BRG1相关因子155对乙型肝炎病毒X蛋白翻译后水平的调控及意义

Hepatitis B virus X protein (HBx) is one of the most important pathogenic factors of hepatitis B virus (HBV) whose expression level is closely correlated with the pathogenicity of HBV. However, the mechanisms underlying the regulation of HBx protein level remain to be fully elucidated. Our prior work has demonstrated the interaction of BRG1-associated factor 155 (BAF155) with HBx by using CytoTrap yeast two-hybrid screening and GST- pull down assay. BAF155 could inhibit the degradation of its binding partner proteins through ubiquitin-dependent and -independent proteasomal pathway. This study will first confirm the interaction of BAF155 with HBx and identify their binding regions by co-immunoprecipitation (Co-IP). Then, the effect of BAF155 on HBx protein level will be evaluated by over-expression and siRNA knock-down of BAF155, and mechanisms by which BAF155 regulates HBx protein expression will be explored by both ubiquitin reporter assay and Co-IP. Lastly, the effects of BAF155-HBx interaction on the biological function, e.g., cell proliferation, apoptosis, et al, of HBx will be investigated. This study will aid in a better understanding of HBV pathogenicity through elucidating the regulation of HBx by BAF155 at protein level.

乙型肝炎病毒（Hepatitis B Virus，HBV）X蛋白（HBx protein）表达水平高低与HBV致病性密切相关，但其蛋白水平的调控机制尚未完全阐明。我们前期利用酵母双杂交与GST-pull down显示BRG1相关因子155（BRG1-associated factor 155，BAF155）与HBx存在相互作用。BAF155能通过泛素依赖和泛素非依赖途径抑制与其结合的蛋白降解。本研究首先通过免疫共沉淀等实验明确HBx与BAF155的相互作用及各自结合区域；随后探讨过表达及RNA干扰抑制BAF155表达对HBx蛋白水平的影响，并以泛素过表达及免疫共沉淀探讨其机制，最后探讨BAF155通过对HBx蛋白水平的调控对细胞增殖、凋亡等的影响。本研究旨在阐明BAF155对HBx蛋白翻译后水平的调控及机制，所获结果将有助于更深入了解HBV致病机制。

乙型肝炎病毒X蛋白（hepatitisbvirus X protein，HBx）是一种具有多种生物学功能的反式激活因子，在乙型肝炎病毒（hepatitisbvirus，HBV）复制和肝癌发生发展中起着重要作用。HBx半衰期短，主要由泛素依赖的蛋白酶体降解途径调节。本项目的前期研究工作使用CytoTrap酵母双杂交筛选系统，获得与HBx相互作用的蛋白BAF155（BRG1相关因子155，BRG1-associated factor 155；又名染色质重塑因子BAF155），BAF155已被证明通过减弱蛋白酶体降解来稳定SWI/SNF染色质重塑复合物的其他成员，本研究提出BAF155也可能有助于以蛋白酶体依赖的方式稳定HBx蛋白。本项目研究结果表明，BAF155通过与20S蛋白酶体亚基PSMA7竞争结合HBx，来保护HBx免受泛素非依赖性蛋白酶体降解。BAF155过表达增加了HBx蛋白水平，而siRNA介导的内源性BAF155的敲除降低了HBx蛋白水平。BAF155增加HBx的稳定性和稳态水平可归因于抑制泛素非依赖的和PSMA7介导的蛋白质降解。此外，BAF155的过表达增强了HBx的反式激活功能，激活了原癌基因的表达，抑制了肝癌细胞的克隆形成。这些结果提示BAF155可能在HBx的泛素非依赖性降解中起重要作用，并可能参与HBV相关性肝癌的发病机制。