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Mechanistic Insights into flavivirus NS5-mediated STAT2 Suppression

黄病毒 NS5 介导的 STAT2 抑制的机制见解

基本信息

批准号：
10581481
负责人：
Rong Hai
金额：
$ 38.88万
依托单位：
UNIVERSITY OF CALIFORNIA RIVERSIDE
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-03-15 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10581481
关键词：
Affect Antiviral Agents Attenuated Vaccines Binding Binding Proteins Biochemical Cells Complex Cryoelectron Microscopy Data Dengue Virus Development Epidemic Evaluation Family Flavivirus Flavivirus Infections Gene Expression Goals Host Defense Human In Vitro Infection Integration Host Factors Interferons Knowledge Link Mediating Molecular Mutation Nonstructural Protein Pathogenicity Pathway interactions Physiological Process Proteins Public Health RNA Role Signal Pathway Signal Transduction Species Specificity Specificity Stat2 protein Techniques Testing Therapeutic Vaccines Viral Viral Genes Viral Pathogenesis Viral Vaccines Virus Virus Diseases X-Ray Crystallography Zika Virus antagonist global health in vivo inhibitor insight interferon-stimulated gene factor 3 member molecular recognition multicatalytic endopeptidase complex novel novel therapeutic intervention novel therapeutics prevent prophylactic protein degradation response ubiquitin-protein ligase vector-borne virology

项目摘要

The family of flavivirus consists of over 90 vector-borne, single-stranded RNA-containing viruses, including Dengue virus (DENV) and Zika virus (ZIKV), which cause major epidemics among humans and pose a serious threat to global public health. No vaccines or antivirals exist to prevent or treat infections caused by DENV, ZIKV, and some other flaviviruses. To establish infection, flaviviruses need to overcome the antiviral state induced by type 1 interferon (IFN-1), the first line of host defense. In this regard, flaviviruses have encoded several antagonists to suppress IFN responses. For example, the nonstructural NS5 proteins of DENV, ZIKV, and some other flaviviruses have been shown to be potent suppressor of IFN signaling, targeting different steps of the IFN signaling pathway. Like DENV, ZIKV NS5 protein bind human signal transducer and activator of transcription 2 (hSTAT2) protein and trigger its proteasomal degradation, albeit using different downstream mechanisms. To elucidate the mechanistic basis of flavivirus NS5-mediated hSTAT2 suppression, we propose to provide structural insight into the ZIKV NS5-hSTAT2 and DENV NS5-hSTAT2 complexes, which, in turn, will guide interrogation of the consequence(s) of the flavivirus NS5-hSTAT2 interactions in proteasome-mediated degradation of hSTAT2 and suppression of IFN signaling. Toward this goal, we will use structural, biochemical, molecular, cellular and virology approaches to investigate the structural basis of the ZIKV NS5-hSTAT2 and DENV NS5-hSTAT2 interactions and their functional consequence. In Aim 1, we will establish the structural basis of the ZIKV-hSTAT2 interaction by using X-ray crystallography and cryo-electron microscopy and validate our observations with mutational and in vitro pull-down analyses. In Aim 2, we will examine the ZIKV NS5-hSTAT2 interaction at a cellular level and investigate the functional consequence of the ZIKV NS5-hSTAT2 interaction through evaluation of the mutational effects of ZIKV NS5 on hSTAT2 degradation, IFN response and viral infection. The results of these studies will provide critical structural and functional insights into the virus- and species-specific ZIKV NS5-hSTAT2 interaction, thereby establishing a mechanistic link between flavivirus NS5 proteins, hSTAT2 degradation, suppression of the IFN response and viral infection. Results from the proposed studies will ultimately benefit development of novel antivirals and live vaccines against flaviviruses infection.

黄病毒家族由超过90种载体传播的、含有单链RNA的病毒组成，包括登革热病毒（DENV）和寨卡病毒（ZIKV），它们在人类中引起重大流行病并对全球公共卫生构成严重威胁。没有疫苗或抗病毒药物可以预防或治疗由DENV、ZIKV和一些其他黄病毒引起的感染。为了建立感染，黄病毒需要克服由1型干扰素（IFN-1）诱导的抗病毒状态，这是宿主的第一道防线。在这方面，黄病毒已编码了几种拮抗剂来抑制IFN应答。例如，DENV、ZIKV和一些其他黄病毒的非结构NS 5蛋白已显示为IFN信号传导的有效抑制剂，靶向IFN信号传导途径的不同步骤。与DENV一样，ZIKV NS 5蛋白结合人信号转导和转录激活因子2（hSTAT 2）蛋白并触发其蛋白酶体降解，尽管使用不同的下游机制。为了阐明黄病毒NS 5介导的hSTAT 2抑制的机制基础，我们提出提供对ZIKV NS 5-hSTAT 2和DENV NS 5-hSTAT 2复合物的结构洞察，这反过来将指导对黄病毒NS 5-hSTAT 2相互作用在蛋白酶体介导的hSTAT 2降解和IFN信号传导抑制中的结果的询问。为了实现这一目标，我们将使用结构、生物化学、分子、细胞和病毒学方法来研究ZIKV NS 5-hSTAT 2和DENV NS 5-hSTAT 2相互作用的结构基础及其功能后果。在目标1中，我们将通过使用X射线晶体学和冷冻电子显微镜建立ZIKV-hSTAT 2相互作用的结构基础，并通过突变和体外下拉分析验证我们的观察结果。在目的2中，我们将在细胞水平上检查ZIKV NS 5-hSTAT 2相互作用，并通过评价ZIKV NS 5对hSTAT 2降解、IFN应答和病毒感染的突变作用来研究ZIKV NS 5-hSTAT 2相互作用的功能后果。这些研究的结果将为病毒和物种特异性ZIKV NS 5-hSTAT 2相互作用提供关键的结构和功能见解，从而建立黄病毒NS 5蛋白、hSTAT 2降解、IFN应答抑制和病毒感染之间的机制联系。从拟议的研究结果将最终有利于新的抗病毒药物和抗黄病毒感染的活疫苗的开发。