蛋白酪氨酸磷酸酶PTP-PEP与14-3-3τ蛋白互作调控T细胞活化的机制研究

蛋白酪氨酸磷酸酶PTP-PEP是T细胞活化的负调节因子，其功能异常能引起多种自体免疫性疾病。但PTP-PEP对T细胞活化的调节机制尚未得到更深入的研究。本项目发现PTP-PEP可调控TCR表达，并采用酵母双杂交系统发现了PTP-PEP新的互作蛋白—14-3-3τ，明确了PTP-PEP P2结构域对二者互作至关重要。PTP-PEP可调控14-3-3τ的酪氨酸磷酸化。在T细胞中，14-3-3τ可与PTP-PEP底物c-Cbl结合，而c-Cbl为TCR降解过程的重要调控因子。据此推测，14-3-3τ作为桥梁协助PTP-PEP与c-cbl接触，并精准调控c-Cbl酪氨酸磷酸化与TCR的表达，影响活化信号传递，避免T细胞过度激活而诱发自体免疫性疾病。本项目拟解析PTP-PEP与14-3-3τ互作对TCR表达、T细胞信号传导及细胞因子分泌等功能的影响，从新角度阐明PTP-PEP调控T细胞活化的机制。

PTP-PEP is a member of proline-, glutamic acid-, serine- and threonine-rich (PEST) family of protein tyrosine phosphatase (PTP), known as proline-enriched phosphatase (PEP). The C-terminal of PTP-PEP contains four proline-rich motif domains, P1, P2, P3 and P4, which are associated with protein interaction. P1 motif binds to Src homology 3 (SH3) domain of protein tyrosine kinase Csk to form complex and suppresses phosphorylation of Csk. Originally, the researchers expressed a substrate-trapping mutant PTP-PEP C227S, in pervanadate-treated T cells and led to the identification of Lck, Fyn (Src homology family protein tyrosine kinases, PTKs), TCR-CD3ζ, and Zap-70 (ζ-associated protein kinase 70) as primary substrates of PTP-PEP, which played the important roles in TCR signaling pathway. Tons of experiments demonstrated that PTP-PEP inhibited TCR signaling by dephosphorylation of the TCR signaling downstream regulators, to avoid the excessive activation of the immune system and protect the body. In recent years, most studies have focused on that a single nucleotide polymorphism (SNP) of PTP-PEP/PTPN22 gene can lead to various Autoimmune Diseases. However, there was no in-depth analysis about PTP-PEP on its potential regulatory mechanism of T cell activation. In the initial stage of our study, we found that the expression of T cell receptor (TCR) increased significantly. Furthermore, we identified a new partner of PTP-PEP, which was the 14-3-3τ protein by using the yeast two-hybrid system. We discovered that PTP-PEP interacted with 14-3-3τ via P2 motif. In addition, PTP-PEP dephosphorylated 14-3-3τ. 14-3-3τ plays a crucial role in the regulation of T cell activation and signal transduction, suggesting that PTP-PEP may regulate T cell activation via the interaction with 14-3-3τ. In T cells, the 14-3-3τ bind to c -Cbl that is the substrates of PTP-PEP and is an important regulators of TCR degradation processes. Therefore, it is speculated that 14-3-3τ as a bridge to assist PTP-PEP contact with c-Cbl, accurately regulate the tyrosine phosphorylation and TCR expression, thereby affecting activation signaling and avoiding excessive activation of T cells to induce autoimmune diseases. In this study, we plan to investigate the regulation mechanism of T cell receptor expression, T cell signaling and cytokine secretion by their interaction. Overall, we aim to illuminate the biological function of PTP-PEP on regulating T cell activation, and provide some effective and safe options to treat autoimmune disease from the current study.