Tetraspanin CD151 Is a Negative Regulator of Fc εRI-Mediated Mast Cell Activation Hiam Abdala-Valencia, Paul J. Bryce, Robert P. Schleimer, Joshua B. Wechsler, Lucas F. Loffredo, Joan M. Cook-Mills, This information is current as Chia-Lin Hsu and Sergejs Berdnikovs of September 26, 2021. J Immunol 2015; 195:1377-1387; Prepublished online 1 July 2015; doi: 10.4049/jimmunol.1302874 http://www.jimmunol.org/content/195/4/1377 Downloaded from Supplementary http://www.jimmunol.org/content/suppl/2015/07/01/jimmunol.130287 Material 4.DCSupplemental http://www.jimmunol.org/ References This article cites 63 articles, 28 of which you can access for free at: http://www.jimmunol.org/content/195/4/1377.full#ref-list-1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision by guest on September 26, 2021 • No Triage! Every submission reviewed by practicing scientists • 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 © 2015 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Tetraspanin CD151 Is a Negative Regulator of Fc«RI-Mediated Mast Cell Activation Hiam Abdala-Valencia,* Paul J. Bryce,* Robert P. Schleimer,* Joshua B. Wechsler,† Lucas F. Loffredo,* Joan M. Cook-Mills,* Chia-Lin Hsu,* and Sergejs Berdnikovs* Mast cells are critical in the pathogenesis of allergic disease due to the release of preformed and newly synthesized mediators, yet the mechanisms controlling mast cell activation are not well understood. Members of the tetraspanin family are recently emerging as mod- ulators of Fc«RI-mediated mast cell activation; however, mechanistic understanding of their function is currently lacking. The tetra- spanin CD151 is a poorly understood member of this family and is specifically induced on mouse and human mast cells upon Fc«RI aggregation but its functional effects are unknown. In this study, we show that CD151 deficiency significantly exacerbates the IgE- mediated late phase inflammation in a murine model of passive cutaneous anaphylaxis. Ex vivo, Fc«RI stimulation of bone marrow– derived mast cells from CD1512/2 mice resulted in significantly enhanced expression of proinflammatory cytokines IL-4, IL-13, and Downloaded from TNF-a compared with wild-type controls. However, Fc«RI-induced mast cell degranulation was unaffected. At the molecular signaling level, CD151 selectively regulated IgE-induced activation of ERK1/2 and PI3K, associated with cytokine production, but had no effect on the phospholipase Cg1 signaling, associated with degranulation. Collectively, our data indicate that CD151 exerts negative regulation over IgE-induced late phase responses and cytokine production in mast cells. The Journal of Immunology, 2015, 195: 1377–1387. he high-affinity receptor for IgE (FcεRI) is a principal type allergic reactions, whereas mast cell–mediated late phase http://www.jimmunol.org/ mast cell receptor mediating immune responses in allergic reactions and IgE-induced chronic allergic inflammatory processes T diseases (1). Crosslinking of IgE-bound FcεRI by Ags are mainly dependent on the de novo production of inflammatory activates downstream signal transduction pathways, resulting in mediators (9, 10). At the same time, receptors bearing ITIM and mast cell degranulation and de novo synthesis of cytokines (2–5). ITAM motifs, protein tyrosine kinases, protein and lipid phospha- Proximal signaling through FcεRI involves phosphorylation of tases, adaptors, and ubiquitin ligases provide a diverse regulatory ITAMs within the FcεRI b and g subunits by the Src-family protein network to achieve the desired response and limit a persistent or tyrosine kinases Lyn, spleen tyrosine kinase (Syk), and Fyn (6, 7). excessive “outside–in” signaling for mast cell activation (11–20). In particular, activation of Syk is indispensable for FcεRI-mediated Members of the tetraspanin family are classically recognized as mast cell activation (5). This tyrosine kinase signaling induces two “passive” facilitators that function as scaffolds in the assembly of by guest on September 26, 2021 principal downstream signaling cascades: the phospholipase Cg1 signaling complexes at the cell membrane (21). Only recently, tet- (PLCg1)–protein kinase C (PKC)–Ca2+ cascade, which is required raspanins have started to emerge as “active” signaling molecules for degranulation and the release of preformed mediators stored in modulating outside–in signals for cellular activation. For example, the mast cell’s cytoplasmic granules, and the Ras-Raf1-ERK1/2 tetraspanin CD9 negatively regulates LPS-induced macrophage ac- cascade, which is critical for de novo synthesis of cytokines (7). tivation and lung inflammation (22). It is also reported that macro- Additionally, there are complementary pathways for amplification phages from CD9 and CD81 null or CD9/CD81 double knockout and maintenance of degranulation and cytokine production. The mice show enhanced in vitro formation of multinucleated giant cells, PI3K-dependent complementary pathway involved in degranulation which are known to contribute to inflammatory tissue damage is mediated via the recruitment of Btk kinase, as well as subsequent through increased secretion of matrix metalloproteinases in vivo (23). amplification and maintenance of PLCg1-mediated latent calcium In B cells, the tetraspanin CD37 has been shown to possess inhibitory signals. Amplification of cytokine/chemokine production is regu- functions upon ligation with an anti–CD37 small modular immu- lated by PI3K via an independent pathway mediated by PDK1 and nopharmaceutical (24). In fibroblasts, CD151 has been reported to Akt signaling (8). The degranulation event is crucial for immediate- negatively regulate the adhesion-dependent activation of Ras (25). Mast cells constitutively express several members of the tetra- spanin family, although the function of these molecules in mast cell *Division of Allergy and Immunology, Department of Medicine, Northwestern Uni- ε versity Feinberg School of Medicine, Chicago, IL 60611; and †Division of Gastro- Fc RI-mediated signaling is largely unknown (26). Interaction of enterology, Hepatology and Nutrition, Department of Pediatrics, Northwestern tetraspanins with FcεRI in mast cells has been demonstrated in two University Feinberg School of Medicine, Chicago, IL 60611 previous studies using the RBL-2H3 mast cell line (27, 28). In both Received for publication October 24, 2013. Accepted for publication June 2, 2015. studies, Abs against the tetraspanins CD63 or CD81 inhibited Address correspondence and reprint requests to Dr. Sergejs Berdnikovs, Northwestern in vitro and in vivo FcεRI-mediated mast cell degranulation, without University, Department of Medicine, Division of Allergy and Immunology, 240 East Huron affecting FcεRI-mediated Ca2+ mobilization or total tyrosine phos- Avenue, M-302, Chicago, IL 60611. E-mail address: [email protected] phorylation levels (27, 28). CD63 is a diagnostic marker in allergic The online version of this article contains supplemental material. diseases (29, 30), and the granular isoform of CD63 has also been Abbreviations used in this article: BMMC, bone marrow–derived mast cell; FDR, false discovery rate; GEO, Gene Expression Omnibus; HSA, human serum albumin; reported as a molecular marker of degranulated human mast cells PCA, passive cutaneous anaphylaxis; PKC, protein kinase C; PLCg1, phospholipase (31). Recently, it was demonstrated that the tetraspanin CD63 is Cg1; qPCR, quantitative PCR; SCF, stem cell factor; Syk, spleen tyrosine kinase; required for IgE-mediated mast cell degranulation and anaphylactic WT, wild-type. response in mice, although the role of CD63 in the mechanisms that Copyright Ó 2015 by The American Association of Immunologists, Inc. 0022-1767/15/$25.00 regulate degranulation was not defined (32). Tetraspanin CD9 has www.jimmunol.org/cgi/doi/10.4049/jimmunol.1302874 1378 CD151 NEGATIVELY REGULATES MAST CELL ACTIVATION been recently reported as a regulator of mast cells chemotaxis, where of the transcriptomics of mast cell activation (P.J. Bryce, unpublished micro- aggregation of CD9 blocked Ag- and IL-13–induced chemotaxis of array data). Briefly, BMMCs from WT mice were plated on six-well plates at 3 6 bone marrow–derived mast cells (BMMCs) (33). In contrast to other a density of 5 10 cells per well. Cells were either unstimulated or coated with 1 mg/ml DNP-IgE overnight, followed by 0.5 mg/ml DNP–human serum tetraspanins, CD151 was reported to be specifically induced upon albumin (HSA) stimulation. Cells were collected 4 h after stimulation and IgE/Ag crosslinking of FcεRI receptors in umbilical cord–derived RNA was extracted using an RNeasy kit (Qiagen). Samples were processed human cells (34). However, the functional significance of expression using Illumina bead array technology by the Genomics Core Facility at of this tetraspanin in mast cells is not known. Northwestern University. Bioinformatics analysis was done using GeneSpring