LDLRAD3 Receptor Interaction with Venezuelan Equine Encephalitis Virus

项目摘要 甲型病毒是Togaviridae家族的蚊子传播的正链包膜RNA病毒， 在人类中引发全球性疾病。目前，尚无抗病毒药物或获得许可的疫苗用于治疗或 预防任何甲型病毒感染。我们最近使用了全基因组CRISPR/Cas9筛查来鉴定 细胞表面分子LDLRAD3作为委内瑞拉马的一种新的高度保守的进入受体 脑炎病毒(VEEV)是一种新出现的病原体，能够导致人类和 其他脊椎动物。小鼠或人LDLRAD3的基因编辑导致VEEV感染减少 相反，LDLRAD3的异位表达导致感染增加。LDLRAD3 直接与VEEV病毒粒子结合，增强病毒附着和内化到细胞中。遗传学研究 提示LDLRAD3的结构域1(D1)是支持VEEV感染的必要条件和充分条件。我们假设 VEEV对LDLRAD3的参与将解释感染、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协作、互动项目的主要目标是 实验室将确定LDLRAD3促进甲型病毒进入细胞的精确机制(S)，以 获得关于LDLRAD3如何与病毒粒子上的尖峰蛋白接合的高分辨率结构洞察，并 确定LDLRAD3在VEEV体内致病过程中的细胞类型特异性作用。这项提案中的实验 将定义VEEV生物学的基本方面，这些方面增强了我们对感染和细胞嗜性的理解。 这些信息可能有助于干扰LDLRAD3的小分子或生物制品的发展 与VEEV尖峰蛋白的相互作用，这可能构成未来改善疾病的治疗方法的基础 这种新出现的高致病性甲型病毒。

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.