Structural and Functional Characterization of the Ebola Virus Replication Complex

埃博拉病毒复制复合体的结构和功能表征

• 批准号: 9312729
• 负责人: Gaya K. Amarasinghe
• 金额: $ 274.29万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-07 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9312729
• 关键词: Address; Adverse effects; Affect; Africa; Attenuated; Biochemistry; Biological Assay; Biological Testing; Biology; Case Study; Categories; Cessation of life; Collaborations; Complex; Cryoelectron Microscopy; DNA-Directed RNA Polymerase; Data Analyses; Development; Disease; Disease Outbreaks; Ebola virus; Elements; Ensure; Epidemic; Europe; Evaluation; Event; Filoviridae; Filovirus; Goals; Health; Human; Human Resources; Immunology; In Vitro; Inflammation; Integration Host Factors; Knowledge; Light; Link; Manuscripts; Maps; Molecular; Monoclonal Antibodies; Nucleoproteins; Pathogenesis; Pathogenicity; Production; Program Research Project Grants; Proteins; Proteome; Proteomics; RNA; RNA Editing; RNA Interference; RNA Viruses; RNA analysis; RNA chemical synthesis; RNA-Directed RNA Polymerase; Reagent; Recording of previous events; Regulation; Reporting; Research; Research Infrastructure; Research Personnel; Research Project Grants; Resolution; Resources; Signal Transduction; Specificity; Structure-Activity Relationship; Technology; Testing; Therapeutic; Time; Training; Trans-Activators; Treatment Efficacy; Variant; Viral; Viral Genome; Viral Hemorrhagic Fevers; Viral Proteins; Virus; Virus Replication; Work; X-Ray Crystallography; antiviral immunity; biodefense; biophysical analysis; biosafety level 4 facility; genome-wide; global health; health economics; high throughput screening; in vivo; in vivo Model; innovation; insight; multidisciplinary; mutant; new therapeutic target; novel; pathogen; programs; structural biology; synthetic antibodies; therapeutic development; viral RNA; virology

Abstract: Project Overview for Structural and Functional Studies of Ebola Virus RNA synthesis Ebolaviruses (EBOVs) are non-segmented negative-strand RNA viruses (NNSVs) and Category A Priority biodefense pathogens that cause frequent lethal hemorrhagic fever, including the devastating and ongoing outbreak in West Africa. Approved anti-EBOV therapeutics are lacking. EBOV replicates with high efficiency in vivo while effectively evading host innate antiviral immunity. The unrestrained replication and associated inflammation leads to death. A complete understanding of EBOV pathogenesis therefore requires understanding how the viral RNA dependent RNA polymerase (RDRP) complex interacts with host factors. The overarching goal of this program project proposal is to understand the mechanisms of RDRP function by defining the interactions among its components and with the host proteome and by defining signals that regulate its function. Toward this goal Project 1 will identify cis- and trans-acting factors that impact EBOV RDRP activity. Project 2 will define a structural basis for the EBOV RDRP complex, identify regulatory mechanisms, and determine species and strain specificities, and Project 3 will use genome-wide RNAi and proteomic strategies to identify host factors that modulate RDRP function. Project 1 provides functional context into which a subset of Project 3 “hits” can be interpreted, while Project 3 will likely identify host factors relevant to the findings of Project 1. Project 2 will provide a structural framework that will be used to integrate findings from both Projects 1 and 3. Together, these studies will provide mechanistic insights into how protein-protein and protein-vRNA interactions control EBOV RDRP function. Research projects are supported by an Administrative Core, a Protein Production and Protein Interaction Core, and critically, a Biosafety Level 4 (BSL4) Core. Core B will generate unique reagents, including monoclonal and synthetic antibodies. Core C (BSL4) will provide unique capabilities to obtain materials from and perform tests using wild type and mutant EBOVs as well as pathogenesis evaluation. The work will be performed by highly productive and collaborative investigators with expertise in every aspect of the proposed studies, including biochemistry, EBOV pathogenesis, high throughput screening and data analysis, immunology, proteomics, structural biology, and virology. These studies will provide unprecedented insights into the components of the EBOV RDRP and in its interaction with the host as well as species and strain differences that affect the RDRP complex. We also expect to identify specific viral and host factor interactions as novel therapeutic targets.

摘要：埃博拉病毒 RNA 合成结构和功能研究项目概述 埃博拉病毒 (EBOV) 是不分段负链 RNA 病毒 (NNSV)，A 类优先级 引起频繁致命性出血热的生物防御病原体，包括毁灭性和持续性的 西非爆发。缺乏批准的抗埃博拉病毒疗法。 EBOV 高效复制 体内同时有效逃避宿主先天抗病毒免疫。无限制的复制和相关的 炎症导致死亡。因此，全面了解埃博拉病毒发病机制需要 了解病毒 RNA 依赖性 RNA 聚合酶 (RDRP) 复合物如何与宿主因子相互作用。 该计划项目提案的总体目标是通过以下方式了解 RDRP 功能的机制： 定义其成分之间以及与宿主蛋白质组之间的相互作用，并通过定义信号 调节其功能。为实现这一目标，项目 1 将确定影响 EBOV 的顺式和反式作用因素 RDRP 活动。项目 2 将定义 EBOV RDRP 综合体的结构基础，确定监管 机制，并确定物种和菌株特异性，项目 3 将使用全基因组 RNAi 和 确定调节 RDRP 功能的宿主因子的蛋白质组学策略。项目 1 提供功能环境 其中可以解释项目 3“命中”的子集，而项目 3 可能会识别相关的宿主因素 项目 1 的调查结果。项目 2 将提供一个结构框架，用于整合调查结果 来自项目 1 和 3。 总之，这些研究将为蛋白质-蛋白质如何 蛋白质-vRNA 相互作用控制 EBOV RDRP 功能。研究项目由一个 行政核心、蛋白质生产和蛋白质相互作用核心，以及最重要的生物安全 4 级 (BSL4) 核心。 Core B 将生成独特的试剂，包括单克隆抗体和合成抗体。核心C (BSL4) 将提供独特的能力，从野生型和突变体中获取材料并进行测试 EBOV 以及发病机制评估。这项工作将由高生产力和协作性的人员来完成 具有拟议研究各方面专业知识的研究人员，包括生物化学、埃博拉病毒 发病机制、高通量筛选和数据分析、免疫学、蛋白质组学、结构生物学和 病毒学。这些研究将为 EBOV RDRP 的组成部分及其实施提供前所未有的见解。 与宿主的相互作用以及影响 RDRP 复合物的物种和菌株差异。我们也 期望将特定的病毒和宿主因子相互作用确定为新的治疗靶点。