猪流行性腹泻病毒通过宿主内吞分选转运复合体系统感染宿主的分子机制研究

猪流行性腹泻病毒（PEDV）引起猪流行性腹泻，病死率高，给全球养猪业造成巨大的经济损失。PEDV依赖内体/溶酶体转运，内体转运依赖内吞分选转运复合体（ESCRT）。ESCRT系统中关键连接组分Nedd4、Hrs、Tsg101、Alix、Vps4参与多种病毒感染过程。在前期工作中，我们通过沉默ESCRT组分Tsg101、Alix、Hrs，使PEDV复制受到抑制，Tsg101、Alix与PEDV编码蛋白S、ORF3存在共定位。在此基础上，我们拟构建Nedd4、Hrs、Tsg101、Alix、Vps4及PEDV编码蛋白表达载体，通过免疫共沉淀、激光共聚焦等技术筛选与病毒编码蛋白互作的连接组分。从病毒入胞、复制过程出发，揭示Nedd4、Hrs、Tsg101、Alix、Vps4在PEDV感染宿主细胞中的作用，阐明PEDV劫持ESCRT系统建立感染的机制，为进一步解析PEDV感染机制奠定基础。

Porcine epidemic diarrhea virus (PEDV) is the causative agent of porcine epidemic diarrhea (PED), which has high mortality and causes huge economic losses to the global pig industry. PEDV transport depends on endosome/lysosome, while endosome transport depends on the endocytic sorting and transport complex (ESCRT). The key connection components Nedd4, Hrs, Tsg101, Alix, Vps4 in the ESCRT system are involved in a variety of virus infection processes. We previously found that PEDV replication was inhibited by silencing Tsg101, Alix and Hrs, and that Tsg101 and Alix co-localized with PEDV protein S and ORF3. Considering these observations, we plan to construct the expression vectors of Nedd4, Hrs, Tsg101, Alix, Vps4 and PEDV proteins, and screen the linkers interactions between viral proteins and the ESCRT components by co-immunoprecipitation and laser confocal microscopy. We will reveal the roles of Nedd4, Hrs, Tsg101, Alix, and Vps4 in viral entry and replication within host cells and elucidate the mechanism by which PEDV hijacks the ESCRT system to establish infection, which will lay the foundation for further analysis of PEDV infection mechanism.