Role of nonstructural proteins in pestivirus assembly

描述（由申请人提供）：丙型肝炎病毒（HCV）引起慢性肝脏感染，通常导致肝硬化和肝癌。由于HCV在细胞培养中无法有效复制，了解其生命周期的某些方面变得复杂。特别是，对HCV病毒粒子的组装机制知之甚少。丙型肝炎病毒在进化上与鼠疫病毒密切相关，后者在细胞培养中生长效率高。我们已经鉴定出具有组装新缺陷的突变鼠疫病毒，这揭示了非结构蛋白在病毒粒子组装中的前所未有的作用。我们假设非结构蛋白在鼠疫病毒和肝病毒的组装中起作用。本研究将以鼠疫病毒系统为模型，分析病毒粒子组装的机制和非结构蛋白在此过程中的重要作用。我们将利用遗传、生化和结构方法来识别和表征非结构蛋白在病毒粒子组装中的作用。具体而言，我们建议：

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) causes chronic liver infections, often leading to cirrhosis and carcinoma. Understanding certain aspects of its life cycle is complicated by the failure of HCV to replicate efficiently in cell culture. In particular, little is known about the mechanism of assembly of HCV virions. HCV is evolutionarily closely related to pestiviruses, which grow efficiently in cell cultures. We have identified mutant pestiviruses with a novel defect in assembly that reveals an unprecedented role of nonstructural proteins in virion asssembly. We hypothesize that nonstructural proteins play a role in pestivirus and hepacivirus assembly. The research proposed here will exploit a pestivirus system as a model to analyze the mechanisms virion assembly and the essential role of non-structural proteins in this process. We will utilize genetic, biochemical and structural approaches to identify and characterize the role of nonstructural proteins in virion assembly. Specifically we propose to: 1. Map critical sites of the pestivirus nonstructural polyprotein involved in virion assembly. 2. Identify and characterize pestivirus proteins involved in the assembly process using a proteomic approach. 3. Characterize the subcellutar localization of pestivirus assembly sites involving nonstructural proteins. 4. Screen the pestivirus nonstructural polyprotein to identify additional loci involved in virion assembly. This research will expand our knowledge on the novel role of nonstructural proteins in pestivirus assembly. This information may help delineate the assembly of HCV when cell culture systems for efficient HCV replication become available.