atrashenok, N.V., Palilova, A.M., and Lyul'kina, E.I. 1968. Mitochondrial structure in Tsuchiya, T., Toriyama, K., Yoshikawa, M., Ejiri, S.-I., and ovules of hybrid corn obtained by crossing sterile Hinata, K. 1995. Tapetum-specific expression of the lines with fertility-restoring lines. Doklady Biol. Sci. gene for an endo-B-l,3-glucanase causes male steril182:536-538. ity in transgenic tobacco. Plant Cell Physiol. 36:487^194. Turcich, M.P., Hamilton, D.A., and Mascarenhas, J.P. 1993. Isolation and characterization of pollen-specific Tsukaya, H., Takahashi, T., Naito, S., and Komeda, Y. maize genes with sequence homology to ragweed 1993. Floral organ-specific and constitutive expresallergens and pectate Iyases. Plant Mol. Biol. sion of an Arabidopsis thaliana heat-shock 23:1061-1065. HSP18.2::GUS fusion gene is retained even after homeotic conversion of flowers by mutation. Mol Turcich, M.P., and Mascarenhas, J.P. 1994. PREM-1, a putative maize retroelement has LTR (long terminal Gen. Genet. 237:26-32. repeat) sequences that are preferentially transcribed Tully, R.E., and Beevers, H. 1976. Protein bodies of castor in pollen. Sex. Plant Reprod. 7:2-11. bean endosperm. Isolation, fractionation, and the characterization of protein components. Plant Turcich, M.P., Bokhari-Riza, A., Hamilton, D.A., He, C, Physiol. 58:710-716. Messier, W., Stewart, C.-B., and Mascarenhas, J.P. 1996. PREM-2, a copia-type retroelement in maize is Turner, N.E., Thanh, V.H., and Nielsen, N.C. 1981. expressed preferentially in early microspores. Sex. Purification and characterization of mRNA from Plant Reprod. 9:65-74. soybean seeds. Identification of glycinin and (3-conglycinin precursors. /. Biol. Chem. 256:8756-8760. Turcich, M.P., Hamilton, D.A., Yu, X., and Mascarenhas, Tupy, J. 1959. Callose formation in pollen tubes and J.P. 1994. Characterization of a pollen-specific gene incompatibility. Biol. Plant. 1:192-198. from Tradescantia paludosa with an unusual cysteine grouping. Sex. Plant Reprod. 7:201-202. Tupy, J. 1961. Investigation of free amino-acids in cross-,
References
653
of seed oil bodies from diverse species. Plant Physiol. Turgut, K., Barsby, T., Craze, M., Freeman, J., Hodge, R., Paul, W., and Scott, R. 1994. The highly expressed 101:267-276. tapetum-specific A9 gene is not required for Tzen, J.T.C., and Huang, A.H.C. 1992. Surface structure male fertility in Brassica napus. Plant Mol. Biol. and properties of plant seed oil bodies. /. Cell Biol. 24:97-104. 117:327-335. Turley, R.B., and Trelease, R.N. 1990. Development and Tzen, J.T.C., Lai, Y.-K., Chan, K.-L., and Huang, A.H.C. 1990. Oleosin isoforms of high and low molecular regulation of three glyoxysomal enzymes during weights are present in the oil bodies of diverse seed cotton seed maturation and growth. Plant Mol. Biol. 14:137-146. species. Plant Physiol. 94:1282-1289. Turnham, E., and Northcote, D.H. 1982. The use of acetylCoA carboxylase activity and changes in wall com- Ueda, K., Miyamoto, Y., and Tanaka, 1.1990. Fusion studposition as measures of embryogenesis in tissue ies of pollen protoplasts and generative cell protocultures of oil palm (Elaeis guineensis). Biochem. J. plasts in Lilium longiflorum. Plant Sci. 72:259-266. 208:323-332. Ueda, K., and Tanaka, I. 1994. The basic proteins of male gametic nuclei isolated from pollen grains of Lilium Tuschall, D.M., and Hannah, L.C. 1982. Altered maize longiflorum. Planta 192:446-452. endosperm ADP-glucose pyrophosphorylases from revertants of a shrunken-2-dissociation allele. GeneticsUeda, K., and Tanaka, I. 1995a. The appearance of male gamete-specific histones gH2B and gH3 during 100:105-111. pollen development in Lilium longiflorum. Devel Biol. Twell, D. 1992. Use of a nuclear-targeted P-glucuronidase 169:210-217. fusion protein to demonstrate vegetative cell-specific gene expression in developing pollen. Plant ]. Ueda, K., and Tanaka, 1.1995b. Male gametic nucleus-spe2:887-892. cific H2B and H3 histones, designated gH2B and gH3, in Lilium longiflorum. Planta 197:289-295. Twell, D. 1994. The diversity and regulation of gene expression in the pathway of male gametophyte Ueda, T., and Messing, J. 1991. A homologous expression system for cloned zein genes. Theor. Appl. Genet. development. In Molecular and Cellular Aspects of 82:93-100. Plant Reproduction, ed. R.J. Scott and A.D. Stead, pp. 83-135. Cambridge: Cambridge University Press. Ueda, T., Waverczak, W., Ward, K., Sher, N., Ketudat, M., Schmidt, R.J., and Messing, J. 1992. Mutations of the Twell, D. 1995. Diphtheria toxin-mediated cell ablation in 22- and 27-kD zein promoters affect transactivation developing pollen: vegetative cell ablation blocks by the opaque-2 protein. Plant Cell 4:701-709. generative cell migration. Protoplasma 187:144-154. Twell, D., Yamaguchi, J., and McCormick, S. 1990. Pollen- Ueng, P., Galili, G., Sapanara, V., Goldsbrough, P.B., Dube, specific gene expression in transgenic plants: coordiP., Beachy, R.N., and Larkins, B.A. 1988. Expression nate regulation of two different tomato gene of a maize storage protein gene in Petunia plants is promoters during microsporogenesis. Development not restricted to seeds. Plant Physiol. 86:1281-1285. 109:705-713. Uknes, S., Dincher, S., Friedrich, L., Negrotto, D., Twell, D., Klein, T.M., Fromm, M.E., and McCormick, S. Williams, S., Thompson-Taylor, H., Potter, S., Ward, 1989a. Transient expression of chimeric genes delivE., and Ryals, J. 1993. Regulation of pathogenesisered into pollen by microprojectile bombardment. related protein-la gene expression in tobacco. Plant Plant Physiol. 91:1270-1274. Cell 5:159-169. Twell, D., Patel, S., Sorensen, A., Roberts, M., Scott, R., Ulrich, T.U., Wurtele, E.S., and Nikolau, B.J. 1990. Draper, J., and Foster, G. 1993. Activation and develSequence of EMB-1, an mRNA accumulating specifically in embryos of carrot. Nucl. Acids Res. 18:2826. opmental regulation of an Arabidopsis anther-specific promoter in microspores and pollen of Nicotiana Umbach, A.L., Lalonde, B.A., Kandasamy, M.K., Nasrallah, J.B., and Nasrallah, M.E. 1990. tabacum. Sex. Plant Reprod. 6:217-224. Immunodetection of protein glycoforms encoded by Twell, D., Wing, R., Yamaguchi, J., and McCormick, S. two independent genes of the self-incompatibility 1989b. Isolation and expression of an anther-spemultigene family of Brassica. Plant Physiol. cific gene from tomato. Mol. Gen. Genet. 93:739-747. 217:240-245. Twell, D., Yamaguchi, J., Wing, R.A., Ushiba, J., and Umbeck, PR, and Gengenbach, B.G. 1983. Reversion of male-sterile T-cytoplasm maize to male fertility in McCormick, S. 1991. Promoter analysis of genes that tissue culture. Crop Sci. 23:584-588. are coordinately expressed during pollen development reveals pollen-specific enhancer sequences Unger, E., Parsons, R.L., Schmidt, R.J., Bowen, B., and and shared regulatory elements. Genes Devel. Roth, B.A. 1993. Dominant negative mutants of 5:496-507. opaque2 suppress transactivation of a 22-kD zein promoter by opaque2 in maize endosperm cells. Tyagi, A.K., Rashid, A., and Maheshwari, S.C. 1979. High Plant Cell 5:831-841. frequency production of embryos in Datura innoxia from isolated pollen grains by combined cold treat- Ursin, V.M., Yamaguchi, J., and McCormick, S. 1989. Gametophytic and sporophytic expression of antherment and serial culture of anthers in liquid medium. specific genes in developing tomato anthers. Plant Protoplasma 99:11-17. Cell 1:727-736. Tyagi, A.K., Rashid, A., and Maheshwari, S.C. 1981. Promotive effect of polyvinylpolypyrrolidone on Usha, S.V. 1965. In vitro pollination in Antirrhinum majus pollen embryogenesis in Datura innoxia. Physiol. L. Cum Sci. 34:511-513. Plant. 53:405-406. Usmanov, P.D., and Miiller, A. 1973. Use of the embryo Tykarska, T. 1979. Rape embryogenesis. II. Development test for the analysis of embryonic lethals induced by of embryo proper. Ada Soc. Bot. Polon. 48:391-421. the irradiation of the pollen grains of Arabidopsis Tzen, J.T.C., Cao, Y.-Z., Laurent, P., Ratnayake, C, and thaliana (L.) Heynh. Sov. Genet. 6:894-902. Huang, A.H.C. 1993. Lipids, proteins, and structure Utsumi, S., Kitagawa, S., Katsube, T., Higasa, T, Kito, M.,
654
References
Takaiwa, R, and Ishige, T. 1994. Expression and van Eldik, G.J., Wingens, M., Ruiter, R.K., van Herpen, accumulation of normal and modified soybean M.M.A., Schrauwen, J.A.M., and Wullems, G.J. 1996. Molecular analysis of a pistil-specific gene expressed glycinins in potato tubers. Plant Sci. 102:181-188. in the stigma and stylar cortex of Solanum tuberosum. Uwate, W.J., and Lin, J. 1980. Cytological zonation of Plant Mol. Biol. 30:171-176. Primus avium L. pollen tubes in vivo. J. Ultrastr. Res. 71:173-184. van Engelen, F.A., de Jong, A.J., Meijer, E.A., Kuil, C.W., Uwate, W.J., and Lin, J. 1981a. Tissue development in the Meyboom, J.K., Dirkse, W.G., Booij, H., Hartog, M.V., Vandekerckhove, J., de Vries, S.C., and van stigma of Primus avium L. Ann. Bot. 47:41-51. Kammen, A. 1995. Purification, immunological charUwate, W.J., and Lin, J. 1981b. Development of the stigacterization and cDNA cloning of a 47 kDa glycomatic surface of Primus avium L., sweet cherry. Am. ]. protein secreted by carrot suspension cells. Plant Bot. 68:1165-1176. Mol. Biol. 27:901-910. Uwate, W.J., Lin, J., Ryugo, K., and Stallman, V. 1982. Cellular components of the midstylar transmitting van Engelen, F.A., Hartog, M.V., Thomas, T.L., Taylor, B., Sturn, A., van Kammen, A., and de Vries, S.C. 1993. tissue of Primus avium. Can. J. Bot. 60:98-104. The carrot secreted glycoprotein gene EP1 is expressed in the epidermis and has sequence homology to Valenta, R., Duchene, M., Pettenburger, Kv Sillaber, C, Brassica S-locus glycoproteins. Plant J. 4:855-862. Valent, P., Bettelheim, P., Breitenbach, M., Rumpold, H., Kraft, D., and Scheiner, O. 1991. Identification of van Engelen, F.A., Sterk, P., Booij, H., Cordewener, J.H.G., profilin as a novel pollen allergen; IgE autoreactivity Rook, W., van Kammen, A., and de Vries, S.C. 1991. in sensitized individuals. Science 253:557-560. Heterogeneity and cell type-specific localization of a cell wall glycoprotein from carrot suspension cells. Vallade, J. 1970. DeVeloppement embryonnaire chez un Plant Physiol. 96:705-712. Petunia hybrida hort. Compt. Rend. Acad. Sci. Paris 270D:1893-1896. van Geyt, J., D'Halluin, K., and Jacobs, M. 1985. Induction of nuclear and cell divisions in microspores of sugVallade, J. 1972. Structure et fonctionnement du arbeet {Beta vulgaris L.). Z. Pflanzenziichtg 95:325-335. me'risteme lors de la formation de la jeune racine primaire chez un Petunia hybrida hort. Compt. Rend. van Heel, W.A. 1981. A S.E.M.-investigation on the develAcad. Sci. Paris 274D:1027-1030. opment of free carpels. Blumea 27:499-522. Vallade, J. 1980. Donn£es cytologiques sur la proembryo- van Herpen, M.M.A., de Groot, P.F.M., Schrauwen, genese du Petunia; interet pour une interpretation J.A.M., van den Heuvel, K.J.P.T, Weterings, K.A.P., morphoge'ne'tique du de'veloppement embryonand Wullems, G.J. 1992. In-vitro culture of tobacco naire. Bull. Soc. Bot. Fr. Actual. Bot. 127:19-37. pollen: gene expression and protein synthesis. Sex. Plant Reprod. 5:304-309. Vallade, J., Cornu, A., Essad, S., and Alabouvette, J. 1978. Niveaux de DNA dans les noyaux zygotiques chez van Herpen, M.M.A., Reijnen, W.H., Schrauwen, J.A.M., le Petunia hybrida hort. Bull. Soc. Bot. Fr. Actual. Bot. de Groot, P.F.M., Jager, J.W.H., and Wullems, G.J. 1989. Heat-shock proteins and survival of germinat125:253-258. ing pollen of Lilium longiflorum and Nicotiana van Aelst, A.C., Theunis, C.H., and van Went, J.L. 1990. tabacum. J. Plant Physiol. 134:345-351. Freeze-fracture studies on isolated sperm cells of Spinacia oleracea L. Protoplasma 153:204-207. van Hoist, G.-J., and Clarke, A.E. 1986. Organ-specific arabinogalactan-proteins of Lycopersicon peruvianum van Aelst, A.C., and van Went, J.L. 1991. The ultrastruc(Mill) demonstrated by crossed electrophoresis. ture of mature Papaver dubium L. pollen grains. Ada Bot. Neerl. 40:319-328. Plant Physiol. 80:786-789. van Aelst, A.C., and van Went, J.L. 1992. Ultrastructural van Lammeren, A.A.M. 1986. A comparative ultrastrucimmuno-localization of pectins and glycoproteins in tural study of the megagametophytes in two strains Arabidopsis thaliana pollen grains. Protoplasma of Zea mays L. before and after fertilization. Agr. 168:14-19. Univ. Wageningen Papers 86-1:1-37. van Aelst, A.C., Mueller, T, Dueggelin, M., and van Lammeren, A.A.M. 1988a. Structure and function of Guggenheim, R. 1989. Three-dimensional observathe microtubular cytoskeleton during endosperm tions on freeze-fractured frozen hydrated Papaver development in wheat: an immunofluorescence dubium pollen with cryo-scanning electron study. Protoplasma 146:18-27. microscopy. Ada Bot. Neerl. 38:25-30. van Lammeren, A.A.M. 1988b. Observations on the strucvan Aelst, A.C., Pierson, E.S., van Went, J.L., and Cresti, tural development of immature maize embryos {Zea mays L.) during in vitro culture in the presence or M. 1993. Ultrastructural changes of Arabidopsis thaliana pollen during final maturation and rehydraabsence of 2,4-D. Ada Bot. Neerl. 37:49-61. tion. Zygote 1:173-179. van Lammeren, A.A.M., Bednara, J., and Willemse, M.T.M. 1989. Organization of the actin cytoskeleton van Boxtel, J., Berthouly, M., Carasco, C, Dufour, M., and during pollen development in Gasteria verrucosa Eskes, A. 1995. Transient expression of (3-glu(Mill.) H. Duval visualized with rhodamine-phalcuronidase following biolistic delivery of foreign loidin. Planta 178:531-539. DNA into coffee tissues. Plant Cell Rep. 14:748-752. van Driessche, E., Smets, G., Dejaegere, R., and Kanarek, van Lammeren, A.A.M., Keijzer, C.J., Willemse, M.T.M., L. 1981. The immuno-histochemical localization of and Kieft, H. 1985. Structure and function of the lectin in pea seeds (Pisum sativum L.). Planta microtubular cytoskeleton during pollen develop153:287-296. ment in Gasteria verrucosa (Mill.) H. Duval. Planta 165:1-11. van Eldik, G.J., Vriezen, W.H., Wingens, M., Ruiter, R.K., van Herpen, M.M.A., Schrauwen, J.A.M., and van Marrewijk, G.A.M., Bino, R.J., and Suurs, L.C.J.M. Willlems, G.J. 1995. A pistil-specific gene of Solanum 1986. Characterization of cytoplasmic male sterility tuberosum is predominantly expressed in the stylar in Petunia hybrida. I. Localization, composition and cortex. Sex. Plant Reprod. 8:173-179. activity of esterases. Euphytica 35:77-88.
References
655
van Overbeek, ]., Conklin, M.E., and Blakeslee, A.F. 1942. van Went, J.L., and Linskens, H.F. 1967. Die Entwicklung des sogenannten "Fadenapparates" im Embryosack Cultivation in vitro of small Datura embryos. Am. ]. von Petunia hybrida. Ziichter 37:51-56. Bot. 29:472-477. van Overbeek, ]., Siu, Rv and Haagen-Smit, A.J. 1944. van Went, J.L., and Willemse, M.T.M. 1984. Fertilization. In Embryology of Angiosperms, ed. B. M. Johri, pp. Factors affecting the growth of Datura embryos in 273-317. Berlin: Springer-Verlag. vitro. Am. J. Bot. 31:219-224. van Rensburg, J.G.J., and Robbertse, P.J. 1988. Seed devel- van der Donk, J.A.W.M. 1974a. Differential synthesis of RNA in self- and cross-pollinated styles of Petunia opment of Ornithogalum dubium, with special referhybrida L. Mol. Gen. Genet. 131:1-8. ence to fertilization and the egg apparatus. S. Afr. ]. Bot. 54:196-202. van der Donk, J.A.W.M. 1974b. Synthesis of RNA and protein as a function of time and type of pollentubevan Rooijen, G.J.H., and Moloney, MM. 1994. Plant seed style interaction in Petunia hybrida L. Mol. Gen. Genet. oil bodies as carriers for foreign proteins. 134:93-98. Biotechnology 13:72-77. van Rooijen, G.J.H., Terning, L.I., and Moloney, M.M. van der Donk, J.A.W.M. 1975. Recognition and gene expression during the incompatibility reaction in 1992. Nucleotide sequence of an Arabidovsis thaliana oleosin gene. Plant Mol. Biol. 18:1177-1179. Petunia hybrida L. Mol. Gen. Genet. 141:305-316. van Rooijen, G.J.H., Wilen, R.W., Holbrook, L.A., and van der Geest, A.H.M., Frisch, D.A., Kemp, J.D., and Hall, T.C. 1995. Cell ablation reveals that expression from Moloney, M.M. 1992. Regulation of accumulation of the phaseolin promoter is confined to embryogenemRNAs encoding a 20-kDa oil-body protein in sis and microsporogenesis. Plant Physiol. microspore-derived embryos of Brassica napus. Can. 109:1151-1158. ]. Bot. 70:503-508. van Tunen, A.J., Hartman, S.A., Mur, L.A., and Mol, van der Geest, A.H.M., Hall, G.E., Jr., Spiker, S., and Hall, J.N.M. 1989. Regulation of chalcone flavanone isoT.C. 1994. The p-phaseolin gene is flanked by matrix merase (CHI) gene expression in Petunia hybrida: the attachment regions. Plant J. 6:413-423. use of alternative promoters in corolla, anthers and van der Klei, H., van Damme, J., Casteels, P., and pollen. Plant Mol. Biol. 12:539-551. Krebbers, E. 1993. A fifth 2S albumin isoform is present in Arabidopsis thaliana. Plant Physiol. van Tunen, A.J., Mur, L.A., Brouns, G.S., Rienstra, J.-D., 101:1415-1416. Koes, R.E., and Mol, J.N.M. 1990. Pollen- and antherspecific chi promoters from Petunia: tandem promoter van der Kley, F.K. 1954. Male sterility and its importance in breeding heterosis varieties. Euphytica 3:117-124. regulation of the MA gene. Plant Cell 2:393^01. van Tunen, A.J., Mur, L.A., Recourt, K., Gerats, A.G.M., van der Krol, A.R., Brunelle, A., Tsuchimoto, S., and Chua, N.-H. 1993. Functional analysis of Petunia floand Mol, J.N.M. 1991. Regulation and manipulation ral homeotic MADS box gene pMADSl. Genes Devel. of flavonoid gene expression in anthers of Petunia: 7:1214-1228. the molecular basis of the Po mutation. Plant Cell van der Leede-Plegt, L.M., van den Ven, B.C.E., Bino, 3:39-48. R.J., van der Salm, T.P.M., and van Tunen, A.J. van Tuyl, J.M., Marcucci, M.C., and Visser, T. 1982. Pollen 1992. Introduction and differential use of various and pollination experiments. VII. The effect of pollen promoters in pollen grains of Nicotiana glutinosa treatment and application method on incompatibiland Lilium longiflorum. Plant Cell Rep. 11:20-24. ity and incongruity in Lilium. Euphytica 31:613-619. van Went, J., and Cresti, M. 1988a. Pre-fertilization degen- van der Maas, H.M., de Jong, E.R., van Aelst, A.C., eration of both synergids in Brassica campestris Verhoeven, H.A., Krens, F.A., and van Went, J.L. ovules. Sex. Plant Reprod. 1:208-216. 1994. Cytological characterization of isolated sperm cells of perennial ryegrass (Lolium perenne L.). van Went, J., and Cresti, M. 1988b. Cytokinesis in Protoplasma 178:48-56. microspore mother cells of Impatiens sultani. Sex. van der Maas, H.M., Zaal, M.A.C.M., de Jong, E.R., Krens, Plant Reprod. 1:228-233. F.A., and van Went, J.L. 1993a. Isolation of viable egg van Went, J., and Cresti, M. 1989. Cytoplasmic differenticells of perennial ryegrass (Lolium perenne L.). ation during tetrad formation and early microspore Protoplasma 173:86-89. development in Impatiens sultani. Protoplasma van der Maas, H.M., Zaal, M.A.C.M., de Jong, E.R., van 148:1-7. Went, J.L., and Krens, F.A. 1993b. Optimization of van Went, J.L. 1970a. The ultrastructure of the synergids isolation and storage of sperm cells from pollen of of Petunia. Acta Bot. Neerl. 19:121-132. perennial ryegrass {Lolium perenne L.). Sex. Plant van Went, J.L. 1970b. The ultrastructure of the egg and Reprod. 6:64-70. central cell of Petunia. Acta Bot. Neerl. 19:313-322. van Went, J.L. 1970c. The ultrastructure of the fertilized van der Meer, I.M., Stam, M.E., van Tunen, A.J., Mol, J.N.M., and Stuitje, A.R. 1992. Antisense inhibition of embryo sac of Petunia. Acta Bot. Neerl. 19:468-480. flavonoid biosynthesis in Petunia anthers results in van Went, J.L. 1981. Some cytological and ultrastructural male sterility. Plant Cell 4:253-262. aspects of male sterility in Impatiens. Acta Soc. Bot. van der Meer, Q.P., and van Bennekom, J.L. 1969. Effect of Polon. 50:248-252. van Went, J.L. 1984. Unequal distribution of plastids durtemperature on the occurrence of male sterility in ing generative cell formation in Impatiens. Theor. onion (Allium cepa L.). Euphytica 18:389-394. van der Meer, Q.P., and van Bennekom, J.L. 1973. Appl. Genet. 68:305-309. Gibberellic acid as a gametocide for the common van Went, J.L., and Gori, P. 1989. The ultrastructure of onion (Allium cepa L.). Euphytica 22: 239-243. Capparis spinosa pollen grains. /. Submicros. Cytol. Path. 21:149-156. van der Meer, Q.P., and van Dam, R. 1979. Gibberellic acid van Went, J.L., and Kwee, H.-S. 1990. Enzymatic isolation as a gametocide for cole crops. Euphytica 28:717-722. of living embryo sacs of Petunia. Sex. Plant Reprod. van der Plas, L.H.W., de Gucht, L.P.E., Bakels, R.H.A., and 3:257-262. Otto, B. 1987. Growth and respiratory characteristics
656
References
of batch and continuous cell suspension cultures Vasil'ev, A.E. 1970. Infrastructure of the stigmatoid cells derived from fertile and male sterile Petunia hybrida. in Lilium. Soviet Plant Physiol. 17:1035-1044. ]. Plant Physiol. 130:449^60. Vaughn, K.C., DeBonte, L.R., Wilson, K.G., and Schaffer, G.W. 1980. Organelle alteration as a mechanism for van der Pluijm, J., and Linskens, H. F. 1966. Feinstruktur maternal inheritance. Science 208:196-198. der Pollenschlauche im Griffel von Petunia. Ziichter 36:220-224. Vazart, B. 1970. Morphogenese du sporoderme et participation des mitochondries a la mise en place de la van der Pluijm, J.E. 1964. An electron microscopic investiprimexine dans le pollen de Linum usitatissimum L. gation of the filiform apparatus in the embryo sac of Torenia fournieri. In Pollen Physiology and Fertilization, Compt. Rend. Acad. Sci. Paris 270D:3210-3212. ed. H.F. Linskens, pp. 8-16. Amsterdam: North- Vazart, B. 1971. Infrastructure de microspores de Nicotiana Holland Publishing Co. tabacum L. susceptibles de se deVelopper en embryoi'des apres excision et mise en culture des antheres. Vancanneyt, G., Sonnewald, U., Hofgen, R., and Willmitzer, L. 1989. Expression of a patatin-like proCompt. Rend. Acad. Sci. Paris 272D:549-552. tein in the anthers of potato and sweet pepper flow- Vazart, B., and Vazart, J. 1965. Infrastructure de 1'ovule de ers. Plant Cell 1:533-540. lin, Linum usitatissimum L. Les cellules du sac embryVance, V.B., and Huang, A.H.C. 1987. The major protein onnaire. Compt. Rend. Acad. Sci. Paris 261:3447-3450. from lipid bodies of maize. Characterization and Vazart, B., and Vazart, J. 1966. Infrastructure du sac structure based on cDNA cloning. /. Biol. Chem. embryonnaire du lin (Linum usitatissimum L.). Rev. 262:11275-11279. Cytol. Biol. Veget. 29:251-266. Vance, V.B., and Huang, A.H.C. 1988. Expression of lipid Vazart, J. 1969. Organisation et ultrastructure du sac body protein gene during maize seed development. embryonnaire du lin (Linum usitatissimum L.). Rev. Spatial, temporal, and hormonal regulation. /. Biol. Cytol. Biol. Veget. 32:227-240. Chem. 263:1476-1481. Vedel, F., and Mathieu, C. 1983. Physical and gene mapping of chloroplast DNA from normal and cytoplasVanDerWoude, W.J., and Morre, D.J. 1968. Endoplasmic mic male sterile (radish cytoplasm) lines of Brassica reticulum-dictyosome-secretory vesicle associations in pollen tubes of Liliwn longiflorum Thunb. Proc. napus. Curr. Genet. 7:13-20. Indiana Acad. Sci. 77:164-170. Vedel, F., Mathieu, C, Lebacq, P., Ambard-Bretteville, F., VanDerWoude, W.J., Morre, D.J., and Bracker, C.E. 1971. Remy, R., Pelletier, G., Renard, M., and Rousselle, P. 1982. Comparative macromolecular analysis of the Isolation and characterization of secretory vesicles in cytoplasms of normal and cytoplasmic male sterile germinated pollen of Lilium longiflorum. J. Cell Sci. 8:331-351. Brassica napus. Theor. Appl. Genet. 62:255-262. Varagona, M.J., Schmidt, R.J., and Raikhel, N.V. 1991. Veen, H. 1963. The effect of various growth-regulators on Monocot regulatory protein opaque-2 is localized in embryos of Capsella bursa-pastoris growing in vitro. the nucleus of maize endosperm and transformed Acta Bot. Neerl. 12:129-171. tobacco plants. Plant Cell 3:105-113. Veit, B., Schmidt, R.J., Hake, S., and Yanofsky, M.F. 1993. Maize floral development: new genes and old Vasil, I.K. 1957. Effect of kinetin and gibberellic acid on mutants. Plant Cell 5:1205-1215. excised anthers of Allium cepa. Phytomorphology 7:138-149. Vellanoweth, R.L., and Okita, T.W. 1993. Analysis of Vasil, I.K. 1959. Cultivation of excised anthers in vitro nuclear proteins interacting with a wheat a/peffect of nucleic acids. /. Expt. Bot. 10:399-408. gliadin seed storage protein gene. Plant Mol. Biol. 22:25-41. Vasil, I.K. 1967. Physiology and cytology of anther development. Biol. Rev. 42:327-373. Venema, G., and Koopmans, A. 1962. A phase-contrast microscopic study of pollengrain germination, Vasil, I.K., and Johri, M.M. 1964. The style, stigma and nuclear movement and pollentube mitosis in pollen tube -1. Phytomorphology 14:352-369. Vasil, V, and Hildebrandt, A.C. 1965. Differentiation of Tradescantia virginiana. Cytologia 27:11-24. tobacco plants from single, isolated cells in micro- Venkateswarlu, J., and Rao, G.R. 1958. A contribution to cultures. Science 150:889-892. the life-history of Rubia cordifolia Linn. /. Indian Bot. Vasil, V., and Vasil, I.K. 1981. Somatic embryogenesis and Soc. 37:442-454. plant regeneration from suspension cultures of pearl Vennigerholz, F. 1992. The transmitting tissue in millet (Pennisetum amerkanum). Ann. Bot. 47:669-678. Brugmansia suaveolens: immunocytochemical localization of pectin in the style. Protoplasma 171:117-122. Vasil, V, and Vasil, I.K. 1982a. The ontogeny of somatic embryos of Pennisetum amerkanum (L.) K. Schum. I. Vergara, M.R., Biasini, G., Lo Schiavo, F., and Terzi, M. 1982. Isolation and characterization of carrot cell In cultured immature embryos. Bot. Gaz. mutants resistant to a-amanitin. Z. Pflanzenphysiol. 143:454-465. 107:313-319. Vasil, V., and Vasil, I.K. 1982b. Characterization of an embryogenic cell suspension culture derived from Vergara, R., Verde, F., Pitto, L., Lo Schiavo, F., and Terzi, cultured inflorescences of Pennisetum amerkanum M. 1990. Reversible variations in the methylation (pearl millet, Gramineae). Am. J. Bot. 69:1441-1449. pattern of carrot DNA during somatic embryogenesis. Plant Cell Rep. 8:697-700. Vasil, V, Castillo, A.M., Fromm, M.E., and Vasil, I.K. 1992. Herbicide resistant fertile transgenic wheat Vergne, P., and Dumas, C. 1988. Isolation of viable wheat plants obtained by microprojectile bombardment of male gametophytes of different stages of developregenerable embryogenic callus. Biotechnology ment and variations in their protein patterns. Plant 10:667-674. Physiol. 88:969-972. Vasil, V., Srivastava, V., Castillo, A.M., Fromm, M.E., and Vergne, P., Riccardi, F., Beckert, M., and Dumas, C. 1993. Vasil, I.K. 1993. Rapid production of transgenic Identification of a 32-kDa anther marker protein for wheat plants by direct bombardment of cultured androgenic response in maize, Zea mays L. Theor. immature embryos. Biotechnology 11:1553-1558. Appl. Genet. 86:843-850.
References
657
Verma, D.C., and Dougall, D.K. 1978. DNA, RNA and Populus alba and P. nigra. ]. Cell Sci. 87:483-490. protein content of tissue during growth and embryo- Viotti, A., Sala, E., Alberi, P., and Soave, C. 1978. genesis in wild-carrot suspension cultures. In Vitro Heterogeneity of zein synthesized in vitro. Plant Sci. 14:183-191. Lett. 13:365-375. Verma, S.C., Malik, R., and Dhir, I. 1977. Genetics of the Vishnyakova, M.A., and Willemse, M.T.M. 1994. Pollen-pistil interaction in wheat. Ada Bot. Neerl. incompatibility system in the crucifer Ernca sativa L. 43:51-64. Proc. Roy. Soc. Land. 196B:131-159. Vernon, D.M., and Meinke, D.W. 1994. Embryogenic Vithanage, H.I.M.V., Gleeson, P.A., and Clarke, A.E. 1980. The nature of callose produced during self-pollinatransformation of the suspensor in twin, a polyemtion in Secale cereale. Planta 148:498-509. bryonic mutant of Arabidopsis. Devel Biol. Vithanage, H.I.M.V., and Heslop-Harrison, J. 1979. The 165:566-573. pollen-stigma interaction: fate of fluorescentVernon, D.M., and Meinke, D.W. 1995. Late embryo-defeclabelled pollen-wall proteins on the stigma surface tive mutants of Arabidopsis. Devel Genet. 16:311-320. in rye (Secale cereale). Ann. Bot. 43:113-114. Verwoert, I.I.G.S., van der Linden, K.H., Walsh, M.C., Nijkamp, H.J.J., and Stuitje, A.R. 1995. Modification Vithanage, H.I.M.V., and Knox, R.B. 1976. Pollen-wall proteins: quantitative cytochemistry of the origins of of Brassica napus seed oil by expression of the intine and exine enzymes in Brassica oleracea. ]. Cell Escherichia coli fabH gene, encoding 3-ketoacyl-acyl Sci. 21:423-435. carrier protein synthase III. Plant Mol. Biol. Vithanage, H.I.M.V., and Knox, R.B. 1977. Development 27:875-886. and cytochemistry of stigma surface and response to Vicente, O., Benito Moreno, R.M., and Heberle-Bors, E. self and foreign pollination in Helianthus animus. 1991. Pollen cultures as a tool to study plant develPhytomorphology 27:168-179. opment. Cell Biol. Rev. 25:295-306. Vielle, J.P., Burson, B.L., Bashaw, E.C., and Hussey, M.A. Vithanage, H.I.M.V., and Knox, R.B. 1979. Pollen development and quantitative cytochemistry of exine and 1995. Early fertilization events in the sexual and intine enzymes in sunflower, Helianthus annuus L. aposporous egg apparatus of Pennisetum ciliare (L.) Ann. Bot. 44:95-106. Link. Plant ]. 8:309-316. Vijayaraghavan, M.R., and Bhat, U. 1983. Synergids Vithanage, H.I.M.V., and Knox, R.B. 1980. Periodicity of pollen development and quantitative cytochemistry before and after fertilization. Phytomorphology of exine and intine enzymes in the grasses Lolium 33:74-84. perenne L. and Phalaris tuberosa L. Ann. Bot. Vijayaraghavan, M.R., Jensen, W.A., and Ashton, M.E. 45:131-141. 1972. Synergids of Aquilegia formosa - their histochemistry and ultrastructure. Phytomorphology Vithanage, H.I.M.V, Howlett, B.J., Jobson, S., and Knox, R.B. 1982. Immunocytochemical localization of 22:144-159. water-soluble glycoproteins, including group 1 allerVijayaraghavan, M.R., and Prabhakar, K. 1984. The gen, in pollen of ryegrass, Lolium perenne, using ferendosperm. In Embryology of Angiosperms, ed. B.M. ritin-labelled antibody. Histochem. J. 14:949-966. Johri, pp. 319-376. Berlin: Springer-Verlag. Vijayaraghavan, M.R., and Shukla, A.K. 1977. Absence of Vivekananda, J., Drew, M.C., and Thomas, T.L. 1992. callose around the microspore tetrad and poorly Hormonal and environmental regulation of the cardeveloped exine in Pergularia daemia. Ann. Bot. rot lea-dass gene Dc3. Plant Physiol. 100:576-581. 41:923-926. Vodkin, L.O. 1981. Isolation and characterization of mesVilardell, J., Goday, A., Freire, M.A., Torrent, M., senger RNAs for seed lectin and Kunitz trypsin Martinez, M.C., Tome, J.M., and Pages, M. 1990. inhibitor in soybeans. Plant Physiol. 68:766-771. Gene sequence, developmental expression, and pro- Vodkin, L.O., and Raikhel, N.V. 1986. Soybean lectin and tein phosphorylation of RAB-17 in maize. Plant Mol. related proteins in seeds and roots of Le+ and Le" Biol. 14:423-432. soybean varieties. Plant Physiol. 81:558-565. Vilardell, J., Marti'nez-Zapater, J.M., Goday, A., Arenas, C., Vodkin, L.O., Rhodes, PR., and Goldberg, R.B. 1983. cA lectin gene insertion has the structural features of a and Pages, M. 1994. Regulation of the rafcl7 gene transposable element. Cell 34:1023-1031. promoter in transgenic Arabidopsis wild-type, ABAdeficient and ABA-insensitive mutants. Plant Mol. Voelker, T., Sturm, A., and Chrispeels, M.J. 1987. Differences in expression between two seed lectin Biol. 24:561-569. alleles obtained from normal and lectin-deficient Vilardell, J., Mundy, J., Stilling, B., Leroux, B., Pla, M., beans are maintained in transgenic tobacco. EMBO ]. Freyssinet, G., and Pages, M. 1991. Regulation of the maize rabl7 gene promoter in transgenic heterolo6:3571-3577. gous systems. Plant Mol. Biol. 17:985-993. Voelker, T.A., Herman, E.M., and Chrispeels, M.J. 1989. In vitro mutated phytohemagglutinin genes expressed Villanueva, V.R., Mathivet, V, and Sangwan, R.S. 1985. in tobacco seeds: role of glycans in protein targeting RNA, proteins and polyamines during gametoand stability. Plant Cell 1:95-104. phytic and androgenetic development in Nicotiana tabacum and Datura innoxia. Plant Growth Reg. Voelker, T.A., Staswick, P., and Chrispeels, M.J. 1986. 3:293-307. Molecular analysis of two phytohemagglutinin Villar, M., Gaget, M., Rougier, M., and Dumas, C. 1993. genes and their expression in Phaseolus vidgaris cv. Pollen-pistil interactions in Popnlus: p-galactosidase Pinto, a lectin-deficient cultivar of the bean. EMBO /. activity associated with pollen-tube growth in the 5:3075-3082. crosses P. nigra X P. nigra and P. nigra X P. alba. Sex. Vogt, T., Pollak, P., Tarlyn, N., and Taylor, L.P. 1994. Pollination- or wound-induced kaempferol accumuPlant Reprod. 6:249-256. lation in Petunia stigmas enhances seed production. Villar, M., Gaget, M., Said, C., Knox, R.B., and Dumas, C. Plant Cell 6:11-23. 1987. Incompatibility in Populus: structural and cytochemical characteristics of the receptive stigmas of von Allmen, J.-M., Rottmann, W.H., Gengenbach, B.G.,
658
References
Harvey, A.J., and Lonsdale, D.M. 1991. Transfer of Walker, J.C., and Zhang, R. 1990. Relationship of a putamethomyl and HmT-toxin sensitivity from Ttive receptor protein kinase from maize to the Scytoplasm maize to tobacco. Mol. Gen. Genet. locus glycoproteins of Brassica. Nature 345:743-746. 229:405-412. Walker, K.A., and Sato, S.J. 1981. Morphogenesis in callus tissue of Medicago sativa: the role of ammonium ion von Arnold, S., and Hakman, I. 1988. Regulation of somatic embryo development in Picea abies by in somatic embryogenesis. Plant Cell Tissue Organ abscisic acid (ABA). /. Plant Physiol. 132:164-169. Cult. 1:109-121. Vonder Haar, R.A., Allen, R.D., Cohen, E.A., Nessler, C.L., Wallace, J.C., Lopes, M.A., Paiva, E., and Larkins, B.A. and Thomas, T.L. 1988. Organization of the sun1990. New methods for extraction and quantitation flower US storage protein gene family. Gene of zeins reveal a high content of y-zein in modified 74:433-443. opaque-2 maize. Plant Physiol. 92:191-196. Voss, R., Schumann, K., and Nagl, W. 1992. Wallace, N.H., and Kriz, A.L. 1991. Nucleotide sequence of a cDNA clone corresponding to the maize Phytohemagglutinin gene expression during seed Globulin-2 gene. Plant Physiol. 95:973-975. development of the runner bean, Phaseolus coccineus. Plant Mol. Biol. 20:781-790. Walling, L., Drews, G.N., and Goldberg, R.B. 1986. Transcriptional and post-translational regulation of Vroemen, C, Langeveld, S., Mayer, U., Ripper, G., soybean seed protein mRNA levels. Proc. Natl Acad. Jiirgens, G., van Kammen, A., and de Vries, S.C. Sci. USA 83:2123-2127. 1996. Pattern formation in the Arabidopsis embryo revealed by position-specific lipid transfer protein Walsh, D.J., Matthews, J.A., Denmeade, R., and Walker, gene expression. Plant Cell 8:783-791. M.R. 1989. Cloning of cDNA coding for an allergen of cocksfoot grass (Dactylis glomerata) pollen. Int. Vujicic, R., Radojevic, L., and Neskovic, M. 1976. Orderly Arch. Allergy Appl. Immunol. 90:78-83. arrangement of ribosomes in the embryogenic callus tissue of Coryhis avellana L. /. Cell Biol. 69:686-692. Walter, C, Smith, D.R., Connett, M.B., Grace, L., and White, D.W.R. 1994. A biolistic approach for the transfer and expression of a gusA reporter gene in Wadsworth, G.J., and Scandalios, J.G. 1989. Differential embryogenic cultures of Pinus radiata. Plant Cell Rep. expression of the maize catalase genes during kernel 14:69-74. development: the role of steady-state mRNA levels. Devel Genet. 10:304-310. Walters, D.A., Vetsch, C.S., Potts, D.E., and Lundquist, R.C. 1992. Transformation and inheritance of a Wagner, G., and Hess, D. 1973. In yifro-Befruchtungen bei hygromycin phosphotransferase gene in maize Petunia hybrida. Z. Pflanzenphysiol. 69:262-269. plants. Plant Mol. Biol. 18:189-200. Wagner, V.T., Dumas, C, and Mogensen, H.L. 1989. Morphometric analysis of isolated Zea mays sperm. /. Walters, M.S. 1970. Evidence on the time of chromosome pairing from the preleptotene spiral stage in Lilium Cell Sci. 93:179-184. longiflorum "Croft". Chromosoma 29:375-418. Wagner, V.T., Kardolus, J.P., and van Went, J.L. 1989. Isolation of the lily embryo sac. Sex. Plant Reprod. Walters, M.S. 1985. Meiosis readiness in Lilium. Can. }. 2:219-224. Genet. Cytol. 27:33-38. Wagner, V.T., and Mogensen, H.L. 1988. The male germ Walthall, E.D., and Brady, T. 1986. The effect of the susunit in the pollen and pollen tubes of Petunia hybrida: pensor and gibberellic acid on Phaseolus vidgaris ultrastructural, quantitative and three-dimensional embryo protein synthesis. Cell Diffn 18:37-44. features. Protoplasma 143:101-110. Wan, Y., and Lemaux, P.G. 1994. Generation of large numbers of independently transformed fertile barley Wagner, V.T., Cresti, M., Salvatici, P., and Tiezzi, A. 1990. plants. Plant Physiol. 104:37-48. Changes in volume, surface area, and frequency of nuclear pores on the vegetative nucleus of tobacco Wan, Y., Sorensen, EX., and Liang, G.H. 1988. Genetic pollen in fresh, hydrated, and activated conditions. control of in vitro regeneration in alfalfa (Medicago Planta 181:304-309. sativa L.). Euphytica 39:3-9. Wagner, V.T., Song, Y.C., Matthys-Rochon, E., and Dumas, Wandelt, C, and Feix, G. 1989. Sequence of a 21 kd gene from maize containing an in-frame stop codon. Nucl. C. 1989. Observations on the isolated embryo sac of Acids Res. 17:2354. Zea mays L. Plant Sci. 59:127-132. Walbot, V. 1971. RNA metabolism during embryo devel- Wang, C, Rathore, K.S., and Robinson, K.R. 1989. The responses of pollen to applied electrical fields. Devel opment and germination of Phaseolus vulgaris. Devel Biol. 136:405-410. Biol. 26:369-379. Walbot, V. 1972. Rate of RNA synthesis and tRNA end- Wang, C.-C, Chu, C.-C, Sun, C.-S., Wu, S.-H., Yin, K.-C, and Hsu, C. 1973. The androgenesis in wheat labeling during early development of Phaseolus. (Triticum aestivum) anthers cultured in vitro. Sci. Planta 108:161-171. Sinica 16:218-222. Walbot, V. 1973. RNA metabolism in developing cotyledons of Phaseolus vulgaris. New Phytol. 72:479^183. Wang, C.-S., Walling, L.L., Eckard, K.J., and Lord, E.M. 1992a. Patterns of protein accumulation in developWalbot, V., and Dure, L., III. 1976. Developmental bioing anthers of Lilium longiflorum correlate with histochemistry of cotton seed embryogenesis and germilogical events. Am. J. Bot. 79:118-127. nation. VII. Characterization of the cotton genome. /. Mol. Biol. 101:503-536. Wang, C.-S., Walling, L.L., Eckard, K.J., and Lord, E.M. 1992b. Immunological characterization of a tapetal Walbot, V, Brady, T, Clutter, M., and Sussex, I. 1972. protein in developing anthers of Lilium longiflorum. Macromolecular synthesis during plant embryoPlant Physiol. 99:822-829. geny: rates of RNA synthesis in Phaseolus coccineus embryos and suspensors. Devel Biol. 29:104-111. Wang, C.-S., Walling, L.L., Eckard, K.J., and Lord, E.M. 1993. Characterization of an anther-specific glycoproWalburg, G., and Larkins, B.A. 1986. Isolation and charactein in Lilium longiflorum. Am. ]. Bot. 80:1155-1161. terization of cDNAs encoding oat 12S globulin mRNAs. Plant Mol. Biol. 6:161-169. Wang, F.-Y, Chen, K.-M., Wang, X.-Z., Zhou, W.-L., and
References
659
responsive promoter by the inverse polymerase Ni, J.-F. 1995. Two-dimensional gel electrophoresis chain reaction and its characterization in transgenic study on endosperm protein of the new line tobacco. Plant J. 3:191-201. obtained by introducing DNA of sorghum into wheat. Chinese ]. Bot. 7:2-8. Watanabe, M., Shiozawa, H., Isogai, A., Suzuki, A., Takeuchi, T., and Hinata, K. 1991. Existence of S-glyWang, H., and Cutler, A.J. 1995. Promoters from kinl and cor6.6, two Arabidopsis thaliana low-temperature- and coprotein-like proteins in anthers of self-incompatiABA-inducible genes, direct strong p-glucuronidase ble species of Brassica. Plant Cell Physiol. expression in guard cells, pollen and young devel32:1039-1047. oping seeds. Plant Mol. Biol. 28:619-634. Watanabe, M., Takasaki, T., Toriyama, K., Yamakawa, S., Isogai, A., Suzuki, A., and Hinata, K. 1994. A high Wang, H., Wu, H.-M., and Cheung, A.Y. 1993. degree of homology exists between the protein Development and pollination regulated accumulaencoded by SLG and the S receptor domain encoded tion and glycosylation of a stylar transmitting tissuespecific proline-rich protein. Plant Cell 5:1639-1650. by SRK in self-incompatible Brassica campestris L. Plant Cell Physiol. 35:1221-1229. Wang, H., Wu, H.-M., and Cheung, A.Y. 1996. Pollination induces mRNA poly(A) tail-shortening and cell Waterkeyn, L., and Bienfait, A. 1970. On a possible function of the callosic special wall in Ipomoea purpurea deterioration in flower transmitting tissue. Plant }. (L.) Roth. Grana 10:13-20. 9:715-727. Wang, H.L., Offler, C.E., and Patrick, J.W. 1995. The cellu- Watson, C.V., Nath, J., and Nanda, D. 1977. Possible mitolar pathway of photosynthate transfer in the develchondrial involvement in mechanism of male steriloping wheat grain. II. A structural analysis and ity in maize (Zea mays L.). Biochem. Genet. histochemical studies of the pathway from the crease 15:1113-1124. phloem to the endosperm cavity. Plant Cell Environ. Webb, M.C., and Gunning, B.E.S. 1990. Embryo sac devel18:373-388. opment in Arabidopsis thaliana. I. Megasporogenesis, including the microtubular cytoskeleton. Sex. Plant Wang, H.L., Patrick, J.W., Offler, C.E., and Wang, X.-D. 1995. The cellular pathway of photosynthate transfer Reprod. 3:244-256. in the developing wheat grain. III. A structural Webb, M.C., and Gunning, B.E.S. 1991. The microtubular analysis and physiological studies of the pathway cytoskeleton during development of the zygote, from the endosperm cavity to the starchy proembryo and free-nuclear endosperm in endosperm. Plant Cell Environ. 18:389^07. Arabidopsis thaliana (L.) Heynh. Planta 184:187-195. Wang, S.-Z., and Esen, A. 1986. Primary structure of a pro- Webb, M.C., and Gunning, B.E.S. 1994. Embryo sac develline-rich zein and its cDNA. Plant Physiol. 81:70-74. opment in Arabidopsis thaliana. II. The cytoskeleton Wang, T.L., Cook, S.K., Francis, R.J., Ambrose, M.J., and during megagametogenesis. Sex. Plant Reprod. Hedley, C.J. 1987a. An analysis of seed development 7:153-163. in Pisum sativum. VI. Abscisic acid accumulation. /. Weber, M. 1988. Formation of sperm cells in Galium mollugo Expt. Bot. 38:1921-1932. (Rubiaceae), Trichodiadema setuliferum (Aizoaceae), and Wang, T.L., Hadavizideh, A., Harwood, A., Welham, T.J., Avena saliva (Poaceae). Plant Syst. Evol. 161:53-64. Harwood, W.A., Faulks, R., and Hedley, C.L. 1990. Weber, M. 1989. Ultrastructural changes in maturing An analysis of seed development in Pisum sativum. pollen grains of Apium nodiflorum L. (Apiaceae), with XIII. The chemical induction of storage product special reference to the endoplasmic reticulum. mutants. Plant Breed. 105:311-320. Protoplasma 152:69-76. Wang, T.L., Smith, CM., Cook, S.K., Ambrose, M.J., and Weber, M. 1992. The formation of pollenkitt in Apium nodHedley, C.L. 1987b. An analysis of seed development iflorum (Apiaceae). Ann. Bot. 70:573-577. in Pisum sativum. III. The relationship between the r Weber, M. 1994. Stigma, style, and pollen tube pathway in locus, the water content and the osmotic potential of Smyrnium perfoliatum (Apiaceae). Int. ]. Plant Sci. seed tissues in vivo and in vitro. Ann. Bot. 59:73-80. 155:437-444. Wang, Y.-C, and Janick, J. 1986. In vitro production of Weber, M., and Frosch, A. 1995. The development of the jojoba liquid wax by zygotic and somatic embryos. /. transmitting tract in the pistil of Hacquetia epipactis Am. Soc. Hort. Sci. 111:798-806. (Apiaceae). Int. ]. Plant Sci. 156:615-621. Wang, Z.-Y., Wu, Z.-L., Xing, Y.-Y, Zheng, F.-G., Guo, X.- Wedderburn, F., and Richards, A.J. 1990. Variation in L., Zhang, W.-G., and Hong, M.-M. 1990. Nucleotide within-morph incompatibility inhibition sites in hetsequence of rice waxy gene. Nucl. Acids Res. 18:5898. eromorphic Primula L. Neiv Phytol. 116:149-162. Wang, Z.-Y, Zheng, F.-Q., Shen, G.-Z., Gao, J.-P., Snustad, Weeden, N.F., and Gottlieb, L.D. 1979. Distinguishing D.P., Li, M.-G., Zhang, J.-L., and Hong, M.-M. 1995. allozymes and isozymes of phosphoglucoisoThe amylose content in rice endosperm is related to merases by electrophoretic comparisons of pollen the post-transcriptional regulation of the waxy gene. and somatic tissues. Biochem. Genet. 17:287-296. Plant J. 7:613-622. Weeden, N.F., and Gottlieb, L.D. 1980. Isolation of cytoWarmke, H.E., and Lee, S.-L.J. 1977. Mitochondrial plasmic enzymes from pollen. Plant Physiol. 66:400-403. degeneration in Texas cytoplasmic male-sterile corn anthers. /. Hered. 68:213-222. Weeks, J.T., Anderson, O.D., and Blechl, A.E. 1993. Rapid production of multiple independent lines of fertile Warmke, H.E., and Lee, S.-L.J. 1978. Pollen abortion in T transgenic wheat (Triticum aestivum). Plant Physiol. cytoplasmic male-sterile corn (Zea mays): a sug102:1077-1084. gested mechanism. Science 200:561-563. Warmke, H.E., and Overman, M.A. 1972. Cytoplasmic Wehling, P., Hackauf, B., and Wricke, G. 1994. Phosphorylation of pollen proteins in relation to male sterility in Sorghum. I. Callose behavior in ferself-incompatibility in rye (Secale cereale L.). Sex. tile and sterile anthers. /. Hered. 63:102-108. Warner, S.A.J., Scott, R., and Draper, J. 1993. Isolation of Plant Reprod. 7:67-75. an asparagus intracellular PR gene (AoPRl) wound- Weigel, D., and Meyerowitz, E.M. 1993. Activation of flo-
660 ral homeotic genes in Arabidopsis. Science 261:1723-1726. Weigel, D., Alvarez,J., Smyth, D.R., Yanofsky, M.F., and Meyerowitz, E.M. 1992. LEAFY controls floral meristem identity in Arabidopsis. Cell 69:843-859. Weinand, U., Briischke, C, and Feix, G. 1979. Cloning of double stranded DNAs derived from polysomal mRNA of maize endosperm: isolation and characterisation of zein clones. Nucl. Acids Res. 6:2707-2715. Weisenseel, M.H., and Jaffe, L.F. 1976. The major growth current through lily pollen tubes enters as K+ and leaves as H+. Planta 133:1-7. Weisenseel, M.H., Nuccitelli, R., and Jaffe, L.F. 1975. Large electrical currents traverse growing pollen tubes. /. Cell Bioi 66:556-567. Weiss, C.A., Huang, H., and Ma, H. 1993. Immunolocalization of the G protein a subunit encoded by a GPA1 gene in Arabidopsis. Plant Cell 5:1513-1528. Weissinger, A.K., Timothy, D.H., Levings, C.S., HI, and Goodman, M.M. 1983. Patterns of mitochondrial DNA variation in indigenous maize races of Latin America. Genetics 104:365-379. Weissinger, A.K., Timothy, D.H., Levings, C.S., III, Hu, W.W.L., and Goodman, M.M. 1982. Unique plasmidlike mitochondrial DNAs from indigenous maize races of Latin America. Proc. Natl Acad. Sci. USA 79:1-5. Welk, M., Millington, W.F., and Rosen, W.G. 1965. Chemotropic activity and the pathway of pollen tube in lily. Am. J. Bot. 52:774-781. Welsh, J.R., and Klatt, A.R. 1971. Effects of temperature and photoperiod on spring wheat pollen viability. Crop Sci. 11:864-865. Wemmer, T., Kaufmann, H., Kirch, H.-H., Schneider, K., Lottspeich, F., and Thompson, R.D. 1994. The most abundant soluble basic protein of the stylar transmitting tract in potato (Solanum tuberosum L.) is an endochitinase. Planta 194:264-273. Wenck, A.R., Conger, B.V., Trigiano, R.N., and Sams, C.E. 1988. Inhibition of somatic embryogenesis in orchardgrass by endogenous cytokinins. Plant Physiol. 88:990-992. Weng, W.-M., Gao, X.-S., Zhuang, N.-L., Xu, M.-L., and Xue, Z.-T. 1995. The glycinin A3B4 mRNA from wild soybean Glycine soja Sieb. et. Zucc. Plant Physiol. 107:665-666. Wenzel, M., Gers-Barlag, H., Schimpl, A., and Riidiger, H. 1993. Time course of lectin and storage protein biosynthesis in developing pea {Pisum sativum) seeds. Biol. Chem. Hoppe-Seyler 374:887-894. Werr, W., Frommer, W.-B., Maas, C, and Starlinger, P. 1985. Structure of the sucrose synthase gene on chromosome 9 of Zea mays L. EMBO J. 4:1373-1380. Weselake, R.J., Pomeroy, M.K., Furukawa, T.L., Golden, J.L., Little, D.B., and Laroche, A. 1993. Developmental profile of diacylglycerol acyltransferase in maturing seeds of oilseed rape and safflower and microspore-derived cultures of oilseed rape. Plant Physiol. 102:565-571. Weselake, R.J., Taylor, D.C., Pomeroy, M.K., Lawson, S.L., and Underhill, E.W. 1991. Properties of diacylglycerol acyltransferase from microspore-derived embryos of Brassica napus. Phytochemistry 30:3533-3538. Wessler, S.R., and Varagona, MJ. 1985. Molecular basis of mutations at the waxy locus of maize: correlation with the fine structure genetic map. Proc. Natl Acad. Sci. USA 82:4177-4181.
References
West, M.A.L, Yee, K.M., Danao, J., Zimmerman, J.L., Fischer, R.L., Goldberg, R.B., and Harada, J.J. 1994. LEAFY COTYLEDON1 is an essential regulator of late embryogenesis and cotyledon identity in Arabidopsis. Plant Cell 6:1731-1745. Weterings, K., Reijnen, W., van Aarssen, R., Korstee, A., Spijkers, J., van Herpen, M., Schrauwen, J., and Wullems, G. 1992. Characterization of a pollen-specific cDNA clone from Nicotiana tabacum expressed during microgametogenesis and germination. Plant Mol. Biol. 18:1101-1111. Weterings, K., Reijnen, W., Wijn, G., van de Heuvel, K., Appeldoorn, N., de Kort, G., van Herpen, M., Schrauwen, J., and Wullems, G. 1995. Molecular characterization of the pollen-specific genomic clone NTPg303 and in situ localization of expression. Sex. Plant Reprod. 8:11-17. Wetherell, D.F., and Dougall, D.K. 1976. Sources of nitrogen supporting growth and embryogenesis in cultured wild carrot tissue. Physiol. Plant. 37:97-103. Wetherell, D.F., and Halperin, W. 1963. Embryos derived from callus tissue cultures of the wild carrot. Nature 200:1336-1337. Wetzel, C.L.R., and Jensen, W.A. 1992. Studies of pollen maturation in cotton: the storage reserve accumulation phase. Sex. Plant Reprod. 5:117-127. Wever, G.H., and Takats, S.T. 1971. Isolation and separation of S-competent and S-incompetent nuclei from Tradescantia pollen grains. Expt. Cell Res. 69:29-32. Weymann, K., Urban, K., Ellis, D.M., Novitzky, R., Dunder, E., Jayne, S., and Pace, G. 1993. Isolation of transgenic progeny of maize by embryo rescue under selective conditions. In Vitro Cell Devel Biol. 29P:33-37. Wheatley, W.G., Marsolais, A.A., and Kasha, K.J. 1986. Microspore growth and anther staging in barley anther culture. Plant Cell Rep. 5:47-49. Wheeler, C.T., and Boulter, D. 1967. Nucleic acids of developing seeds of Vicia faba L. J. Expt. Bot. 18:229-240. Whipple, A.P., and Mascarenhas, J.P. 1978. Lipid synthesis in germinating Tradescantia pollen. Phytochemistry 17:1273-1274. White, D.W.R., and Williams, E. 1976. Early seed development after crossing of Trifolium semipilosum and T. repens. New Zealand ]. Bot. 14:161-168. White, O.E. 1914. Studies of teratological phenomena in their relation to evolution and the problems of heredity. I. A study of certain floral abnormalities in Nicotiana and their bearing on theories of dominance. Am. J. Bot. 1:23-36. Whitmore, F.W., and Kriebel, H.B. 1987. Expression of a gene in Pinus strobus ovules associated with fertilization and early embryo development. Can. ]. For. Res. 17:408-412. Wiberg, E., Rahlen, L., Hellman, M., Tillberg, E., Glimelius, K., and Stymne, S. 1991. The microsporederived embryo of Brassica napus L. as a tool for studying embryo-specific lipid biogenesis and regulation of oil quality. Theor. Appl. Genet. 82:515-520. Wienand, U., Langridge, P., and Feix, G. 1981. Isolation and characterization of a genomic sequence of maize coding for a zein gene. Mol. Gen. Genet. 182:440-444. Wilde, H.D., Nelson, W.S., Booij, H., de Vries, S.C., and Thomas, T.L. 1988. Gene-expression programs in embryogenic and non-embryogenic carrot cultures. Planta 176:205-211.
References
661
1973. The activity of the nucleolus organising region Wilen, R.W., Mandel, R.M., Pharis, R.P., Holbrook, L.A., and the origin of cytoplasmic nucleoloids in meioand Moloney, M.M. 1990. Effects of abscisic acid and cytes of Lilium. Protoplasma 77:79-93. high osmoticum on storage protein gene expression in microspore embryos of Brassica napus. Plant Williams, E., and White, D.W.R. 1976. Early seed development after crossing of Trifolium ambiguum and T. Physiol. 94:875-881. repens. New Zealand J. Bot. 14:307-314. Wilen, R.W., van Rooijen, G.J.H., Pearce, D.W., Pharis, R.P., Holbrook, L.A., and Moloney, M.M. 1991. Effects of Williams, E.G. 1987. Interspecific hybridization in pasture jasmonic acid on embryo-specific processes in legumes. Plant Breed. Rev. 5:237-305. Brassica and Linum oilseeds. Plant Physiol. 95:399-405.Williams, E.G., Clarke, A.E., and Knox, R.B., ed. 1994. Genetic Control of Self-Incompatibility and Reproductive Wilkins, T.A., and Raikhel, N.V. 1989. Expression of rice Development in Flowering Plants. Dordrecht: Kluwer lectin is governed by two temporally and spatially Academic Publishers. regulated mRNAs in developing embryos. Plant Cell 1:541-549. Williams, E.G., and Heslop-Harrison, J. 1979. A comparison of RNA synthetic activity in the plasmodial and Willcox, M.C., Reed, S.C, Burns, J.A., and Wynne, J.C. secretory types of taperum during the meiotic inter1991. Effect of microspore stage and media on anther val. Phytomorphology 29:370-381. culture of peanut (Arachis hypogaea L.). Plant Cell Williams, E.G., Knox, R.B., and Rouse, J.L. 1982. Tissue Organ Cult. 24:25-28. Pollination sub-systems distinguished by pollen Willemse, M.T.M. 1972. Morphological and quantitative tube arrest after incompatible interspecific crosses in changes in the population of cell organelles during Rhododendron (Ericaceae). /. Cell Sci. 53:255-277. microsporogenesis of Gasteria verrncosa. Acta Bot. Williams, E.G., Knox, R.B., Kaul, V., and Rouse, J.L. 1984. Neerl. 21:17-31. Post-pollination callose development in ovules of Willemse, M.T.M. 1981. Polarity during megasporogeneRhododendron and Ledum (Ericaceae): zygote special sis and megagametogenesis. Phytomorphology wall. /. Cell Sci. 69:127-135. 31:124-134. Willemse, M.T.M., and Bednara, J. 1979. Polarity during Williams, E.G., Ramm-Anderson, S., Dumas, C., Mau, megasporogenesis in Gasteria verrucosa. PhytoS.L., and Clarke, A.E. 1982. The effect of isolated components of Primus avium L. styles on in vitro morphology 29:156-165. growth of pollen tubes. Planta 156:517-519. Willemse, M.T.M., and de Boer-de Jeu, M.J. 1981. Megasporogenesis and early megagametogenesis. Williamson, J.D., and Quatrano, R.S. 1988. ABA-regulation of two classes of embryo-specific sequences in Acta Soc. Bot. Polon. 50:111-120. mature wheat embryos. Plant Physiol. 86:208-215. Willemse, M.T.M., and Franssen-Verheijen, M.A.W. 1986. Stylar development in the open flower of Gasteria ver- Williamson, J.D., Quatrano, R.S., and Cuming, A.C. 1985. Em polypeptide and its messenger RNA levels are rncosa (Mill.) H. Duval. Acta Bot. Neerl. 35:297-309. modulated by abscisic acid during embryogenesis in Willemse, M.T.M., and Kapil, R.N. 1981a. Antipodals of wheat. Eur. ]. Biochem. 152:501-507. Gasteria verrucosa (Liliaceae) - An ultrastructural Williamson, J.D., and Scandalios, J.G. 1992. Differential study. Acta Bot. Neerl. 30:25-32. Willemse, M.T.M., and Kapil, R.N. 1981b. On some anomresponse of maize catalases to abscisic acid: Vpl alies during megasporogenesis in Gasteria verrucosa transcriptional activator is not required for abscisic acid-regulated Catl expression. Proc. Natl Acad. Sci. (Liliaceae). Acta Bot. Neerl. 30:439-447. USA 89:8842-8846. Willemse, M.T.M., Plyushch, T.A., and Reinders, M.C. 1995. In vitro micropylar penetration of the pollen Williamson, J.D., Galili, G., Larkins, B.A., and Gelvin, S.B. 1988. The synthesis of a 19 kilodalton zein protein in tube in the ovule of Gasteria verrucosa (Mill.) H. transgenic Petunia plants. Plant Physiol. 88:1002-1007. Duval and Lilium longiflorum Thunb.: conditions, Willing, R.P., Bashe, D., and Mascarenhas, J.P. 1988. An attraction and application. Plant Sci. 108:201-208. analysis of the quantity and diversity of messenger Willemse, M.T.M., and Reznickova, S.A. 1980. Formation RNAs from pollen and shoots of Zea mays. Theor. of pollen in the anther of Lilium. I. Development of Appl. Genet. 75:751-753. the pollen wall. Acta Bot. Neerl. 29:127-140. Willemse, M.T.M., and van Lammeren, A.A.M. 1988. Willing, R.P., and Mascarenhas, J.P. 1984. Analysis of the Structure and function of the microtubular complexity and diversity of mRNAs from pollen cytoskeleton during megasporogenesis and embryo and shoots of Tradescantia. Plant Physiol. 75:865-868. sac development in Gasteria verrucosa (Mill.) H. Willing, R.R., and Pryor, L.D. 1976. Interspecific hybridisDuval. Sex. Plant Reprod. 1:74-82. ation in poplar. Theor. Appl. Genet. 47:141—151. Willemse, M.T.M., and van Went, J.L. 1984. The female Wilms, H.J. 1980a. Ultrastructure of the stigma and style gametophyte. In Embryology of Angiosperms, ed. B.M. of spinach in relation to pollen germination and Johri, pp. 159-196. Berlin: Springer-Verlag. pollen tube growth. Acta Bot. Neerl. 29:33-47. Willemse, M.T.M., and Vletter, A. 1995. Appearance and Wilms, H.J. 1980b. Development and composition of the spinach ovule. Acta Bot. Neerl. 29:243-260. interaction of pollen and pistil pathway proteins in Gasteria verrucosa (Mill.) H. Duval. Sex. Plant Reprod. Wilms, H.J. 1981a. Ultrastructure of the developing 8:161-167. embryo sac of spinach. Acta Bot. Neerl. 30:75-99. Williams, B., and Tsang, A. 1991. A maize gene expressed Wilms, H.J. 1981b. Pollen tube penetration and fertilizaduring embryogenesis is abscisic acid-inducible and tion in spinach. Acta Bot. Neerl. 30:101-122. highly conserved. Plant Mol. Biol. 16:919-923. Wilms, H.J., Leferink-ten Klooster, H.B., and van Aelst, A.C. 1986. Isolation of spinach sperm cells: 1: Ultrastructure Williams, E., and Heslop-Harrison, J. 1975. Membraneand three-dimensional construction in the mature bounded cisternae associated with the chromosomes pollen grain. In Biotechnology and Ecology of Pollen, ed. at first meiotic metaphase in microsporocytes of D.L. Mulcahy, G.B. Mulcahy and E. Ottaviano, pp. Rhoeo spathacea. Protoplasma 86:285-289. 307-312. New York: Springer-Verlag. Williams, E., Heslop-Harrison, J., and Dickinson, H.G.
662
References
Wilson, D.R., and Larkins, B.A. 1984. Zein gene organiza- Woodard, J.W. 1958. Intracellular amounts of nucleic tion in maize and related grasses. /. Mol. Evol. acids and protein during pollen grain growth in 20:330-340. Tradescantia. J. Biophys. Biochem. Cytol. 4:383-390. Wilson, G.F., Rhodes, A.M., and Dickinson, D.B. 1973. Woodcock, C.L.F., and Bell, P.R. 1968a. The distribution of Some physiological effects of viviparous genes up, deoxyribonucleic acid in the female gametophyte of and vp5 on developing maize kernels. Plant Physiol. Myosurus minimus. Histochemie 12:289-301. 52:350-356. Woodcock, C.L.F., and Bell, P.R. 1968b. Features of the Wilson, H.J., Israel, H.W., and Steward, F.C. 1974. ultrastructure of the female gametophyte of Morphogenesis and the fine structure of cultured Myosurus minimus. ]. Ultrastr. Res. 22:546-563. carrot cells. /. Cell Sci. 15:57-73. Woodward, J.R., Bacic, A., Jahnen, W., and Clarke, A.E. 1989. N-linked glycan chains on S-allele-associated Wilson, H.M. 1977. Culture of whole barley spikes stimulates high frequencies of pollen calluses in individglycoproteins from Nicotiana alata. Plant Cell 1:511-514. ual anthers. Plant Sci. Lett. 9:233-238. Wilson, H.M., Mix, G., and Foroughi-Wehr, B. 1978. Early Woodward, J.R., Craik, D., Dell, A., Khoo, K.-H., Munro, microspore divisions and subsequent formation of S.L.A., Clarke, A.E., and Bacic, A. 1992. Structural microspore calluses at high frequency in anthers of analysis of the N-linked glycan chains from a stylar Hordeum vulgare L. /. Expt. Bot. 29:227-238. glycoprotein associated with expression of self-incompatibility in Nicotiana alata. Glycobiology 2:241-250. Wing, R.A., Yamaguchi, J., Larabell, S.K., Ursin, V.M., and McCormick, S. 1989. Molecular and genetic charac- Worrall, D., Hird, D.L., Hodge, R., Paul, W., Draper, J., terization of two pollen-expressed genes that have and Scott, R. 1992. Premature dissolution of the sequence similarity to pectate lyases of the plant microsporocyte callose wall causes male sterility in pathogen Erwinia. Plant Mol. Biol. 14:17-28. transgenic tobacco. Plant Cell 4:759-771. Wintz, H., Chen, H.-C, Sutton, C.A., Conley, C.A., Wright, D.J., and Boulter, D. 1972. The characterisation of Cobb, A., Ruth, D., and Hanson, M.R. 1995. vicilin during seed development in Vicia faba (L.). Expression of the CMS-associated urfS sequence in Planta 105:60-65. transgenic petunia and tobacco. Plant Mol. Biol. Wright, S.Y., Suner, M.-M., Bell, P.J., Vaudin, M., and 28:83-92. Greenland, A.J. 1993. Isolation and characterization of male flower cDNAs from maize. Plant ]. 3:41-49. Wise, R.R, Fliss, A.E., Pring, D.R., and Gengenbach, B.G. 1987. urfli-T of T cytoplasm maize mitochondria Wrobel, B., and Bednarska, E. 1994. Nuclear DNA content encodes a 13 kD polypeptide. Plant Mol. Biol. and ultrastructure of secretory cells of Vicia faba L. 9:121-126. stigma. Acta Soc. Bot. Polon. 63:139-145. Wise, R.R, Pring, D.R., and Gengenbach, B.G. 1987. Wu, F.S., and Murry, L.E. 1985a. Changes in protein and Mutation to male fertility and toxin insensitivity in amino acid content during anther development in Texas (T)-cytoplasm maize is associated with a fertile and cytoplasmic male sterile Petunia. Theor. frameshift in a mitochondrial open reading frame. Appl. Genet. 71:68-73. Proc. Natl Acad. Sci. USA 84:2858-2862. Wu, F.S., and Murry, L.E. 1985b. Proteolytic activity in Wobus, U., Baumlein, H., Bassiiner, R., Heim, U., Jung, R., anther extracts of fertile and cytoplasmic male sterMuntz, K., Saalbach, G., and Weschke, W. 1986. ile Petunia. Plant Physiol. 79:301-305. Characteristics of two types of legumin genes in the Wu, H.-M., Wang, H., and Cheung, A.Y. 1995. A pollen tube growth stimulatory glycoprotein is deglycosyfield bean {Vicia faba L. var. minor) genome as lated by pollen tubes and displays a glycosylation revealed by cDNA analysis. FEBS Lett. 210:74-80. gradient in the flower. Cell 82:395-403. Wochok, Z.S. 1973a. Microtubules and multivesicular bodies in cultured tissues of wild carrot: changes Wu, H.-M., Zou, J., May, B., Gu, Q., and Cheung, A.Y. during transition from the undifferentiated to the 1993. A tobacco gene family for flower cell wall proembryonic condition. Cytobios 7:87-95. teins with a proline-rich domain and a cysteine-rich domain. Proc. Natl Acad. Sci. USA 90:6829-6833. Wochok, Z.S. 1973b. DNA synthesis during development of adventive embryos of wild carrot. Biol. Plant. Wu, S.-C, Bogre, L., Vincze, E., Kiss, G.B., and Dudits, D. 15:107-111. 1988. Isolation of an alfalfa histone H3 gene: structure and expression. Plant Mol. Biol. 11:641-649. Wochok, Z.S., and Wetherell, D.F. 1972. Restoration of declining morphogenetic capacity in long term tis- Wu, X.-L., and Zhou, C. 1990. Nuclear divisions of isosue cultures of Daucus carota by kinetin. Experientia lated, in vitro cultured generative cells in Hemerocallis 28:104-105. minor Mill. Acta Bot. Sinica 32:577-581. Wodehouse, R.P. 1935. Pollen Grains, Their Structure, Wu, X.-L., and Zhou, C. 1991a. A comparative study on Identification and Significance in Science and Medicine. methods for isolation of generative cells in various angiosperm species. Acta Biol. Expt. Sinica 24:15-23. New York: McGraw-Hill Book Co. Woittiez, R.D., and Willemse, M.T.M. 1979. Sticking of Wu, X.-L., and Zhou, C. 1991b. Fusion experiments of isopollen on stigmas: the factors and a model. lated generative cells in several angiosperm species. Phytomorphology 29:57-63. Acta Bot. Sinica 33:897-904. Wong, K.C., Watanabe, M., and Hinata, K. 1994a. Protein Wu, Y., and Zhou, C. 1988. Observation on enzymatically profiles in pin and thrum floral organs of distylous isolated-embryo sacs in Adenophora axilliflora: fertilAverrhoa carambola L. Sex. Plant Reprod. 7:107-115. ization, zygote and endosperm. Acta Bot. Sinica Wong, K.C., Watanabe, M., and Hinata, K. 1994b. 30:210-212. Fluorescence and scanning electron microscopic Wu, Y, and Zhou, C. 1990. An ultrastructural study on study on self-incompatibility in distylous Averrhoa pollen protoplasts and pollen tubes germinated from them in Gladiolus gandavensis. Acta Bot. Sinica carambola L. Sex. Plant Reprod. 7:116-121. 32:493^198. Woodard, J.W. 1956. DNA in gametogenesis and embryogeny in Tradescantia. J. Biophys. Biochem. Cytol. Wu, Y., and Zhou, C. 1992. An ultrastructural study on triggering of cell division in young pollen protoplast 2:765-776.
References
663
during development. /. Expt. Bot. 46:687-694. culture of Hemerocaliis fulva L. Ada Bot. Sinica 34:20-25. Xu, N., Coulter, K.M., and Bewley, J.D. 1990. Abscisic acid and osmoticum prevent germination of developing Wu, Y., Zhou, C, and Koop, H.-U. 1993. Enzymatic isolaalfalfa embryos, but only osmoticum maintains the tion of viable nucelli at the megaspore mother cell synthesis of developmental proteins. Planta stage and in developing embryo sacs in Nicotiana 182:382-390. tabacum. Sex. Plant Reprod. 6:171-175. Wu, Y., Haberland, G., Zhou, C, and Koop, H.-U. 1992. Xu, N., Coulter, K.M., Krochko, J.E., and Bewley, J.D. 1991. Somatic embryogenesis, formation of morphoMorphological stages and storage protein accumulagenetic callus and normal development in zygotic tion in developing alfalfa (Medicago sativa L.) seeds. embryos of Arabidopsis thaliana in vitro. Protoplasma Seed Sci. Res. 1:119-125. 169:89-96. Xu, S.-X. (S.Y. Zee). 1991. Changes in microtubule cytoskeleton of generative cells during mitosis and Wulff, H.D. 1935. Galvanotropismus bei Pollencytokinesis. Chinese J. Bot. 3:77-80. schlauchen. Planta 24:602-608. Wunderlich, R. 1954. Uber das Antherentapetum mit Xu, S.-X. (S.Y. Zee), Zhu, C, and Hu, S.-Y. 1990. Changes besonderer Berucksichtigung seiner Kernzahl. in the organization of microtubules during generative cell division and sperm formation in Lilium. Ada Osterr. Z. Bot. 101:1-63. Bot. Sinica 32:821-826. Wurtele, E.S., and Nikolau, B.J. 1992. Differential accumulation of biotin enzymes during carrot somatic Xu, S.-X. (S.Y. Zee), Zhu, C, and Hu, S.-Y. 1993. embryogenesis. Plant Physiol. 99:1699-1703. Microtubule-cytoskeleton changes during generative cell division and sperm formation in the pollen Wurtele, E.S., Wang, H., Durgerian, S., Nikolau, B.J., and tubes of Zantedeschia aethiopica. Chinese J. Bot. Ulrich, T.H. 1993. Characterization of a gene that is 5:18-22. expressed early in somatic embryogenesis of Daucus carota. Plant Physiol. 102:303-312. Xu, X., and Li, B. 1994. Fertile transgenic indica rice plants obtained by electroporation of the seed embryo cells. Xiao, C.-M., and Mascarenhas, J.P. 1985. High temperaPlant Cell Rep. 13:237-242. ture-induced thermotolerance in pollen tubes of Xu, Z.H., Huang, B., and Sunderland, N. 1981. Culture of Tradescantia and heat-shock proteins. Plant Physiol. barley anthers in conditioned media. /. Expt. Bot. 78:887-890. 32:767-778. Xu, B., Grun, P., Kheyr-Pour, A., and Kao, T.-H. 1990a. Xue, J., Pihlgren, U., and Rask, L. 1993. Temporal, cell-speIdentification of pistil-specific proteins associated cific, and tissue-preferential expression of myrosiwith three self-incompatibility alleles in Solarium chanase genes during embryo and seedling coense. Sex. Plant Reprod. 3:54-60. development in Sinapis alba. Planta 191:95-101. Xu, B., Mu, J., Nevins, D.L., Grun, P., and Kao, T.-H. Xue, Y., Carpenter, R., Dickinson, H.G., and Coen, E.S. 1990b. Cloning and sequencing of cDNAs encoding 1996. Origin of allelic diversity in Antirrhinum S two self-incompatibility associated proteins in locus RNases. Plant Cell 8:805-814. Solatium chacoense. Mol. Gen. Genet. 224:341-346. Xue, Y., Collin, S., Davies, D.R., and Thomas, CM. 1994. Differential screening of mitochondrial cDNA Xu, R, Yao, Y.-X., Fu, C.-S., and Li, Z.-L. 1993. Studies on libraries from male-fertile and cytoplasmic malethe development of pollen wall in Pyrus betulaefolia sterile sugar-beet reveals genome rearrangements at Bge. (Rosaceae). Chinese ]. Bot. 5:41-46. atp6 and atpA loci. Plant Mol. Biol. 25:91-103. Xu, H., Davies, S.P., Kwan, B.Y.H., O'Brien, A.P., Singh, M., and Knox, R.B. 1993. Haploid and diploid Xue, Z.-T., Xu, M.-L., Shen, W., Zhuang, N.L., Hu, W.-M., expression of a Brassica campestris anther-specific and Shen, S.C. 1992. Characterization of a Gy4 gene promoter in Arabidopsis and tobacco. Mol. Gen. glycinin gene from soybean Glycine max cv. Forrest. Genet. 239:58-65. Plant Mol. Biol. 18:897-908. Xu, H., Knox, R.B., Taylor, P.E., and Singh, M.B. 1995a. XuHan, X., and van Lammeren, A.A.M. 1993. Bcpl, a gene required for male fertility in Arabidopsis. Microtubular configurations during the cellularizaProc. Natl Acad. Sci. USA 92:2106-2110. tion of coenocytic endosperm in Ranunculus sceleraXu, H., Theerakulpisut, P., Taylor, P.E., Knox, R.B., Singh, tus L. Sex. Plant Reprod. 6:127-132. M.B., and Bhalla, P.L. 1995b. Isolation of a gene pref- XuHan, X., and van Lammeren, A.A.M. 1994. erentially expressed in mature anthers of rice (Oryza Microtubular configurations during endosperm sativa L.). Protoplasma 187:127-131. development in Phaseolus vulgaris. Can. }. Bot. 72:1489-1495. Xu, N., and Bewley, J.D. 1991. Sensitivity to abscisic acid and osmoticum changes during embryogenesis of alfalfa (Medicago sativa).}. Expt. Bot. 42:821-826. Yabe, N., Takahashi, T., and Komeda, Y. 1994. Analysis of tissue-specific expression of Arabidopsis thaliana Xu, N., and Bewley, J.D. 1995a. Embryos of alfalfa HSP90-family gene HSP81. Plant Cell Plnjsiol. (Medicago sativa L.) can synthesize storage proteins 35:1207-1219. after the completion of germination, in response to abscisic acid and osmoticum, if maturation drying is Yadegari, R., de Paiva, G.R., Laux, T., Koltunow, A.M., prevented. Planta 196:469-476. Apuya, N., Zimmerman, J.L., Fischer, R.L., Harada, J.J., and Goldberg, R.B. 1994. Cell differentiation and Xu, N., and Bewley, J.D. 1995b. Temporal and nutritional morphogenesis are uncoupled in Arabidopsis raspfactors modulate responses to abscisic acid and berry embryos. Plant Cell 6:1713-1729. osmoticum in their regulation of storage protein synthesis in developing seeds of alfalfa (Medicago Yaklich, R.W., and Herman, E.M. 1995. Protein storage vacuoles of soybean aleurone cells accumulate a sativa L.). /. Expt. Bot. 46:675-686. unique glycoprotein. Plant Sci. 107:57-67. Xu, N., and Bewley, J.D. 1995c. The role of abscisic acid in germination, storage protein synthesis and desicca- Yaklich, R.W., Vigil, E.L., Erbe, E.F., and Wergin, W.P. 1992. The fine structure of aleurone cells in the soytion tolerance in alfalfa (Medicago sativa L.) seeds, as bean seed coat. Protoplasma 167:108-119. shown by inhibition of its synthesis by fluridone
664
References
Yakovlev, M.S., and Yoffe, M.D. 1957. On some peculiarimicroscopic study on in vitro parthenogenesis in ties in the embryogeny of Paeonia L. Phytomorphology sunflower. Sear. Plant Reprod. 2:154-166. 7:74-82. Yan, H., Yang, H.-Y., and Jensen, W.A. 1991a. Yamada, Y. 1965. Studies on the histological and cytologiUltrastructure of the micropyle and its relationship cal changes in the tissues of pistil after pollination. to pollen tube growth and synergid degeneration in Jap. J. Bot. 19:69-82. sunflower. Sex. Plant Reprod. 4:166-175. Yamagata, H., and Tanaka, K. 1986. The site of synthesis Yan, H., Yang, H.-Y, and Jensen, W.A. 1991b. and accumulation of rice storage proteins. Plant Cell Ultrastructure of the developing embryo sac of sunPhysiol. 27:135-145. flower (Helianthus annuus) before and after fertilizaYamagata, H., Sugimoto, T., Tanaka, K., and Kasai, Z. tion. Can. J. Bot. 69:191-202. 1982. Biosynthesis of storage proteins in developing Yan, L., Wang, X., Teng, X., Ma, Y, and Liu, G. 1986. Actin rice seeds. Plant Physiol. 70:1094-1100. and myosin in pollens and their role in the growth of Yamagata, H., Tamura, K., Tanaka, K., and Kasai, Z. 1986. pollen tubes. Chinese Sci. Bull. 31:267-272. Cell-free synthesis of rice prolamin. Plant Cell Yang, C, Xing, L., and Zhai, Z. 1992. Intermediate filaPhysiol. 27:1419-1422. ments in higher plant cells and their assembly in a cell-free system. Protoplasma 171:44-54. Yamaguchi, H., and Kakiuchi, H. 1983. Electrophoretic analysis of mitochondrial DNA from normal and Yang, H., Zhou, C, Cai, D., Yan, H., Wu, Y., and Chen, X. male sterile cytoplasms in rice. Jap. J. Genet. 1986. In vitro culture of unfertilized ovules in 58:607-611. Helianthus annuus L. In Haploids of Higher Plants In Yamaguchi-Shinozaki, K., Mundy, J., and Chua, N.-H. Vitro, ed. H. Hu and H. Yang, pp. 182-191. Beijing: 1989. Four tightly linked rab genes are differentially China Academic Publishers. expressed in rice. Plant Mol. Biol. 14:29-39. Yang, H.-Y., and Zhou, C. 1979. Experimental researches Yamaguchi-Shinozaki, K., Mino, M, Mundy, J., and Chua, on the two pathways of pollen development in N.-H. 1990. Analysis of an ABA-responsive rice gene Oryza sativa L. Ada Bot. Sinica 21:345-351. promoter in transgenic tobacco. Plant Mol. Biol. Yang, H.Y., and Zhou, C. 1982. In vitro induction of hap15:905-912. loid plants from unpollinated ovaries and ovules. Yamakawa, S., Shiba, H., Watanabe, M., Shiozawa, H., Theor. Appl. Genet. 63:97-104. Takayama, S., Hinata, K., Isogai, A., and Suzuki, A. Yang, H.-Y., and Zhou, C. 1984. Observations on megas1994. The sequences of S-glycoproteins involved in porogenesis and megagametophyte development in self-incompatibility of Brassica campestris and their Paulownia sp. and Sesamum indicum by enzymatic distribution among Brassicaceae. Biosci. Biotech. maceration technique. Acta Bot. Sinica 26:355-358. Biochem. Japan 58:921-925. Yang, H.-Y., and Zhou, C. 1989. Isolation of viable sperms Yamakawa, S., Watanabe, S., Hinata, K., Suzuki, A., and from pollen of Brassica napus, Zea mays and Secale Isogai, A. 1995. The sequences of S-receptor kinases cereale. Chinese J. Bot. 1:80-84. (SRK) involved in self-incompatibility and their Yang, L.J., Barratt, D.H.P., Domoney, C, Hedley, C.L., and homologies to S-locus glycoproteins of Brassica Wang, T.L. 1990. An analysis of seed development in campestris. Biosci. Biotech. Biochem. Japan 59:161-162. Pisum sativum. X. Expression of storage protein genes in cultured embryos. /. Expt. Bot. 41:283-288. Yamakawa, S., Watanabe, M., Isogai, A., Takayama, S., Satoh, S., Hinata, K., and Suzuki, A. 1993. The cDNA Yano, M., Isono, Y., Satoh, H., and Omura, T. 1984. Gene sequence of NS3-glycoprotein from Brassica analysis of sugary and shrunken mutants of rice, campestris and its homology to related proteins. Plant Oryza sativa L. Jap. J. Breed. 34:43-49. Cell Physiol. 34:173-175. Yano, M., Okuno, K., Kawakami, J., Satoh, H., and Omura, T. 1985. High amylose mutants of rice, Oryza Yamamoto, S., Nishihara, M., Morikawa, H., Yamauchi, D., and Minamikawa, T. 1995. Promoter analysis of sativa L. Theor. Appl. Genet. 69:253-257. seed storage protein genes from Canavalia gladiata Yanofsky, M.F., Ma, H., Bowman, J.L., Drews, G.N., D.C. Plant Mol. Biol. 27:729-741. Feldmann, K.A., and Meyerowitz, E.M. 1990. The Yamauchi, D., and Minamikawa, T. 1986. In vivo studies protein encoded by the Arabidopsis homeotic gene agamous resembles transcription factors. Nature on protein synthesis in developing seeds of 346:35-39. Canavalia gladiata D.C. Plant Cell Physiol. 27:1033-1041. Yao, J.-L., Wu, J.-H., Gleave, A.P., and Morris, B.A.M. 1996. Transformation of Citrus embryogenic cells Yamauchi, D., and Minamikawa, T. 1987. Synthesis of using particle bombardment and production of canavalin and concanavalin A in maturing Canavalia transgenic embryos. Plant Sci. 113:175-183. gladiata seeds. Plant Cell Physiol. 28:421^30. Yamauchi, D., and Minamikawa, T. 1990. Structure of the Yenofsky, R.L., Fine, M., and Liu, C. 1988. Isolation and gene encoding concanavalin A from Canavalia gladicharacterization of a soybean (Glycine max) lipoxygeata and its expression in Escherichia coli cells. FEBS nase-3 gene. Mol. Gen. Genet. 211:215-222. Lett. 260:127-130. Yeung, E.C. 1980. Embryogeny of Phaseolus: the role of the suspensor. Z. Pflanzenphysiol. 96:17-28. Yamauchi, D., Nakamura, K., Asahi, T., and Minamikawa, T. 1988. cDNAs for canavalin and concanavalin A Yeung, E.C, and Brown, D.C.W. 1982. The osmotic envifrom Canavalia gladiata seeds. Nucleotide sequence ronment of developing embryos of Phaseolus vulof cDNA for canavalin and RNA blot analysis of garis. Z. Pflanzenphysiol. 106:149-156. canavalin and concanavalin A mRNAs in develop- Yeung, E.C., and Cavey, M.J. 1988. Cellular endosperm ing seeds. Em. J. Biochem. 170:515-520. formation in Phaseolus vulgaris. I. Light and scanning electron microscopy. Can. J. Bot. 66:1209-1216. Yamaura, A. 1933. Karyologische und embryologische Studien iiber einige Bambws-Arten. Bot. Mag. Tokyo Yeung, E.C, and Clutter, M.E. 1978. Embryogeny of Phaseolus coccineus: growth and microanatomy. 47:551-555. Protoplasma 94:19^0. Yan, H., Yang, H.-Y, and Jensen, W.A. 1989. An electron
References
665
5:169-181. Yeung, E.C., and Clutter, M.E. 1979. Embryogeny of Phaseolus coccineus: the infrastructure and develop-Yu, H.-S., and Russell, S.D. 1993. Three-dimensional ultrament of the suspensor. Can. ]. Bot. 57:120-136. structure of generative cell mitosis in the pollen tube of Nicotiana tabacum. Eur. ]. Cell Biol. 61:338-348. Yeung, E.C., and Meinke, D.W. 1993. Embryogenesis in angiosperms: development of the suspensor. Plant Yu, H.-S., and Russell, S.D. 1994a. Populations of plastids Cell 5:1371-1381. and mitochondria during male reproductive cell maturation in Nicotiana tabacum L.: a cytological Yeung, E.C., and Sussex, I.M. 1979. Embryogeny of basis for occasional biparental inheritance. Planta Phaseolus coccineus: the suspensor and the growth of 193:115-122. the embryo-proper in vitro. Z. Pflanzenphysiol. 91:423-433. Yu, H.-S., and Russell, S.D. 1994b. Male reproductive cell development in Nicotiana tabacum: male germ unit Yeung, E.C., Thorpe, T.A., and Jensen, C.J. 1981. In vitro associations and quantitative cytology during sperm fertilization and embryo culture. In Plant Tissue maturation. Sex. Plant Reprod. 7:324-332. Culture. Methods and Applications in Agriculture, ed. T.A. Thorpe, pp. 253-271. New York: Academic Yuffd, A.M., de Garcia, E.G., and Nieto, M.S. 1994. Press. Comparative study of protein electrophoretic patYibrah, H.S., Manders, G., Clapham, D.H., and von terns during embryogenesis in Coffea arabica cv Arnold, S. 1994. Biological factors affecting transient Catimor. Plant Cell Rep. 13:197-202. transformation in embryogenic suspension cultures Yunes, J.A., Neto, G.C., da Silva, M.J., Leite, A., Ottoboni, of Picea abies. J. Plant Physiol. 144:472-478. L.M.M., and Arruda, P. 1994. The transcriptional Yin, Y, and Beachy, R.N. 1995. The regulatory regions of activator opaque2 recognizes two different target the rice tungro bacilliform virus promoter and intersequences in the 22-kD-like a-prolamin gene. Plant acting nuclear factors in rice (Oryza sativa L.). Plant ]. Cell 6:237-249. 7:969-980. Ylstra, B., Busscher, J., Franken, ]., Hollman, P.C.H., Mol, Zachgo, S., de Andrade Silva, E., Motte, P., Trobner, W., J.N.M., and van Tunen, A.J. 1994. Flavonols and ferSaedler, H., and Schwarz-Sommer, Z. 1995. tilization in Petunia hybrida: localization and mode of Functional analysis of the Antirrhinum floral homeotic DEF1CIENS gene in vivo and in vitro by action during pollen tube growth. Plant ]. 6:201-212. using a temperature-sensitive mutant. Development Ylstra, B., Touraev, A., Benito Moreno, R.M., Stoger, E., 121:2861-2875. van Tunen, A.J., Vicente, O., Mol, J.N.M., and Heberle-Bors, E. 1992. Flavonols stimulate develop- Zaki, M., and Dickinson, H. 1995. Modification of cell development in vitro: the effect of colchicine on ment, germination, and tube growth of tobacco anther and isolated microspore culture in Brassica pollen. Plant Physiol. 100:902-907. napus. Plant Cell Tissue Organ Cult. 40:255-270. Yokota, E., McDonald, A.R., Liu, B., Shimmen, T, and Palevitz, B.A. 1995. Localization of a 170 kDa myosin Zaki, M., and Kuijt, J. 1994. Ultrastructural studies on the heavy chain in plant cells. Protoplasma 185:178-187. embryo sac of Viscum minimum. II. Megagametogenesis. Can. ]. Bot. 72:1613-1628. Yoo, J., and Jung, G. 1995. DNA uptake by imbibition and expression of a foreign gene in rice. Physiol. Plant. Zaki, M., and Kuijt, J. 1995. Ultrastructural studies on the 94:453-459. embryo sac of Viscum minimum. I. MegasporoYou, R., and Jensen, W.A. 1985. Ultrastructural observagenesis. Protoplasma 185:93-105. tions of the mature megagametophyte and the fertil- Zaki, M.A.M. and Dickinson, H.G. 1990. Structural ization in wheat (Triticum aestivum). Can. ]. Bot. changes during the first divisions of embryos result63:163-178. ing from anther and free microspore culture in Brassica napus. Protoplasma 156:149-162. Youle, R.J., and Huang, A.H.C. 1981. Occurrence of low molecular weight and high cysteine containing albu- Zaki, M.A.M., and Dickinson, H.G. 1991. Microsporemin storage proteins in oilseeds of diverse species. derived embryos in Brassica: the significance of diviAm. J. Bot. 68:44-48. sion symmetry in pollen mitosis I to embryogenic development. Sex. Plant Reprod. 4:48-55. Young, E.G., and Hanson, M.R. 1987. A fused mitochondrial gene associated with cytoplasmic male sterility ZiSrsky, V., RihovA, L., and Tupy, J. 1987. Interference of pollen diffusible substances with peroxidase catis developmentally regulated. Cell 41:41-49. alyzed reaction. Plant Sci. 52:29-32. Young, L.C.T., and Stanley, R.G. 1963. Incorporation of tritiated nucleosides thymidine, uridine and cytidine Zarsky, V., Capkova, V., HrabetovS, E., and TupyJ. 1985. Protein changes during pollen development in in nuclei of germinating pine pollen. Nucleus Nicotiana tabacum L. Biol. Plant. 27:438-444. 6:83-90. Yu, H.-S., Hu, S.-Y, and Russell, S.D. 1992. Sperm cells in Zarsky, V., Garrido, D., Eller, N., Tupy, J., Vicente, O., Schoffl, F., and Heberle-Bors, E. 1995. The expression pollen tubes of Nicotiana tabacum L.: three-dimenof a small heat shock gene is activated during inducsional reconstruction, cytoplasmic diminution, and tion of tobacco pollen embryogenesis by starvation. quantitative cytology. Protoplasma 168:172-183. Plant Cell Environ. 18:139-147. Yu, H.-S., Hu, S.-Y, and Zhu, C. 1989. Ultrastructure of sperm cells and the male germ unit in pollen tubes Zarsky, V, Garrido, D., Rfhova, L., Tupy, J., Vicente, O., and Heberle-Bors. E. 1992. Derepression of the cell of Nicotiana tabacum. Protoplasma 152:29-36. cycle by starvation is involved in the induction of Yu, H.-S., Huang, B.-Q., and Russell, S.D. 1994. tobacco pollen embryogenesis. Sex. Plant Reprod. Transmission of male cytoplasm during fertilization 5:189-194. in Nicotiana tabacum. Sex. Plant Reprod. 7:313-323. Yu, H.-S., and Russell, S.D. 1992. Male cytoplasmic Zee, S.Y. 1992. Confocal laser scanning microscopy of diminution and male germ unit in young and microtubule organizational changes in isolated genmature pollen of Cymbidium goeringii: a 3-dimenerative cells of Allemanda neriifolia during mitosis. sional and quantitative study. Sex. Plant Reprod. Sex. Plant Reprod. 5:182-188.
666
References
Zee, S.Y., and Aziz-Un-Nisa. 1991. Mitosis and micro- Zhang, H.-Q., Croes, A.F., and Linskens, H.F. 1982. tubule organizational changes in isolated generative Protein synthesis in germinating pollen of Petunia: cells of Allemanda neriifolia. Sex. Plant Reprod. role of proline. Planta 154:199-203. 4:132-137. Zhang, H.-Q., Bohdanowicz, ]., Pierson, E.S., Li, Y.-Q., Zee, S.-Y., and O'Brien, T.P. 1971. Aleurone transfer cells Tiezzi, A., and Cresti, M. 1995. Microtubular organiand other structural features of the spikelet of millet. zation during asymmetrical division of the generaAust. ]. Biol. Sci. 24:391-395. tive cell in Gagea lutea. J. Plant Res. 108:269-276. Zee, S.-Y., and Siu, I.H.P. 1990. Studies on the ontogeny of Zhang, J.Z., Gomez-Pedrozo, M., Baden, C.S., and the pollinium of a massulate orchid (Peristylus spiHarada, J.J. 1993. Two classes of isocitrate lyase ranthes). Rev. Palaeobot. Palynol. 64:159-164. genes are expressed during late embryogeny and postgermination in Brassica napus L. Mol. Gen. Genet. Zee, S.-Y, Wu, S.C., and Yue, S.B. 1979. Morphological 238:177-184. and SDS-polyacrylamide gel electrophoretic studies of pro-embryoid formation in the petiole explants of Zhang, J.Z., Laudencia-Chingcuanco, D.L., Comai, L., Li, Chinese celery. Z. Pflanzenphysiol. 95:397-403. M., and Harada, J.J. 1994. Isocitrase lyase and malate synthase genes from Brassica napus L. are active in Zee, S.-Y, and Ye, X.L. 1995. Changes in the pattern of pollen. Plant Physiol. 104:857-864. organization of the microtubular cytoskeleton during megasporogenesis in Cymbidium sinense. Zhang, J.Z., Santes, CM., Engel, M.L., Gasser, C.S., and Protoplasma 185:170-177. Harada, J.J. 1996. DNA sequences that activate isocitrate lyase gene expression during late embryogeneZeijlemaker, F.C.J. 1956. Growth of pollen tubes in vitro sis and during postgerminative growth. Plant and their reaction on potential differences. Acta Bot. Physiol. 110:1069-1079. Neerl. 5:179-186. Zenkteler, M. 1962. Microsporogenesis and tapetal devel- Zhang, S., Chen, L., Qu, R., Marmey, P., Beachy, R., and opment in normal and male-sterile carrots (Daucus Fauquet, C. 1996. Regeneration of fertile transgenic indica (group 1) rice plants following microprojectile carota). Am. ]. Bot. 49:341-348. transformation of embryogenic suspension culture Zenkteler, M. 1967. Test-tube fertilization of ovules in cells. Plant Cell Rep. 15:465-469. Melandrium album Mill, with pollen grains of several species of the Caryophyllaceae family. Experientia Zhang, W., McElroy, D., and Wu, R. 1991. Analysis of rice 23:775. Actl 5' region activity in transgenic rice plants. Plant Cell 3:1155-1165. Zenkteler, M. 1990. In vitro fertilization and wide hybridization in higher plants. Crit. Rev. Plant Sci. Zhang, X.S., and O'Neill, S.D. 1993. Ovary and gameto9:267-297. phyte development are coordinately regulated by auxin and ethylene following pollination. Plant Cell Zenkteler, M., Maheswaran, G., and Williams, E.G. 1987. 5:403-418. In vitro placental pollination in Brassica campestris Zhao, J.-P., Simmonds, D.H., and Newcomb, W. 1996. and B. napus. ]. Plant Physiol. 128:245-250. Induction of embryogenesis with colchicine instead Zenkteler, M., Misiura, E., and Guzowska, 1.1975. Studies of heat in microspores of Brassica napus L. cv. Topas. on obtaining hybrid embryos in test tubes. In Form, Planta 198:433-439. Structure and Function in Plants, ed. H.Y. Mohan Ram, J.J. Shaw and C.K. Shaw, pp. 180-187. Meerut: Zhao, Y, Leisy, D.J., and Okita, T.W. 1994. Tissue-specific expression and temporal regulation of the rice Sarita Prakashan. glutelin Gt3 gene are conferred by at least two spaZenkteler, M., and Nitzsche, W. 1984. Wide hybridization tially separated ris-regulatory elements. Plant Mol. experiments in cereals. Theor. Appl. Genet. 68:311-315. Biol. 25:429^136. Zenkteler, M., and Nitzsche, W. 1985. In vitro culture of ovules of Triticum aestiviim at early stages of embryo- Zheng, Y, He, M., Hao, S., and Huang, B. 1992. The ultragenesis. Plant Cell Rep. 4:168-171. structural evidence on the origin of protein bodies in the rough endoplasmic reticulum of developing Zhan, X.-Y, Wu, H.-M., and Cheung, A.Y. 1996. Nuclear cotyledons of soybean. Ann. Bot. 69:377-383. male sterility induced by pollen-specific expression Zheng, Y.-Z., He, M.-Y, Hu, A.-L., and Hao, S. 1990. of a ribonuclease. Sex. Plant Reprod. 9:35-43. Patterns of the transition of vacuoles into protein Zhang, R, and Boston, R.S. 1992. Increases in binding probodies in developing cotyledon cells of soybean. tein (BiP) accompany changes in protein body morActa Bot. Sinica 32:97-102. phology in three high-lysine mutants of maize. Zheng, Z., Kawagoe, Y, Xiao, S., Li, Z., Okita, T, Hau, Protoplasma 171:142-152. T.L., Lin, A., and Murai, N. 1993. 5' distal and proxiZhang, G., Gifford, D.J., and Cass, D.D. 1993. RNA and mal cis-acting regulator elements are required for protein synthesis in sperm cells isolated from Zea mays L. pollen. Sex. Plant Reprod. 6:239-243. developmental control of a rice seed storage protein glutelin gene. Plant}. 4:357-366. Zhang, G., Campenot, M.K., McGann, L.E., and Cass, D.D. 1992a. Flow cytometric characteristics of sperm Zheng, Z., Sumi, K., Tanaka, K., and Murai, N. 1995. The bean seed storage protein fJ-phaseolin is synthecells isolated from pollen of Zea mays L. Plant Physiol. sized, processed, and accumulated in the vacuolar 99:54-59. type-II protein bodies of transgenic rice endosperm. Zhang, G., Williams, CM., Campenot, M.K., McGann, Plant Physiol. 109:777-786. L.E., and Cass, D.D. 1992b. Improvement of longevity and viability of sperm cells isolated from Zhong, H., Bolyard, M.G., Srinivasan, C, and Sticklen, pollen of Zea mays L. Plant Physiol. 100:47-53. M.B. 1993. Transgenic plants of turfgrass (Agrostis palustris Huds.) from microprojectile bombardment Zhang, G., Williams, CM., Campenot, M.K., McGann, of embryogenic callus. Plant Cell Rep. 13:1-6. L.E., Cutler, A.J., and Cass, D.D. 1995. Effects of calcium, magnesium, potassium and boron on sperm Zhong, H., Zhang, S., Warkentin, D., Sun, B., Wu, T., Wu, R., and Sticklen, M.B. 1996. Analysis of the funccells isolated from pollen of Zea mays L. Sex. Plant tional activity of the 1.4-kb 5-region of the rice actin Reprod. 8:113-122.
References
667
embryogenesis and analysis of peroxidase in cul1 gene in stable transgenic plants of maize (Zea mays tured lettuce (Lactuca sativa L.) cotyledons. Ann. Bot. L.). Plant Sci. 116:73-84. 69:97-100. Zhou, C. 1987a. A study of fertilization events in living embryo sacs isolated from sunflower ovules. Plant Zhu, C, and Liu, B. 1990. Microtubules in generative and Sci. 52:147-151. sperm cells of Amaryllis pollen tubes. Chinese J. Bot. 2:1-6. Zhou, C. 1987b. Cell-biological studies on artificially isolated generative cells from angiosperm pollen. Ada Zhu, C, Hu, S., Xu, L., Li, X., and Shen, J. 1980. Ultrastructure of sperm cell in mature pollen grain Bot. Sinica 29:117-122. of wheat. Sri. Sinica 23:371-379. Zhou, C. 1988. Isolation and purification of generative cells from fresh pollen of Viciafaba L. Plant Cell Rep. Zhu, J.-K., Damsz, B., Kononowicz, A.K., Bressan, R.A., 7:107-110. and Hasegawa, P.M. 1994. A higher plant extracellular vitronectin-like adhesion protein is related to the Zhou, C. 1989a. A study on isolation and culture of pollen translational elongation factor-la. Plant Cell protoplasts. Plant Sci. 59:101-108. 6:393^04. Zhou, C. 1989b. Cell divisions in pollen protoplast culture Zhu, Q., Doemer, P.W., and Lamb, C.J. 1993. Stress inducof Hemerocallis fulva L. Plant Sci. 62:229-235. tion and developmental regulation of a rice chitinase Zhou, C, and Wu, X.-L. 1990. Mass isolation and purifipromoter in transgenic tobacco. Plant J. 3:203-212. cation of generative cells from pollen grains. Ada Zhu, T., Ma, Z.-L., and Li, W.-T. 1990. Ultrastructural Bot. Sinica 32:404-406. localization of adenosine triphosphatase activity in Zhou, C, and Yang, H.-Y. 1980. Anther culture and androstigma cells of Populus lasiocarpa and its changes durgenesis of Hordeum vulgare L. Ada Bot. Sinica 22:211-215. ing development. Ada Bot. Sinica 32:91-96. Zhou, C, and Yang, H.-Y. 1982. Enzymatic isolation of Zhu, X, Mogensen, H.L., and Smith, S.E. 1990. Generative cell composition and its relation to male plastid embryo sacs in angiosperms: isolation and microinheritance patterns in Medicago sativa. Protoplasma scopical observation on fixed materials. Ada Bot. Sinica 24:403-407. 158:66-72. Zhou, C, and Yang, H.Y. 1985. Observations on enzymat- Zhu, T., Mogensen, H.L., and Smith, S.E. 1991. Quantitative cytology of the alfalfa generative cell ically isolated, living and fixed embryo sacs in sevand its relation to male plastid inheritance patterns eral angiosperm species. Planta 165:225-231. in three genotypes. Theor. Appl. Genet. 81:21-26. Zhou, C, and Yang, H.Y. 1991. Microtubule changes during the development of generative cells in Zhu, T., Mogensen, H.L., and Smith, S.E. 1992. Heritable paternal cytoplasmic organelles in alfalfa sperm Hippeastrum vittatum pollen. Sex. Plant Reprod. 4:293-297. cells: ultrastructural reconstruction and quantitative cytology. Eur. J. Cell Biol. 59:211-218. Zhou, C, Yang, H.-Y, and Xu, S.-X. 1990. Fluorescence microscopic observations on actin filament distribu- Zhu, T., Mogensen, H.L., and Smith, S.E. 1993. tion in corn pollen and Gladiolus pollen protoplasts. Quantitative, three-dimensional analysis of alfalfa Ada Bot. Sinica 32:657-662. egg cells in two genotypes: implications for biparental plastid inheritance. Planta 190:143-150. Zhou, C, Zee, S.Y., and Yang, H.Y. 1990. Microtubule organization of in situ and isolated generative cells Zhu, Z.-P., Shen, R.-J., and Tang, X.-H. 1980a. Studies on in Zephyranthes grandiflora Lindl. Sex. Plant Reprod. the developmental biology of embryogenesis in higher plants. II. Biochemical changes during 3:213-218. embryogeny in Oryza sativa L. Ada Phytophysiol. Zhou, C, Orndorff, K., Allen, R.D., and DeMaggio, A.E. Sinica 6:141-148. 1986a. Direct observations on generative cells isolated from pollen grains of Haemanthus katherinae Zhu, Z.-P, Shen, R.-J., and Tang, X.-H. 1980b. Studies on Baker. Plant Cell Rep. 5:306-309. the developmental biology of embryogenesis in higher plants. III. Kinetic changes of nucleic acids Zhou, C, Orndorff, K., Daghlian, C.P., and DeMaggio, and protein during embryogenesis of wheat A.E. 1988. Isolated generative cells in some (Triticum vulgaris L). Ada Bot. Sinica 22:122-126. angiosperms: a further study. Sex. Plant Reprod. 1:97-102. Zhu, Z.-P, Shen, R.-J., and Tang, X.-H. 1989. The biosynZhou, C, Yang, H., Tian, H., Liu, Z., and Yan, H. 1986b. In thesis of storage protein and changes of free amino acids content during seed development of rice. vitro culture of unpollinated ovaries in Oryza sativa L. In Haploids of Higher Plants In Vitro, ed. H. Hu and Chinese]. Bot. 1:123-130. H. Yang, pp. 165-181. Beijing: China Academic Zhu, Z.-Q., Sun, J.-S., and Wang, J.-J. 1978. Cytological Publishers. investigation on androgenesis of Triticum aestivum. Ada Bot. Sinica 20:6-12. Zhou, G.Y., Weng, J., Gong, Z., Zhen, Y., Yang, W., Shen, W., Wang, Z., Tao, Q., Huang, J., Qian, S., Lin, G., Zhukova, G.Y., and Sokolovskaya, T.B. 1977. Ultrastructure of antipodals of Aconitum napellus L. Ying, M., Xue, D., Hong, A., Xu, Y, Chen, S., and Duan, X. 1988. Molecular breeding of agriculture. A (Ranunculaceae) embryo sac before fertilization. Bot. Zh. 62:1600-1611. technique for introducing exogenous DNA into Ziebur, N.K., Brink, R.A., Graf, L.H., and Stahmann, M.A. plants after self pollination. Sri. Agr. Sinica 21:1-6. Zhou, J.-Y. 1980. Pollen dimorphism and its relation to 1950. The effect of casein hydrolysate on the growth the formation of pollen embryos in anther culture in vitro of immature Hordeum embryos. Am. ]. Bot. of wheat (Triticum aestivum). Ada Bot. Sinica 37:144-148. 22:117-121. Zimmerman, J.L. 1993. Somatic embryogenesis: a model Zhou, X., and Fan, Y.-L. 1993. The endosperm-specific for early development of higher plants. Plant Cell expression of a rice prolamin chimeric gene in trans5:1411-1423. genic tobacco plants. Transgen. Res. 2:141-146. Zimmerman, J.L., Apuya, N., Darwish, K., and O'Carroll, Zhou, X., Han, Y, Yang, W., and Xi, T. 1992. Somatic C. 1989. Novel regulation of heat shock genes during
668
References
carrot somatic embryo development. Plant Cell interaction in Brassica. III. Hydration of the pollen 1:1137-1146. grains. /. Cell Sri. 76:321-336. Zonia, L.E., and Tupy, J. 1995. Lithium treatment of Zubkova, M., and Sladky, Z. 1975. The possibility of Nicotiana tabacum microspores blocks polar nuclear obtaining seeds following placental pollination in migration, disrupts the partitioning of membranevitro. Biol. Plant. 17:276-280. associated Ca2+, and induces symmetrical mitosis. zur Nieden, U., Manteuffel, R., Weber, E., and Neumann, Sex. Plant Reprod. 8:152-160. D. 1984. Dictyosomes participate in the intracellular Zou,J., Abrams, G.D., Barton, D.L., Taylor, D.C., Pomeroy, pathway of storage proteins in developing Viciafaba cotyledons. Eur. J. Cell Biol. 34:9-17. M.K., and Abrams, S.R. 1995. Induction of lipid and oleosin biosynthesis by (+)-abscisic acid and zur Nieden, U., Neumann, D., Bucka, A., and Nover, L. its metabolites in microspore-derived embryos 1995. Tissue-specific localization of heat-stress of Brassica napus L. cv. Reston. Plant Physiol. proteins during embryo development. Planta 108:563-571. 196:530-538. Zou, J.-T., Zhan, X.-Y., Wu, H.-M., Wang, H., and Cheung, zur Nieden, U., Neumann, D., Manteuffel, R., and Weber, A.Y. 1994. Characterization of a rice pollen-specific E. 1982. Electron microscopic immunocytochemical gene and its expression. Am. ]. Bot. 81:552-561. localization of storage proteins in Vicia faba seeds. Eur. J. Cell Biol. 26:228-233. Zuberi, M.I., and Dickinson, H.G. 1985. Pollen-stigma
Index A-B chromosome interchange 400 ABA-deficient (aba) mutant 433 ABA-INSENSITIVE (AB1) gene, (abi) mutant 433, 455 Abelmoschus esculentus 408 aberrant testa shape (ats) mutant 156 abscisic acid (ABA) 9,10,114,117,134,135, 381, 387, 388, 400-402, 406, 411, 426-433, 436, 451-453, 463, 464, 473, 482, 489, 492, 493, 495, 520, 521 Abutilon megapotamicum 71 Acacia retinodes 84,185, 253 ACC (see 1-aminocyclopropane-l-carboxylic acid) Acetabularia mediterranea 352 acetohydroxyacid synthase 520 acetolactate synthase 40 acetyl choline esterase 188 acetyl-coA carboxylase 488 acetylesterase 380 Achillea millefolium 170 acid phosphatase 34, 37, 44, 70-72, 92, 93,137,194, 219, 230, 247, 380, 520 Aconitum sp. 170 A. napellus 170 A. vitlparia 170,171 acridine orange 260 actin, actin gene 45, 47, 78,102,105,107,109,117, 214-217, 227, 229, 234-238, 240, 241, 295, 307, 326, 406, 426, 495 actin-binding factor 118 actin corona 308 actin-depolymerizing factor (ADF) 109, 111 actinomycin-D 53, 54, 221, 224, 225, 260, 517 activator (ac), mobile or transposable element 345, 369 actomyosin 215, 237, 238, 241 acyl-ACP thioesterase 451, 462 acyl carrier protein (ACP), ACP gene 104,116,197, 200, 451, 462, 464 adenine phosphoribosyl transferase 134 adenosine triphosphatase (ATPase) 56,142,176,187 S-adenosylmethionine decarboxylase 136, 491 adenosine diphosphate-glucose pyrophosphorylase 335, 336 gene 117 Fj-adenosine triphosphate synthase a-subunit gene (ATP-A) 41,146,147,149,150 P-subunit 99 subunit-6 gene (ATP-6) 107, 147, 148-150 subunit-9 gene (ATP-9) 41,107,144,145 adenylate cyclase 187, 270 ADF-like proteins 216 adventive embryogenesis 3,14,176 Aesculus sp. 502 A. californicus 329
A. hippocastanum 513 A. woerlitzensis 329 African blood lily (see Haemanthus katherinae) African violet (see Saintpaulia ionantha) AGAMOUS (AG) gene, ag mutant 20-25,153, 156 AGAMOUS-LIKE (AGL) gene 20, 22, 23, 25, 425 Agapanthus umbellatus 51, 214, 240 Agave americana 96 A. attenuata 99 A. parryi 164,166 Agrobacterium sp. 13, 20, 525-528, 530 A. tumefaciens 10, 20, 39,117, 227, 351, 398, 413, 414, 454, 479, 497, 525-528, 531 A. rhizogenes 122, 132, 426, 496, 531 agroinfection 527 Agropyron trichophorum 149 Agrostemma githago 74 Agrostis palustris 530 AINTEGUMENTA (ANT) gene, ant mutant 156,174 alanine amino transferase 479 Albizzia sp. 503 albumin 336, 337, 341, 438, 441, 444, 450, 459, 460-462 alcohol dehydrogenase (ADH), ADH gene 19, 91-93,104, 107,116,117, 219, 334, 372, 426 aldehyde dehydrogenase 138 aldolase 77 Alectra vogelii 367 aleurone cells 323, 324, 327, 333-335, 339, 340, 342, 347, 348, 350, 352, 353, 355, 384, 400, 401 aleurone layer (see aleurone cells) alfalfa 80, 200, 454, 460, 473, 478, 484, 495 alfin 441, 449 Alisma lanceolatum 378, 379 A. plantago-aquatka 378, 379 alkaline phosphatase 40, 380 Allamanda neriifolia 85, 88 Allanblackia parviflora 329 allergenic proteins (see allergens) allergens 72, 73, 83,106, 108,109, 117-118 Allium sp. 168 A. ammophilum 168 A. angulosum 168 A. cepa 58, 81,133,134,160,168 A. chamaemoly 86 A. nutans 168 A. pidchellwn 168, 170 A. schoenoprasum mitochondrial transcripts of 150 A. tuberosum 164 A. ursinum 170 Alocasia macrorrhiza 88 Aloe ciliaris 214
669
670 A.jucunda 80 A. secundiflora 80 A. vera 128 Alopecurus myosuroides 126 A. pratensis 236, 298 Alstroemeria sp. 235 Alyssum maritimum 378 oc-amanitin -resistant cells 497 Amaranthus hypochondriacus 445, 450 Amaryllis sp. 70 A. belladonna 83, 235, 241 A. vittata 88,187,190, 214 Ambrosia sp. 72 A. artemisiifolia (see also ragweed) 70, 72,108,109 A. elatior 70 A. psilostachya 70 A. tenuifolia 70, 205 A. trifida 70,118, 203 1-aminocyclopropane-l-carboxylic acid (ACC) oxidase 207 synthase 207 Ammophila arenaria 170 amygdalin hydrolase 412, 413 amylase (see a -amylase) a-amylase, a-amylase gene 70,137, 219, 340, 401, 527 a-amylase inhibitor 436 amylose extender (ae) mutant 92, 336 Ananas sp. 253 androecium 19 androgenesis 317, 500 Aneilema sp. 182,186,188 Anemarrhena asphodeloides 35 Anemone sp. 502 A. canadensis 507 Angiospermae (see angiosperms) angiosperms 1-9,11,13,14,17, 25, 29, 36, 41, 42, 48, 56, 60-62, 64, 65, 70, 73, 81, 82, 86-91,100,104,118, 119,120-123,129,151,152,154,158-160,162, 163,166,168,172-174,176-178,181-183,185, 195, 214, 222, 226, 229, 232, 234, 240, 244, 245, 251, 268, 292-299, 301, 308, 309, 311, 313, 317, 321, 322, 326, 328, 337, 361-363, 367, 371, 376, 377, 379, 393, 394, 395, 407, 408, 411, 432, 437, 441, 459, 464, 467, 470, 494, 500, 501, 521, 531 Anoda crispa 72 anther 2, 4-8,10,13,14, 42,49, 51, 53, 63, 72, 73, 75, 77, 86, 90, 91, 94-96, 99,101,102,104-108,111-114,116, 120-138,142,145,146, 150,151,181,182,192, 202, 211, 244, 245, 247-249, 257, 258, 264, 277, 279, 284, 289, 472, 501-510, 512, 519, 526, 528 allometric relations in 25-26 cell lineages of 28 culture 4,10, 48, 57-60, 75, 501-522, 526, 528 determination 17-25 development 25-41 wall 5, 8, 26-28, 30-33, 49, 80,102,112,123,124,129, 130 anther locule (see microsporangium) anther-specific genes 29-30, 41, 60, 90, 104, 111, 115, 116
Index anthocyanin, anthocyanin genes 340, 368, 400, 401, 434, 526 Anthophyta 1 anthranilate synthetase, anthranilate synthetase gene 200, 497 Anthurium sp. 67 antipodals 2, 3,162,163,172-175,177, 317 structure of 169-171 Antirrhinum sp. 24, 25 Antirrhinum majus (see also snapdragon) 8, 19, 20, 37, 122, 123,132,153,172,198, 291, 303, 314 floral homeotic mutants in 24-25 antisense construct (see antisense RNA) antisense RNA 11,12, 20, 21, 39^1,106, 111, 122,132, 200, 282, 288, 395, 449, 494 APETALA (AP) gene, ap mutant 20-25,153,156 aphidicolin 446 Apium nodiflorum 35, 83 apocytochrome-6 gene (COB) 107,146,149 apomict 322, 408 apomictic species (see apomict) apomixis 176-178 apple 223, 290, 291, 329 Aptenia cordifolia 188 Aquilegia formosa 18,153,166 A. vulgaris 166 Arabidopsis 7, 8, 25, 40, 41 63,107,116,117,123,127-129, 131,134,135,150,153-156,160,164,167, 169-172,174,192,197, 200, 202, 204-206, 227, 228, 276, 278, 283, 284, 305, 322, 352, 358, 359, 361, 362, 364, 370, 372, 376, 382, 384, 390, 398, 399, 402, 419^26, 429, 432, 433, 442, 450, 452, 453, 459, 461^63, 505, 527 embryo-lethal mutants in 413-415 floral homeotic mutants in 19-24 transgenic 21, 23, 24,112,114,116,117,132,156,197, 200, 228, 283, 369, 426, 429, 434, 456, 459 A. thaliana 7 arabinogalactan 9, 83,102,110,187,191,194-196, 200, 208, 232, 311, 478, 484, 486 Arabis cypria 271 Arachis sp. 502 A. hypogaea 442, 506 Aranda sp. 25 arcelin 436 archegonium 163, 407 archesporial cell (see archesporial initial) archesporial initial 5, 26, 62,123,156-159,177 archesporium (see archesporial initial) Argemone mexicana 157,173, 314 A. ochroleuca 314 arginine decarboxylase 491 Armeria maritima 249 Arum italicum 35 A. maculatum 170, 326 Asclepias exaltata 304 ascorbate oxidase 110, 111, 227 ascorbic acid 77,175, 413 asexual embryogenesis (see adventive embryogenesis) asexual reproduction 3, 176
Index asparaginase gene 426 Asparagus sp. 502 A. officinalis 116, 531 Aspergillus oryzae 41 Asphodelus tenuifolius 325 Aster tripolium 214, 225 H+-ATPase, H+-ATPase gene 116, 227, 239 Atropa belladonna 512 autoradiography 6,11, 29, 31, 35-38, 44, 48, 51-53, 76, 86, 94-96, 99,100,170, 202, 222, 232, 239, 326, 333, 379, 380, 516 auxin 3,10,18,132,133, 207, 329, 364, 381, 385, 420, 426, 468, 471, 472, 475-478, 482, 484, 486^88, 492, 493, 497, 504, 506, 507, 527 role in somatic embryogenesis 480-481 auxin transport 364, 365 auxotroph 414, 417 Avena coleoptile test 480 Avena sativa (see also oat) 32, 33, 87, 298, 337 avenin 337, 339, 350, 355 Averrhoa carambola 250, 251 avocado (see Persea americana) B-A translocation 92, 415 Bacillus amyloliquefaciens 41 B. thuringiensis 529 bamboo (see Simocalamus latiflora) BAR gene 528-530 barley 86, 87,117,122,126,128,129,131,134-137, 205, 296, 299, 300, 305, 307, 309-312, 317, 323-326, 329, 331, 333, 336, 339-342, 347, 348, 355, 385-387, 389, 392, 407,425, 427, 430, 432, 435,458, 484, 486, 488, 503, 506, 508, 509, 511, 515, 518, 520, 528-530 hordein genes of 352-353 storage protein gene expression in 347 BARNASE gene 41, 42,132, 204 BARSTAR gene 132 basic region/helix-loop-helix protein 455 bean 30,117,146,151,197, 440, 442, 443, 454, 455, 529 beet root 80, 304, 317 BELL (BEL-1) locus, bel-1 mutant 156,174 l,2-benz-isoxazol-3-acetic acid 480 benzylaminopurine 451, 507, 529 Berberis vulgaris 66 Bertholletia excelsa (see also Brazil nut) 450 Beta sp. 503 B. vulgaris (see also beet root) 80,128,130, 149,164,166, 167,169,170, 294, 305, 507 Betula pendula 493 B. verrucosa (see also birch) 106,117 Bidens cervicata 124 biolistic approaches (see microprojectile bombardment) biolistic methods (see microprojectile bombardment) Bipolaris (Helminthosporium) maydis 138 birch 106,117 boron in pollen germination 210 Bougainvillea spectabilis 65 Brassica sp. 264, 267, 272, 273, 275, 276, 281, 284, 288, 292, 298
671 B. alboglabra 487 B. campestris 40, 41, 65, 92,104,106,112,127,129,132, 148,164,166,167,169,172, 226, 258, 261, 266, 274-277, 279, 280, 282, 294, 297, 300, 301, 305, 308, 315, 358, 359, 380, 462, 464, 502, 504, 520 B. juncea 364, 390, 502, 529 B. napus (see also rapeseed) 21, 25, 35, 40, 41, 64, 68, 71, 72, 79, 81, 82, 87,104-107,110-112,114,116, 118,125,131-135,147,148,197, 204, 227, 264, 275-284, 294, 298, 300, 311, 363, 373, 378, 387, 412, 422, 423, 425, 428, 438, 441, 445, 446, 449-453, 459-464, 482, 495, 502, 503, 506, 509, 510, 512, 515, 518-520, 528 cybrids of 147,148 ogura (ogu) cms line of 133,134 transgenic 40, 41, 283, 423, 452 B. nigra 502 B. oleracea 34, 35, 64, 71, 72,102,104,116,127,134,137, 185,188, 201-203, 205, 211, 254, 260-264, 266, 273-284, 294, 297, 300 B. pekinensis 502 B. rapa 118 Brazil nut 450, 460, 462 BRITTLE (BT) gene, bt mutant 332, 335, 336 broad bean 150, 458 Brugmansia (Datura) suaveolens 192,194, 257 bud pollination 261, 263 Bupleurum dianthifolium 160 Burmannia pusilla 367, 368 Butomus umbellatus 35, 382 Ca2+ 81, 138,191, 202, 206, 231, 242, 254, 260, 270, 287, 290-292, 302-304, 306, 308, 316, 391, 433, 507 as second messenger 290 -binding proteins 118 gradient in pollen tube 238-240 in pollen mitosis 77 in pollen germination 210, 217-218, in pollen recognition 34 in pollen tube entry 168 in somatic embryogenesis 478 -modulated phosphatase 433 cabbage (see Brassica oleracea) cacao 253, 481 Caesalpinia japonica 65 caffeine 86 Cajanus sp. 502 C. cajan 127,131 calcium-dependent calmodulin-independent protein kinase (CDPK) gene 106,109 Calcium Green-1 290, 291 Calendula officinalis 166 callase, callase gene 40, 63,125,128,131 callose 35, 42, 46, 61-65, 67, 74, 78, 84, 86, 89,125-128, 131,158,159,177, 220, 228, 229, 232, 254-256, 264, 270, 271, 298, 307, 358, 476 callus-specific proteins 488 calmodulin 109,191, 217, 239, 240, 316, 477, 478 Calocedrus decurrens 458 Caltha palustris 170
672 Camellia japonica 224, 234 C. sasanqua 234 C. sinensis 234 CaMV (see cauliflower mosaic virus) canal cells 192,195-197 secretion products of 195-197 canary grass (see Phalaris tuberosa) Canavalia sp. 441 C. ensiformis 460 C. gladiata 445, 446, 449, 460, 461 canavalin, canavalin gene 441, 449, 460, 461 Canna sp. 34 C. generalis 43, 65 Cannabis sativa (see also hemp) 68 Capparis spinosa 83, 241 Capsella bursa-pastoris 156,158,160,164-166,168-170, 174, 306, 307, 309, 358, 359, 361-365, 371, 372, 374, 376, 377, 382, 383, 385, 386, 390, 391, 393, 398, 442 C. grandiflora 257 Capsicum sp. 502 C. annuum (see also pepper) 31,128,130,133,134, 506, 510 caraway 482 Cardamine pratensis 257 Carica sp. 502 C. papaya (see also papaya) 529, 531 carnation (see Dianthus caryophyllus) carotene 340, 400 carpel (see also pistil) 2, 19-25, 122, 123, 133, 152, 156, 161, 192, 258 determination 153-154 polypeptides of 19 carrot 3, 10, 99,126, 341, 352, 364, 370, 374, 425, 427, 468-472, 474-499 somatic embryogenesis in 470-472 transgenic cell line of 491, 496 Carthamus tinctorius 462 Carum carvi (see also caraway) 482 cassava (see Manihot esculenta) Castilleia sp. 81, 83 castor bean 339-341 CAT (see chloramphenicol acetyltransferase) catalase, catalase gene 107, 219, 333, 334, 401, 412, 431, 435 Catananche caerulea 298 cauliflower mosaic virus (CaMV) promoter 21, 39,117, 132, 352-354, 369, 401, 426, 457, 462, 527-529 cDNA clones 20, 25, 29, 30, 39, 40, 56, 63, 73,105,106, 108-112,116-118,155,197-199, 262, 263, 273, 274, 276, 277, 279, 280, 284-291, 323, 331, 335, 336, 342, 345-347, 350, 351, 353, 355, 395, 401, 403, 405, 407, 411, 425, 427-431, 435, 446, 447, 449, 450, 453-455, 458-460, 462, 463, 491-493, 495, 521 cDNA library 8, 9, 11,12, 90, 285, 395, 411, 439 from anthers 29, 30, 39, 40,118 from carrot cells 494 from egg cells 174 from embryos 411, 428-430
Index from flowers 20, 25, 40, 198, 200 from microsporocytes 55-56 from mitochondria 140,149 from pistil 197, 198 from pollen 12, 40,104, 111, 117 from pollen embryoids 521 from pollen tubes 111, 227 from proembryogenic masses 494 from seedlings 422 from somatic embryos 495 from stamens 37 from stigma 197, 273, 276, 277, 279 from style 284, 288 cDNA probes (see cDNA clones) cell-cycle control protein kinase gene (CDC-2) 423, 426 cell-free translation 11,19, 221, 222, 225, 260, 287, 338, 342, 343, 347, 405-407, 409, 411, 446, 447, 487-489, 491, 495, 517, 518 cellulase 11, 208, 219, 232, 300 Cenchrus ciliaris 530 central cell 2,162,163,165-167,170,172,174-176, 293, 300, 306, 308, 309, 311, 312, 315, 316, 321-325, 361, 382, 383 structure of 168-169 Cerastium arvense 314 Ceratozamia sp. 470 Ceras canadensis 488 chalaza 6,154,157-163,165,166,170,171, 173-176, 304, 306-308, 312, 325, 326, 328, 330, 332, 333, 358, 359, 361, 366, 368, 376-378, 382-384, 397 chalazal end (see chalaza) chalazal half (see chalaza) chalazal part (see chalaza) chalazal pole (see chalaza) chalazal proliferating tissue 383 chalazogamy 304 chalcone flavanone isomerase, chalcone flavanone isomerase gene 41,106,115, 227, 426 chalcone synthase, chalcone synthase gene 30, 41,121, 132 Chamaenerion angustifolium 156 Chara 238 chasmogamous flowers 246 Cheiranthus cheiri 261 chemotropism 302, 303 chick pea (see Cicer arietinum) chicory 166, 489 chimeric gene 8,10,12, 21, 39, 41,112,115,116,122,125, 132,148, 350, 352, 353, 355, 403, 415, 423, 455, 459, 461-464, 496, 527, 529, 530 Chinese celery (see Brassica alboglabra) chitinase, chitinase gene 191,199, 347, 353, 484 chloramphenicol 221, 224 chloramphenicol acetyltransferase (CAT), CAT gene 12, 13,116, 352, 353 2-chloroethyl-trimethyl ammonium chloride (CCC) 134 chlorophyll a/b-binding protein (CAB) gene 116, 424 Chlorophytum comosum 80, 84 C. elatum 84 chloroplast DNA
Index role in male sterility 150-151 chlorotetracycline 217, 239, 240 chlorpromazine 217 Chorisia chodatii 253 C. speciosa 253 chromosome puffs 379-381 Chrysanthemum alpinum 170 Cicer arietinum 428, 445, 527 Cichorium intybus 166, 476, 489-491 Cirsium arvense 123 C. oleraceum 124 C. palustre 127 C. spinosissimum 127 cis-acting elements 56,112, 114-116, 282, 353, 354, 439, 450, 454, 457, 461, 464 cis-acting sequences (see cis-acting elements) ris-regulatory domain (see cis-acting elements) Cistanche tubulosa 367 citrate synthase 412 Citrullus lanatus 185,189 Citrus sp. 253, 330, 502, 503 C. aurantifolia 124 C. aumntium 484, 486 C. jambhiri 374 C. /imon 34, 68,190,195 C. reticulata X C. paradisi 531 C. smewsis 476, 481, 487 Clarkia dudleyana 92 C. unguiculata 219 clathrin-coated vesicles 242 CLAV54TA3 (OV-3) gene 420 cleft coleoptile 397 cleistogamous flowers 129, 246 cleistogamy 246 Clivia miniata 170, 223, 235, 303 C. nobilis 302 cms 121,122, 125,127,128, 130,131,133-151, 510 CO 2 effect on incompatibility 258 Cobaea scandens 69 Cochlearia danica 384 coconut (see Cocos nucifera) coconut milk 329, 389, 390, 468, 473-475, 501, 503 coconut water (see coconut milk) Cocos sp. 502 C. nucifera 329, 385, 511 Coffea sp. 503, 531 C. arabica 489 coffee (see Coffea arabica) cofilin 111 Coix lacryma-jobi 316, 337, 346, 355 coixin 337, 355 colcemid (see colchicine) colchicine 50, 54, 66, 67, 75, 78, 79, 85,114, 217, 236, 237, 241, 503, 512 coleoptile 366, 367, 383, 397, 398, 415-417, 424, 425, 435, 436 coleorhiza 366, 367, 424, 425, 435, 436 Collomia grandiflora 246 Colocasia esculenta 386
673 Commelina sp. 182, 186, 188 C. erecta 186 complementary DNA (see cDNA) concanavalin-A (con-A), con-A gene 187, 201, 220, 265, 266, 268, 441, 449, 460, 461 conglycinin, conglycinin gene 403, 441, 444, 445, 448, 449, 456, 457 p-conglycinin (see conglycinin) connective 17,18, 25, 28, 30 Convallaria majalis 80, 83 convicilin, convicilin gene 441, 444, 447, 458 cordycepin 224 Cordylanthus sp. 81, 83 coriander (see Coriandrum sativum) Coriandrum sp. 503 C. sativum 341 coronatine 128 Corydalis cava 170 C. nobilis 170 Corylus avellana 211, 238, 476 Cosmos bipinnatus 44, 65, 66, 70-72, 74, 75, 203, 257, 264 cotton 45, 81,115,151,190, 206, 296, 306-309, 311, 373, 382, 386, 387, 393, 405-407, 409, 427-432, 447, 450, 451, 453, 460, 493, 527, 529 cotyledon 1, 2, 6, 9, 277, 362-370, 372-375, 377, 380, 381, 383, 384, 386, 387, 398, 402, 405-409, 411-414, 417, 419, 420, 422, 424, 426-428, 434, 436-438, 440-453, 455, 460, 462, 464, 470-472, 475, 477, 481, 488, 510, 528, 529 4-coumarate: coenzyme A ligase gene 29, 30, 116,197 Crambe abyssinica 442 Craterostigma plantagineum 116, 427 Crepis capillaris 168, 305 C.foetida257 C. tectorum 164,166,167,169 Crinum defixum 187,190 Crocus chrysanthus 74, 186, 205, 271 C. longiflorus 95 C. vernus 70, 71 cross-incompatibility 14, 244, 245, 269, 271, 316 Crotalaria retusa 192 cruciferin, cruciferin gene 363, 434, 441, 445, 449-452, 459, 520 Cryptomeria japonica 72,108 cucumber 18, 110,153 Cucumis sp. 393 C. melo 59,126,127 C. sativus (see also cucumber) 18, 329, 389, 390 Cucurbita sp. (see also pumpkin) 92, 442, 446 C. maxima 125, 131, 389, 390 C. moschata 223, 389, 390 C. pepo 36, 63,110, 317 cuticle 10, 71, 73, 74, 172, 183, 185-189, 205, 206, 246, 254, 271, 374, 425, 484 cuticle-pellicle 249 cutin (see cuticle) cutinase 71, 72, 74, 205, 219, 224, 269 Cycfls sp. 470 cycloheximide 50, 221, 223, 224, 231, 251 -resistant cells 488, 497
674 suppression of self-incompatibility by 260 Cymbidium sinense 156 Cyphomandra betacea 298 Cypripedium californicum 45 C. fasciculatum 78 C. insigne 358 Cyrtandra pendula 84 cysteine proteinase 155 Cylinus hypocistis 155,166 cytochalasin 214-216, 231, 234, 236, 237, 241 cytochrome oxidase 137,138 cytochrome oxidase-c subunit genes (COX-1, COX-U) 41, 143-145,147,149,150 cytochrome P450 monooxygenase 155 cytokinesis-defective (cyd) mutant 417 cytokinin 3,134, 381, 385, 386, 391, 471, 472, 474, 506, 507, 527 role in somatic embryogenesis 481 cytoplasmic male sterility (see cms) cytotoxic gene (see diphtheria toxin chain-A gene) Dactylis glomerala 118,129, 474, 489 DAPI (see 4'6-diamidino-2-phenylindole) date palm 86, 488 Datura sp. 502 D. innoxia 86, 385, 501-504, 507-510, 512, 514, 516, 518, 520, 527, 528 D. metel 102, 507, 508, 510, 511, 514 D. stramonium 182, 329, 358, 385, 389, 390 D. tatula 389, 390 Daucus aureus 164 D. carota (see also carrot) 3,130,164, 312, 425 mitochondrial transcripts of 150 D. muricatus 164, 312, 313 deaminases 77 defective endosperm-B30 (de*-B30) mutant 344, 346 defective in pollen-pistil interactions (pop) mutant 204 defective kernel (dek) mutant 332, 333, 415, 416 defective seed (de-17) mutant 332 defense-related genes 434-438 DEFIC1ENS (DEF) gene, def mutant 20, 24, 25,123 dehydrins 426, 428 Dendrathema grandiflora 531 Dendrobium sp. 156,157 deoxyadenosine 49, 50 2-deoxy-2-glucose 254 desiccation tolerance gene expression related to 426-434 4'6-diamidino-2-phenylindole (DAPI) 81,101,173, 310 7,8-diaminopelargonic acid aminotransferase 415 Dianthus caryophyllus 208 D. serotinns 314 Dichanthium aristatum 133 (2,4-dichlorophenoxy) propanoic acid 472 2,4-dichlorophenoxyacetic acid (2,4-D) 18,140, 316, 385, 468, 471^75, 478-483, 488, 491, 493, 495, 497, 498, 501, 507, 529 dicots 1, 8, 9,13, 29, 32, 52, 61, 114, 115,183,194, 325, 353, 362, 363, 366, 367, 383, 393, 415, 427-430, 469, 470, 502, 503, 525, 528-531
Index Dicotyledonae (see dicots) dicotyledons (see dicots) Dicranostigma franchetianum 314 dicyclohexylcarbodiimide (DCCD) 142 Digitalis sp. 502, 503 D. lanata 482, 484, 489, 491, 493 a-difluoromethylarginine 491 difluoromethylorni thine 491 digitonin 138 2,4-dinitrophenol 221 Dipcadi montanum 157 diphtheria toxin chain-A (DT-A) gene 12, 84,116,122, 204, 283, 459 Diplotaxis erucoides 378, 382 D. tenuifolia 205 dissociation (ds) transposable element 335 distyly 250, 251 features of 245-247 DNA polymerase 77, 86 DNA replication (see DNA synthesis) DNA sequences 108-112,140-151, 273-275, 277, 279, 350-356, 423, 427, 429, 430, 453^64 DNA sequencing 12, 39 DNA synthesis 159,173 during haploid pollen mitosis 76-77 during microsporocyte meiosis 43, 47-52 during pollen development 76-77 during pollen embryogenesis 516 in embryos 373-374 in the developing endosperm 326-327 in the generative cell 86 in the suspensor 379-380 in the vegetative cell 86 DNA-unwinding protein ("U-protein") 54 DNA uptake 526-527 DNase 48, 77,173 Doritis pulcherrima 45, 47 dormancy-related genes 432-434 dot-blot hybridization 12, 29, 30,107,132, 263, 347, 446-450
double fertilization 2, 6,181, 208, 293, 298, 299, 301, 309, 311, 314, 317, 321, 322, 356, 399 nuclear fusions (karyogamy) during 308-311 Downingia bacigalupi 375 D. pulchella 375 Drosophila melanogaster 7,107, 228 dynein 238 early methionine-labeled (EM) protein, EM gene 401, 402, 427, 429-431, 463, 493, 494 Echinochloa utilis 324 Echinocystis macrocarpa 329 egg 2-6, 9, 81,152,162,163,165-169,172-178, 243, 293, 297-301, 305-317, 357-359, 361, 382, 393, 395, 396, 398, 399, 407, 408, 498, 501, 505, 507, 526, 527, 531 macromolecules of 173-174 eggplant (see Solanum melongena) Eichhornia paniculata 249 electroporation 13,114, 352, 401, 525, 526, 528, 530
Index elongation factor 1 (EF1, EF-loc) 57, 344, 494 Elymus rectisetus 177 E. scabrus 177 embellum 304 embryo 1-3, 5, 6, 9,10,13,14, 40, 111, 166,168,176,177, 245, 269, 271, 311-315, 317, 321-324, 327, 329-334, 336, 337, 340, 347, 350, 352, 353, 355-358, 369-382, 394, 395, 428, 437, 467-474, 477, 478, 486, 487, 492, 493, 495, 497, 498, 525, 527-531 biotin autotroph 414 nutrition of the 382-384 types 361-363, 367-368 embryo axis 367, 369, 374, 411, 412, 416, 417, 419, 424, 431, 436-438, 444, 445, 450, 452, 461, 464, 529 embryo culture 329, 331, 384-394, 416-417, 420, 432, 446, 451-453 embryo development (see embryogenesis) embryo-lethal genes, embryo-lethal mutants 398, 413-417, 420 embryo mother cell (see embryoid mother cell) embryo pattern mutants 417-422 embryo sac 2-6, 9,152,156,159,162-164,166-178,181, 209, 217, 252, 253, 257, 293, 297, 299, 301, 304-309, 311, 313-317, 321, 323, 325-330, 361, 376, 381-384, 389, 397, 399, 407, 527 culture of 392-393 isolation of 172-173 embryo-specific genes, embryo-specific proteins 9, 404, 408, 425, 435 EMBRYO-SPECIFIC (EMB) locus, emb mutant 417, 418 embryo storage lipids genes of 462-464 origin of 443 synthesis of 445-446 transcripts of 450-451 embryo storage proteins genes of 451-462 mRNA changes associated with 446-450 origin of 441-443 regulation of synthesis of 443-445, 451-453 embryo transformation 527-531 embryogenesis (see also pollen embryogenesis, somatic embryogenesis) 6, 7, 9,10,13,14,173,176, 244, 314, 322, 325, 357, 358, 362, 379, 384, 386-388, 393, 394, 395, 467, 501, 525, 527 cell lineage during 358, 365, 368-370 cellular aspects of 371-374 comparative 363-367 gene expression patterns during 404-413 genes involved in 413-422 genetic control of 395—404 tissue differentiation during 374-376 embryogenic callus (see embryogenic cells) embryogenic cells 3,10,13,14, 401, 469, 471, 472, 474-480, 482, 484, 485, 487, 489, 491^93, 499, 525, 526, 528, 530, 531 embryogenic pollen 10, 501-504, 506, 508, 509, 515-520, 528 embryogenic pollen grains (see embryogenic pollen)
675 embryoid (see also pollen embryoid) 3, 4,10, 467-484, 486-512, 514-518, 520-522, 525, 528, 530, 531 embryoid mother cell 475-477, 479, 511, 514, 516 EMBRYOLESS (EML-1) gene, eml-1 mutant 417 embryology experimental 14 molecular 13,14 embryonic axis (see embryo axis) embryonic genes, embryonic proteins 487-490, 522 endo-p-l,4-glucanase 198 endochitinase 116,198, 484, 486 endocytosis 242 endomitosis 36,177,185 endopolyploid (see endopolyploidy) endopolyploidy 504, 512 endoreduplication 6,168,170,185, 326, 327, 332, 356, 409 in embryo cells 373-374 in suspensor cells 379-380 in tapetal cells 31, 36 endosperm 2, 5-7,13,14, 40,104,163,168,170,171,177, 253, 269, 293, 311, 314, 315, 321-356, 360, 367, 368, 376, 378, 379, 384, 386, 389, 390, 393, 396^01, 415-417, 419, 425, 437, 454, 458, 460, 464, 470, 477, 497, 527, 529, 530 cellular 6, 322, 323, 325, 328, 329, 333, 337 chemical composition of 329 development 322-326 free-nuclear, nuclear 6, 322, 323-325, 328, 329, 333 haustoria 328 helobial 322, 325, 328 mutants 92, 332-336 role in embryo nutrition 382-383 storage reserves of 9,10, 40, 321-323, 325, 327, 331-356, 440, 441 endosperm balance number 330-331 endosperm box 349-351, 353, 354 endosperm culture 342, 345 endosperm gene expression 333-334 endosperm nucleus 2, 6,168, 321-323, 325, 326, 328, 330, 333, 356, 382, 399, 527 endothecial cells (see endothecium) endothecium 27-31, 60,112,129 endothelium 154,163,175, 330 Endymion non-scriptus 81, 88, 214, 236 enoyl-ACP-reductase 451, 462 enoyl-coenzyme-A (coA) hydratase 412 5-enolpyruvylshikimate-3-phosphate synthase 116 Ephedra sp. 311 E. trifurca 309, 311 epiblast 366, 367 epicotyl 367, 368 Epidendrum scutella 164,166,169,170, 307, 359 Epilobium sp. 80 male sterility in 150 E. angustifolium 229 E. hirsutum 150 E. montanum 150 E. palustre 160, 307 epiphysis 363 episomal DNA143
676 epithelium 477 Eranthis hiemalis 398 Eriocaulon robusto-brownianum 368 E. xeranthemum 368 Eruca saliva 157, 257, 379, 391 Erwinia sp.109 £. chrysanthemi 108 Erysimum capitulum 271 Escherichia coli 55, 141,142, 278, 281, 290, 345, 354 Eschscholzia californica 314 esterase 34, 70, 72, 93, 136,194, 201, 205, 219, 247, 259, 479, 487, 520 ethrel 128,135, 507, 510 ethylene 11, 207, 482, 492, 507 ethylene-response-binding protein 156 ettin (ett) mutant 192 eucalypt (see Eucalyptus rhodantha) Eucalyptus globulus 529 E. rhodantha 211 Euphorbia dulcis 81, 298 E. helioscopia 169, 382 exine 2, 5, 8, 31, 33-35, 39, 61-63, 70-72, 89, 91,117,118, 121,126,128,129, 201, 203, 204, 218, 246, 247, 249, 257, 263, 264, 501, 507, 510, 511, 515 inclusions of 68-69 proteins of 34, 73-74 structure and formation of 64-68 exocytosis 188,189, 242, 476 exotoxin-A 122 extensin 102, 198, 200, 232 extensinlike protein (see extensin) extracellular proteins in somatic embryogenesis 483-486 fackel (fk) mutant 419, 420 Fagus sylvatica 442 Farsetia hamiltonii 157 fasciated mutant 153-154 FASS (FS) gene,/s mutant 419-421 Fe2+/ascorbate-dependent oxidase 198 female gametogenesis (see megagametogenesis) female gametophyte (see embryo sac) female germ unit 2 female sporogenesis (see megasporogenesis) ferritin 185 fertility restorer genes 122,138-140,142,143,146-149 fertilization 2, 3, 6,13, 14, 75, 82,121,162,163,166-170, 172-174,176, 181, 197, 198, 244, 245, 252, 254, 257, 269, 271, 293, 294, 298-300, 303, 305-308, 311-317, 322, 329, 332, 333, 357-359, 361, 369, 382, 383, 396, 398, 399, 407, 408, 413, 414, 498, 526, 527 fertilization-independent endosperm (fie) mutant 322 Festuca arundinacea 530 F. rubra 530 Feulgen reaction 5, 36,173, 326, 379 fiddlehead (fdh) mutant 202 filament 2,17, 18, 25, 31, 470 filiform apparatus 5,166-168,172,175, 304-307, 314, 360 filter hybridization 12
Index first haploid microspore mitosis (see first haploid mitosis) first haploid mitosis 30, 36, 39, 61, 75, 86, 89, 92, 94, 95, 97,100-102,105-107,110,112, 114, 311, 506, 510 514-516, 519, 522 DNA synthesis during 76-77 ultrastructural changes during 78-82 first haploid pollen mitosis (see first haploid mitosis) FITC (see fluorescein isothiocyanate) flax (see Linum usitatissimum) FLORAL BINDING PROTEIN (FBP) gene 154 floral meristem 2, 4, 7, 8,17-25, 28, 58, 60,152-154 floral iflo) mutant 20 floret (see flower) floury (fl) mutant 332, 344, 346, 416 flower 1, 2, 4, 5, 7, 8, 17-25, 31, 37, 40, 48, 56-60, 63, 68, 104, 120-123,128,132-134, 147,149, 151-154, 166,181,182,188,190-192,194, 196-198, 200, 201, 204, 207, 208, 244-247, 249, 251-253, 258, 259, 261-264, 266, 268, 272, 283, 287, 288, 292, 305, 306, 313, 317, 398, 407, 503, 508, 510, 526, 527 flowering plants 1, 4, 6, 10, 13, 42, 75,152, 245, 525 fluridone 400, 432, 452 fluorescein isothiocyanate (FITC) 72,101, 231 fluphenazine 217 Forsythia intermedia 188, 247, 251 Fragaria sp. 503 Freesia sp. 503 Fritillaria imperialis 80 fruit 1, 2, 6 ripening 10,11 fruit fly (see Drosophila melanogaster) functional megaspore 157-163, 173,177 fungal endopolygalacturonase 379 fungal toxins effect on mitochondria 138,141,142 funiculus 154, 175,176, 304, 328, 384, 428 Fusarium sp. 71 FUSCA (FUS) gene,fus mutant 434 G-box 454, 455 G-box/C-box 455, 463 Gagea lutea 88 galactomannan 334 P-galactosidase 92,104 Galanthus nivalis 241 Galium mollugo 87, 298 gametocides 128,132,135 gametoclonal variation 504 Gasteria sp. 253 G. verrucosa 34, 35, 45, 46, 66, 80,156, 157,160,170-172, 192, 206, 217, 304 Gaudinia fragilis 256 GC box (see G-box/C-box) gene action 7,10, 67, 90-92, 94, 97, 100,102,104, 106, 111, 112,114-117,120,121,123,130,133-135,137,151, 159, 223-224, 243, 248, 249, 257, 258, 298, 395, 415, 420, 422, 443, 486, 487, 489, 492, 497, 514
Index during anther dehiscence 129 during endosperm ontogeny 331-334 during meiosis 125-128 in radiation-induced mutants 397-399 in spontaneous mutants 399-404 in wide hybrids 396-397 postmeiotic 128-129 gene activation (see gene action) gene activity (see gene action) gene-cytoplasmic male steriles 121,122 gene expression 7-12,14, 56, 62, 84, 90-92, 94,111-115, 117-120,153,154,197, 221, 225, 226, 228, 245, 253, 254, 259, 260, 271, 276-278, 280, 322, 334, 340, 354-356, 363, 364, 367, 369, 374, 392, 395, 440, 471, 487, 500, 516 during anther development 28-30, 36-41 during embryo storage protein synthesis 443-453 during embryogenesis 404-413, 422-426 during endosperm storage protein synthesis 342-350 during female gametogenesis 173-174 during pollen development 104-108 during pollen embryogenesis 520-521 during somatic embryogenesis 492-498 in self-incompatible plants 281-284, 287-288, 290 in the floral meristem 20-25 in the tapetum 36-41 gene transfer 10,12 General Control Nonderepressible motif (GCN4) 349, 353 generative cell 2, 3, 5, 6, 61, 62, 73, 75, 78-86, 91, 94-97, 99-101,104,106,107,112,114,119,181, 212, 214, 215, 217, 221, 222, 224, 229, 235 238, 240, 241, 295, 296, 298, 299, 301, 307, 311, 312, 399, 507, 510, 516-520, 521 division of 87-89 embryogenic division of 511-515 isolation of 89 nuclear pores in 86-87 generative pole microtubule system 78-79 geneticin resistance 530 genie male steriles 121,122,124,125,131,134-136 genomic library 12, 28,107,109, 277, 402 Gentiana acaulis 35 Gerbera sp. 503 G. hybrida 200 G. jamesonii 300, 317 germin 486 germless mutant 415 Gibasis karwinskyana 35 G. venustula 35, 65 gibberellic acid (GA) 18,19, 58,133,134,137, 306, 329, 340, 381, 385, 386, 391, 393, 433, 481, 482 gibberellin (see gibberellic acid) Ginkgo biloba 458, 504 ginseng (see Panax ginseng) Gladiolus sp. 503 Gladiolus gandavensis 70,185,187,196, 201, 205, 216, 300, 301
677 gliadin 337, 338, 342, 348, 349, 351, 353, 355 GLOBOSA (GLO) gene, glo mutant 24, 25,123 globulin-like protein 348 globulins, globulin genes 336, 337, 339, 341, 342, 350, 352, 355, 400, 402, 407, 441, 444, 450, 457, 458-460 a-globulins (see globulins) glucan hydrolases 259 (3-glucan synthetase 234 (l,3)-p-glucan (see also callose) 35, 232, 234, 242 (l,3)-p-glucanase (see also callase) 40,198 glucose dehydrogenase 430 glucose-6-phosphatase 194 p-glucuronidase (GUS), GUS gene 12,13, 28, 30, 39-41, 56,111-117,132, 227, 282, 283, 284, 289, 352, 354, 369, 401, 423, 426, 429, 452, 454, 455, 459-461, 463, 464, 479, 493, 496, 497, 520, 526-529, 531 glutamate dehydrogenase 259 glutamic dehydrogenase 487 glutamic-oxaloacetic transaminase 92, 93 glutamine synthetase 117, 227 glutathione 48, 77 glutelin 336, 337, 339, 341, 342, 344, 350, 351, 355 glutenin 337, 348, 349, 353, 354, 529 glyceraldehyde-phosphate dehydrogenase, glyceraldehyde-phosphate dehydrogenase gene 77, 219, 334 Glycine max (see also soybean) 56,127,128,156,160,164, 166,167,169,187,193, 305, 325, 358, 359, 373, 378, 382, 383, 387, 424, 441, 442, 446, 463, 531 G. soja 456 glycinin, glycinin gene 374, 403, 441, 444, 445, 448, 449, 456, 457 glycocalyx 65 glycosylrransferases 442 Gnetum sp. 311 Gnetutn gnemon 309 GNOM (GN) gene, gn mutant, GN protein 419, 420, 422 Gossypium sp. 502 G. anomalum 122 G. hirsutum (see also cotton) 45, 82, 83,105,109,122,161, 163,166,169,183,193, 206, 213, 294, 305, 325, 358, 359, 361, 369, 372, 373, 387, 412, 424, 428, 445, 446, 463, 531 G. thurberi 122 grape 482, 484, 485 GTP-binding protein (G-protein) 384, 492 gurke (gu) mutant 419, 420 GUS (see P-glucuronidase) gymnosperms 2,14, 33,108, 222, 226, 228, 309, 311, 337, 363, 407, 467, 481, 494, 495 pollen embryogenesis in 504 somatic embryogenesis in 470 transformation in 531 gynoecium 19,153, 207 gynogenesis 317 Hacquetia epipactis 188 Haemanthus katherinae 80, 85, 325, 326, 383 haploid embryogenesis 500
678 haploid microspore mitosis (see first haploid mitosis) haploid mitosis (see first haploid mitosis) Haynaldia villosa 338 hazel (see Corylus avellana) heat-shock proteins (HSP), HSP genes 56, 102-104,107, 108,117, 200, 228, 346, 411, 412, 423, 445, 494, 495, 498, 518-520, 522 Hebe salicifolia 127 H. townsoni 126 Hela cells 352 helianthinin, helianthinin gene 441, 450, 460, 461 Helianthus sp. 502, 503 H. annuiis (see also sunflower) 30, 65, 66, 68, 72, 104, 111, 128,130,156,164, 166, 169, 170, 172, 205, 306, 307, 325, 358, 359, 371, 412, 423, 441, 445, 452, 453, 460, 463 Helleborus foetidus 33, 34, 36, 65, 68, 78, 80, 214, 241, 298 Hemerocallis sp. 253 7-7. fulva 208, 219, 301 7-7. minor 301 hemp 68,126 henbane (see Hyoscyamus niger) heterostyly 245, 248, 251 Hevea sp. 502 Hevea brasiliensis 126, 482, 491 hexokinase 334 hexose phosphate isomerase 334 Hibiscus aculeatus 330, 397 H. costatus 330, 358, 397 H. furcellatus 330, 358, 397 H. rosa-sinensis 72 Hieracium 177 Hippeastrum belladonna 53, 80,100 H. vittatum 85, 298, 301 Hippuris vulgaris 67 histone 489 H2 100,101, H3 gene 28, 29, 37,174, 424, 429, 495 H4 gene 424 of pollen 100,101 synthesis during meiosis 53 homeobox domain (see homeobox gene) homeobox gene 367, 423, 495, 496 homeotic mutants 7, 8,123,153 in Antirrhinum majus 24-25 in Arabidopsis 19-24 hordein, hordein gene 337, 341, 342, 347, 351, 352, 355, 458 hordenin 337 Hordeum bulbosum 322, 373, 408 H. distichum 170, 373, 408, 412, 512 H. vulgare (see also barley) 86, 92,128,160,164,170,171, 185,294,316, 322,324, 337, 343, 348,358, 371-373, 387, 393, 396-398, 408, 463, 505, 512, 513 horse chestnut (see Aesculus woerlitzensis) Hosta sp. 80 Hyacinthoides non-scripta 45 Hyacinthus orientalis 84, 86, 88, 95, 99, 214, 241 hydrolases from pollen 219, 224
Index 3-hydroxy-3-methylglutaryI coenzyme A reductase (HMC) gene 116,197 hydroxyacyl-coA dehydrogenase 412 hydroxyproline 102,194, 227, 423, 424 hydroxyproline-rich protein (see hydroxyproline) hygromycin resistance 527, 530 Hymenocallis occidentalis 80 Hyoscyamus sp. 502 H. niger 29, 37, 38, 80, 95, 96, 99,100, 222, 502, 506-509, 511, 512, 514, 516, 517, 519 Hyphaene natalensis 329 hypocotyl 2,13, 362, 363, 365-370, 372, 375, 385, 414, 419, 426, 450 hypophysis 363, 364, 366, 369, 371, 376, 420, 421, 468, 470-472, 474, 475, 477, 483 hypoploids 92-94 hypostase 175, 382 Iberis amara 34 7. semperflorens 72 7. sempervirens 72, 74, 264 immunoglobulin heavy-chain-binding protein (BiP) 337, 346 Impatiens balsamina 84 7. glandulifera 80, 309 7. palmata 191 7. roylei 328 7. sultani 45, 126 7. walleriana 80, 235 in situ accumulation (see in situ hybridization) in situ hybridization 11, 20-22, 24, 26, 28-30, 37, 40, 44, 95,112,146,158,173,174,197, 226, 263, 281, 288, 309, 344, 359, 363, 379, 414, 423, 424, 429, 437, 446, 449, 493, 494, 506 in situ localization (see in situ hybridization) incongruity 271 indeterminate gametophyte (ig) mutant 331 indoleacetic acid (IAA) 18,133, 208, 259, 306, 364, 381, 385, 390, 393, 468, 474, 480, 481, 497, 501, 507 indolebutyric acid (IBA) 507 integument 154,156,161,167,172,174-177, 304, 323, 328, 330, 361, 382, 383 integumentary tapetum (see endothelium) intermediate filaments 78 interspecific incompatibility 74 intine 61, 63, 67, 74, 80, 83, 84,109,110,117, 211, 218, 220, 264, 513, 515 development of 69-70 proteins of 34, 71-73 intraovarian pollination 313-314 intraspecific incompatibility 74 invertase 219, 224, 332 in vitro fertilization 300, 315-317 in vitro translation (see cell-free translation) ionophore A23187 239, 507 Ipomoea sp. 503 7. purpurea 67, 378 7. trifida 291 7ns pallida 130
Index I. pseudacorus 235, 237 isocitrate lyase (ICL), ICL gene 107,116, 412, 422, 423 isoflavone reductase gene 200 N6(A-isopentenyl) adenine (2iP) 471, 481, 507 isoperoxidase 487 isopropyl-N-phenylcarbamate 85 isozymes 92, 93,136, 333-335, 372, 412, 446, 487-488, 520 Jacranda sp. 502 Japanese cedar (see Cryptomeria japonica) Japanese pear (see Pyrus serotina) ]asione montana 169, 358, 359 jasmonate (see jasmonic acid) jasmonic acid 135, 432, 452, 493, 520 Juglans regia 329, 531
for embryo growth 391 in pollen 225, 239 traffic in embryoids 478 kaempferol 190,191, 204 kafirin 337, 355 kanamycin resistance 526, 527, 531 Kentucky blue grass (see Poa pratensis) kernel mutants 331-333, 344 KEULE (KEU) gene 419 kinesin 238 kinetin 18,19, 58, 59,134,153, 386, 391, 393, 471^73, 501, 507 KNOLLE (KN) gene 419, 420, 422 KNOPF (KNF) gene 419 KNOTTED-1 (KN-1) gene 367, 420, 495 Kunitz trypsin inhibitor (KTI), KTI gene 108, 363-364, 375, 403, 404, 437 lactic dehydrogenase 37 Lactuca saliva (see also lettuce) 317, 362, 373 Lagenaria vulgaris 157 Lagerstroemia speciosa 358 Larix sp. 470 Larix X eurolepis 477 late embryogenesis abundant (LEA) genes, LEA proteins 9,116, 347, 388, 402, 414, 426, 434, 436, 453, 489, 492-494 ABA effects on 431^32 from embryos of dicots 427—429 from embryos of monocots 429-431 Lathyrus odomtus 125, 126 L. sativus 531 LEA genes, LEA proteins (see late embryogenesis abundant) LEAFY COTYLEDON (LEC) gene, lee mutant 434 LEAFY (LFY) gene, Ify mutant 23, 24 lectin, lectin gene 187, 201, 220, 265, 403, 404, 435-438, 441^43, 462 Ledebouria socialis 35, 68 Ledum groenlandicum 358 legumin, legumin gene 381, 383, 441, 443, 444, 446-452, 457-459 legumin-box 457-460, 462
679 leguminlike proteins (see legumin) lemon (see Citrus limon) Lens culinare 28 leptotene protein (L-protein) 49 lettuce 317, 488 leucine aminopeptidase 198, 219 leucine zipper DNA-binding protein (bZIP) (see leucine zipper protein) leucine zipper protein 56, 345, 346, 350, 354, 429, 430, 431, 455 LEUN1G (LUG) gene, lug mutant 20, 21 Lilium hybrid 55 Lilium sp. 37, 44, 48, 50, 52, 54, 56, 63, 64, 67, 68, 77, 88, 253, 302 L. auratum 219, 236, 302 L. candidum 53, 58, 80, 95,156,157,160 L. davidii 185,190 L. formosanum 399 L. henryi 36, 37, 45, 47, 48, 51, 66, 67, 96,101 L. hybrida 35, 45 L. japonicum 302 L. leucanthum 302 L. longiflorum (see also lily) 25, 34, 35-37, 43-45, 47-49, 51-54, 56, 57, 59, 64-70, 75-77, 80, 89, 94-96, 98,100-102,104, 111, 114,117,156,159,160, 168,190,192,195,196, 214-217, 221, 223, 225, 227-230, 232-240, 242, 253, 255, 259, 260, 264, 265, 268, 296, 302, 306, 315 L. martagon 80 L. regale 169,185,190,195,196, 228, 302 L. rubrum 241 L. speciosum 48-51, 53, 54 L. superbum 302 L. tigrinum 48 lily 25, 26, 37, 44, 49-52, 54-56, 76, 97, 111, 210, 211, 214, 218, 220-222, 224, 225, 233, 234, 239, 240, 303, 317 Limnophyton obtusifolium 361 Limonium sp. 249 L. vulgare 250 Linaria bipartita 328 Linum sp. 246, 247 L. austriacum 331 L. cateri 67 L. catharticum 164,169, 359 L. grandiflorum 69,182,187, 218, 246, 247, 249-251 L. perenne 331 L. pubescens 246 L. usitatissimum 66,131,164,166,169, 305, 307, 359, 387, 390, 391 lipid body membrane protein 492 lipid transfer protein, lipid transfer protein gene 10, 39, 40,197, 200, 352, 355, 414, 425, 434, 484, 486 lipoxygenase 137, 446, 452 Litchi sp. 502 Lobelia erinus 81, 224 Lolium perenne (see also ryegrass) 71, 72, 80, 81, 300, 530 mitochondrial transcripts of 150 Lonicera japonica 45
600 Lotus corniculatus 259 transgenic 117, 227 L-phosphinothricin (L-PPT), L-PPT gene 528-530 luciferase gene 352, 436 Luculia gratissima 250 Luffa sp. 502 L. cylindrica 237, 510, 512 L echinata 512 Lupinus sp. 481 L. albus 442 L, angustifolius 446 L. arboreus 426 L. luteus 329, 389, 411 L. mutabilis 127,129 Lychnis alba 205, 220 Lycium barbarum 508 L. chinense 508 Lycopersicon sp. 270, 502 L. esculentum (see also tomato) 18, 33, 36, 43, 45, 92,122, 128-130,150,153,185,193, 270, 412, 505 anther cDNA clones of 30,104 L. peruvianum 153, 154,188,189,192,194, 213, 229, 235, 255, 261, 262 267, 270, 289, 290, 329, 330, 505 L. pimpinellifolium 329, 330 Lythrum sp. 248 L. junceum 247, 248, 250 L. salicaria 250 Macadamia integrifolia 169 macerozyme 300 MADS protein, MADS-box gene 24, 25,154, 425 Magnolia denudata 45, 47 M. salicifolia 458 M. soulangeana 45 M. tripetala 45, 47 maize 19, 25, 28, 30, 36, 37, 40, 47, 76, 91, 92, 94, 97-99, 105-110,112,114-116,122-128,131,133-136, 147,151,154,159,171-174, 216, 218, 219, 226-228, 232, 278, 299-302, 304, 309, 314-317, 323, 324, 326, 327, 329-336, 338, 340, 341-346, 348, 351-353, 355, 367, 369, 385, 386, 389, 392, 399-402, 409-411, 420, 423-426, 428-433, 445, 450-452, 460, 463, 476, 477, 486-488, 492, 495, 503, 505, 508, 511, 526-530 cms-C 122,137-140,143 cms-S 122,137-140,142-144 cms-T 122,136-143 embryo-lethal mutants in 415-417 storage protein gene expression in 342-347 transgenic 117, 426, 530 zein genes of 350-352 malate dehydrogenase 219, 412 malate synthase (MS), MS gene 107,116, 412, 422, 423 male gametogenesis (see microgametogenesis) male gametophyte 2,14, 60, 68, 89,178, 265, 283, 293, 399, 526 male germ unit 2, 296-299, 317 male sporogenesis (see microsporogenesis) male sterile lines (see male sterility)
Index male sterile mutants (see male sterility) male steriles (see male sterility) male sterility 8, 41,120-132, 244 amino acid changes associated with 135-136 effects of photoperiod on 133 effects of temperature on 133 hormonal effects on 133-135 role of mitochondrial genome in 8, 137-151 role of mitochondrial proteins in 140-142, 146 protein changes associated with 136 Malus sp. 503 M. communis 193 M. domestica (see also apple) 223, 225, 231, 234, 235, 265, 268 Malvaviscus arboreus 70, 72 mandelnitrile lyase 412, 413 Manihot esculenta 127, 531 mannopine synthase gene 117 Matteuccia struthiopteris 441 matteuccin 441 medicagin 441, 449 Medicago sativa (see also alfalfa) 80, 87,109, 131, 158,174, 195, 299, 312, 378, 383, 387, 388, 397, 441, 445, 449, 452, 475, 477, 479, 482, 491, 494, 495 somatic embryogenesis in 472 M. scutellata 378, 397 megagametogenesis 7, 9,152,154,155,162,172-174, 177, 178, 293, 525 megagametophyte (see also embryo sac) 152,154,158, 162, 168, 170, 172, 173, 177, 178, 407 nutrition of 174-176 megagametophytic tissue 450 megaspore 2,158-163,176 megaspore mother cell (see megasporocyte) megasporocyte 2,156-162,174,177 megasporogenesis 5,13,152, 154,155,157-160,162,174, 177,178 meiosis 2, 8, 61-67, 76, 90-92, 97,105-107,123-128, 130, 131,133-136,158-160,162,174,177, 257, 284, 478, 513, 527 endonuclease activity during 50 in microsporocytes 17, 28, 29, 32-60 molecular biology of 47-57 organelle dedifferentiation during 44-45,159-160 meiosis-specific genes 55-57 meiotic division (see meiosis) meiotic histone 53 meiotic mutants 47, 159 Melandrium album 314 M. rubrum 314 meiotin-1 53-54 melon (see Cucumis melo) Mercurialis annua 134 mericarp 471 meromyosin 235 mesogamy 304 metallothionein-like protein 411 methomyl 138,141,142 6-methylaminopurine 386
Index methylglyoxal bis(guanylhydrazone) 491 6-methylpurine 260 5-methyltryptophan -resistant cells 480-481, 497 MICKEY (MIC) gene 419 microfilaments (see also actin) 45, 47, 78, 79, 83, 85, 86, 107,127, 128,157, 172, 214-216, 229, 234-237, 241, 242, 295, 296, 307, 326, 515 microgametogenesis 7, 8, 62, 89, 90,100,101,119,120, 293, 525 microinjection 3,13, 526, 528 microprojectile bombardment 10,13, 112,117, 227, 340, 352,525,526, 528-531 micropylar end (see micropyle) micropyle 6, 154,156, 160-163, 165-168,172-176, 304, 306, 312, 325, 328, 330, 359, 361, 364-366, 368, 375, 377, 380, 382-384, 469 microsporangium 17, 20, 28, 31-34, 42, 62, 67, 68, 72, 77, 83, 92,123-125,127-131, 283, 501, 508, 510 microspore 2,17, 26, 28, 31, 32, 34, 35, 38-42, 44, 45, 47, 48, 50, 59-72, 75-81, 86, 89, 92, 94, 95, 97, 99,102, 104,106,107,110,112,114-116,123,125-132, 134,136,137,146, 159, 257, 283, 284, 301, 392, 500, 504, 506, 509, 510, 512, 516, 518-521, 526, 528 microspore mother cell (see microsporocyte) microsporocyte 2,17, 26-29, 32-39, 41-67, 75, 76, 90-92, 97, 104,106, 107,123-128,130, 131,133-137,146, 159, 257, 284 culture of 58 microsporogenesis 5,13,17, 25-27, 31, 32, 34, 37, 41, 58, 59,104,107,108, 112,120-123,128,129,132,133, 151,159 ultrastructural changes during 42-47 microtubules (see also tubulin) 32, 34, 35, 45^7, 66, 69, 78-80, 83-85, 87-89,107,127,128, 156,157, 171, 172, 216, 217, 229, 235-238, 241, 294, 295, 297, 306, 307, 325, 326, 358, 361, 477, 515 preprophase band of 45, 78, 79, 158, 325, 361 middle layers 27, 28, 31, 32, 39,124,137 miniature seed (mn) mutant 332 minichromosome maintenance (MCM-1) gene 20, 24 Mirabilis jalapa 81, 309 mitochondria in male sterility 137-151 inheritance of 80-81 mitomycin-C 50 mitotic cyclin gene 426 molecular hybridization 11, 374, 379 cDNA-poly(A)-RNA hybridization 98-99, 405, 407 cDNA-RNA reassociation 447 DNA-DNA hybridization 48 DNA-rRNA hybridization 380, 487 RNA-DNA hybridization 104, 405-406 solution hybridization 453 monensin 231, 338, 443 Monocotyledonae (see monocots) monocotyledons (see monocots) monocots 1, 9, 10, 32, 52, 61,114,183,194, 325, 327, 353,
681 362-364, 366, 367, 393, 415, 429, 470, 502, 503, 525, 528, 530 monopteros (mp) mutant 419-421 Monotropa hypopiiys 359 M. uniflora 80, 358, 359, 362, 367, 368 Monotropsis odorata 207 morphactin 123 mRNA (see also poly(A)-RNA, stored mRNA) 6, 8, 9-12, 20, 22, 23, 26, 29, 30, 37, 39, 40, 49, 52-56, 95, 97-99, 102,104,106,107,109-111,117, 118,132, 145,173,174,198, 200, 206, 221, 224-226, 243, 263, 273, 278, 279, 284, 285, 288, 334, 338, 339, 342-350, 352, 359, 376, 389, 402^09, 411, 412, 414, 422-433, 435-437, 442, 446^54, 456, 457, 479, 487, 489, 491^95, 509, 515, 517, 518, 520, 521 mucronate (me) mutant 344, 346 multigene family 105, 276, 277, 280, 285, 350, 351, 353, 355, 427, 431, 437, 450, 453, 454, 456, 458, 462 Musa sp. 530 Muscari racemosum 86 mutase 334 mutator (mu) transposon 333, 415 MYB motif 352 MYC motif 352 myosin 83, 215, 237, 238, 240, 241 myosinlike proteins (see myosin) Myosurus minimus 164,173 myrosinase, myrosinase gene 438 NADH dehydrogenase 520 NADH dehydrogenase subunit-3 (NADS) gene 145 NADPH-dependent aldose reductase 430 Najas marina 240 naphthaleneacetic acid (NAA) 58, 386, 420, 451, 472, 473, 481, 507 napin, napin gene 122, 363, 441, 445, 449-452, 459, 460, 495, 520 narbonin 449, 460 Narcissus biflorus 510 N. pseudonarcissus 214, 215, 217, 220, 235, 241, 302, 303 neomycin phosphotransferase (NPTII) 527, 529-531 Nerium oleander 375 Nicotiana sp. 259, 272, 273, 286, 288, 292, 330, 362, 502 N. alata 19, 81,191,192, 194,195,197-199, 210, 214, 219, 223, 225, 232, 234, 235, 237, 238, 245, 259, 261, 263, 267, 281, 282, 284-289, 302 N. debneyi 19 N. forgetiana 245 N. gossei 227, 526 N. glutinosa 114, 117, 508 N. langsdorfii 19, 288 N. plumbaginifolia 116, 288, 354, 489, 493 transgenic 458 N. rustica 114,117, 314, 359, 398, 503 N. sylvestris 99,188,192, 206, 207, 225, 234, 236, 237, 511, 512 N. tabacum (see also tobacco) 3,18,19, 52, 63, 75, 80, 81, 86-88, 95, 97, 102, 104, 105, 108, 114, 117, 122,
682 153,164,166,167,172,185,187,192,193,195, 198, 200, 210, 214, 215, 217, 219, 223, 225-228, 232, 233, 235-238, 240, 282, 286, 288, 294, 296, 298, 299, 305, 308, 312, 314, 358, 361, 412, 423, 461, 502-504, 506, 508-512, 519, 520, 528 transgenic 283, 458, 459 nicotinamide adenine dinucleotide (NAD +) 138 nifedipine 217, 239 Nigellla damascena 36 nigericin 443 Nitella 238 nitrate reductase, nitrate reductase gene 487, 520 NO APICAL MERISTEM (NAM) gene, nam mutant 420 nodulation (NOD) genes 484 nonembryogenic callus (see nonembryogenic cells) nonembryogenic cells 3, 10, 476, 477, 480, 482, 484, 485, 487, 491, 492, 495, 496 nopaline synthase (NOS) gene 39, 227, 353, 354, 401, 527 norflurazon 63 Northern blot 12, 20, 29, 37, 57,104-107,110,112,114, 140,198, 226, 227, 262, 263, 273, 279, 285, 287, 288, 348, 402, 422, 423, 429, 431, 435, 446, 447, 449, 451, 493, 521 Nothapodytes foetida 328 NPTII (see neomycin phosphotransferase) nucellar cap 382 nucellar cells (see nucellus) nucellar tissues (see nucellus) nucellus 2, 3, 9,154,156-158,161-163,168,170-172,174, 175, 177, 253, 304, 323, 324, 328, 330, 333, 361, 382, 383, 393, 470, 474, 481, 486 nuclear male sterility 121 nuclease-C 81 nucleases, from pollen 219 nucleoside diphosphate kinase 224 nucleoside diphosphate sugar transferase 335 Nymphaea alba 45 oat 32, 326, 337, 339, 340, 342, 350, 355, 385, 386, 530 storage protein gene expression in 350 obturator 304 Oenothera sp. 503 0. biennis 158, 329, 396 O. hookeri 80,128,131 O. lamarckiana 160,164, 329, 396 O. longiflora 303 O. murkata, 329, 396 O. organensis 104-106,108,110, 206, 218, 219, 226, 227, 253, 264, 265 okadaic acid 279 Olea europaea 32, 33, 35, 44, 69,185,193 oleosin, oleosin gene 40, 111, 341, 425, 434, 443, 445, 450-452, 462-464, 489, 492, 520 oligomycin 221, 260 olive (see Olea europaea) oncus (see Zwischenkorper) onion (see Allium cepa) Onoclea sensibilis 427 OPAQUE (OP) gene, op mutant, OP protein 332, 344-347
Index OP-2 MODIFIER gene 346 Ophiocephalus sp. 81, 83 Ophrys lutea 82 orbicules 33-35 orchard grass (see Dactylis glomemta) Orchis maculata 158 ornithine decarboxylase, ornithine decarboxylase gene 136, 491 Ornithogalum caudatum 157,161, 166,185,195, 304, 306 O. virens 45, 88, 224 Orobanche aegyptiaca 367 Orthocarpus sp. 81, 83 Oryza sativa (see also rice) 25,104,125,164,166,169,174, 305, 327, 337, 341, 358, 371, 372, 387, 408, 424, 501, 503 oryzenin 337 oryzin 337 osmotin gene 116, 227 ovary 2, 7,14,104,152,181,192,196, 205, 207-209, 250, 251, 253, 257-259, 268, 270, 282-284, 287, 301-304, 313-315, 317, 332, 376, 408, 471 culture of 392-393 ovulata (ovu) mutant 24 ovule 2, 4, 7, 9,13,14,18,104,122,133,152, 153,159,161, 163, 172-177,181,191,192, 207-209, 251, 253, 257, 282, 293, 303-307, 313-317, 322, 323, 328, 330-333, 348, 359, 365, 376, 379, 381, 382, 384, 387-389, 395-398, 402, 407, 408, 428, 432, 467, 474, 488 ATPase activity in 176 culture of 392-393 development of 154-158 ovule-mediated transformation 526-527 Oxalis sp. 68, 248 P-particles 230, 233 P-pollen 510 pachytene DNA (P-DNA) 50, 51 pachytene-small-nuclear DNA (psn-DNA) 51 pachytene-small-nuclear RNA (psn-RNA) 51 Paeonia sp. 70, 313, 363, 510 P. anomala 363 P. lactiflora 114,117 P. moutan 363 P. tenuifolia 37, 52, 53, 95, 99 P. vnttmanniana 363 palindrome 114, 426 palindromic sequence (see palindrome) Panax ginseng 527 Papaver sp. 272, 273, 292, 503 P. dubium 87 P. nudicaule 314, 359 P. rhoeas 170, 253, 260, 265, 268, 290, 291, 313, 314 P. somniferum 314, 325, 393, 508 papaya 529 transgenic 529 Paphiopedilum spicerianum 160 parietal cell 5, 26, 27,131,156,157 Parietaria officinalis 83 Parkinsonia aculeata 80
Index parsley 29, 30,197 Partheniutn argentatum 257 parthenogenesis 176, 408 particle acceleration (see microprojectile bombardment) particle bombardment (see microprojectile bombardment) Paspalum orbiculare 304 patatin 31,117 pathogenesis-related proteins 198, 200, 347 pattern-forming genes 417-422 PCR (see polymerase chain reaction) pea 34,117,122,123,126, 200, 383-385, 402, 403, 405, 407, 409, 417, 424, 428, 432, 437, 440, 442^44, 446-449, 451, 453, 458, 459, 481, 489 disease-resistant response proteins of 420 peanut (see Arachis hypogaea) pear (see Pyrus communis) pearl millet 123, 528 pectate lyase 108, 226 pectin esterase 110, 111, 226 pectinase 108,110, 208, 219, 232, 300 Pelargonium sp. 503 P. hortorum 80 P. zonale 80, 82,165 pellicle 74,187-188, 201, 270 Peltophorum pterocarpum 506 Pennisetum americanum (see also pearl millet) 123,125, 126, 473 male sterility in 150 P. ciliare 170 P. glaucum 167, 303 male sterility in 150 P. typhoides 205, 206 P. typhoideum 230 pepper 31,122 perianth 20, 58 pericarp 174, 412 periclinal chimeras 153 peroxidase 93,194,219, 225, 251, 381,482,483,486,487, 520 role in self-incompatibility 259 peroxidase-labeled con-A 279, 485 Persea americana 189,193 petal 2, 7,17-25,104,106,122,123,153,154, 202, 261, 268 Petroselinum crispum (see also parsley) 29,116 Petunia sp. 10, 81,116,117,144-146,151, 204, 267, 273, 288, 289, 292, 420, 502, 516, 526 somatic hybrids in 144,145 transgenic 117,121,145,154, 227 P. axillaris 59, 261, 267, 315 P. hybrida 8, 25, 34, 36, 41, 59,102,106,115,117, 125,130, 135-137,144,145,154,164,166,169,183, 186-192, 206, 218, 219, 223-225, 227, 229, 233-235, 237, 255, 258, 259-261, 263, 265, 267, 288, 289, 296, 298, 299, 303, 306-308, 315, 359, 371, 373, 426, 456, 508, 526 po mutation in 115 transgenic 40,132, 204, 352, 426, 457 P. inflata 105,106, 111, 198, 207, 227, 261, 265, 288-290 transgenic 111, 289
683 P. parodii 136,144,145 P. violacea 314 Phalaenopsis sp. 45, 47,155, 207 P. equestris 78, 79 Phalaris coerulescens 268, 291 P. minor 201 P. tuberosa 70, 75, 203 Pharbitis sp. 503 P. nil 188, 191 phaseolin, phaseolin gene 117,122, 352, 429, 436, 441, 442, 448, 453-456, 462 (5-phaseolin (see phaseolin) Phaseolus sp. 377, 379, 396, 502 P. acutifolius 192, 396 P. atropurpureus 128,130 P. coccineus 377-382, 391, 392, 396, 408, 411, 436 P. htnatus 396 P. vulgaris (see also bean) 30,117,127,146, 325, 326, 352, 372-374, 378-381, 388, 396, 404, 408, 409, 411, 413, 436, 441, 442, 445, 446, 448, 452, 455, 458 Phaseolus vulgaris ABAI-3-like factor (PVALF) gene 455 phenylalanine ammonia lyase (PAL), PAL gene 30,117, 137,197 Phleum nodosum 127 Phoenix dactylifera (see also date palm) 86 phosphoglucoisomerase 92, 93, 219 phosphoglucomutase 93 phosphorylation activity 47,138, 204, 221, 260, 270, 278, 279, 287, 290, 431, 433, 434 photosystem II quinone-binding protein (PSB-A) gene 107, 424 phragmoplast 45, 47, 61, 79, 80, 89, 325, 326, 515 Phyllosticta maydis 138 Physalis ixocarpa 257 phytase 224 phytic acid 224, 234, 258 phytochrome, phytochrome gene 117,133, 426 phytohaemagglutinin (PHA) 220, 404, 436, 437 Picen sp. 470 P. abies 117, 227, 228, 445, 470, 482, 484, 491, 531 P. glauca 442, 445, 446, 450, 477, 484, 494, 531 P. glauca/P. engelmatmii 445, 450, 491 P. marina 477, 531 Picris echioides 166 pigeon pea (see Cajanus cajan) pin-formed (pin-1) mutant 364 pin morph 182,185,197, 218, 246, 247, 249-251 Finns sp. 470 P. banksiana 33, 66, 68, 75 P. caribaea 484, 486 P. densiflora 235, 236 P. monticola 445 P. pinaster 227, 228 P. ponderosa 222 P. radiata 531 P. strobus 407 P. taeda 222, 226, 228 P. thunbergii 116, 235, 236, 411 pistil 2, 7, 19, 192, 200, 202, 245, 251, 252, 257-259, 261,
684
Index
263, 265-269, 271, 272, 277, 279, 281, 283, 285, 510, 513, 514, 516, 521 pollen embryogenesis 4,10,13,14, 317, 357, 500-516, 302, 303, 314, 315 abortion 19 520, 521 pistil-specific genes 197-199 genotypic effects on 505-506 PISTILLATA (PI) gene, pi mutant 20, 21, 23-25,123, 153 pollen embryoid 474, 503, 507, 515, 517, 518, 520, 528, 531 Pisum sp. 502 transformation mediated by 528 P. arvense 373 pollen germination 2, 8, 63, 69, 73, 74, 83, 97,102,108, 109,119,121,188,195, 202-206, 208, 210-217, P. sativum (see also pea) 34, 80,122,127,159,160, 235, 373, 231, 234, 239-241, 245, 247, 249-255, 258, 260, 378, 411, 412, 423, 428, 441, 442, 443, 446, 458, 264, 265, 266, 268, 270, 271, 283, 299, 301, 302, 462, 527 314, 315, 399, 516 placenta 153,154,175, 304, 306, 307, 315, 328, 332 biochemistry of 220-226 placental pollination 315 efflux of metabolites during 217-220 Plantago lanceolata 134 gene expression during 226-228 male sterility in 150 in heterostylous plants 249-250 plasmatubules 206, 207, 234 pollen grain (see pollen) plastids pollen isozymes 92 inheritance of 80-82, 229, 294, 311 plastocyanin gene 200 pollen-mediated transformation 526-527 PLENA (PLE) gene, pie mutant 24,153 pollen mother cell (see microsporocyte) pollen recognition 8,14, 34, 36, 63, 64, 68, 69, 73-75,102, Plumbagella sp. 306 P. micrantha 168, 306, 308 109, 111, 181, 187,188,191, 201-208 Plumbago sp. 306 during cross-incompatibility 269-271 P. capensis 168 during self-incompatibility 251-268, 279-292 P. zeylanka 80,164-166,168, 169,171-173, 294, 296-301, gene for 277-279 Pollen RetroElement Maize (PREM) 109,110 306-308, 310, 312 pollen-specific genes 8,12, 30, 90,118,119, 226, 227, 232, plumular pole (see plumule) plumule 362, 367, 368, 400 290 Poa annua 68 early 105-106 P. pratensis 118, 385 expression of 105-108, 112 Podocarpus macrophyllus 75 isolation of 104-105 polar fusion nucleus 2,162,163, 293, 298, 299, 309, 311, late 105-106, 111, 117 promoter analysis of 112-117 312, 321, 322, 333 structure of 108-112 polarity 9, 359, 395, 419 of egg 165,173-174 pollen-stigma interaction (see pollen recognition) of embryoids 478-479, 515 pollen tube 5, 7-9,14, 61, 69, 73-75, 82-89,102,105,108, of embryos 364, 370, 420, 422 109, 119,121,168,181, 185, 187,191,192, of megaspores 160-161 194-198, 200-211, 245, 247, 249-260, 264, 265, of microspore nucleus 78 267-271, 280, 282-284, 286-296, 298-302, of organelles 80 311-315, 399, 408 of pollen 66, 78 calcium gradient in 238-240 of somatic embryos 478-479 cytoplasmic streaming of 234 of zygote 361 cytoskeleton of 234-238 pollen 2-10, 12-14,17, 28-30, 33-41, 57, 59, 61, 62, 65, gene expression during growth of 226-228 growth 240-242 68-75, 78, 80-87, 89-123,125-135,140,142, 146, 151,181,182,185,187,188,191,195,198, growth in heterostylous plants 249-251 201-210, 229-231, 234, 235, 238, 241-261, guidance into embryo sac 302-308 263-272, 276, 277, 279-284, 286-292, 294, 296, structure of 228-231 298-303, 313-315, 332, 333, 396, 399, 400, 413, transformation mediated by 526, 527, 531 415, 500-505, 511, 513-515, 518-521 wall, structure of 231-234 abortion 106, 111, 120,122, 128-132,135,137,150 pollen wall 2, 5, 45, 61, 62, 83, 84, 87, 89, 93,108,110,117, cytoskeleton of 214-217 125, 128,131,132, 136, 203, 515 dimorphism 91, 509 adaptive significance of 73-75 embryogenic competence of 509-510 assembly of 63-70 germination pores (apertures, germ pores, germinaof hypoploids 93 tion sites) of 63-67, 69, 70, 72, 73, 76, 80, 83, 94, proteins of 70-74 211, 214-217, 220, 264 pollenkitt 35, 36, 41, 68, 70, 257 pollen callus 501-512 pollination 4, 8,14, 73-75, 91,108,177,181, 182, 187, 188, pollen culture 191,196-198, 200-209, 217, 219, 225, 244-255, 257-261, 263-265, 268-272, 276, 277, 283, 286, for embryogenic induction 501-504, 506, 508, 509,
Index 287, 299, 303, 305, 306, 313-315, 317, 322, 330, 332, 334, 341, 342, 346, 369, 396, 400, 417, 423, 526, 527, 531 poly(A)-RNA 11,12, 20, 26, 28-30, 37, 40, 49, 50, 52, 55, 57, 97, 98,158,174,197,198, 221-223, 225, 273, 284, 343, 368, 403, 407, 411, 422, 428-430, 446, 493, 494, 521 polyamines 136, 210, 379 in pollen embryogenesis 519-520 in somatic embryogenesis 490-492 polyembryony 311, 315 polygalacturonase 11,108-110,115, 226 Polygonum capitatum 176 P. divarication 163 polymerase chain reaction (PCR) 9,12,13,107,118,174, 277, 279 polynucleotide kinase 50 polynucleotide ligase 50, 77 polyploid (see polyploidy) polyploidy 36,170, 326, 356, 420 polyspermy 316, 317 polytene (see polyteny) polytenic (see polyteny) polyteny 31, 36,168,170, 328, 379-381 Poncirus sp. 502 Pontederia cordata 247, 249, 250 P. rotundifolia 247 P. sagittata 247, 250, 251 ponticulus 304 poplars 74 Populus sp. 270, 317, 503 P. alba (see also poplars) 70, 74, 75, 270 P. deltoides (see also poplars) 70, 74, 75, 92, 270, 295, 298, 305, 311 P. lasiocarpa 188 P. maximowiczii 92 P. nigra 92 P. tremula 67, 68 P. tremuloides 270 P. trichocarpa 270 porogamy 304 postament 175,176 potato 25, 30, 31, 107, 116, 197, 270, 506 transgenic 117, 456 precocious germination 386-389, 406, 428, 432, 434, 451, 452, 473, 474, 482 Primula sp. 250, 502 P. acaidis 251 P. obconica 247, 250 P. officinalis 202 P. sinensis 250 P. veris 250 P. vulgaris 185,193, 247, 250 pro-orbicules 33 pro-Ubisch bodies 32 Proboscidea louisianica 166, 305 proembryo 313, 322, 329, 362, 363, 365-371, 375-377, 383, 384, 386, 393, 397-399, 415, 417-^19, 424, 530 culture of 389-392
685 proembryogenic mass 471, 475, 476, 478-481, 484, 486-488, 492-194 profilin 102,106,109,117-118 prolamins 336-339, 341, 342, 350, 352, 355 prolifera (prl) mutant 174 proline-hydroxyproline-rich proteins 198 promoter analysis 90 of embryo storage protein genes 453-464 of endosperm storage protein genes 350-356 of pollen-specific genes 112-117 of self-incompatibility genes 281-284 prophase chromosome reduction (see reductional grouping) protease 70, 387, 487 protein kinase, protein kinase gene 54, 109, 111, 227, 240, 260, 270, 277-279, 287, 327, 429, 434, 482, 484, 492, 495 protein phosphatase 482 protein receptor kinase (see protein kinase) protein synthesis and metabolism during endosperm development 333-334 during meiosis 53-55 during pollen development 99-104 during pollen embryogenesis 518-520 during pollen germination and pollen tube growth 223-226 following cross- and self-pollinations 260 for vesicle production 231 in developing embryos 411-413 in suspensor 380-381 in tapetum 37 protoplasts 3, 4,10,13,14, 34, 65, 78,125, 345, 353, 401, 402, 435, 454, 455, 470, 475, 477, 478, 499, 525 from embryo sac cells 172 from microspores 301 from microsporocytes 59 from pollen 70, 75, 85, 89,100,101,114, 214-217, 235, 236 fusion of 137,144,148-150 tapetal 34 prunasin hydrolase 412, 413 prunin 450 Prunus amygdalus 450, 451 P. armeniaca 267 P. avium 80,188,193, 229, 230, 235, 267, 268 P. cerasws 267 P. persica 267, 329 P. serotina 412 PSB-A gene (see photosystem II quinone-binding protein) Pseudomonas aeruginosa 122 P. syringae 128 Pseudotsuga sp. 470 P. menziesii 450, 458 Psidium sp. 503 Psophocarpus sp. 502 Pulmonaria affinis 250 pumpkin 110 pyrophosphorylase 334, 335 Pyrus communis 84, 211, 212, 214-217, 219, 232, 235, 236, 241
686 P. serotina 193,194, 268, 290, 291 pyruvate decarboxylase 107 pyruvate orthophosphate dikinase (PPDK) 334, 346
Index
355, 367, 402, 407, 409, 411, 417, 419, 425, 426, 431, 432, 435, 454, 477, 487, 489, 493, 501, 503, 511, 512, 527, 529, 530 anther development in 26-28 quercetin 251 glutelin genes of 355-356 Quercus gambelii 164,166,176, 305, 308, 325, 358-361, 371 male sterility in 148 quiescent center 376, 424, 425 storage protein gene expression in 350 Quinchamalium chilense 175 transgenic 28,117,197, 336, 350, 355, 426, 527 rice tungro bacilliform virus (RTBV) 117,197 Ricinus communis (see also castor bean) 339 RAB gene (see responsive to ABA gene) radicle 365, 367, 368, 372, 385, 386, 424, 435, 436, 451, 478 RNA helicases 111 RNA polymerase 53, 409, 497 radicular pole (see radicle) RNA synthesis and metabolism radish 35, 409, 411, 431, 459 ragweed 72,118 during meiosis 44, 52-53 Rana catesbeiana 501 during pollen development 94-97 R. pipiens 501 during pollen embryogenesis 516-518 Ranunculus macranthus 80 during pollen germination and pollen tube growth R. repens 153, 222-223 R. sceleratus 170, 323, 325, 474, 477 during somatic embryogenesis 487 rapeseed 21,148,151, 427, 440, 445, 459, 462 following cross- and self-pollinations 259 Raphanus sp. 147,148 in developing embryos 408—410 R. caudatus 384 in generative cell 94-95 R. landra 384 in suspensor 379-381 R. raphanistrum 193,205 in tapetum 37 R. sativus (see also radish) 35, 36, 68, 73, 74,147,148,190, in vegetative cell 94-95 205, 254, 261, 264, 384, 428, 446, 449, 450, 459, RNase 41, 51, 70,117,132, 195, 268, 286, 288-292 502 ROL gene (see root loci gene) ogura (ogu) cms line of 147 root 2, 3,13, 28,102,104, 117, 362-367, 369-371, 374-376, reassociation protein ("recA-protein") 54-56 385-387, 397, 398, 400, 414, 417,419-421, 423-425, receptacle (seefloralmeristem) 431, 433, 435, 454, 459, 468-472, 475, 476, 479, reduced endosperm (re) mutant 332 501, 515, 527, 531 reductional grouping 478 root loci (ROL) gene 122, 132, 426, 496, 497 ROOTLESS (RTL) gene 419 responsive to ABA (RAB) gene 402, 427, 428, 431, 493 restitution nucleus 36,177 Rottboellia exaltata 133 restriction fragment length polymorphism (RFLP) 277, rubber (see Hevea brasiliensis) Rubia cordifolia 175 312, 414 RFLP (see restriction fragment length polymorphism) Rubisco (see ribulose-l,5-bisphosphate carboxylase) Rhinanthus minor 328 rugosus (r) locus 402, 403 Rhizobium leguminosarum 484 Rumex acetosa 25 Rhizophora mangle 375, 402 run-off transcription 39, 342, 422, 436, 446, 448, 451, 461 Rhododendron sp. 80, 271, 299, 358 Ruscus aculeatus 191, 202 R./orh(nei207 rutin 251 RY-repeats 457, 463 R. laetum 84, 86, 229, 294-296, 298 R. macgregoriae 84, 294-296, 298, 300 rye 48, 87,159, 203, 260, 329, 338, 340-342, 355, 503, 504, Rho family GTPases 235 527 ryegrass 71,117,118 Rhoeo discolor 52, 53 R. spathacea 35-37, 44, 53 Ribes rubrum 52 S-associated proteins (see S-locus-specific glycoproteins) ribitol dehydrogenase 430 S-family proteins (see S-locus-specific glycoproteins) ribosomal DNA (rDNA) 44,170, 487 S-gene (see sterility gene) ribosomal RNA (rRNA), rRNA gene 41, 44, 52, 174, 222, S-LOCUS ANTHER (SLA) gene 279 223, 286, 373, 379, 380, 408, 409, 487 S-LOCUS RECEPTOR KINASE (SRK) gene 277-281, 291 ribosome S-LOCUS-RELATED (SLR) gene 276, 277, 280, 281, 283, 284, 484 changes during meiosis 44, 52 ribulose-l,5-bisphosphate carboxylase (Rubisco) 107, S-locus-specific glycoproteins (SLG), SLG genes 108,122, 151, 413 261, 262, 264-268, 273-282, 284, 285, 289, 291, large subunit (RBC-L) gene 57,107, 424 484 small subunit (RBC-S) gene 424 S-RNase (see stylar RNase) rice 25, 27, 29, 37, 40, 41, 95,105,109,115-117,123,125, Saccharum bengalense 264 126,133, 227, 317, 324, 326, 329, 334, 336, 39-342, S. officinarum (see also sugarcane) 126
Index safflower (see Carthamus tinctorius) Sagina procumbens 362 Sagittark sagittaefolia 366 Saintpaulia ionantha 176 Salix hiitnilis 68 Sambucus sp. 502 S. nigra 87 Saponaria officinalis 64 Saxifraga granulata 362 Scilla bifolia 170 S. peruviana 81 S. sibirica 169 scutellum 366, 367, 369, 371, 383, 386, 387, 397, 416, 417, 419, 423-425, 430, 431, 435, 436, 445, 473, 477, 527, 529, 530 SDS-PAGE 11,12, 43, 53, 70, 99,100,102,103,140,150, 219, 225, 226, 237, 261, 262, 268, 284, 301, 341, 343, 405, 443, 485, 488, 490, 518 Secale cereale (see also rye) 48, 58, 59,126, 130,185,189, 202, 203, 211, 222, 230, 237, 298, 300, 322, 337, 371, 389, 396 secalin 337, 338, 341, 342, 355 Sechium edule 389 secreted proteins role in somatic embryogenesis 482-486 seed 1-3, 6, 9,14,19,120,121,135,152,176,177, 244, 245, 260, 261, 263, 265, 271, 313-315, 323, 325, 328, 329, 331, 334, 341, 352-354, 356, 367, 368, 373, 382-384, 386-389, 393-399, 402-405, 409, 411-415, 422, 423, 426-429, 432-434, 436, 437, 440-442, 444, 446, 449, 450, 452, 454-456, 458, 459-462, 464, 468, 525 synthetic 3, 499 seed embryo 369, 370, 384-386, 399, 405-407, 482, 484, 489, 490, 492-495, 498, 520, 527-529 selective gene amplification 326-327, 348, 373-374, 380, 446, 486 self-incompatibility 6, 8,14, 74, 75, 121,195,197,198, 201, 244, 245, 248, 249, 251-254, 257-263, 265, 268, 269, 271-273, 276, 277, 279-284, 286-292, 315, 484 acquisition of 261, 263 heteromorphic 245-251 homomorphic 251-269 relation to host-pathogen responses 292 sepal 2, 7,17-22, 24, 25,122,123,153,154, 202 sepaloidea (sep) mutant 24, 123 Sequoia sp. 470 serum response factor (SRF) gene 20, 24 Sesamum indicnm 270 S. mulayanum 270 Setaria httescens 324 sexual reproduction 1,17,176, 201, 208, 293, 531 -SH compounds 47, 48, 77 shed pollen culture 503-504, 506 shikimate dehydrogenase 93 shoot (see shoot apex) shoot apex 2-5, 7,13, 24, 99,104, 364-370, 372, 374, 375, 385, 397, 400, 416, 417, 419, 420, 423-425, 449, 468, 472, 473, 479, 501, 527, 529 shoot apical meristem (see shoot apex)
687 shoot bud (see shoot apex) SHOOT MERISTEMLESS (STM) gene 419, 420 shoot pole (see shoot apex) short integuments (sin) mutant 156,174 shrunken endosperm expressing xenia (sex) mutant 333 shrunken endosperm (se and seg) mutant 333 SHRUNKEN (SH) gene, s/i mutant 332, 335, 336, 401 signal transduction 102, 111, 201, 204, 208, 210, 217, 227, 240, 260, 270, 277, 279, 281, 287, 290, 292, 322, 384, 400, 401, 429, 430, 434, 482, 507 Silene alba 67 S. latifolia 25 S. pendula 64-66, 68 S. schafta 314 S. tatarka 314 Simmondsia chinensis 446 Simocalamus latiflora 506 Sinapis alba 25, 37, 39, 373, 411, 438, 442, 445, 452, 502 slot-blot assay (see dot-blot hybridization) slot-blot hybridization (see dot-blot hybridization) SMALL POLLEN (SP-1) gene 91 Smyrnium perfoliatum 188 snapdragon 8, 25 Solanum sp. 269, 270, 510 S. acaule 150 S. carolinense 507 S. chacoense 80, 267, 270, 289 S. melongena 45 S. nigrum 164,166,167,169 S. phureja 257 S. pinnatisectum 257 S. soukupii 270 S. stentomum 257 S. surattense 510, 512 S. tuberosum (see also potato) 25, 29, 31, 80,126,150,188, 198, 199, 263, 267, 289, 290 S. verrucosum 270 somatic embryo (see somatic embryogenesis) somatic embryo-mediated transformation 530-531 somatic embryogenesis 3,10,13,14, 357, 364, 370, 385, 399, 467-470, 499, 501, 522, 525, 527, 530, 531 biochemistry of 486-492 cell biology and physiology of 475-486 cell lineage during 469 direct 474 from callus 471-474 from protoplasts 475 gene expression during 492-498 recurrent 474-475 transgenic 531 somatoplastic sterility 330 Sophora flavescens 379 sorghum 65, 336, 339-342, 355, 411, 527, 529 Sorghum bicolor (see also sorghum) 65, 68, 103,122,125, 130,133,147, 228, 316, 324, 337, 339 S. vulgare 130,131,135,136 Southern blot 12,13, 39, 105,144, 273, 276, 285, 286, 350, 374, 453, 526-528 Southern hybridization (see Southern blot) soybean 56,117,123,128, 131,193, 227, 323, 364, 374, 375,
688 382, 384, 387, 403-405, 407, 409, 424, 429, 432, 437, 438, 440, 442, 444-449, 452, 453, 456, 457, 462, 481, 488, 489, 495, 496, 529 transgenic 529 soybean embryo factor-3 (SEF-3) 457 Spathoglottis plicata 368
Index of endosperm 350-356 of somatic embryos 494-495 storage protein manipulation 529 storage proteins (see also embryo storage proteins) endosperm - storage reserves of 6, 321-323, 325, 327, 363, 372-374, 381, 383, 384, 388, 399, 400, 402-404, 411, 414, 422, 429, 432-434, 436, 437, 489, 490 of embryos 441-464, 522 of endosperm 334-342 stored mRNA in the egg 407-408 of embryos 411 of pollen grains 97, 221-223 sequence complexity of 97-99
sperm 2, 3, 5, 6, 61, 62, 80-82, 87, 89,100,109,119,152, 163, 166,168,177,178,181, 209, 229, 241, 243, 255, 293-296, 303-305, 307-317, 333, 357, 399, 408, 526 isolation of 299-301 sperm cells (see sperm) sperm dimorphism 296, 297, 299, 309, 317 sperm recognition 311 spikelet culture 503 spinach 164, 300, 312 Striga gesnerioides 367, 368 Spinacia oleracea (see also spinach) 164,166,167, 169,193, stylar RNase {S-RNase) gene, protein 286-290 stylar secretion 193-197 294, 298, 305, 309 Spironema fragrans 185 style 2, 7-9, 91,104,108,129,152,175, 181-183,185,187, Spodoptera frugiperda 454 188,190-202, 204-209, 217, 218, 232, 240, 242, 243, 245-253, 255, 258-265, 267-270, 272, 273, sporogenous cells 5, 26-28, 31, 33, 34, 36, 40, 47, 53, 58, 281-288, 291, 300-305, 308, 313, 314, 527 121-124,128-131,135,136,151,156,157, 526 sporogenous tissues (see sporogenous cells) genes expressed in 197-201 sporopollenin 5, 33, 41, 61, 63, 65, 73, 89,128,131-133, succinic dehydrogenase 137, 224, 372 sucrose synthase 334-336 136, 211, 515 synthesis of 67-68 SUCROSE SYNTHASE (SUS) gene from maize 335 stamen 2, 7,17-25, 37, 91,121-123,133,134,136,137, sucrose synthase gene from potato 107 sucrose synthetase 412 151-153, 245-249, 258 sugarcane 126, 530 polypeptides of 19 sugary {su) mutant 332, 336 starch-branching enzyme 403, 424 sunflower 30, 41,105,106, 111, 134, 137,149, 317, 352, stearoyl-acyl carrier protein (ACP) desaturase, stearoyl369, 411, 428, 429, 441, 450, 454, 461 ACP desaturase gene 40,116, 451, 462 SUPERMAN (SUP) gene, sup mutant 20, 23,153,156 Stellaria media 157,169,173, 325, 361, 377 sterility gene (S-gene) 7, 8, 245, 248, 253, 254, 257, 258, superoxide dismutase 107,116, 435 suppressor mutator (spm) mobile element 345 261, 266-268, 271-273, 276, 281, 287-290 suspensor 6, 312, 313, 331, 362-366, 371, 372, 376, 383, stigma 2, 7-9,14, 34, 36, 73, 74,102,104, 121, 129,152, 393, 397-399, 408, 411, 414, 421, 425, 426, 469, 478 181-183,185,187-192,194,195,197,198, 201-209, 218, 237, 242-244, 246-256, 258, nuclear cytology of 379-380 260-271, 272-274, 276-284, 287, 290, 291, 302, physiology and biochemistry of 380-382 role in the growth of cultured embryos 389, 391 303, 313, 414 anatomy of 182-187 ultrastructure of 377-379 senescence of 191-192 SUSPENSOR (SUS) gene, sus mutation 398 stigma-specific genes 197-201 suspensorlike structure 469, 511 sweet cherry (see Prunus avium) stigmatic exudate 182,186,187,193, 202, 205, 209, 249 synaptonema (see synaptonemal complex) composition of 190-191 synaptonemal complex 43, 49, 50, 52, 54, 56,158 secretion of 188-190 synergids 2, 3, 5,162,163,169,170, 172-175,177,178, types of 182-183 297, 311, 314, 315, 317, 360 stigmatic papillae 73, 74,156,182,183,185-191,197, 201-206, 247, 250, 254, 255, 258, 260, 264, 271, role in fertilization 304-309 272, 279-284, 287 structure of 166-168 Stipa elmeri 164,166,169 syntaxin 422 stomium 28, 31, 60,129 transcript accumulation in 30 Tabebuia caraiba 253 storage compounds (see storage proteins) T. ochracea 253 storage lipids 443, 445-446, 450-451, 462-464, 520 Tagetes erecta 122 storage materials (see storage proteins) tapetal cells (see tapetum) storage products (see storage proteins) taperum 5, 8, 26-29, 31-33, 42, 53, 58, 60, 63, 67, 68, 72, storage protein genes 74, 75, 77, 99,102,112,116,117,124,125,133, expression of 342-350, 520 135-137,142,151, 257, 263, 264, 283, 284, 501, of embryos 453-462 508, 509
Index amoeboid (periplasmodial) 34, 35, 37,130 cytology of 36-37 DNA accumulation in 36 DNA transfer from 34 gene expression in 37-41 glandular (secretory) 34, 35, 37 role in pollen sterility 129-132 structure-function relationships of 31-36 ultrastructure of 32 tapetum-specific genes 37-41,122, 125 taro (see Colocasia esculenta) tassel seed (ts) mutant 19 TATA-box 114, 289, 349, 351, 352, 427, 455-459, 461, 463 TATA element (see TATA-box) TATA sequence (see TATA-box) T-DNA (see transferred DNA) temperature-sensitive cell lines 497-498 test-tube fertilization 314-315 tetrad 31, 32, 34, 35, 37, 38, 40, 44-^6, 53, 57, 61-69, 71, 75, 78, 89, 100,102,104,106, 123,125-128,130, 131, 133,134,137,158-162,177, 506 Theobroma cacao (see also cacao) 253, 257, 438, 446, 460 thiamine pyrophosphatase 37 thiol endopeptidase 29 thiolase 412 thionin 30, 198, 339, 341, 342 y-thionin (see thionin) thioredoxins 111, 291 2-thiouracil 210 thrum morph 182,187, 218, 246, 247, 249-251 thymidine kinase 77 Tillandsia caput-medusae 80, 82 tissue culture (see also under anther, embryo culture, ovary, ovule, pollen culture, zygote) 3, 4, 10, 13, 14, 18,19, 25, 57-60, 75,134-136,140, 141,144, 153,172, 314-317, 321, 329, 338, 345, 364, 370, 381, 400, 402, 407, 408, 414, 525, 527-530 tobacco 3, 4, 8,10, 18, 25, 29-31, 3 9 ^ 1 , 63, 77, 78, 86, 88, 97,102, 104,106-108, 111, 112,114-116, 132,134, 141,149,151, 153,176, 197,198, 200, 208, 222, 223, 227, 230, 236, 239, 281, 282, 303, 304, 312, 317, 352, 353, 355, 383, 432, 437, 454-457, 459, 461, 462, 468, 503, 505, 506, 508-511, 513, 514,516, 518, 520, 525, 528 cDNA clones from 29-31 cybrid (somatic hybrid) 137, 149 transgenic 21, 30, 3 9 ^ 2 , 75, 84, 111, 112,114-117, 125,126,132,145,197,198, 200, 204, 227, 228, 281-283, 288, 290, 352-355, 403, 404, 423, 426, 427, 429, 431, 433, 436, 452, 454-464, 493, 525 tomato 11,18, 25, 31, 37, 39, 93, 103,105,106, 108-110, 112,113, 115,117, 122,123,131, 133,134,136, 153,197,198, 200, 226, 290 GA-deficient mutant of 134 stamenless-2 (sl-2) mutant of 18,133, 136,137,153 transgenic 112, 113, 198, 423 Torenia fournieri 163, 166,167, 308 totipotency 4, 468, 470, 498, 521 Tradescantia sp. 86, 182 T. bracteata 34, 65, 69, 78, 84, 86
689 T. paludosa 37, 45, 48, 53, 58, 76, 79, 80, 86, 94-99,104-106, 109, 115, 173, 214, 221-225, 234, 309, 373 T. reflexa 84, 213 T. virginiana 34, 35, 66, 86, 88, 89, 214, 230, 231, 235, 237, 239, 241, 295, 296 trans-acting factors 9,113,114, 280, 334, 344, 349, 426, 431, 439 transcription factor 20, 23, 40, 111, 114,116,156, 345, 346, 350, 354, 355, 401, 423, 425, 429, 433, 434, 455, 496 transcription regulator (see transcription factor) transfer cell 69, 83, 84,167,169,175,186,195, 234, 323, 324, 335, 378, 382, 384 transferase 336 transferred DNA (T-DNA) 13, 20,122,127, 398, 413, 426, 434, 454, 479, 497, 526-528 transient assay 112, 113, 115-117, 353-355, 454, 455, 458 transmitting tissue 185,188,192,193,198-200, 204-207, 232, 259, 270, 280-284, 287, 288, 301, 303 transposon mutagenesis 7, 8, 24, 92,122,150, 333, 345, 369, 400, 415, 417 transposon tagging (see transposon mutagenesis) 2,4,5-trichlorophenoxyacetic acid 472 2,4,6-trichlorophenoxyacetic acid (2,4,6-T) 480 Trichodiadema setuliferum 87, 298 trichome 185, 203, 206 Trifolium sp. 257 T. ambiguum 330, 396 T. medium 396 T. pratense 192,194, 489 T. repens 330, 396, 477 T. rubens 489 T. semipilosum 396 Trigonella foenum-graecum 223 Trillium sp. 48 T. erectum 44, 47, 48, 53, 54, 58, 76, 77, 94 triosephosphate isomerase 92, 93 Tripsacum laxum 126 tristyly 249 features of 247-248 Triticale 170, 309, 322, 512 triticin 337, 348, 353 Triticum X Hordeum 530 Triticum sp. 322, 337 T. aestivum (see also wheat) 9, 26, 45, 51, 65, 66, 68, 76, 80, 149, 164,165,167, 169,170, 192, 214, 300, 308, 316, 322, 325, 337, 349, 361, 364, 366, 367, 372, 387, 390, 393, 396, 435, 452, 521 T. durum 337, 399, 445, 446 T. timopheevi 149 T. vulgare 337, 408 Tropaeolum majus 186, 219, 224, 378, 380, 381 tryphine 35, 36, 68, 70, 74, 201, 204, 257, 264 trypsin inhibitors 30 tubulin (a, P, and y) 45, 57, 88, 99,107,116, 156, 217, 227, 235-237, 406, 495 Tulbaghia violacea 36, 48, 52, 76, 80, 81, 94, 99 tulip (see Tulipa gesneriana) Tulipa sp. 33, 35 T. gesneriana 53, 78, 234 tumor-inducing (Ti) plasmid 13, 20, 39
690 tunicamycin 260, 442, 483, 484 tunicate (tu) mutant 19 twin (twn) mutant 398 Typha latifolia 212, 214, 221, 223, 225
Index
cms in 149 gliadin and glutenin genes of 353-354 histone H3 gene from 28 ribosomal RNA probe of 174 storage protein gene expression in 347-350 ubiquitin 102,116,195, 460, 528 tapetal disintegration in 130 Ubisch bodies 33 transformation of 526-530 Ulmus sp. 503 transgenic 530 uniconazol 433 wheat germ agglutinin (WGA) 435, 436 unilateral incompatibility 269-271 wide hybridization 329-331, 395-397 UNIVERSAL FLORAL ORGANS (UFO) gene, ufo mutant wild soybean (see Glycine soja) 20, 23, 24 WUSCHEL (WUS) gene 419 uridine diphosphoglucose dehydrogenase 224 X-bodies 309 Vanda sp. 174, 361, 368 Xenopus sp. 352 X. laevis 259, 265, 338, 442 V. sanderiana 373 vegetative cell 2, 5, 6, 61, 62, 70, 75, 78-84, 92, 94-97, xyloglucan 334 99-102,112, 114,119,181, 202, 211, 212, 214-217, xyloglucan endotransglycosylase 484 220-222, 229, 235, 238, 240, 241, 255, 295-301, yeast 174, 349, 351, 353, 354, 458 307-310, 507, 516-519, 521 DNA metabolism of 86 cell division cycle phosphoprotein (p34cdc2) 495 embryogenic division of 510-515 expression of protein in 141 isolation of 89 secretory protein 422 nuclear pores in 86-87, 214 verapamil 217 Z-Iayer (see Zwischenkorper) Verbascum phlomoides 189 Zamia sp. 470 Viciafaba (see also broad bean) 150,172,185,187, 206, 303, Zantedeschia aethiopica 88 323, 372, 373, 378, 381-383, 408, 413, 441, 442, Zea mays (see also maize) 19, 26, 32, 36, 45, 52, 53, 65, 66, 444, 446, 449, 452, 458, 459, 462 68, 92, 99,102-104,107,122,130,156,157,159, 160,163,165,166,169,170,174,183,185, 216, V. narbonensis 449, 460 vicilin, vicilin gene 381, 383,441,443,444, 446-452,458-460 219, 221, 294, 300, 305, 316, 323, 337, 340, 344, 345, 359-361, 367, 369, 387, 392, 393, 411, 412, vicilin-box 458, 461 Vigna sp. 502 423, 424, 445, 452, 515 cms-C 122,130 V. radiata 411 cms-S 122,130 V. sinensis 323, 378, 382, 383 cms-T 122, 130 V. unguiculata 66,192, 411, 445, 527 inflorescence culture of 59 V. vexillata 65, 66 zeatin 134, 329, 391, 471, 481, 482, 507 virulence (VIR) gene 13 zeatin riboside 134, 391, 507 Viscum minimum 156,164,167,169 Vitis sp. 502, 531 zein, zein genes 337-339, 341-343, 350, 352, 355, 460 V. berlandieh X V. rupestris 531 characterization of 350-352 V. rupestris 531 expression of 342-347, 351-352 Zepkyranthes sp. 393 V. vinifera (see also grape) 188, 373, 491, 531 Z. Candida 301 vitronectin 200, 303 Z. grandiflora 85, 236 vitronectinlike protein (see vitronectin) Z. rosea 157 VIVIPAROUS (VP) genes, vp mutants, VP protein 399-402, 433, 455 zinc finger domains 111 Zwischenkorper 63, 211 vivipary 399, 400, 402, 432 zygote 1-3, 6, 9, 82,163,168,173, 253, 293, 309, 310-316, 322, 329, 357, 362-364, 366, 367, 372-374, 377, walnut (see Juglans regia) 382-384, 389, 391, 394, 396-399, 407, 420, watermelon (see Citrullus lanatus) 467-469, 471, 478, 498, 525-527, 531 WAXY (WX) gene, wx mutant 92,117, 332, 335, 530 culture of 392-393 expression of 336 ultrastructure of 358-361 Western blot 12, 238, 273, 423, 485 wheat 9, 29, 56,102,128,135-138,148,150,151, 296, 298, zygotene DNA (zyg-DNA) 49-51 zygotene RNA (zyg-RNA) 49-50 305, 315, 317, 323, 324, 326, 329, 334, 336-340, zygotic embryogenesis 10,13,14, 357, 467, 469, 470, 478 342, 347, 348, 352, 354, 355, 382, 385, 387, 389, zygotic embryos 467^71, 478, 494, 495, 501, 520, 521, 401, 411, 427-432, 435, 436, 453, 473, 482, 486, 531 488, 493, 503, 506, 509-512, 516
