增殖细胞核抗原通过其DNA结合结构域促进乙肝病毒rcDNA向cccDNA的转化  

PCNA Confers the Conversion of Hepatitis B Virus Relaxed Circular DNA to Covalently Closed Circular DNA by Its DNA Binding Domain

作　　者：袁颖 冯巾晏 赵丽娜[1] 杨光 赵曼 袁红凤 贠昊林 刘姿娴 李悦国[2] 张晓东[1] YUAN Ying;FENG Jin-Yan;ZHAO Li-Na;YANG Guang;ZHAO Man;YUAN Hong-Feng;YUN Hao-Lin;LIU Zi-Xian;LI Yue-Guo;ZHANG Xiao-Dong(Department of Cancer Research,College of Life Sciences,Nankai University,Tianjin 300071,China;Clinical Laboratory,Tianjin Medical University Cancer Institute and Hospital,National ClinicalResearch Cancer for Cancer,Tianjin 300060,China)

机构地区：[1]南开大学生命科学学院肿瘤研究室,天津300071 [2]天津医科大学肿瘤研究所医院临床实验室,国家肿瘤临床研究中心,天津300060

出　　处：《中国生物化学与分子生物学报》2021年第9期1197-1204,共8页Chinese Journal of Biochemistry and Molecular Biology

基　　金：南开大学中央高校基本科研业务费( No.63191741) 资助。

摘　　要：慢性乙型肝炎病毒(HBV)感染是造成肝癌的主要原因,消除病毒基因组中稳定的共价闭合环状DNA(cccDNA)是乙肝治疗的主要难点。HBV cccDNA的形成需要填充单链区域和闭合松弛环状DNA(rcDNA)。我们之前已报道,增殖细胞核抗原(PCNA)参与了HBV cccDNA的形成。然而,HBV rcDNA向cccDNA转化的分子机制尚不清楚。本研究旨在探讨PCNA参与从HBV rcDNA到HBV cccDNA的转化过程的分子机制。结果显示,CRISPR/Cas9系统敲除PCNA能显著阻止HBV rcDNA向cccDNA的转化,而过表达PCNA能够有效地拯救这一现象(P<0.001)。敲除PCNA可显著减缓HBV rcDNA向cccDNA转化的动力学(P<0.01)。在HBV rcDNA向cccDNA转化的过程中,PCNA的DNA结合域是必需的(P<0.01)。由此获得结论,PCNA通过其DNA结合结构域促进乙肝病毒从rcDNA向cccDNA的转化。在临床上,PCNA可能成为抗病毒治疗的新靶点。Chronic hepatitis B virus(HBV)infection is a primary cause for liver cancer.And the main challenge of curing hepatitis B is the elimination of the stable covalently closed circular DNA(cccDNA)of the viral genome.The formation of HBV cccDNA requires the filling of single-stranded region and the ligation of nicks in relaxed circular DNA(rcDNA)strands.Previously,our group reported that proliferating cell nuclear antigen(PCNA)was involved in the formation of HBV cccDNA.However,the underlying mechanism of the conversion of HBV rcDNA to cccDNA is poorly understood.In the present study,we aim to explore the mechanism by which PCNA contributes to the conversion of HBV rcDNA to cccDNA.Our data showed that PCNA was involved in the process of HBV rcDNA repair.The knockout of PCNA by the CRISPR/Cas9 system remarkably blocked the conversion of HBV rcDNA to cccDNA,while the ectopic expression of PCNA could effectively rescue the event(P<0.001).Knockout of PCNA significantly slowed down the conversion kinetics of HBV rcDNA to cccDNA(P<0.01).Mechanically,the DNA binding domain of PCNA was required for the process of HBV rcDNA repair to cccDNA(P<0.01).Thus,we conclude that PCNA confers the conversion of HBV rcDNA to cccDNA by its DNA binding domain.Clinically,PCNA might serve as a novel target for antiviral therapy.

关 键 词：乙型肝炎病毒 闭合松弛环状DNA 共价闭合环状DNA 增殖细胞核抗原 DNA结合域 

分 类 号：R73[医药卫生—肿瘤]