NFAT1在抗RNA病毒天然免疫应答中的调控作用及机制研究

RIG-I like receptor signaling plays an important role in recognizing viral RNA and initiating antiviral innate immune responses. RLR signaling is finely tuned to ensure the appropriate duration and strength to prevent damaging inflammation to the host. Therefore, the regulation of RLR signaling is a hot topic in the frontiers of innate immunity. NFAT is a family of transcription factors with diversified functions. The roles of NFAT in the immune system have been extensively studied mainly in development, differentiation and activation of lymphocytes. However, little attention was paid to the roles of NFAT in antiviral innate immunity. Our preliminary findings showed that the expression of NFAT1 was markedly increased in macrophages following RNA virus VSV infection. Furthermore, NFAT1 knockdown significantly suppressed VSV-triggered production of IFN-β and IL-6 in macrophages while overexpression of NFAT1 markedly increased VSV-triggered production of IFN-β and IL-6 and promoted RIG-I-, IPS-1-induced IFN-β and ISRE luciferase activity, suggested that NFAT1 is involved in the positive regulation of RNA virus-triggered innate immune response. The primary aim of this study is to further investigate the regulatory function of NFAT1 in antiviral innate immunity. We will further observe the immunomodulating roles of NFAT1 and explore the targets of NFAT1 and the underlying molecular mechanisms mainly through co-immunoprecipitation, mass spectrum and infectious disease model, which will provide a new therapeutic approach for viral infection.

RIG-I样受体信号在识别病毒RNA和激活抗病毒天然免疫应答中发挥重要作用，对其信号通路的调控是控制炎症强度的关键，也是天然免疫领域研究的热点。NFAT是一类功能多样的转录因子，目前对其在免疫系统中的研究主要集中于淋巴细胞活化、分化发育方面，但其在抗病毒天然免疫领域的相关研究甚少。预实验发现RNA 病毒VSV感染可上调巨噬细胞中NFAT1蛋白的表达，干扰NFAT1显著抑制巨噬细胞中VSV诱导的IFN-β和IL-6产生，而过表达NFAT1可促进VSV诱导的IFN-β和IL-6产生以及RIG-I或IPS-1依赖的IFN-β、ISRE报告基因活性，提示NFAT1可能正向调控RNA病毒触发的天然免疫应答。本研究拟在体内外探讨NFAT1在抗病毒天然免疫中的调控功能及机制，利用免疫共沉淀-质谱、感染模型等手段确证NFAT1的免疫调节功能，发现其作用的靶分子和作用机制，为抗病毒免疫治疗提供新思路和靶点。

Ⅰ型干扰素是机体参与抗病毒免疫反应的关键效应分子。I型干扰素的产生虽然能有效的抗病毒, 但是过量或者是长时间的干扰素反应也会导致病毒感染中的免疫抑制与炎症反应, 从而使疾病恶化，因此对Ⅰ型干扰素产生的调控机制的研究是当前研究的热点问题。NFAT是一类功能多样的转录因子，其在抗病毒天然免疫中的效应一直未得到深入的研究。本项目通过Real-time PCR、Western-blot、共聚焦显微镜、质谱分析、免疫共沉淀以及染色质免疫共沉淀等技术深入研究了NFAT1对RNA病毒诱导的I型干扰素产生的影响及其作用机制。我们的研究发现，RNA 病毒VSV感染可上调巨噬细胞中NFAT1蛋白的表达，并使NFAT1蛋白主要定位于细胞浆中。NFAT1能够促进I 型干扰素与干扰素诱导基因的产生，抑制病毒的复制。在RNA病毒感染的过程中, NFAT1调控RIG-I的翻译后修饰, 从而影响RIG-I的降解, 进而通过RIG-I-MAVS信号促进I 型干扰素的产生。NFAT1可能是通过招募去泛素化酶USP14介导RIG-I的稳定性，其具体机制在进一步深入研究中。通过以上研究，我们对NFAT1在抗RNA病毒天然免疫应答中的调控作用及机制有了更深入的了解，希望能为炎症性疾病的防治提供新靶点。