Structural basis for RIG-I and MDA-5 mediated antiviral response

我们阐述了RIG-I样蛋白的激活、调节和抑制的结构和生化机制 抗病毒干扰素反应中的解旋酶。RIG-I和相关MDA5蛋白检测病毒RNA和 启动一系列信号转导来刺激先天免疫。病毒对抗自身的分子基础 RIG-I和MDA5对RNA的区别还不清楚，但对于理解内在的重要意义 我们细胞的抗病毒功能。我们使用X射线结晶学、小角X射线散射相结合的方法 和生化技术，以了解RIG-I如何识别病毒RNA模式、如何 在模式识别过程中使用RIG-I对ATP的结合和水解，最后，病毒是如何 蛋白质抑制剂会干扰模式识别和激活。基于现有的X射线衍射晶体 我们的目标是在第一个目标中推导出RIG-I的解旋酶结构域。这一结构将引导 用病毒模式分析RIG-I的激活机制。我们将致力于派生出分子 ATPase和ATPase识别5‘三磷酸RNA和双链RNA的决定因素 RIG-I的调节域。我们将讨论RIG-I的调节域以及ATPase域是如何 在机械上联系在一起，并检验了RIG-I将几种模式集成到一个活跃的“信号”中的假设 关于“构象。然后我们将致力于得出这一信号的详细的分子和机制图” 关于使用多学科和协作方法构建RIG-I。这一方法将包括 依赖于ATP的模式识别与RIG-I的翻译后修饰之间的关联 解决这些翻译后修饰是否表现出“信号打开”的构象。最后，我们会 目的揭示病毒如何通过使用一种针对MDA5的蛋白质类抑制剂来中和MDA5信号。 总而言之，预期的结果将促进我们对具体和校对模式的理解 RIG-I类似解旋酶在分子机制和原子水平上识别病毒RNA。

We address the structural and biochemical mechanisms of activation, regulation and inhibition of RIG-I like helicases in the antiviral interferon response. RIG-I and the related MDA5 protein detect viral RNA and initiate a signal transduction cascade to stimulate innate immunity. The molecular basis for virus versus self RNA differentiation by RIG-I and MDA5 is not understood, but of central importance to understand intrinsic antiviral functions of our cells. We use a combination of X-ray crystallography, small angle X-ray scattering and biochemical techniques to understand key principles of how RIG-I recognizes viral RNA patterns, how ATP binding and hydrolysis by RIG-I is used in the process of pattern recognition and finally, how viral protein inhibitors interfere with pattern recognition and activation. Based on existing, X-ray diffracting crystals we aim at deriving in the first aim a structure of the helicase domain of RIG-I. This structure will guide the analysis of the mechanism of activation of RIG-I by viral patterns. We will aim at deriving molecular determinants for the recognition of 5' triphosphate RNA as well as double stranded RNA by the ATPase and regulatory domains of RIG-I. We will address how regulatory domains as well as ATPase domain of RIG-I are mechanistically linked and test the hypothesis that RIG-I integrates several patterns into an active "signal on" conformation. We will then aim at deriving a detailed molecular and mechanistic picture of this "signal on" conformation of RIG-I using a multidisciplinary and collaborative approach. This apprach will include the interlink between ATP dependent pattern recognition and posttranslational modification of RIG-I and we will address whether these posttranslational modifications manifest the "signal on" conformation. Finally, we will aim at revealing how viruses counteract MDA5 signalling by using a proteinaceous inhibitor against MDA5. All in all, the expected outcome will advance our understanding of the specific and proofread pattern recognition of viral RNA by RIG-I like helicases at the molecular mechanistic and atomic level.