Regulation of RIG-I signaling and viral immune evasion by ufmylation

摘要 I型和III型干扰素可抑制RNA病毒感染。感染细胞通过激活信号产生干扰素 通过模式识别受体，如RIG-I。RIG-I和下行信令必须高度协调 有效的抗病毒反应。因此，这些过程由几个翻译后的 修改(PTM)。这些PTM对于RIG-I的激活和最终终止都是必不可少的。 发信号。虽然RIG-I信号是通过泛素化和磷酸化来协调的，但其机制 其他的PTM，如泛素样修饰物，可能调节RIG-I信号，但在很大程度上是未知的。我们的预赛 数据显示，泛素样修饰物被称为ufm化，调节参与rig-I信号转导的多种蛋白质。 以获得最佳的干扰素诱导以应对病毒感染。此外，我们的数据表明，ufmyl化被 病毒逃避宿主细胞内的先天免疫反应。因此，这项建议的目标是 确定超甲基化如何调节细胞内对病毒感染和病毒逃避的先天免疫反应。 根据我们的初步数据，中心假设是ufmyl化调节宿主和病毒的功能。 调节抗病毒先天免疫和病毒逃避的蛋白质。根据我们的初步数据，这一假设 将通过追求以下三个具体目标进行测试：1)定义UFl1，ufmyl化E3连接酶， 促进RIG-I的激活；2)了解关键蛋白质超甲基化的分子机制 在RIG-I信号通路中下调其功能和信号；3)决定登革病毒如何 免疫逃逸的超甲基化结合系统。在目标1中，UFl1通过其分子机制 诱导RIG-I的激活以响应RNA病毒感应将被定义。在目标2中，机制和 超甲基化如何调节RIG-I途径中信号蛋白的功能的动力学将被确定。在……里面 目的3，DENV蛋白的超甲基化促进DENV免疫逃避的功能，在人类细胞中 线条和原生单元格，将被确定。综上所述，本申请中建议的工作将是 具有重大意义和创新性，因为它将把一种新的免疫调节功能归因于ufmyl化，有助于 我们对其基本功能的了解，并揭示了抗病毒天然免疫的一种新的控制(ufm化)。 此外，这项工作将提供对宿主引导的过程的理解，该过程被病毒用于免疫 逃避，以促进病毒复制。总体而言，这项工作将定义一个新的PTM来协调RIG-I信令 途径，这将提高我们对抗病毒免疫和调节先天免疫途径的认识 这将增加对最终可能导致的失调干扰素的机制原因的理解 在自身免疫性疾病中。它还将定义一种新的黄病毒免疫逃避机制，这种机制将具有 对限制其感染的治疗和疫苗策略的影响。

ABSTRACT Type I and III interferons (IFN) restrict RNA virus infection. Infected cells produce IFN through signaling activated by pattern recognition receptors such as RIG-I. Both RIG-I and downstream signaling must be highly coordinated for efficient antiviral responses. As such, these processes are regulated by several post-translational modifications (PTMs). These PTMs are essential for both the activation and eventual termination of RIG-I- signaling. While RIG-I-signaling is coordinated by ubiquitination and phosphorylation, the mechanisms by which other PTMs, such as ubiquitin-like modifiers, may regulate RIG-I-signaling are largely unknown. Our preliminary data reveal that the ubiquitin-like modifier called ufmylation regulates multiple proteins involved in RIG-I signaling for optimal IFN induction in response to viral infection. Further, our data suggest that ufmylation is utilized by viruses to evade the host intracellular innate immune response. Therefore, the goal of this proposal is to determine how ufmylation regulates the intracellular innate immune response to virus infection and viral evasion. Based on our preliminary data, the central hypothesis is that ufmylation modulates the function of host and viral proteins to regulate antiviral innate immunity and viral evasion. Guided by our preliminary data, this hypothesis will be tested by pursuing the following three specific aims: 1) Define how UFL1, the ufmylation E3 ligase, promotes the activation of RIG-I; 2) Understand the molecular mechanism by which ufmylation of a key protein in the RIG-I signaling pathway downregulates its function and signaling; 3) Determine how dengue virus co-opts the ufmylation conjugation system for immune evasion. In Aim 1, the molecular mechanisms by which UFL1 induces the activation of RIG-I in response to RNA virus sensing will be defined. In Aim 2, the mechanism and dynamics of how ufmylation regulates the function of a signaling protein in RIG-I pathway will be determined. In Aim 3, the function by which ufmylation of a DENV protein promotes DENV immune evasion, both in human cell lines and in primary cells, will be determined. Taken together, the work proposed in this application will be significant and innovative because it will attribute a novel immune regulatory function to ufmylation, contribute to our understanding of its basic functions, and uncover a novel control (ufmylation) of antiviral innate immunity. Additionally, this work will provide understanding of a host-directed process that is utilized by viruses for immune evasion to facilitate viral replication. Overall, this work will define a new PTM that coordinates the RIG-I signaling pathway, which will improve our knowledge of both antiviral immunity and regulation of innate immune pathways that will lead to increased understanding of the mechanistic causes of dysregulated IFN that can ultimately result in autoimmune disease. It will also define a new mechanism of immune evasion by flaviviruses that will have implications for therapeutic and vaccine strategies to limit their infection.