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孤儿核受体hB1F (NR5A2 ,也称之为LRH 1、FTF或CPF)在胆汁酸合成代谢相关基因、乙型肝炎病毒基因和一些肝特异性基因的表达调控中起着非常重要的作用。为了认识hB1F转录激活的分子机制 ,对核受体辅抑制子SMRT在其中的作用进行了研究。利用与GAL4 DBD融合的hB1F所进行的报告基因分析发现 ,SMRT能够以剂量依赖的方式显著地抑制hB1F的反式激活能力 ,但对融合蛋白质的表达没有影响。而且 ,SMRT也能够明显地抑制hB1F响应的乙肝病毒增强子II 核心启动子的活性 ,对增强子II的点突变分析表明这种抑制作用主要是由hB1F介导的。共转染实验显示 ,SMRT在多种细胞中都表现出抑制作用。有意思的是 ,哺乳动物细胞的双杂交与体外的GST下拉 (pull down)分析都表明hB1F与SMRT之间不存在直接的相互作用。实验数据首次证实 ,辅抑制子SMRT可能通过间接的方式特异地抑制孤儿核受体hB1F的转录活性。
The orphan nuclear receptor hB1F (NR5A2, also known as LRH1, FTF or CPF) plays a very important role in the regulation of the expression of bile acid anabolic genes, hepatitis B virus genes and some liver-specific genes. In order to understand the molecular mechanism of hB1F transcriptional activation, the role of nuclear receptor co-repressor SMRT in it was studied. Reporter gene analysis using hB1F fused to GAL4 DBD found that SMRT significantly inhibited the transactivation ability of hB1F in a dose-dependent manner but had no effect on the expression of the fusion protein. Moreover, SMRT also significantly inhibited hB1F-responsive hepatitis B enhancer II core promoter activity, and point mutation analysis of enhancer II showed that this inhibition was primarily mediated by hB1F. Co-transfection experiments showed that SMRT showed inhibition in a variety of cells. Interestingly, both two-hybrid and mammalian GST pull down assays in mammalian cells indicate that there is no direct interaction between hB1F and SMRT. Experimental data for the first time confirmed that co-suppressor SMRT may indirectly inhibit the orbital nuclear receptor hB1F transcriptional activity.