IL-27 Robert A. Kastelein* and Stefan Pflanz DNAX Research Institute, 901 California Ave, Palo Alto, CA 94304, USA * corresponding author tel: (650) 496-1271, fax: (650) 496-1200, e-mail:
[email protected] DOI: 10.1006/rwcy.2002.0331.
SUMMARY Interleukin 27 (IL-27) is a heterodimeric cytokine that consists of a four helix bundle cytokine-like subunit, termed p28, and a soluble cytokine receptor-like subunit, EBI3 (Epstein±Barr virus-induced gene 3). The p28 subunit has homology with other singlechain cytokines, whereas EBI3 is related to the extracellular domain of the hematopoietic cytokine receptor family. As such, IL-27 is a member of a small family of heterodimeric cytokines that includes IL-12 and IL-23. Efficient expression and secretion of the IL-27 heterodimer requires coexpression of p28 and EBI3. Both subunits are produced by activated antigen-presenting cells (APCs) and the presence of both is required for the biologic activity of IL-27. Cellular targets include naõÈ ve CD4+ T cells and NK cells (Pflanz et al., 2002). The hematopoietic cytokine receptor WSX-1/TCCR, together with at least one additional ± yet to be identified ± receptor subunit(s) is believed to mediate the biologic activity of IL-27.
BACKGROUND
Discovery The p28 subunit of IL-27 was discovered by a computational approach. Sequence databases were searched with a consensus sequence extracted from an alignment of the IL-6 family of long-chain cytokines. Transient mammalian expression of p28 indicated that this protein was expressed, but not
Cytokine Reference
secreted despite the presence of a signal peptide. The similarity of p28 to IL-12 p35 suggested that a partner protein might be required for secretion. Among members of the IL-12 p40-related cytokine receptors EBI3 was identified as the partner protein for p28. Coexpression of p28 and EBI3 led to secretion of biologically active heterodimeric IL-27.
Alternative names The subunits of IL-27 are p28 and EBI3. The designation p28 represents the approximate molecular weight of this IL-27 subunit.
Structure Not yet determined. IL-27 is likely to resemble the structure of IL-12 with the exception that EBI3 ± unlike IL-12 p40 ± does not possess an N-terminal Ig-like domain.
Main activities and pathophysiological roles IL-27 drives proliferation of naõÈ ve human and mouse CD4+ T cells in a CD3-dependent manner. It synergizes with IL-12 to trigger production of high levels of IFN in these cells. In synergy with IL-12 and IL-2 it can also lead to increased production of IFN in NK cells. Pathophysiological roles of IL-27 have not been determined at this point.
Copyright # 2002 Published by Elsevier Science Ltd
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Robert A. Kastelein and Stefan Pflanz
GENE AND GENE REGULATION
Accession numbers Human p28: NM_145659 Mouse p28: NM_145636 Human EBI3: NM_005755 Mouse EBI3: NM_015766
Figure 1 Amino acid sequences for human and mouse p28 and human and mouse EBI3. Human p28 MGQTAGDLGW VRGQAHRFAE GGLGTQGRWT LPGALGSALQ PQP
RLSLLLLPLL SHLPGVNLYL NMERMQLWAM GPAQVSWPQL
LVQAGVWGFP LPLGEQLPDV RLDLRDLQRH LSTYRLLHSL
RPPGRPQLSL SLTFQAWRRL LRFQVLAAGF ELVLSRAVRE
QELRREFTVS SDPERLCFIS NLPEEEEEEE LLLLSKAGHS
LHLARKLLSE TTLQPFHAPL EEEEEERKGL VWPLGFPTLS
RLSLLLLPLL GVNLDLLPLG EQLWAMRLDL SQVSWPQLLY
LVQAGSWGFP YHLPNVSLTF RDLHRHLRFQ TYQLLHSLEL
TDPLSLQELR QAWHHLSDSE VLAAGFKCSK VLSRAVRDLL
REFTVSLYLA RLCFLATTLR EEEDKEEEEE LLSLPRRPGS
RKLLSEVQGY PFPAMLGGLG EEEEEKKLPL AWDS
LWASCPPCSG AARGHSWPCL PPEGVRLSPL RAVRPRARYY
RKGPPAALTL QQTPTSTSCT AERQLQVQWE VQVAAQDLTD
PRVQCRASRY ITDVQLFSMA PPGSWPFPEI YGELSDWSLP
PIAVDCSWTL PPAPNSTSPV PYVLNVTAVH PWGSSSSFVP FSLKYWIRYK RQGAARFHRV ATATMSLGK
LWASRSPGYT TQQQSQPCLQ PEGVRLRTAG SKPHAKYCIQ
ETALVALSQP RSPQASRCTI QRLQVLWHPP VSAQDLTDYG
RVQCHASRYP PDVHLFSTVP ASWPFPDIFS KPSWSLPGQV
VAVDCSWTPL QAPNSTRSTS YMLNVTAVHP GGASSSLLAF LKYRLRYRRR GASHFRQVGP ESAPHKP
Mouse p28
Chromosome location EBI3 is on human chromosome 19p13 and mouse chromosome 17. P28 is on human chromosome 16p11 and mouse chromosome 7.
Cells and tissues that express the gene Expression of mRNA encoding the p28 subunit is restricted to APCs from the monocyte/macrophage lineage. Highest levels of EBI3 mRNA are also found in cells from the monocyte/macrophage lineage as well as in placental tissue.
MGQVTGDLGW VHSFAESRLP TQGTWTSSER GALGGPNQVS
Human EBI3 MTPQLLLALV SFIATYRLGM FITEHIIKPD GPIEATSFIL
Mouse EBI3 MSKLLFLSLA FIATYRLGVA VAERIIKPDP IEATTFTLRN
Mouse p28
PROTEIN
Accession numbers Human p28: AAM34498 Mouse p28: AAM34499 Human EBI3: NP_005746 Mouse EBI3: NP_056581
Sequence See Figure 1.
Description of protein
The full open reading frame encodes 234 amino acid residues, including a predicted 28 amino acid residue N-terminal signal peptide. The protein belongs to the family of long-chain four helix bundle cytokines (other members include CNTF, oncostatin M, IL-6, IL-11, IL-12 p35, etc.). One potential site for N-linked glycosylation is present (N85). Human EBI3 The full open reading frame encodes 229 amino acid residues, including a predicted 20 amino acid residue N-terminal signal peptide. The protein belongs to the family of soluble cytokine receptor-like molecules (other members being sCNTFR , sIL-6R , CLF-1, IL-12 p40, etc.). Two potential sites for N-linked glycosylation are present (N55 and N105).
Human p28
Mouse EBI3
The full open reading frame encodes 243 amino acid residues, including a predicted 28 amino acid residue N-terminal signal peptide. The protein belongs to the family of long-chain four helix bundle cytokines (other members include CNTF, oncostatin M, IL-6, IL-11, IL-12 p35, etc.). No potential sites for N-linked glycosylation are present.
The full open reading frame encodes 227 amino acid residues, including a predicted 18 amino acid residue N-terminal signal peptide. The protein belongs to the family of soluble cytokine receptor-like molecules (other members being sCNTFR , sIL-6R , CLF-1, IL-12 p40, etc.). Two potential sites for N-linked glycosylation are present (N54 and N104).
IL-27 3
Important homologies The closest sequence homologs of p28 protein are IL-11, NNT-1/CLC, and CNTF; the closest homologs of EBI3 protein are CLF-1 and IL-12 p40.
CELLULAR SOURCES AND TISSUE EXPRESSION
Cellular source that produce Expression of the p28 subunit seems restricted to antigen-presenting cells from the monocyte/macrophage lineage. High levels of EBI3 are also found in cells from the monocyte/macrophage lineage as well as in placental tissue.
RECEPTOR UTILIZATION IL-27 binds to the hematopoietic cytokine receptor WSX-1/TCCR. However, this receptor by itself is not sufficient for IL-27-mediated signal transduction. At least one additional receptor subunit remains to be determined.
IN VITRO ACTIVITIES
In vitro findings IL-27 stimulates proliferation of naõÈ ve CD4+ T cells in a CD3-dependent manner. It synergizes with IL-12
to trigger production of high levels of IFN in these cells. In synergy with IL-12 and IL-2 it can also lead to increased production of IFN in NK cells.
Bioassays used NaõÈ ve T cells can be used to assay IL-27. Purified naõÈ ve mouse and human T cells (CD45RBhigh and CD45RA, respectively) can be cultured with platecoated anti-CD3 mAb and soluble anti-CD28 mAb and a neutralizing anti-IL-2 mAb (to eliminate stimulation due to CD3-dependent autocrine IL-2 production). The cells will show dose-dependent proliferation in the presence of IL-27.
ACKNOWLEDGEMENTS The above authors thank DNAX Research Institute and Schering-Plough.
Reference Pflanz, S., Timans, J. C., Cheung, J., Rosales, R., Kanzler, H., Gilbert, J., Hibbert, L., Churakova, T., Travis, M., Vaisberg, E., Blumenschein, W. M., Mattson, J. D., Wagner, J. L., To, W., Zurawski, S., McClanahan, T. K., Gorman, D. M., Bazan, J. F., de Waal Malefyt, R., Rennick, D., and Kastelein, R. A. (2002). IL-27, a heterodimeric cytokine composed of EBI3 and p28 protein, induces proliferation of naõÈ ve CD4+ T cells. Immunity 16, 779±790.
