Department of Biomedical Sciences, University at Albany School of Public Health, Albany, NY 12201-0509, USA.
Computational Biology Branch, National Center for Biotechnology Information, National Library of Medicine, NIH, Bethesda, MD 20814, USA.
Nucleic Acids Res. 2020 May 7;48(8):4244-4255. doi: 10.1093/nar/gkaa165.
Transcription of eukaryotic mRNA-encoding genes by RNA polymerase II (Pol II) begins with assembly of the pre-initiation complex (PIC), comprising Pol II and the general transcription factors. Although the pathway of PIC assembly is well established, the mechanism of assembly and the dynamics of PIC components are not fully understood. For example, only recently has it been shown that in yeast, the Mediator complex normally occupies promoters only transiently, but shows increased association when Pol II promoter escape is inhibited. Here we show that two subunits of TFIID, Taf1 and Taf4, similarly show increased occupancy as measured by ChIP upon depletion or inactivation of Kin28. In contrast, TBP occupancy is unaffected by depletion of Kin28, thus revealing an uncoupling of Taf and TBP occupancy during the transcription cycle. Increased Taf1 occupancy upon Kin28 depletion is suppressed by depletion of TBP, while depletion of TBP in the presence of Kin28 has little effect on Taf1 occupancy. The increase in Taf occupancy upon depletion of Kin28 is more pronounced at TFIID-dominated promoters compared to SAGA-dominated promoters. Our results support the suggestion, based on recent structural studies, that TFIID may not remain bound to gene promoters through the transcription initiation cycle.
真核生物 mRNA 编码基因由 RNA 聚合酶 II（Pol II）转录起始，首先组装前起始复合物（PIC），包括 Pol II 和一般转录因子。尽管 PIC 组装途径已得到充分证实，但组装机制和 PIC 组件的动力学仍未完全了解。例如，直到最近才发现，在酵母中，中介体复合物通常仅短暂占据启动子，但当 Pol II 启动子逃避被抑制时，其结合增加。在这里，我们发现 TFIID 的两个亚基 Taf1 和 Taf4 在 Kin28 耗尽或失活时，如 ChIP 测量所示，其占有率同样增加。相比之下，TBP 占有率不受 Kin28 耗尽的影响，因此在转录循环中揭示了 Taf 和 TBP 占有率的解偶联。Kin28 耗尽时 Taf1 占有率的增加被 TBP 耗尽所抑制，而 Kin28 存在时 TBP 的耗尽对 Taf1 占有率几乎没有影响。与 SAGA 主导的启动子相比，Kin28 耗尽时 Taf 占有率的增加在 TFIID 主导的启动子上更为明显。我们的结果支持了基于最近结构研究的建议，即 TFIID 可能不会在整个转录起始循环中保持与基因启动子的结合。
