食源性诺如病毒主要衣壳蛋白VP1参与病毒复制的机制研究

Noroviruses are the major causative etiological of foodborne outbreaks of acute gastroenteritis. Our understanding of the pathogenicity of noroviruses has lagged behind due to the inability of cultivate human noroviruses in vitro cell culture. As is kwown, viral caspid proteins play a critical role in viral infection and replication. Our previous study demonstrates that the major caspid protein VP1 of norovirus has a direct interaction with the other viral moleculars. Hence the scientific hypothesis is that the capsid protein VP1 of norovirus can regulate the viral replication by interaction with viral proteins or RNA elements, and hence shapes the biological properties and evolution of norovirus. To confirm the hypothesis, replicons of norovirus will be constructed using reverse genetics tools. Luciferase-based reporter gene assays will be used to investigate the precise role of VP1 in the replication of norovirus. Besides, we will determine the key sites that are crucial to the function of VP1 by mutational analysis. Furthermore, Co-IP and CLIP-seq methods will be applied to screen the viral proteins or RNA elements that VP1 targets. These results will unveil the mechanism how VP1 regulation the replication of norovirus. The implementation of this project will contribute to our understanding the role of VP1 in the pathogenicity of norovirus, and provide new strategies for virus prevention and exploring new drug targets against norovirus, and promoting the food safety-risk prevention and control in our county.

诺如病毒是引起急性胃肠炎的重要食源性病毒，由于缺乏稳定的体外细胞培养体系，对其致病机制缺乏深刻认识。衣壳蛋白在病毒感染和复制中扮演着重要角色，前期研究中我们发现诺如病毒主要衣壳蛋白VP1与其它病毒分子间存在相互作用，我们推测：诺如病毒衣壳蛋白VP1通过与维持病毒复制的相关病毒蛋白或RNA元件相互作用，参与调控病毒复制，影响病毒的致病性及进化方向。因此，本项目拟利用反向遗传学技术构建诺如病毒的复制子系统，基于荧光报告系统检测VP1蛋白对诺如病毒复制的影响；并对VP1蛋白进行截短缺失及位点替换，确定VP1蛋白的功能区域；进一步利用Co-IP、CLIP-seq等手段筛选VP1蛋白靶向作用的病毒蛋白或RNA元件，从而揭示VP1蛋白调控病毒复制的分子机制。本项目的开展将进一步阐明VP1蛋白与诺如病毒致病能力的关系，为诺如病毒防控策略的制定和药物靶点挖掘提供新思路，推进我国食品安全风险防控工作。