STAT3 PROMOTES T-CELL SURVIVAL AND INHIBITS IL-2 PRODUCTION THROUGH UP-REGULATION OF CLASS O FORKHEAD TRANSCRIPTION FACTORS Hyun-Mee Oh * , Cheng-Rong Yu * , Nady Golestaneh † , Ahjoku Amadi-Obi * , Yun Sang Lee * , Amarachi Eseonu ‡ , Rashid M. Mahdi * and Charles E. Egwuagu *§ * Molecular Immunology Section, Laboratory of Immunology, National Eye Institute, National Institutes of Health, Bethesda, Maryland, USA; † Georgetown University School of Medicine, Department of Biochemistry and Molecular & Cellular Biology, Washington, D.C; ‡ Department of Biomedical Engineering, Harvard College, Harvard University, Cambridge, Massachusetts Running title: STAT3 Promotes T cell Survival and Inhibits IL-2 Production Conflict of interest: The authors have declared that no conflict of interest exists. ‡ Address correspondence: Dr. Charles E. Egwuagu. Molecular Immunology Section, NEI, National Institutes of Health, 10/10N116, 10 Center Drive, Bethesda, MD 20892; Tel: (301) 496-0049; E-mail: egwuaguc@nei.nih.gov Much is known about the role of STAT3 in regulating differentiation of Interleukin-17- producing Th17 cells but its function in other lymphocyte subsets is not well understood. In this report, we reveal wide-ranging functions of STAT3 in T-cells and provide evidence that STAT3 is convergence point for mechanisms that regulate lymphocyte quiescence and those controlling T-cell activation and survival. We show here that STAT3 inhibits T-lymphocyte proliferation by up-regulating the expression of Class O Forkhead transcription factors, which play essential roles in maintaining T cells in quiescent state. We further show that STAT3 binds directly to FoxO1 or FoxO3a promoter and that STAT3-deficiency resulted in downregulation of the expression of FoxO1, FoxO3a and FoxO-target genes (Iκ B and p27Kip1). Compared to wild-type T cells, STAT3-deficient T cells produced more IL-2, due in part, to marked decrease in Iκ B- mediated sequestration of NF-κ B in the cytoplasm and resultant enhancement of NF- κ B activation. However, the high level of IL-2 production by STAT3-deficient T cells was partially restored to normal levels by over- expressing FoxO1. It is notable that their exaggerated increase in IL-2 production rendered STAT3-deficient lymphocytes more susceptible to activation-induced cell death, suggesting that STAT3 might protect T cells from apoptosis by limiting their production of IL-2 through upregulation of FoxO1/FoxO3a expression. Moreover, we found that STAT3 enhanced survival of activated T cells by up- regulating OX-40 and Bcl-2 while down- regulating FasL and Bad expression, suggesting that similar to role of FoxOs in regulating the lifespan of worms, STAT3 and FoxO pathways converge to regulate lifespan of T lymphocytes. Introduction Production of IL-2 is one of the hallmarks of TCR signaling and largely responsible for T cell proliferation and expansion (1). However, transcription of IL-2 is actively suppressed in resting T cells by T cell quiescence factors (2,3). Members of the FoxO (Forkhead box, class-O) subfamily of Forkhead transcription factors are important T-lymphocyte quiescence factors with broad influence on cell growth and survival. FoxO proteins maintain naïve or resting T cells in quiescent state (G 0 cell cycle phase) by up- regulating the expression of cell cycle inhibitors, such as p27kip1, Gadd45, cyclin E and p130 (4). When the resting T cell encounters its cognate Ag in context of APC, TCR-mediated activation of Ras, PI3K/Akt and mTOR kinases inactivate FoxOs by inducing their phosphorylation and translocation from the nucleus to the cytoplasm through 14-3-3-dependent mechanisms (5). The expulsion from the nucleus relieves the T cell from inhibitory effects of FoxOs and allows cell cycle progression (3,6). FoxOs also regulate T cell quiescence by inducing expression of IκB, a protein that interacts with and sequesters NF-κB in the cytoplasm (7). Thus, by sequestrating NF-κB in the cytoplasm the naïve T cell is deprived of a transcription factor required for transcription of IL-2 (8). IL-2 is a growth and survival factor for T cells (6). It promotes the expression of anti- apoptotic Bcl-2 members while inhibiting pro- apoptotic members (e.g. Bim) through PI3K/AKT- http://www.jbc.org/cgi/doi/10.1074/jbc.M111.253500 The latest version is at JBC Papers in Press. Published on July 5, 2011 as Manuscript M111.253500 Copyright 2011 by The American Society for Biochemistry and Molecular Biology, Inc. by guest on November 14, 2016 http://www.jbc.org/ Downloaded from by guest on November 14, 2016 http://www.jbc.org/ Downloaded from by guest on November 14, 2016 http://www.jbc.org/ Downloaded from