Department of Physiological Chemistry and Netherlands Proteomic Center, University Medical Centre Utrecht, Universiteitsweg 100, 3584 CG Utrecht, Netherlands.
J Cell Sci. 2010 Aug 1;123(Pt 15):2663-71. doi: 10.1242/jcs.064097. Epub 2010 Jul 13.
Gene transcription in mammalian cells is a dynamic process involving regulated assembly of transcription complexes on chromatin in which the TATA-binding protein (TBP) plays a central role. Here, we investigate the dynamic behaviour of TBP by a combination of fluorescence recovery after photobleaching (FRAP) and biochemical assays using human cell lines of different origin. The majority of nucleoplasmic TBP and other TFIID subunits associate with chromatin in a highly dynamic manner. TBP dynamics are regulated by the joint action of the SNF2-related BTAF1 protein and the NC2 complex. Strikingly, both BTAF1 and NC2 predominantly affect TBP dissociation rates, leaving the association rate unchanged. Chromatin immunoprecipitation shows that BTAF1 negatively regulates TBP and NC2 binding to active promoters. Our results support a model for a BTAF1-mediated release of TBP-NC2 complexes from chromatin.
哺乳动物细胞中的基因转录是一个动态过程，涉及在染色质上调节转录复合物的组装，其中 TATA 结合蛋白（TBP）起着核心作用。在这里，我们通过荧光恢复后荧光漂白（FRAP）和使用不同来源的人细胞系的生化测定的组合来研究 TBP 的动态行为。大多数核质 TBP 和其他 TFIID 亚基以高度动态的方式与染色质结合。TBP 的动力学受 SNF2 相关 BTAF1 蛋白和 NC2 复合物的共同作用调节。引人注目的是，BTAF1 和 NC2 都主要影响 TBP 的解离速率，而不改变结合速率。染色质免疫沉淀显示 BTAF1 负调节 TBP 和 NC2 与活性启动子的结合。我们的结果支持了 BTAF1 介导的 TBP-NC2 复合物从染色质释放的模型。
