●

Cutting Edge: Identification of the Orphan Receptor G-Protein-Coupled Receptor 2 as CCR10, a Specific Receptor for the Chemokine ESkine1,2

David I. Jarmin,* Miriam Rits,* Dalena Bota,* Norma P. Gerard,* Gerard J. Graham,† Ian Clark-Lewis,‡ and Craig Gerard3*

(1, 2). Chemokines mediate their functions through their binding to A number of orphan G-protein coupled receptors (GPR) have specific cell-surface receptors, and these receptors are members of been reported as putative chemokine receptors. One previ- the seven transmembrane spanning G protein-coupled-receptor ously reported orphan receptor is an incomplete PCR clone, (GPR)4 superfamily. To date, this interaction appears subfamily called GPR2. Here we report the cloning of full-length human specific, and receptors have been identified for CC chemokines (h)GPR2 and mouse (m)GPR2 cDNAs, and the identification of (CCR1–9), CXC (CXCR1–5), C (XCR1), and CX3C (CX3CR1) GPR2 as a receptor for a novel CC chemokine called ESkine. (1–6). In addition to the known chemokine/receptor pairings, a hGPR2 is expressed at high levels in testis and small intestine, number of orphan GPRs have been reported (7–9). Several che- and at lower levels in other tissues. mGPR2 was expressed at mokines have also been identified, for which the specific recep- high levels in small intestine, colon, lymph nodes, and Peyer’s tor(s) is not yet known (10–12). patches and at lower levels in thymus and spleen. Stimulation One orphan receptor previously reported as having homology to of L1.2/hGPR2 transfectants with hESkine induced their mi- known chemokine receptors is GPR2 (13). The GPR2 gene cloned gration and resulted in intracellular calcium mobilization. previously from human genomic DNA lacked an initiating methi- These results provide evidence that GPR2 is a specific receptor onine, suggesting the existence of an intron in the 5Ј region. No for ESkine. We propose that GPR2 be renamed as CCR10. positive data on the expression of GPR2 has been reported to date. The expression pattern of mGPR2/CCR10 suggests that it may Here we report that GPR2 is a specific receptor for the novel CC by guest on October 1, 2021. Copyright 2000 Pageant Media Ltd. play a role in the homing/trafficking of leukocytes within in- chemokine known as ESkine/ALP/ILC/CTACK (14–17). ESkine testinal and lymphoid environments. The Journal of Immu- induced chemotaxis of the murine pre-B cell line L1.2, when trans- fected with human (h)GPR2. Moreover, calcium flux assays demon- nology, 2000, 164: 3460–3464. strated that hGPR2 and mouse (m)GPR2 transfectants responded to human or murine ESkine. Here we also report the low-level con- hemokines are believed to play a critical role in the traf- stitutive expression of GPR2 RNA in a variety of tissues and, in ficking, homing, and activation of leukocytes, as well as particular, higher levels of mGPR2 RNA expression in the intestine, C mediating a wide range of other functions (1). At present colon, lymph node, Peyer’s patches, spleen, and thymus. Previous reports have indicated that those chemokines and their receptors

https://www.jimmunol.org they are classified into four separate subfamilies on the basis of the spacing of conserved cysteine residues within their N-terminal re- (namely EBI1-ligand chemokine (ELC)/secondary lymphoid tissue

gions: these are known as the CC, CXC, C, and CX3C subfamilies chemokine (SLC)/CCR7, liver- and activation-regulated chemokine (LARC)/CCR6, and thymus-expressed chemokine (TECK)/CCR9), whose expression is relatively lymphoid restricted, appear to play crit- *Ina Sue Perlmutter Laboratory, Children’s Hospital, Harvard Medical School, Bos- ical roles in the homing and localization of leukocytes (18–20). ton, MA 02115; †The Beatson Institute for Cancer Research, Cancer Research Cam- Therefore, we speculate that GPR2 may also play a similar role. Using ‡ Downloaded from paign Laboratories, Glasgow, Scotland, United Kingdom; and Biomed Research Center, University of British Columbia, Vancouver, British Columbia, Canada the current nomenclature rules for chemokine receptors, we propose that GPR2 be designated as CCR10. Received for publication December 13, 1999. Accepted for publication February 2, 2000. The costs of publication of this article were defrayed in part by the payment of page Materials and Methods charges. This article must therefore be hereby marked advertisement in accordance Cloning and sequencing of human and mouse GPR2 cDNAs with 18 U.S.C. Section 1734 solely to indicate this fact. 1 This work was supported in part by a grant from The Wellcome Trust, U.K. The full-length cDNA for hGPR2 was cloned from a CEMx 174 cell line (053125; to D.I.J.), National Institutes of Health Grant HL39579 (to C.G.), and by the (T cell/B cell hybrid) cDNA library, while the mGPR2 cDNA was cloned Rubenstein/Cable Fund at the Perlmutter Laboratory. from a Peyer’s patch cDNA library. The cDNA libraries were constructed 2 The sequences presented in this article have been submitted to GenBank under accession numbers AF215981 (hCCR10), AF215982, and AF215983 (mCCR10). 4 Abbreviations used in this paper: GPR, G-protein-coupled receptor, h, human; m, 3 Address correspondence and reprint requests to Dr. Craig Gerard, Ina Sue Perlmut- mouse; SLC, secondary lymphoid-tissue chemokine, ELC, EBI1-ligand chemokine; ter Laboratory, Enders Suite 144, Children’s Hospital, 320 Longwood Avenue, Bos- LARC, liver- and activation-regulated chemokine; TECK, thymus-expressed ton, MA 02115. E-mail address: [email protected] chemokine.

Copyright © 2000 by The American Association of Immunologists 0022-1767/00/$02.00

● The Journal of Immunology 3461

FIGURE 1. A, Alignment of the predicted amino acid sequences of human and mouse GPR2. Transmembrane domains (TM) are outlined with lines above the corresponding amino acids. The eight additional N-terminal amino acids are indicated with a bold line. B, Schematic of mGPR2 cDNA clones. Two forms of mGPR2 are shown. Potential exons are indicated above boxes

using the Superscript plasmid kit (Life Technologies, Grand Island, NY) twice with RPMI 1640/1% FBS and then used in mobilization studies. according to the manufacturer’s instructions. The cDNA libraries were Chemotaxis assays were also performed on untransfected L1.2 cells, screened with 32P-labeled PCR-derived probe fragments encoding the 5Ј hGPR2/L1.2 transfectants, or mGPR2 transfectants in Costar 5.0-␮m regions of hGPR2 and mGPR2 (human, 5Ј-GTTTCCTGGGCCATTAC-3Ј 24-well cluster transwells (Corning, Corning, NY) as described (21). The and 5Ј-ACAGCGTCGTTGGCCTTC-3Ј; mouse, 5Ј-GTCTCCTGGGGAC number of migrating cells was determined by comparison against a stan- by guest on October 1, 2021. Copyright 2000 Pageant Media Ltd. TTTAC-3Ј and 5[pime]-GCTGAAAGAAAGCGCAGG-3Ј). The murine dard curve derived from a dilution series of a known number of cells, and probe sequence was derived from an incomplete cDNA clone derived from the number of migrated cells was expressed as a percentage of the total a murine Th1 cDNA library (our unpublished data), whose sequence was number of input cells. homologous to the clone previously reported (13). Positive cDNA inserts were subcloned into the pcDNA 3 vector (Stratagene, La Jolla, CA) (hu- man, EcoRI and XbaI; mouse, NotI and XbaI) and sequenced fully. Results and Discussion Comparison of the hGPR2 sequence to known chemokine receptors Chemokines and cells suggests that this orphan receptor may be a putative chemokine re- Chemokines were purchased from PeproTech (Rocky Hill, NJ) or R&D ceptor. We examined the expression of mGPR2 RNA in various Systems (Minneapolis, MN). Mouse and human ESkine were chemically https://www.jimmunol.org mouse tissues and determined that it was expressed at low levels in the synthesized as previously described (14). hGPR2 and mGPR2 were trans- fected into the murine pre-B cell line L1.2 by electroporation, and stable thymus. This data prompted us to further characterize GPR2. transfectants were obtained after G418 selection, as described (21). Cloning of full-length hGPR2 and mGPR2 cDNAs Northern blot analysis Expression of mGPR2 RNA in the thymus suggested that hGPR2 Multiple tissue Northern blots (Clontech Laboratories, Palo Alto, CA) were would be expressed in human T cells. Therefore, we screened a probed with a 32P-labeled 5Ј hGPR2-specific DNA probe, according to the Downloaded from CEMx 174 cell cDNA library and identified several GPR2-like suppliers instructions. Total RNA from 4- to 8-wk-old BALB/c mouse tissues were stored in RNA Later (Ambion, Austin, TX) before their ex- clones. The nucleic sequence of one of these clones encoded an traction using the RNAqueous kit (Ambion). Next, 15 ␮g of each sample additional 8 aa at the amino terminus, compared with the previ- was electrophoresed on a denaturing formaldehyde-agarose gel and blotted ously published sequence of the hGPR2 gene (13) (Fig. 1A). In this onto Hybond N membranes (Amersham Pharmacia Biotech, Piscataway, report, it was speculated that there was an exon in the 5Ј terminal NJ). The membranes were prehybridized using an SDS/phosphate buffer before hybridization with a 32P-labeled 5Ј mGPR2-specific DNA probe, as region of hGPR2, and our data supports this conclusion. described (22). After washing, the blots were exposed to BioMax MR film Further analysis of mGPR2 RNA expression revealed a striking using BioMax MS intensifying screens (Eastman Kodak, Rochester, NJ). level of expression in Peyer’s patches (see below, and Fig. 2B). Therefore, we constructed a cDNA library from murine Peyer’s Chemotaxis and calcium mobilization patches. Two murine clones were identified and sequenced. Con- Calcium mobilization studies were performed on untransfected L1.2 cells, ceptual translation of one of the cDNA clones revealed that the hGPR2/L1.2 transfectants, or mGPR2 transfectants loaded with fura-2AM encoded protein was 8 aa longer than that previously reported in (Molecular Probes, Eugene, OR), as previously described (21). A 2-ml aliquot of cells, containing 4 ϫ 106 cells, was used for each analysis. For the cloning of the human genomic GPR2 clone (13). Fig. 1A shows pertussis toxin sensitivity, cells were stimulated for 2 h with 100 ng/ml an alignment of the predicted amino acid sequences for the hGPR2 pertussis toxin (Sigma, St. Louis, MO), and the cells were then washed and mGPR2 proteins. These appear to be 86% identical at the level 3462 CUTTING EDGE

either human poly(A)ϩ RNA or mouse total RNA (Fig. 2). Anal- ysis of hGPR2 showed a high level of expression in the adult testis and small intestine (Fig. 2A). In addition, relatively high levels of expression were detected in fetal lung and fetal kidney. Weaker expression was also observed in many other adult tissues including spleen, thymus, lymph node, colon, heart, ovary, peripheral blood lymphocytes, and spinal cord. The primary transcript size was ϳ1.4 kb in size, but a second ϳ2 kb transcript was also visible in most of the same tissues. These transcripts are of similar sizes to the two forms of mGPR2, suggesting that they represent alterna- tively spliced forms of hGPR2. In contrast to the high levels ob- served in human testis, mGPR2 expression was not detected in testis. However, high levels of expression were observed in small intestine, colon, lymph node, and Peyer’s patches (Fig. 2B). In addition, weak expression was observed in spleen and thymus. This appeared to be a single transcript of ϳ1.4 kb. Longer expo- sure of murine tissue blots revealed very faint expression levels in other tissues (such as spinal cord and ovary), as well as very faint expression of a second larger size transcript visible in Peyer’s patches and small intestine. The relevance of the discrepancy ob- served between the high level of expression in human testis and the apparent absence of any expression in murine testis is unclear; however, this may simply reflect the different analysis using the more sensitive poly(A)ϩ vs total RNA or that species differences may exist per se. The observation that GPR2 is expressed at mod- erate to strikingly high levels in lymphoid, secondary lymphoid, and intestinal tissues is of particular interest, given the known im- portance of chemokines and their receptors in homing and traf- ficking of leukocytes. It has been previously speculated that CCR9, CCR7, and CCR6 may form a second CC chemokine receptor subfamily (4), as their expression is relatively restricted to lym- phoid and intestinal tissues (20, 24, 25). Like GPR2, these recep- tors also share a common structural feature in that their coding sequences are interrupted by introns. by guest on October 1, 2021. Copyright 2000 Pageant Media Ltd.

ESkine induces migration of L1.2 cells transfected with GPR2 To investigate the potential of known chemokines to function as specific ligands for GPR2, we tested 18 chemokines (see Fig. 3A) for their ability to induce migration of GPR2-expressing L1.2 cell transfectants in chemotaxis assays. Only the addition of human ESkine (hESkine) resulted in the dose-dependent chemotaxis of hGPR2/L1.2 cells, relative to untransfected L1.2 cells (Fig. 3B). https://www.jimmunol.org FIGURE 2. Analysis of hGPR2 and mGPR2 RNA expression. A, Mem- This migration of hGPR2/L1.2 cells in response to hESkine began ϩ ϳ branes with poly(A) RNA from the indicated human tissues were hybrid- at around 10–30 nM and was maximal at 300 nM. ized with a 5Ј hGPR2 probe. Following exposure, blots were stripped and loading levels confirmed with a h␤-actin probe. B, Total RNA from the indicated mouse tissues was hybridized with a mGPR2 probe. Blots were ESkine induces calcium mobilization of GPR2-expressing L1.2 also stripped and loading levels confirmed with a h␤-actin probe. cells

Downloaded from We next investigated the ability of ESkine to mobilize intracellular calcium in GPR2-expressing L1.2 cells. The addition of 100 nM hE- of amino acid sequence. The second clone was incomplete as it Skine to hGPR2/L1.2 cells resulted in a significant calcium flux, and lacked an initiating methionine, but it also had an additional 465 bp this response was dose dependent down to at least 1 nM (Fig. 4A), ϩ at the 3Ј end with a classical poly(A) tail. The first clone repre- although a 10-nM dose appeared to be as effective as 100 nM. The sents a full-length cDNA, composed of at least 3 exons. The sec- addition of 100 nM SLC to hGPR2-expressing L1.2 cells (which ex- ond clone represents an incomplete cDNA with an alternative 3Ј press endogenous CCR7) also resulted in a significant mobilization region. Fig. 1B shows a schematic of the structure of the two (Fig. 4). Murine ESkine was also able to induce a strong calcium flux mouse clones. The hGPR2 gene has previously been reported to be in mGPR2-expressing L1.2 cells (Fig. 4B), as well as in hGPR2/L1.2 localized to chromosome 17q21, next to a human homologue for cells (data not shown). The addition of 100 nM hESkine or mESkine the Drosophila enhancer of zeste gene (13, 23). resulted in desensitization of hGPR2/L1.2 cells to a second stimulus of ESkine of the same dose (Fig. 4C). The addition of SLC did not Analysis of the expression of hGPR2 and mGPR2 result in the desensitization of hGPR2/L1.2 cells to a subsequent hE- The expression of hGPR2 and mGPR2 RNA in various mouse and Skine stimulation and vice versa (Fig. 4D). The response of hGPR2 to human tissues was investigated using Northern blot analysis of hESkine was also pertussis toxin sensitive (Fig. 4E). The Journal of Immunology 3463

FIGURE 3. Migration assays. A, Chemokines tested in migration as- says. B, hESkine-induced migration of L1.2 cells transfected with hGPR2. Mean values of duplicate measurements are shown; bars, SD. Data shown FIGURE 4. ESkine-induced mobilization of intracellular calcium in is representative of three separate experiments. L1.2 cells stably expressing GPR2. A, Dose response of hGPR2/L1.2 cells to hESkine (top three traces). L1.2 cells expressing hGPR2 treated with by guest on October 1, 2021. Copyright 2000 Pageant Media Ltd. SLC as a positive control and untransfected L1.2 cells were stimulated with Taken together, the data we report herein strongly support the 100 nM hESkine (bottom trace). B, mGPR2/L1.2 cells and untransfected L1.2 cells were stimulated with 100 nM mESkine. C, hGPR2/L1.2 cells conclusion that GPR2 is a specific receptor for the novel CC che- were stimulated with 100 nM of either hESkine or mESkine, followed by mokine ESkine, and that this occurs cross-species in both human a second identical stimulation. D, Desensitization experiments. hGPR2/ and murine systems. Currently, 10 CC chemokine receptors have L1.2 cells were stimulated with 100 nM SLC followed by a second stim- been designated, and we propose that GPR2 be redesignated as ulation of 100 nM hESkine (top trace) or vice versa (bottom trace). E, CCR10, according to the guidelines established for chemokine no- Pertussis toxin-sensitivity. hGPR2/L1.2 cells either previously treated with menclature at the 1997 Gordon Conference on Chemokines. We 100 ng/ml pertussis toxin (PTX) (bottom trace) or left untreated (top trace) https://www.jimmunol.org are currently further investigating the nature of the cells expressing were stimulated with 100 nM Eskine. Arrows indicate the time of the GPR2 and how this relates to ESkine expression/function. Inter- additions. Un.L1.2, Untransfected L1.2 control cells. estingly, a large number of ESkine-like expressed sequence tags that are derived from epidermal tissues have been identified; there- fore, it is likely that the skin is a major source of ESkine expres- ceptors exist for ESkine. Interestingly, although ESkine is highly sion. Indeed, during the preparation of this manuscript, ESkine was divergent from other CC chemokines, it is most highly homolo-

Downloaded from independently isolated as CTACK and shown to preferentially at- gous to LARC and TECK, the ligands for CCR6 and CCR9, re- tract skin homing CLAϩ memory T cells (17). However, in addi- spectively (14). Both CCR6 and CCR9 are expressed in lymphoid tion to CLAϩ T cells, ESkine is also functional on activated (but tissues (24, 20), and, in addition, CCR9 is selectively expressed on not naive) CD4ϩ T cells (14). We have been unable to demonstrate intestinal homing T lymphocytes and mucosal lymphocytes. Hope- the expression of mGPR2 in murine skin, either by Northern blot fully, the exact role of CCR10/GPR2 and ESkine will become analysis or by RT-PCR (data not shown). However, this does not more evident following further characterization of the ESkine li- preclude the expression of low levels of GPR2 in a rare circulating gand, identification of the cell types expressing GPR2 and re- population of cells, such as CLAϩ memory T cells. Therefore, the sponding to ESkine, and by the generation of mice lacking the pairing of GPR2 with ESkine is an intriguing one, particularly CCR10/GPR2 gene. given the highly unusual nature of the ligand as recently reported (14, 17) and the strong lymphoid and intestinal expression ob- References served for GPR2 as reported herein. Although ESkine was the only 1. Luster, A. D. 1998. Chemokines—chemotactic cytokines that mediate inflam- chemokine that seemed to function as a specific ligand for GPR2, mation. N. Engl. J. Med. 338:346. 2. Rollins, B. J. 1997. Chemokines. Blood 90:909. we cannot exclude the fact that additional novel ligands for GPR2 3. Tiffany, H. L., L. L. Lautens, J. L. Gao, J. Pease, M. Locati, C. Combadiere, are likely to exist, and indeed it is also likely that additional re- W. Modi, T. I. Bonner, and P. M. Murphy. 1997. Identification of CCR8: a 3464 CUTTING EDGE

human monocyte and thymus receptor for the CC chemokine I-309. J. Exp. Med. 16. Ishikawa-Mochizuki, I., M. Kitaura, M. Baba, T. Nakayama, D. Izawa, T. Imai, 186:165. H. Yamada, K. Hieshima, R. Suzuki, H. Nomiyama, and O. Yoshie. 1999. Mo- 4. Zaballos, A., J. Gutie´rrez, R. Varona, C. Ardavı´n, and G. Ma´rquez. 1999. Iden- lecular cloning of a novel CC chemokine, interleukin-11 receptor ␣-locus che- tification of the orphan chemokine receptor GPR-9–6 as CCR9, the receptor for mokine (ILC), which is located on chromosome 9p13 and a potential homologue the chemokine TECK. J. Immunol. 162:5671. of a CC chemokine encoded by Molluscum contagiosum virus. FEBS Lett. 460: 5. Gunn, M. D., V. N. Ngo, K. M. Ansel, E. H. Ekland, J. G. Cyster, and 544. L. T. Williams. 1998. A B-cell homing chemokine made in lymphoid follicles 17. Morales, J., B. Homey, A. P. Vicari, S. Hudak, E. Oldham, J. Hedrick, R. Orozco, activates Burkitt’s lymphoma receptor-1. Nature 391:799. N. G. Copeland, N. A. Jenkins, L. M. McEvoy, and A. Zlotnik. 1999. CTACK, 6. Yoshida, T., T. Imai, M. Kakizaki, M. Nishimura, S. Takagi, and O. Yoshie. a skin-associated chemokine that preferentially attracts skin-homing memory T 1998. Identification of single C motif-1/lymphotactin receptor XCR1. J. Biol. cells. Proc. Natl. Acad. Sci. USA 96:14470. Chem. 273:16551. 18. Gunn, M. D., S. Kyuwa, C. Tam, T. Kakiuchi, A. Matsuzawa, L. T. Williams, and 7. Fan, P., H. Kyaw, K. Su., Z. Zeng, M. Augustus, K. C. Carter, and Y. Li. 1998. H. Nakano. 1999. Mice lacking expression of secondary lymphoid organ che- Cloning and characterization of a novel human receptor. Biochem. Biophys. Res. mokine have defects in lymphocyte homing and dendritic cell localization. Commun. 243:264. J. Exp. Med. 189:451. 8. Heesen, M., M. A. Berman, A. Charest, D. Housman, C. Gerard, and M. E. Dorf. 19. Dieu, M.-C., B. Vanbervliet, A. Vicari, J.-M. Bridon, E. Oldham, S. Aı¨t-Yahia, 1998. Cloning and chromosomal mapping of an orphan chemokine receptor: F. Brie`re, A. Zlotnik, S. Lebecque, and C. Caux. 1998. Selective recruitment of mouse RDC1. Immunogenetics 47:364. immature and mature dendritic cells by distinct chemokines expressed in different 9. Liao, F., G. Alkhatib, K. W. C. Peden, G. Sharma, E. A. Berger, and J. M. Farber. anatomic sites. J. Exp. Med. 188:373. 1997. STRL33, A novel chemokine receptor-like protein, functions as a fusion 20. Zabel, B. A., W. W. Agace, J. J. Cambell, H. M. Heath, D. Parent, A. I. Roberts, cofactor for both macrophage-tropic and T cell line-tropic HIV-1. J. Exp. Med. E. C. Ebert, N. Kassam, S. Qin, M. Zovko, et al. 1999. Human G protein-coupled 185:2015. receptor GPR-9–6/CC chemokine receptor 9 is selectively expressed on intesti- 10. Hromas, R., H. E. Broxmeyer, C. Kim, H. Nakshatri, K. Christopherson II, nal homing T lymphocytes, mucosal lymphocytes, and thymocytes and is re- M. Azam, and Y-H. H. 1999. Cloning of BRAK, a novel divergent CXC che- quired for thymus-expressed chemokine-mediated chemotaxis. J. Exp. Med. 190: mokine preferentially expressed in normal versus malignant cells. Biochem. Bio- 1241. phys. Res. Commun. 255:703. 21. Lu, B., A. Humbles, D. Bota, C. Gerard, B. Moser, D. Soler, A. D. Luster, and 11. Hieshima, K., T. Imai, M. Baba, K. Shoudai, K. Ishizuka, T. Nakagawa, N. P. Gerard. 1999. Structure and function of the murine chemokine receptor J. Tsuruta, M. Takeya, Y. Sakaki, K. Takatuski, et al. 1997. A novel human CC CXCR3. Eur. J. Immunol. 29:3804. chemokine PARC that is most homologous to macrophage-inflammatory protein- 22. Jarmin, D. I., R. J. B. Nibbs, T. Jamieson, J. S. DeBono, and G. J. Graham. 1999. 1␣/LD78 ␣ and chemotactic for T lymphocytes, but not for monocytes. J. Im- Granulocyte-macrophage colony-stimulating factor and interleukin-3 regulate munol. 159:1140. chemokine and chemokine receptor expression in bone marrow macrophages. 12. Rossi, D. L., S. D. Hurst, Y. Xu, W. Wang, S. Menon, R. L. Coffman, and Exp. Hematol. 27:1735. A. Zlotnik. 1999. Lungkine, a novel CXC chemokine, specifically expressed by 23. Abel, K. J., L. C. Brody, J. M. Valdes, M. R. Erdos, D. R. McKinley, lung bronchoepithelial cells. J. Immunol. 162:5490. L. H. Castilla, S. D. Merajver, F. J. Couch, L. S. Freidman, E. A. Ostermeyer, et 13. Marchese, A., J. M. Docherty, T. Nguyen, M. Heiber, R. Cheng, H. H. Q. Heng, al. 1996. Characterization of EZH1, a human homolog of Drosophila enhancer of L.-C, Tsui, X. Shi, S. R. George, and B. F. O’Dowd. 1994. Cloning of human zeste near BRCA1. Genomics 37:161. genes encoding novel G protein-coupled receptors. Genomics 23:609. 24. Schweickart, V. L., C. J. Raport, R. Godiska, M. G. Byers, R. J. Eddy, 14. Baird, J. W., R. J. B. Nibbs, M. Komai-Koma, J. A. Connolly, K. Ottersbach, T. B. Shows, and P. W. Gray. 1994. Cloning of human and mouse EBI1, a I. Clark-Lewis, F. Y. Liew, and G. J. Graham. 1999. ESkine, a novel ␤-chemo- lymphoid-specific G-protein-coupled receptor encoded on human chromosome kine, is differentially spliced to produce secretable and nuclear targeted isoforms. 17q12–q21. 2. Genomics 23:643. J. Biol. Chem. 274:33496. 25. Baba, M., T. Imai, M. Nishimura, M. Kakizaki, S. Takagi, K. Hieshima, 15. Hromas, R., H. E. Broxmeyer, C. Kim, K. Christopherson II, and Y.-H. Hou. H. Nomiyama, and O. Yoshie. 1997. Identification of CCR6, the specific receptor 1999. Isolation of ALP, a novel divergent murine CC chemokine with a unique for a novel lymphocyte-directed CC chemokine LARC. J. Biol. Chem. carboxy terminal extension. Biochem. Biophys. Res. Commun. 258:737. 272:14893. by guest on October 1, 2021. Copyright 2000 Pageant Media Ltd. https://www.jimmunol.org Downloaded from