CD151 Expression Is Associated with a Hyperproliferative T Cell Phenotype Lillian Seu, Christopher Tidwell, Laura Timares, Alexandra Duverger, Frederic H. Wagner, Paul A. Goepfert, Andrew O. This information is current as Westfall, Steffanie Sabbaj and Olaf Kutsch of September 26, 2021. J Immunol published online 27 September 2017 http://www.jimmunol.org/content/early/2017/09/27/jimmun ol.1700648 Downloaded from Supplementary http://www.jimmunol.org/content/suppl/2017/09/27/jimmunol.170064 Material 8.DCSupplemental 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 • Fast Publication! 4 weeks from acceptance to publication by guest on September 26, 2021 *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 © 2017 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. Published September 27, 2017, doi:10.4049/jimmunol.1700648 The Journal of Immunology CD151 Expression Is Associated with a Hyperproliferative T Cell Phenotype Lillian Seu, Christopher Tidwell, Laura Timares, Alexandra Duverger, Frederic H. Wagner, Paul A. Goepfert, Andrew O. Westfall, Steffanie Sabbaj, and Olaf Kutsch The tetraspanin CD151 is a marker of aggressive cell proliferation and invasiveness for a variety of cancer types. Given reports of CD151 expression on T cells, we explored whether CD151 would mark T cells in a hyperactivated state. Consistent with the idea that CD151 could mark a phenotypically distinct T cell subset, it was not uniformly expressed on T cells. CD151 expression frequency was a function of the T cell lineage (CD8 > CD4) and a function of the memory differentiation state (naive T cells < central memory T cells < effector memory T cells < T effector memory RA1 cells). CD151 and CD57, a senescence marker, defined the same CD282 T cell populations. However, CD151 also marked a substantial CD28+ T cell population that was not marked by CD57. Kinome array analysis demonstrated that CD28+CD151+ T cells form a subpopulation with a distinct molecular baseline and Downloaded from activation phenotype. Network analysis of these data revealed that cell cycle control and cell death were the most altered process motifs in CD28+CD151+ T cells. We demonstrate that CD151 in T cells is not a passive marker, but actively changed the cell cycle control and cell death process motifs of T cells. Consistent with these data, long-term T cell culture experiments in the presence of only IL-2 demonstrated that independent of their CD28 expression status, CD151+ T cells, but not CD1512 T cells, would exhibit an Ag-independent, hyperresponsive proliferation phenotype. Not unlike its reported function as a tumor aggressiveness marker, CD151 in humans thus marks and enables hyperproliferative T cells. The Journal of Immunology, 2017, 199: 000–000. http://www.jimmunol.org/ cell homeostasis is essential to maintain the naive T cell proteins, including kinases (10–15) and phosphatases (16–20). (TN) population, as well as the T cell populations that Misalignment of any of the components that form this highly T constitute our immunological memory (1–4). In healthy complex network can result in increased T cell proliferation, a individuals, T cell homeostasis is tightly controlled through the lowered threshold for Ag activation, or a loss of immune tolerance integration of a wide variety of mechanisms. Homeostatic T cell and the development of autoimmune phenotypes (8, 21, 22). A proliferation is driven by various combinations of cytokines and protein family that has a functional profile compatible with low-affinity interactions with self-antigen (5–8). Accordingly, influencing the T cell activation state via their ability to mobilize by guest on September 26, 2021 known receptors involved in T cell homeostasis include, but are intracellular T cell signaling molecules would be tetraspanins, but certainly not limited to, cytokine receptors such as IL-7R or IL- a potential role of tetraspanins in T cell homeostasis or mis- 15R (9), the TCR, as well as costimulatory molecules such as alignment thereof has not been explored. CD28. At the cellular level, the involved signaling pathways that In mammalians, tetraspanins are a family of 33 different small transduce these signals into the cells involve a plethora of different proteins with 4 transmembrane-spanning domains that are known to regulate cell morphology, motility, invasion, and fusion, and that Department of Medicine, University of Alabama at Birmingham, Birmingham, AL are involved in cell proliferation and apoptosis in a variety of 35294 different cell types (for reviews, see Refs. 23–28). Tetraspanins ORCIDs: 0000-0002-8456-019X (L.S.); 0000-0002-9739-2244 (C.T.); 0000-0001- have an exceptional ability to coordinate cell surface molecules 8886-8354 (L.T.); 0000-0001-8441-5737 (P.A.G.); 0000-0002-0468-4695 (A.O.W.); 0000-0003-4052-6819 (S.S.); 0000-0002-1853-8023 (O.K.). laterally binding to other tetraspanins or to other membrane pro- Received for publication May 4, 2017. Accepted for publication August 28, 2017. teins (integrins, CD2, CD4, CD8, CD5, CD19, CD21, Fc recep- This work was supported by National Institutes of Health (NIH) Grants F32 tors, MHC class I, and MHC class II) (reviewed in Ref. 26) to HL121924 (to L.S.) and R01-AI104499, R56-AI122842, and R21-AI116188 (to form the highly specialized structure known as tetraspanin-enriched O.K.). Parts of this work were performed in the University of Alabama (UAB) Center microdomains (29, 30) or tetraspanin webs (31). Tetraspanins, likely for AIDS Research BSL-3 facilities and by the UAB Center for AIDS Research Flow Cytometry Core/Joint UAB Flow Cytometry Core, which are funded by NIH/ as a function of the cell type, have been described to influence Rac, National Institute of Allergy and Infectious Diseases Grant P30 AI027767 and by PKC, and CDC42 activity (32, 33) or Ras, ERK/MAPK1/2, and NIH Grant 5P30 AR048311. protein kinase B/Akt signaling (34, 35). Also, PKCa or PI4K has The microarray data presented in this article have been submitted to the ArrayEx- press database (https://www.ebi.ac.uk/arrayexpress/) under accession numbers E- been reported to associate closely with certain tetraspanins, includ- MTAB-5978 and E-MTAB-5977. ing CD9 and CD81 (23, 36, 37). Address correspondence and reprint requests to Dr. Lillian Seu and Dr. Olaf Kutsch, Some tetraspanins (CD9, CD37, CD81, CD151) have been Department of Medicine, University of Alabama at Birmingham, BBRB 510, 845 reported to be expressed on T cells, where their function has 19th Street South, Birmingham, AL 35294. E-mail addresses: [email protected] (L.S.) and [email protected] (O.K.) mostly been described using knockout mouse models. With only The online version of this article contains supplemental material. little experimental evidence having been accumulated using human T cells, it nevertheless seems clear that tetraspanins affect T cell Abbreviations used in this article: GO, Gene Ontology; IS, immunologic synapse; NIH, National Institutes of Health; PIN, protein–protein interaction network; TCM, activation in ways similar to costimulation (25, 38–42). We central memory T cell; TEM, effector memory T cell; TEMRA, T effector memory were particularly interested to detail the role of CD151 in T cell RA1 cell; T , naive T cell. N biology, because increased CD151 expression has been associated Copyright Ó 2017 by The American Association of Immunologists, Inc. 0022-1767/17/$35.00 with aggressive cell proliferation and invasiveness for a variety of www.jimmunol.org/cgi/doi/10.4049/jimmunol.1700648 2 CD151 MARKS HYPERRESPONSIVE T CELLS different tumor types, suggesting that its altered expression may an intracellular protein that is exclusively expressed in cycling cells in the be linked to a hyperactivated T cell state. For example, increased late G1,G2, S, and M phases, and accordingly, it is absent in resting cells CD151 expression has been associated with tumor aggressiveness (G0) (57) and is therefore frequently used to detect actively cycling cells. Flow cytometric analysis and cell sorting experiments were performed in breast cancer, endometrial cancer, esophageal squamous cell using an LSR II flow cytometer and FACSAria II instrument, respectively carcinoma, epithelial malignancies, non–small cell lung cancer, (Becton Dickinson). Gates for flow cytometric acquisition and analyses pancreatic ductal adenocarcinoma, colon cancer, or prostate can- were based on “fluorescence-minus-one” controls and single-stain com- cer (43–51). Genetic ablation of CD151 expression has been pensation controls. Data were analyzed using FlowJo software (Tree Star, Ashland, OR). reported to reduce metastatic spread of cancer, providing func- tional evidence that increased CD151 expression may actively T cell proliferation assays contribute to this aggressive phenotype (52). In immunology, For proliferation assays, the T cells were labeled with 1 mM CFSE CD151 has been reported as a negative regulator of FcεRI-mediated (Invitrogen). Cells were plated at 5 3 105 cells per well in 96-well flat- mast cell activation (53). On T cells, CD151 has been shown to bottom plates and incubated for 5 d after addition of IL-2. The extent of stabilize the immunological synapse after T cell activation, and T cell proliferation is proportional to the decrease in CFSE fluorescence, silencing of CD151 blunts IL-2 secretion and expression of the with every cell division decreasing the CFSE fluorescence intensity of the daughter cells by 50%.