抑制伪狂犬病病毒复制的宿主蛋白的筛选与鉴定  

Screening and Identifying of Host Proteins that Inhibit Pseudorabies Virus Replication

作　　者：何文峰 李琛 常洪涛 李隆熙 陈静 杨国庆[1] 刘慧敏[1] HE Wenfeng;LI Chen;CHANG Hongtao;LI Longxi;CHEN Jing;YANG Guoqing;LIU Huimin(College of Life Science,Henan Agricultural University,Zhengzhou 450002,China)

机构地区：[1]河南农业大学生命科学学院,郑州450002

出　　处：《畜牧兽医学报》2023年第3期1177-1186,共10页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金(31902268);2021年度河南省高等学校青年骨干教师培养计划(2021GGJS034)。

摘　　要：本试验旨在利用TMT蛋白组学分析并筛选出抑制伪狂犬病病毒(pseudorabies virus, PRV)复制的关键宿主蛋白。前期通过TMT定量蛋白组学技术筛选出241个差异蛋白,Western blot和RT-qPCR验证4个差异表达的蛋白,结果与蛋白组学结果一致,表明蛋白组学结果真实可信。通过构建4个差异表达蛋白的真核表达载体,Western blot、RT-qPCR、病毒噬斑结果显示这4个宿主蛋白均可抑制PRV的复制;然后通过双荧光素报告基因检测IFN-β启动子活性,结果显示,MCCC1、STRAP促进IFN-β启动子活性效果更为显著;针对MCCC1和STRAP设计siRNA,Western blot和病毒噬斑结果显示,敲低STRAP显著促进PRV的复制。本试验最终筛选出显著影响PRV复制的宿主蛋白STRAP,为研究PRV与宿主互作的分子机制奠定基础。To screen and analyze anti-pseudorabies virus(PRV) host proteins, four differential proteins were verified by Western blot and RT-qPCR based on the results of TMT proteomics. The results were consistent with the previous results of TMT proteomics, indicating that the proteomic analysis was reliable. Eukaryotic expression vectors of four proteins were constructed, and then detected by Western blot, RT-qPCR and viral plaque assays. The results showed that all the four differential proteins could inhibit PRV replication, while the IFN-β promoter activity of MCCC1 and STRAP proteins were more significant by dual-luciferase reporter gene assay. The MCCC1 and STRAP knockdown by siRNA demonstrated that STRAP significantly promoted PRV replication. In conclusion, we found that the screened STRAP protein remarkably inhibited PRV replication, which will build the foundation for the mechanism underlying interactions between PRV-host.

关 键 词：蛋白组学 伪狂犬病病毒 差异蛋白 STRAP 

分 类 号：S852.659.1[农业科学—基础兽医学]