Assembly of Picornaviral Replication Complexes

描述（由申请人提供）：小核糖核酸病毒是一种小的阳性单链RNA病毒家族，每年造成数亿美元的费用，导致各种疾病。这些病毒包括急性甲型肝炎病毒、导致心脏病的柯萨奇B3病毒、导致半数以上普通感冒的鼻病毒以及麻痹性脊髓灰质炎病毒。这些病毒共享一个生命周期，RNA复制和病毒组装发生在来自内质网的囊泡表面组装的大型膜锚定复制复合体中。复制过程是由病毒编码的依赖RNA的RNA聚合酶，即3Dpol蛋白驱动的，它负责所有病毒RNA的合成。本研究项目以脊髓灰质炎病毒和柯萨奇病毒为主要实验系统，重点研究病毒复制中心的结构和组装。我们之前已经解决了这两种病毒的3Dpol蛋白的晶体结构，并阐明了这些蛋白在病毒3CDpro前体蛋白裂解后蛋白水解激活的分子机制。我们正在继续对小核糖核酸复制蛋白的研究，重点关注与3Dpol延伸复合物形成相关的结构变化，并了解聚合酶的突变如何影响病毒RNA的体外和体内复制速率和保真度。该项目的一个新方面集中在膜相关病毒2C和2BC蛋白的结构和功能上，这些蛋白负责宿主细胞膜重排，导致病毒复制复合体在其上组装的囊泡的形成。

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.