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不能在细胞培养中有效复制，因此理解其生命周期的某些方面变得复杂。 特别是，对HCV病毒体的组装机制知之甚少。HCV在进化上与瘟病毒密切相关，瘟病毒在细胞培养物中有效生长。我们已经确定了突变瘟病毒与一个新的缺陷，在大会上，揭示了前所未有的作用，非结构蛋白在病毒粒子assassessment。我们假设非结构蛋白在瘟病毒和肝炎病毒装配中发挥作用。本研究将利用瘟病毒系统作为模型，分析病毒粒子组装的机制以及非结构蛋白在此过程中的重要作用。我们将利用遗传学，生物化学和结构的方法来确定和表征非结构蛋白在病毒体组装中的作用。具体而言，我们建议： 1.参与病毒体装配的瘟病毒非结构多聚蛋白的关键位点图。 2.使用蛋白质组学方法鉴定和表征参与组装过程的瘟病毒蛋白。 3.表征涉及非结构蛋白的瘟病毒装配位点的亚细胞定位。 4.筛选瘟病毒非结构多聚蛋白以鉴定参与病毒体组装的其他基因座。 这项研究将扩大我们对非结构蛋白在瘟病毒组装中的新作用的认识。这些信息可能有助于描绘组装的HCV时，细胞培养系统，有效的HCV复制变得可用。

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.