CCL28 Has Dual Roles in Mucosal Immunity as a Chemokine with Broad-Spectrum Antimicrobial Activity This information is current as Kunio Hieshima, Haruo Ohtani, Michiko Shibano, Dai of September 26, 2021. Izawa, Takashi Nakayama, Yuri Kawasaki, Fumio Shiba, Mitsuru Shiota, Fuminori Katou, Takuya Saito and Osamu Yoshie J Immunol 2003; 170:1452-1461; ; doi: 10.4049/jimmunol.170.3.1452 Downloaded from http://www.jimmunol.org/content/170/3/1452 References This article cites 36 articles, 19 of which you can access for free at: http://www.jimmunol.org/content/170/3/1452.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 26, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2003 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology CCL28 Has Dual Roles in Mucosal Immunity as a Chemokine with Broad-Spectrum Antimicrobial Activity1 Kunio Hieshima,* Haruo Ohtani,2‡ Michiko Shibano,* Dai Izawa,* Takashi Nakayama,* Yuri Kawasaki,* Fumio Shiba,* Mitsuru Shiota,† Fuminori Katou,§ Takuya Saito,¶ and Osamu Yoshie3* CCL28 is a CC chemokine signaling via CCR10 and CCR3 that is selectively expressed in certain mucosal tissues such as exocrine glands, trachea, and colon. Notably, these tissues commonly secrete low-salt fluids. RT-PCR analysis demonstrated that sali- vary glands expressed CCL28 mRNA at the highest levels among various mouse tissues. Single cells prepared from mouse parotid glands indeed contained a major fraction of CD3؊B220low cells that expressed CCR10 at high levels and CCR3 at low levels and responded to CCL28 in chemotaxis assays. Morphologically, these cells are typical plasma cells. By immunohistochemistry, acinar epithelial cells in human and mouse salivary glands were strongly positive for CCL28. Furthermore, human saliva and milk Downloaded from were found to contain CCL28 at high concentrations. Moreover, the C terminus of human CCL28 has a significant sequence similarity to histatin-5, a histidine-rich candidacidal peptide in human saliva. Subsequently, we demonstrated that human and mouse CCL28 had a potent antimicrobial activity against Candida albicans, Gram-negative bacteria, and Gram-positive bacteria. The C-terminal 28-aa peptide of human CCL28 also displayed a selective candidacidal activity. In contrast, CCL27, which is most similar to CCL28 and shares CCR10, showed no such potent antimicrobial activity. Like most other antimicrobial peptides, http://www.jimmunol.org/ CCL28 exerted its antimicrobial activity in low-salt conditions and rapidly induced membrane permeability in target microbes. Collectively, CCL28 may play dual roles in mucosal immunity as a chemoattractant for cells expressing CCR10 and/or CCR3 such as plasma cells and also as a broad-spectrum antimicrobial protein secreted into low-salt body fluids. The Journal of Immunology, 2003, 170: 1452–1461. hemokines are known to play pivotal roles in innate and CCR10 was originally identified as the receptor for CCL27 (also acquired immunity by regulating migration and activa- called IL-11R ␣-locus chemokine or cutaneous T cell-attracting C tion of leukocytes via a group of seven transmembrane G chemokine) (5, 6), whereas CCR3 is the receptor for eotaxin/ protein-coupled receptors (1). In humans, more than 45 members CCL11 and many other chemokines known to act on eosinophils by guest on September 26, 2021 and 18 functional receptors have been identified (1). According to (1). CCL28 transcripts are detected in a variety of tissues but most the arrangement of the amino-terminal conserved cysteine resi- abundantly in trachea, colon, rectum, and exocrine glands such as dues, the chemokines are classified into four subfamilies: CXC, salivary and mammary glands (3, 4). In the colon, CCL28 is se- CC, C, and CX3C (1). Recently, based on the classification of lectively expressed by the epithelial cells (3, 4). Structurally, these four subfamilies, a systematic nomenclature system of the CCL28 is most similar to CCL27, which is expressed highly se- chemokine ligands has been formulated (2). Except for the two lectively in the skin and has been shown to attract cutaneous lym- transmembrane-type chemokines, CX3CL1 and CXCL16, chemo- phocyte Agϩ memory T cells via CCR10 (5–8). Consistently, kines are small (8–14 kDa), mostly cationic polypeptides with two CCL28 has also been shown to attract cutaneous lymphocyte Agϩ to three intramolecular disulfide bonds (1). CCL28 (also called memory T cells via CCR10 and eosinophils via CCR3 among hu- mucosae-associated epithelial chemokine) is a recently described man peripheral blood leukocytes (3). CC chemokine signaling via CCR10 as well as CCR3 (3, 4). Antimicrobial peptides, now known by Ͼ500 in number, are the diverse family of small, mostly cationic polypeptides found widely Departments of *Microbiology and †Gynecology and Obstetrics, Kinki University in all forms of multicellular organisms that exert a broad spectrum School of Medicine, Osaka, Japan; ‡Department of Pathology, Tohoku University of cytotoxic activity against bacteria, fungi, parasites, and envel- § Graduate School of Medicine, Miyagi, Japan; Department of Oral and Maxillofacial oped viruses (9, 10). In mammals, such peptides are particularly Surgery 1, Tohoku University School of Dentistry, Miyagi, Japan; and ¶Department of Biotechnological Science, Kinki University, Wakayama, Japan abundant in the storage granules of phagocytic cells and on the Received for publication August 26, 2002. Accepted for publication November surface of mucosal tissues. Accumulating evidence points out that 20, 2002. chemokines and antimicrobial peptides have substantially overlap- The costs of publication of this article were defrayed in part by the payment of page ping functions (11). For example, several members of the mam- charges. This article must therefore be hereby marked advertisement in accordance malian antimicrobial proteins are capable of attracting leukocytes with 18 U.S.C. Section 1734 solely to indicate this fact. via interactions with selected seven-transmembrane G protein-cou- 1 This work was supported by grants-in-aid and a High-Tech Research Center Grant from the Ministry of Education, Culture, Sports, Science and Technology, Japan, and pled receptors (12–14). Most notably, Yang et al. (15) have re- by Solution Oriented Research for Science and Technology of the Japan Science and cently demonstrated that human ␤-defensins are potent agonists Technology Corporation. for CCR6, the receptor for the chemokine CCL20 (also called liver 2 Current address: Department of Clinical Laboratory Research, National Mito Hos- and activation-regulated chemokine or macrophage inflammatory pital, 3-2-1 Higashihara, Mito, Ibaragi 310-0035, Japan. protein-3␣), which is produced by mucosal epithelial cells and 3 Address correspondence and reprint requests to Dr. Osamu Yoshie, Department of Microbiology, Kinki University School of Medicine, 377-2 Ohno-Higashi, Osaka- epidermal keratinocytes upon proinflammatory stimulations that Sayama, Osaka 589-8511, Japan. E-mail address: [email protected] selectively attracts immature dendritic cells, memory T cells, and Copyright © 2003 by The American Association of Immunologists, Inc. 0022-1767/03/$02.00 The Journal of Immunology 1453 B cells (1, 16). In contrast, some chemokines have been shown to binding to Fc receptors (22). Human embryonic kidney 293T cells were have antimicrobial activity. Krijgsved et al. (17) purified antibac- transiently transfected with the CCL27-Fc and control Fc constructs, and terial proteins stored in the ␣-granules of human platelets and de- CCL27-Fc and control Fc proteins were purified from the culture super- natants by using Protein A-agarose (21). Purified CCL27-Fc and control Fc termined their amino acid sequences. Surprisingly, these proteins were 76 kDa and 55 kDa, respectively, which was in agreement with the turned out to be CXCL7 (59–126) and CXCL7 (44–126), two predicted molecular mass. By using a panel of L1.2 cell lines covering all related chemokine variants processed from a common precursor human and murine CC chemokine receptors (CCR1–10), we confirmed that platelet basic protein and truncated by 2 aa in the C terminus (17). CCL27-Fc specifically bound to human and murine CCR10 (data not shown). Furthermore, Cole et al. (18), by examining a panel of 11 chemo- kines representing all four chemokine subfamilies, demonstrated Preparation of single cells from mouse parotid glands that the three IFN-inducible non-ELR (Glu-Leu-Arg)-motif CXC Female BALB/c mice were purchased from CLEA Japan (Osaka, Japan). chemokines, monokine induced by IFN-␥/CXCL9, IFN-inducible After cervical dislocation, parotid glands were excised from 12- to 16-wk- protein-10/CXCL10, and IFN-inducible T-cell ␣ chemoattractant/ old mice, carefully avoiding