下调细胞P21^(WAF1/CIP1)基因表达可抑制单纯疱疹病毒Ⅱ型复制  

Silencing Cellular P21^(WAF1/CIP1) by Small Interfering RNA Inhibits Herpes Simplex Virus Type Ⅱ Propagation

作　　者：周琦[1] 张浩[1] 王波[1] 彭宜红[1] 屠静[1] 朱萌[1] 徐萍[2] 

机构地区：[1]北京大学医学部病原生物学系,北京100191 [2]北京大学医学部药学院,北京100191

出　　处：《中国生物化学与分子生物学报》2009年第4期339-344,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家高技术发展研究项目863计划(No.2007AA02Z317);国家自然科学基金(No.30470084)资助~~

摘　　要：为了揭示细胞P21蛋白在单纯疱疹病毒Ⅱ型(herpes simplex virus type2,HSV-2)复制中的作用,通过用HSV-2感染和感染前用特异性小干扰RNA(small interfering RNA,siRNA)抑制P21基因表达,应用Western印迹方法检测宿主细胞和病毒蛋白水平,用终点滴定法测定病毒半数组织培养感染量(50%tissue culture infectious dose,TCID50),以及观察感染细胞的细胞病变效应(cytopathic effect,CPE)等3个方面,揭示细胞P21蛋白水平的变化对病毒复制的影响.结果表明,HSV-2在细胞内复制时可引起P21蛋白水平增高;而用特异性siRNA下调细胞P21基因表达时,可显著地抑制HSV-2gB蛋白水平,减少培养细胞上清液中病毒TCID50.提示P21蛋白对HSV-2的复制具有重要的作用.The regulation of cell cycle progression during viral DNA replication has evolved as an important propagation strategy for herpesvirus. Upon infection of the cell, herpesvirus frequently interfers with cellular DNA synthesis, although the exact mechanisms are unclear. P21 is a universal cyclin kinase inhibitor （CKI） that inhibits cell cycle progression. To determine whether P21 is involved in the process of herpes simplex virus type Ⅱ （HSV-2） proliferation, NIH3T3 cells were challenged with one multiplicity of infection （MOI） of HSV-2. The cytopathic effect （CPE） caused by virus replication was observed by microscope, 50% tissue culture infectious dose （ TCID50 ） representing the virus titers was determined by end-point assay, cellular and viral proteins were detected by Western blot. Overall, these results indicated a positive correlation between levels of the virus and P21. Most importantly, the knockdown of P21 by specific small interfering RNA （siRNAs） effectively reduced HSV-2 replication by 90 %, thus providing direct evidence that P21 upregnlation has a significant role for HSV-2 replication. Altogether, our data suggest that cellular P21 is favor for the replication of HSV-2.

关 键 词：单纯疱疹病毒Ⅱ型 P21蛋白 小干扰RNA 

分 类 号：Q936[生物学—微生物学]