一个新的负性调控元件对乙型肝炎病毒X启动子启动作用的调节  

Regulation of the hepatitis B virus X promoter activity by a novel negative regulatory element

作　　者：杨扬[1] 吴莹[1] 张文露[1] 余波[1] 黄爱龙[1] 

机构地区：[1]重庆医科大学病毒性肝炎研究所、教育部感染性疾病分子生物学重点实验室,400010

出　　处：《中华肝脏病杂志》2007年第12期893-896,共4页Chinese Journal of Hepatology

基　　金：国家自然科学基金（30600520）

摘　　要：目的在前期工作已证实HBV基因中的250～453nt为一个新的负性调控元件的基础上，探讨其对HBV X启动子（XP）启动作用的影响。方法构建含250～453nt的HBV X启动子驱动虫荧光素酶表达的质粒pNRE—XP，与表达海肾荧光素酶的质粒RL-TK共转染人肝癌HepG2细胞，用双荧光素酶检测系统检测虫荧光素酶表达，RT—PCR检测虫荧光素酶基因mRNA水平的表达，并与相应对照组进行比较。同时，在含HBV全基因组的质粒LJ196上通过缺失突变的方法除去250～453nt片段，将其与反式提供C、P蛋白的质粒LJ96共转染HepG2细胞，RT—PCR检测X基因mRNA表达，并与对照组比较。结果该负性调控元件存在时，HepG2细胞中虫荧光素酶活性明显低于对照组；逆转录聚合酶链反应中虫荧光素酶基因mRNA表达也低于相应的对照组。在LJ196上突变掉250～453tat后，X基因表达量高于对照组。结论该负性调控元件可下调HBV XP的启动作用，在HBV基因组上缺失掉该片段后XP驱动下的基因表达增加。Objective To learn the effect of hepatitis B virus （HBV） sequence nt250-453 on the HBV X promoter. Methods A plasmid pNRE-XP which contains the NRE and the HBV X promoter was constructed to co-transfect HepG2 cell line with plasmid RL-TK. The firefly luciferase activity and mRNA expression of the firefly luciferase gene were both detected. Then, nt250-453 of HBV was removed from LJ196, which contained HBV full genes. The mutated plasmid LJ196 and plasmid LJ96 which provided core protein and the viral DNA polymerase were used to co-transfect HepG2 cell line. Reverse transcription polymerase chain reaction （RT-PCR） was performed to detect the X gene mRNA level. Results The activity of firefly luciferase and the expression of firefly luciferase gene mRNA were both down-regulated in the presence of the NRE, while the HBV X gene mRNA expression increased as it was removed from the HBV genes. Conclusion This study demonstrates that nt250-453 of HBV acts as a novel negative regulatory element which could suppress the HBV X promoter activity.

关 键 词：肝炎病毒 乙型 启动子 基因 调节 

分 类 号：R686[医药卫生—骨科学]