Assembly of Picornaviral Replication Complexes

描述(申请人提供)：小核糖核酸病毒是一类小的正向单链RNA病毒，每年造成数亿美元的疾病。成员包括急性甲型肝炎病毒，引起柯萨奇B3病毒的心脏病，导致一半以上普通感冒发生的鼻病毒，以及瘫痪的脊髓灰质炎病毒。这些病毒具有相同的生命周期，RNA复制和病毒组装发生在大的膜锚定复制复合体中，这些复制复合体组装在来自内质网的小泡的表面。复制过程是由病毒编码的依赖RNA的RNA聚合酶3Dpol蛋白驱动的，该蛋白负责合成所有病毒RNA。本研究项目的重点是病毒复制中心的结构和组装，在那里我们使用脊髓灰质炎病毒和柯萨奇病毒作为我们的主要实验系统。我们先前已经解决了这两种病毒3Dpol蛋白的晶体结构，并阐明了这些蛋白从病毒3CDpro前体蛋白裂解后被蛋白水解性激活的分子机制。我们正在继续我们对小核糖核酸病毒复制蛋白的研究，重点是与3Dpol延长复合体的形成相关的结构变化，以及了解聚合酶的突变如何在体外和体内影响病毒RNA复制速度和保真度。该项目的一个新方面是关注与膜相关的病毒2C和2BC蛋白的结构和功能，这些蛋白负责宿主细胞膜的重排，导致病毒复制复合体组装在其上的囊泡的形成。

DESCRIPTION (provided by applicant): The picornaviruses are a family of small positive sense single stranded RNA viruses that cause a wide range of diseases at an annual cost well into the hundreds of million dollars. Members include acute hepatitis A virus, the heart disease causing coxsackie B3 virus, rhinoviruses that cause more than half the occurrences of the common cold, and the paralyzing poliovirus. These viruses share a life cycle where RNA replication and viral assembly occurs in large membrane anchored replication complexes assembled on the surfaces of vesicles derived from the endoplasmic reticulum. The replication process is driven by a virally encoded RNA dependent RNA polymerase, the 3Dpol protein, that is responsible for the synthesis of all viral RNA. This research project is focused on the structure and assembly of viral replication centers, where we use poliovirus and coxsackievirus as our main experimental systems. We have previously solved the crystal structures of the 3Dpol proteins from both these viruses and elucidated the molecular mechanism behind the proteolytic activation of these proteins upon cleavage from the viral 3CDpro precursor protein. We are continuing our studies of picornaviral replication proteins by focusing on the structural changes associated with the formation of the 3Dpol elongation complex and on understanding how mutations in the polymerase affect viral RNA replication rate and fidelity both in vitro and in vivo. A new aspect of the project is focused on the structure and function of the membrane associated viral 2C and 2BC proteins that are responsible for host cell membrane rearrangements resulting in the formation of the vesicles upon which the viral replication complexes assemble.