端锚聚合酶TNKS通过泛素连接酶RNF146调控抗病毒天然免疫的机制研究

RIG-I样受体识别病毒RNA后可通过关键接头蛋白VISA引起抗病毒天然免疫反应。该过程对机体抵御RNA病毒感染至关重要，但其调控机制尚不清楚。本项目前期，申请人通过串联亲和纯化及质谱技术，发现端锚聚合酶TNKS1与VISA特异结合。过量表达TNKS1和同源分子TNKS2均与VISA存在相互作用且使其发生PARsylation修饰。TNKSs相关E3连接酶RNF146也与VISA相互作用并对其进行降解。过量表达TNKS1、TNKS2和RNF146均能抑制SeV诱导的IFN-β激活，敲低或敲除TNKS1或TNKS2则具有相反的效果。本项目将利用Tnks1-/-和Tnks2-/-小鼠以及RNF146-/-细胞深入研究TNKSs-RNF146轴在抗病毒天然免疫反应中的作用及机制。该研究将揭示一种抗病毒天然免疫调控的新机制，并证实PARsylation修饰在该过程中的重要意义。

The retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs) sense cytoplasmic viral RNA and initiate innate antiviral response, which is mediated by the central adaptor VISA. How RIG-I-VISA-mediated antiviral response is regulated is enigmatic. In this project, using a biochemical purification approach and MS analysis, we identified TNKS1 as a VISA-associated protein. TNKS1 and its homolog TNKS2 are ubiquitously expressed and known to induce the PARsylation of the substrate proteins. Overexpression of TNKS1 or TNKS2 interacted with and mediated PARsyaltion and degradation of VISA. VISA also interacted with an ADP-ribose-binding E3 ubiquitin ligase, RNF146, which leads to VISA degradation. Overexpression of TNKS1, TNKS2 or RNF146 inhibited SeV-triggered activation of IFN-β, while either knockdown of TNKS1 or TNKS2 had opposite effects. Based on these findings, by using Tnks1-/- and Tnks2-/- mice and RNF146-/- cells, we aims to reveal the physiological functions and mechanisms of TNKSs-RNF146 axis in antiviral innate immune response. Our research will reveal a new regulatory mechanism of attenuating VISA-mediated signaling and highlight a role of PARsylation in innate immune response.