Dissecting the interactions between TCR signaling strength and inhibitory pathways

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PROJECT SUMMARY/ABSTRACT This 5-year K08 training program is designed to facilitate my (Dr. Lin Shen) career development in preparation of my independent research career as a physician-scientist. My long-term goal is to study the pathogenesis and, potentially, the therapy of autoimmune diseases. This proposal focuses on how inhibitory pathways restrain abnormal TCR signaling strength to achieve peripheral tolerance. Central guidance and the training environment will be provided by my primary mentor, Dr. Arthur Weiss, an expert in TCR signaling. Further expertise will be provided by a strong advisory panel of experts in lymphocyte signaling, immune tolerance, autoimmunity and transcriptomic profiling. A training plan with experimental research, didactics, and seminars has been developed to advance me towards my career goals. The extensive resources and support at the UCSF will facilitate my pathway to independence and my long-term goals. Dysregulation of TCR signaling has long been recognized to play important roles in the pathogenesis of autoimmune diseases. However, how inhibitory pathways are engaged by enhanced TCR signaling strength to regulate peripheral tolerance and prevent autoimmunity remains incompletely understood. The cytoplasmic tyrosine kinase ZAP70 plays a requisite role in TCR signaling. Two ZAP70 hypermorphic mutants, the stronger W131A and the weaker R360P, interfere with autoinhibition and have varying effects on TCR signaling. As a consequence of enhanced basal signaling, OTII-TCR transgenic W131A T cells exhibit marked upregulation of inhibitory receptors and acquisition of an anergic phenotype. The ZAP70 R360P mouse mutant that I generated derives from a familial severe autoimmune syndrome. R360P mice failed to develop overt autoimmune disease on a C57BL/6 background but exhibited expansion of regulatory and anergic T cells. Strikingly, OTI-TCR transgenic R360P T cells showed enhanced responses to weak and self-peptides. Introducing deficiency in Cbl-b, a negative regulator for TCR signaling, resulted in enhanced R360P-OTI T cell responses to a weak agonist peptide and reversed functional anergy in W131A-OTII T cells. Therefore, I hypothesize that increased TCR signaling strength renders T cells subject to greater control by inhibitory pathways but may confer upon them greater sensitivity to disruption of these inhibitory pathways. In this proposal, I will take advantage of the R360P and W131A mouse models to study how enhanced TCR signaling strength and inhibitory pathways are coupled together to either maintain or subvert peripheral tolerance. In Aim 1, using a range of biochemical techniques and transcriptomic profiling, I will dissect how TCR signaling strength is dynamically coupled to inhibitory programs. In Aim 2, I will study whether augmented TCR signaling strength promotes sensitivity to the disruption of negative regulatory pathways. This study will provide new mechanistic insights into how these opposing signaling pathways integrate to impact T cell antigen sensitivity and to regulate peripheral tolerance and the quality of immune responsiveness.