摘要
本研究旨在寻找戊型肝炎病毒(hepatitis E virus,HEV)衣壳蛋白ORF2的相互作用蛋白,探讨其在HEV感染中的作用。采用酵母双杂交方法从人肝细胞文库中筛选与HEV ORF2相互作用的蛋白,结果显示CD63与HEV ORF2相互作用。Pull-down实验提示原核表达的ORF2与CD63结合较弱,而免疫共沉淀实验提示真核表达的ORF2能与CD63结合。流式细胞术检测结果显示,HEV易感细胞PLC/PRF/5细胞膜表面的CD63表达水平普遍低于HEV非易感细胞。过表达CD63抑制PLC/PRF/5细胞的HEV感染,而小干扰RNA(small interfering RNA,siRNA)干扰CD63表达则促进HEV感染。结果提示,CD63能与HEV ORF2相互作用,可能抑制HEV感染肝细胞。
The present paper aimed to find out the interacting protein of hepatitis E virus（HEV）capsid protein ORF2 and explore its role in HEV infection.CD63 was identified as a potential HEV ORF2-interacting protein by screening a human liver cDNA library for yeast two-hybrid method. Coimmunoprecipitation and pull-down assays were used to verify the interaction.The expression of cell surface CD63 was detected by flow cytometry in HEV susceptible,non-susceptible and overexpressed cells（PLC/PRF/5）respectively.HEV infectivity was measured through virus infection assay.The results indicated that expression level of CD63 on the cell membrane of HEV susceptible cell lines was lower than that of HEV non-susceptible cell lines.Overexpression of CD63 inhibited the HEV infection of PLC/PRF/5 cells,while siRNA interference of CD63 promoted the HEV infection.The results indicated that CD63 could interact with HEV ORF2,and may inhibit HEV infection in hepatocytes.
出处
《微生物与感染》
2017年第6期348-354,共7页
Journal of Microbes and Infections
基金
国家自然科学基金(81371830)
