LDLRAD3 Receptor Interaction with Venezuelan Equine Encephalitis Virus

项目摘要 甲病毒是披膜病毒科的蚊子传播的正链包膜RNA病毒， 导致全球性的人类疾病。目前，没有抗病毒药物或许可的疫苗可用于治疗或 预防任何甲病毒感染。我们最近使用了一种基于CRISPR/Cas9的全基因组筛选， 细胞表面分子LDLRAD 3作为委内瑞拉马的新型高度保守进入受体 脑炎病毒（VEEV），一种能够在人类中引起致命的神经侵入性疾病的新兴病原体， 其他脊椎动物。小鼠或人LDLRAD 3的基因编辑导致VEEV感染减少， 神经元细胞和LDLRAD 3的异位表达导致感染增加。LDLRAD 3 直接与VEEV病毒粒子结合并增强病毒附着和内化到细胞中。遗传研究 表明LDLRAD 3的结构域1（D1）是支持VEEV感染所必需的且足够的。我们假设 VEEV与LDLRAD 3结合将解释感染、组织靶向和疾病发病机制 发生。钻石，弗里蒙特和惠兰之间的这个合作，互动项目的主要目标 实验室将确定LDLRAD 3促进甲病毒进入细胞的精确机制， 获得关于LDLRAD 3如何与病毒粒子上的刺突蛋白结合的高分辨率结构洞察， 确定LDLRAD 3在体内VEEV发病机制中的细胞类型特异性作用。该提案中的实验 将定义VEEV生物学的基本方面，增强我们对感染和细胞向性的理解。 这些信息可能有助于开发破坏LDLRAD 3的小分子或生物制剂 与VEEV刺突蛋白的相互作用，这可能成为未来改善疾病治疗的基础。 这种新出现的高致病性甲病毒

Project Summary Alphaviruses are mosquito-transmitted, positive-strand enveloped RNA viruses of the Togaviridae family that cause global disease in humans. At present, no antiviral agents or licensed vaccines exist for the treatment or prevention of any alphavirus infections. We recently used a genome-wide CRISPR/Cas9-based screen to identify the cell surface molecule LDLRAD3 as a novel, highly conserved entry receptor for Venezuelan equine encephalitis virus (VEEV), an emerging pathogen capable of causing fatal neuroinvasive disease in humans and other vertebrate animals. Gene editing of mouse or human LDLRAD3 resulted in reduced VEEV infection of neuronal cells, and reciprocally, ectopic expression of LDLRAD3 resulted in increased infection. LDLRAD3 bound directly to VEEV virions and enhanced virus attachment and internalization into cells. Genetic studies indicated that domain 1 (D1) of LDLRAD3 is necessary and sufficient to support VEEV infection. We hypothesize that engagement of LDLRAD3 by VEEV will explain how infection, tissue targeting, and disease pathogenesis occurs. The primary goals of this collaborative, interactive project between the Diamond, Fremont, and Whelan laboratories are to define the precise mechanism(s) by which LDLRAD3 facilitates alphavirus entry into cells, to gain high-resolution structural insight as to how LDLRAD3 engages the spike proteins on the virion, and to determine the cell-type specific role of LDLRAD3 in VEEV pathogenesis in vivo. The experiments in this proposal will define fundamental aspects of VEEV biology that enhance our understanding of infection and cell tropism. This information may facilitate the development of small molecules or biologicals that disrupt LDLRAD3 interaction with VEEV spike proteins, which could form the basis of future therapeutics that ameliorate disease of this emerging and highly pathogenic alphavirus.