Role of FIP200 in RIG-I-mediated innate immunity

项目总结 视黄酸诱导基因I(RIG-I)是识别病毒双链RNA的细胞内传感器 (DsRNA)和5‘三磷酸RNA。在RNA识别之后，RIG-I诱导了I型的强健生产 干扰素(干扰素)。为了防止干扰素的异常表达，RIG-I的激活通过几个步骤进行调节，包括 RIG-I的CARD结构域去磷酸化和寡聚化CARD结构域的磷酸化 在休眠细胞中保持RIG-I不活动。在RNA配体结合时，RIG-I被 蛋白磷酸酶1(PP1)，它允许随后的寡聚激活。然而，PP1有 细胞中有许多底物，是什么决定了PP1对RIG-I的特异性尚不清楚。 此外，去磷酸化如何促进RIG-I齐聚尚不清楚。因此，迫切需要 明确RIG-I去磷酸化和寡聚化的调控机制。澄清： 这些机制不仅将揭示一种新的寄主防御调控机制，还将为寄主防御提供洞察 用于开发新的治疗策略以防止异常的干扰素激活。 我们实验室的长期目标是了解核酸诱导的先天免疫调节机制。 豁免权。这项提案的总体目标是研究RIG-I介导的调节机制 先天免疫力。我们的初步蛋白质组学研究发现RIG-I与FIP200(FAK-家族相互作用)相互作用 200 kDa的蛋白)，这是一种众所周知的自噬蛋白。自噬有一种与生俱来的免疫力 一些自噬基因抑制RIG-I信号通路。然而，令我们惊讶的是，FIP200的不足之处 取消5‘三磷酸RNA和dsRNA诱导的I型干扰素表达，促进RNA病毒复制， 如水疱性口炎病毒(VSV)。有趣的是，FIP200也被称为PP1调节亚单位 131(PPP1R131)，但其在PP1去磷酸化中的作用尚不清楚。基于现有的文献和我们的 初步数据，我们提出了中心假设，即FIP200是dsRNA介导的先天所必需的 调节RIG-I去磷酸化和寡聚化的免疫力。我们将调查以下目标： 目的1：确定FIP200在RIG-I信号通路中作用的分子基础。 目的2：确定FIP200促进RIG-I活化的机制。 目的3：用小鼠模型研究FIP200与RIG-I在体内的相互作用。 干扰素应答的失调会导致对病毒感染抵抗力的降低或因 与过度活跃的免疫系统有关。这项建议研究了FIP200 控制对病毒感染的先天免疫反应。从这个提议中衍生出的机械论概念将 不仅填补了RIG-I调控方面的知识空白，而且为抗病毒治疗提供了见解。

Project summary Retinoic acid-inducible gene I (RIG-I) is an intracellular sensor for recognition of viral double-stranded RNA (dsRNA) and 5` triphosphate RNA. Following RNA recognition, RIG-I induces a robust production of type I interferons (IFN). To prevent aberrant IFN expression, RIG-I activation is regulated by several steps, including dephosphorylation and oligomerization of the CARD domain of RIG-I. Phosphorylation of the CARD domain retains RIG-I inactive in the resting cells. Upon RNA ligand engagement, RIG-I is dephosphorylated by the protein phosphatase 1 (PP1), which permits subsequent oligomerization for activation. However, PP1 has numerous substrates in the cell and what determines the specificity of PP1 towards RIG-I is unknown. Furthermore, how dephosphorylation facilitates RIG-I oligomerization is not clear. Thus, there is a critical need to define the regulatory mechanisms for the dephosphorylation and oligomerization of RIG-I. Elucidation of these mechanisms will not only reveal a novel regulatory mechanism of host defense, but also provide insights for developing novel therapeutic strategies to prevent aberrant IFN activation. The long-term goal of our lab is to understand the regulatory mechanisms of nucleic acid-elicited innate immunity. The overall objective of this proposal is to investigate the regulatory mechanisms for RIG-I-mediated innate immunity. Our pilot proteomics study found that RIG-I interacted with FIP200 (FAK-family interacting protein of 200 kDa), a well-known autophagy protein. Autophagy has a crosstalk with innate immunity and several autophagy genes suppress RIG-I signaling pathway. However, to our surprise, deficiency of FIP200 abolishes 5` triphosphate RNA and dsRNA-induced type I IFN expression and promotes RNA virus replication, such as the vesicular stomatitis virus (VSV). Interestingly, FIP200 is also known as the PP1 regulatory subunit 131 (PPP1R131), but its role in PP1 dephosphorylation is unknown. Based on the existing literature and our preliminary data, we propose the central hypothesis that FIP200 is essential for dsRNA-mediated innate immunity by regulating RIG-I dephosphorylation and oligomerization. We will investigate the following aims: Aim 1: Determine the molecular basis for the role of FIP200 in RIG-I signaling pathway. Aim 2: Determine the mechanisms by which FIP200 promotes RIG-I activation. Aim 3: Determine the in vivo interaction between FIP200 and RIG-I using a mouse model. Dysregulation of IFN responses can result in decreased resistance to viral infection or detrimental effects due to an overactive immune system. This proposal investigates the molecular mechanisms by which FIP200 controls innate immune responses to viral infection. The mechanistic concepts derived from this proposal will not only fill the knowledge gap on RIG-I regulation, but also provide insights for antiviral therapeutics.