泛素特异性肽酶22调控乙型肝炎病毒X蛋白水平及功能  

Ubiquitin-specific peptidase 22(USP22)regulates the level and function of hepatitis B virus X protein

作　　者：周世颖 吴琼 张翼 张璐 林旭 陈婉南 Zhou Shiying;Wu Qiong;Zhang Yi;Zhang Lu;Lin Xu;Chen Wannan(Key Laboratory of Gastrointestinal Cancer of the Ministry of Education,Fujian Provincial Key Laboratory of Tumor Microbiology,School of Basic Medical Sciences,Fujian Medical University,Fuzhou 350122,China)

机构地区：[1]福建医科大学基础医学院,消化道恶性肿瘤教育部重点实验室,福建省肿瘤微生物学重点实验室,福州350122

出　　处：《中华微生物学和免疫学杂志》2022年第6期434-442,共9页Chinese Journal of Microbiology and Immunology

基　　金：福建省自然科学基金(2019J01299);福建省财政厅项目(2019B027);国家自然科学基金(81672031);福建医科大学起航基金(2018QH2005)。

摘　　要：目的探究泛素特异性肽酶22(ubiquitin specific peptidase 22,USP22)与乙型肝炎病毒X蛋白(hepatitis B virus X protein,HBx)相互作用对HBx蛋白水平及其生物学功能的影响。方法通过GST pull-down、免疫共沉淀以及激光共聚焦试验检测HBx与USP22的相互作用。瞬时转染USP22真核表达质粒或干扰USP22表达的siRNA,Western blot检测肝细胞中HBx蛋白表达水平的变化。以环己酰亚胺(cycloheximide,CHX)、蛋白酶体抑制剂MG132(carbobenzoxy-Leu-Leu-leucinal,MG132)分别处理转染后的细胞,检测HBx蛋白的降解速率。进一步通过细胞内泛素化试验检测USP22对HBx泛素化的影响,明确USP22影响HBx蛋白稳定性的具体机制。最后,以双荧光素酶报告试验和平板克隆试验检测USP22对HBx生物学功能的影响。结果USP22与HBx存在相互作用。USP22显著增加HBx蛋白稳定性,并通过去除HBx的泛素化抑制HBx通过蛋白酶体降解,进而增强HBx的生物学功能。结论USP22抑制HBx通过泛素依赖-蛋白酶体途径降解,增强HBx蛋白稳定性并促进HBx功能。Objective To investigate the effects of the interaction between ubiquitin-specific peptidase 22(USP22)and hepatitis B virus X protein(HBx)on the protein level and the biological function of HBx.Methods The interactions between HBx and USP22 were analyzed by GST pull-down,co-immunoprecipitation assay and confocal laser scanning assay.USP22 recombinant plasmids or specific siRNA were transiently co-transfected with HBx plasmids.Western blot were used to detect the protein level of HBx.The half-life and degradation pathway of HBx in the transfected cells treated with cycloheximide(CHX)or proteasome inhibitor MG132 were detected.In vivo ubiquitination assay was used to detect the ubiquitination of HBx with USP22 overexpression.Moreover,dual-luciferase reporter assay and colony formation assay were used to analyze the effects of USP22 on the biological function of HBx.Results USP22 could interact with HBx in vivo and in vitro.USP22 significantly increased the stability of HBx and inhibited the proteasome-mediated degradation of HBx protein by reducing the ubiquitination of HBx,thereby enhancing the biological function of HBx.Conclusions USP22 inhibited HBx protein degradation through ubiquitin-dependent proteasome pathway,thus enhancing the stability and biological function of HBx.

关 键 词：乙型肝炎病毒X蛋白 泛素特异性肽酶22 泛素 蛋白酶体 蛋白降解 

分 类 号：R512.62[医药卫生—内科学]