乙型脑炎病毒NS1蛋白与宿主蛋白互作的分子机制

Japanese encephalitis virus(JEV) is an arbovirus maintained in a zoonotic cycle,which has assumed increasing public health importance. NS1 protein is a critical factor on virus replication, immunology protection, neuroinvasiveness and pathogenic consequence.Until nowadays, the most mechanism of NS1 protein is uncertain.We thus develop a new strategy based on Japanese Encephalitis virus replicon and infections cDNAs labled by Strep-tag. This allows NS1-Host protein complex analysis directly from transfected/infected cells by combining modified tandem affinity chromatography and mass spectrometry analysis, then protein interaction network was established.Through RNAi and over-expressing, the function of interaction proteins will be made more clearly in virus life cycle. By Alanine-Scanning Mutagenesis, the mapped domains of NS1 protein will be evaluated in vitro and in vivo. Altogether, this project will strongly support the following basic study and disease control.

乙型脑炎是一种重要的人兽共患传染病。NS1蛋白在病毒的复制、免疫保护、神经侵染性和致病性方面都有重要作用。迄今为止，NS1蛋白的大部分作用机制还不清楚。本研究拟通过一种新型高效的研究蛋白互作的方法，利用Strep标签标记的JEV复制子及JEV感染性cDNA克隆，通过Pull Down技术和LC-MS/MS分析，鉴定与NS1蛋白互作的宿主蛋白，建立相关的蛋白互作网络；进一步通过针对相关蛋白的RNAi和过表达方式，系统研究相关蛋白在JEV生命周期中的作用；利用丙氨酸扫描诱变技术确定NS1蛋白的互作功能区域，通过对相关功能区域的修饰，在体内和体外评估其功能。揭示乙型脑炎病毒NS1蛋白与宿主蛋白的互作网络和分子机制，为该乙型脑炎的基础研究和综合防控提供科学数据和技术支持。