外源性HCV RDRP对宿主细胞的表观遗传调控及在肝癌发病中的作用

项目名称： 外源性HCV RDRP对宿主细胞的表观遗传调控及在肝癌发病中的作用

项目编号： No.31271355

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 生物科学

项目作者： 于文强

作者单位： 复旦大学

项目金额： 80万元

中文摘要： 细胞内源性的RDRP在植物及线虫中RNA介导的基因沉默和DNA甲基化中发挥重要作用。在人类细胞中siRNA仅能引起靶位点异染色质形成，难于导致DNA甲基化。RITS和RISC复合物组分比较发现人类细胞不存在确定性的RDRP。丙肝病毒RDRP在感染宿主细胞后除复制自身的基因组外，能否替代内源性的RDRP对宿主细胞的表观遗传学修饰状态产生影响呢？本项目通过将高活性HCV RDRP导入宿主细胞后，研究外源性HCV RDRP对宿主细胞表观遗传学修饰的影响；利用TAP和质谱技术解析RDRP复合物组分，探讨其作用机制；生物信息学分析寻找肝癌特异性miRNA在肿瘤抑制基因启动子区域靶位点，验证RDRP在肝癌肿瘤抑制基因沉默中的作用。本项目研究有助于回答两个重要科学问题：（1）siRNA在哺乳动物细胞中难于引起DNA甲基化的分子机制；（2）HCV RDRP在肝癌发病的作用，为肝癌治疗和预防提供新的思路

中文关键词： 表观遗传学；RNA 干扰；RNA依赖型RNA聚合酶；DNA甲基化；肝癌

英文摘要： Endogeneous RDRP plays an important role in RNA-directed gene silencing and DNA methylation in plants and C.elegans.In human cells,siRNA can only cause the formation of heterochromatin on its target sites without detectable DNA methylation.Comparing components of RITS and RISC complexes reveals that there is no convincible evidence which shows that human cells do exist endogenous RDRP. So is there any possibility that exogenous HCV NS5B, in addition to participate in HCV genome replication, has any other function like modifying the epigenetic status of host human cells? In our research project,HCV RDRP with high activity will be transfected to human cells to find out how the exogenous RDRP tunes the epigenetic modifications of host cells. The components of the RDRP complex will be determined by TAP and mass-spectrometry;The target sites of hepatoma-specific miRNA in the promoter of tumor suppressor gene will be identified and the function of RDRP in the tumor suppressor gene silencing will be verified. Conclusions from this project will contribute to answer two fundamental scientific questions:(1)The mechanism by which siRNA rarely induces DNA mathylation in mammalian cells;(2)Role of exogenous HCV RDRP played in pathogenesis of HCC, and shed new light on HCC prevention and treatment.

英文关键词： Epigenetics；RNA interference；RNA-dependent RNA polymerase；Hepatocellular carcinoma；DNA methylation