Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Proc Natl Acad Sci U S A. 2011 May 17;108(20):8385-9. doi: 10.1073/pnas.1019175108. Epub 2011 May 2.
Extra TF(III)C (ETC) sites are chromosomal locations bound in vivo by the RNA polymerase III (Pol III) transcription factor III C (TF(III)C) complex, but are not necessarily associated with Pol III transcription. Although the location of ETC sequences are conserved in budding yeast, and similar sites are found in other organisms, their functions are largely unstudied. One such site, ETC6 in Saccharomyces cerevisiae, lies upstream of TFC6, a gene encoding a subunit of the TF(III)C complex itself. Promoter analysis shows that the ETC6 B-box sequence is involved in autoregulation of the TFC6 promoter. Mutation of ETC6 increases TFC6 mRNA levels, whereas mutation immediately upstream severely weakens promoter activity. A temperature-sensitive mutation in TFC3 that weakens DNA binding of TF(III)C also results in increased TFC6 mRNA levels; however, no increase is observed in mutants of TF(III)B or Pol III subunits, demonstrating a specific role for the TF(III)C complex in TFC6 promoter regulation. Chromatin immunoprecipitation shows an inverse relationship of TF(III)C occupancy at ETC6 versus TFC6 mRNA levels. Overexpression of TFC6 increases association of TF(III)C at ETC6 (and other loci) and results in reduced expression of a TFC6 promoter-URA3 reporter gene. Both of these effects are dependent on the ETC6 B-box. These results demonstrate that the TFC6 promoter is directly regulated by the TF(III)C complex, a demonstration of an RNA polymerase II promoter being directly responsive to a core Pol III transcription factor complex. This regulation could have implications in controlling global tRNA expression levels.
ETC 位点是体内 RNA 聚合酶 III（Pol III）转录因子 III C（TF（III）C）复合物结合的染色体位置，但不一定与 Pol III 转录相关。尽管 ETC 序列在芽殖酵母中的位置是保守的，并且在其他生物中也发现了类似的位点，但它们的功能在很大程度上尚未得到研究。酵母中的 ETC6 就是这样一个位点，它位于 TFC6 基因的上游，TFC6 编码 TF（III）C 复合物的一个亚基。启动子分析表明，ETC6 的 B 框序列参与了 TFC6 启动子的自身调控。ETC6 的突变会增加 TFC6mRNA 的水平，而紧邻上游的突变则严重削弱了启动子的活性。TFC3 的一个温度敏感突变，削弱了 TF（III）C 的 DNA 结合能力，也会导致 TFC6mRNA 水平的增加；然而，在 TF（III）B 或 Pol III 亚基的突变体中没有观察到增加，这表明 TF（III）C 复合物在 TFC6 启动子调控中具有特定的作用。染色质免疫沉淀显示，TF（III）C 在 ETC6 上的占有率与 TFC6mRNA 水平呈反比关系。TFC6 的过表达增加了 TF（III）C 在 ETC6（和其他基因座）上的结合，并导致 TFC6 启动子-URA3 报告基因的表达减少。这两种效应都依赖于 ETC6 的 B 框。这些结果表明，TFC6 启动子直接受到 TF（III）C 复合物的调控，这证明了 RNA 聚合酶 II 启动子可以直接响应核心 Pol III 转录因子复合物。这种调控可能对控制全局 tRNA 表达水平具有重要意义。
