Occlusion of Regulatory Sequences by Promoter Nucleosomes In Vivo
Occlusion of Regulatory Sequences by Promoter Nucleosomes In Vivo
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DOI：
10.1371/journal.pone.0017521
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发表时间：
2011-03-03
期刊：
影响因子：
3.7
通讯作者：
Boeger, Hinrich
中科院分区：
文献类型：
--
作者：
Mao, Changhui;Brown, Christopher R.;Boeger, Hinrich
核小体被认为通过转录因子抑制DNA结合。试图理解核小体在基因表达和调控中的重要性的理论就是基于这一假设。然而，核小体对转录因子与DNA结合的抑制并不完全。相反，进入核小体DNA取决于许多因素，包括转录因子- DNA相互作用的立体化学、核小体结构中热波动的体内动力学以及转录因子的细胞内浓度。因此，体外结合研究必须辅以体内测量。酵母的诱导型PHO 5启动子在这一讨论中发挥了突出的作用。它带有转录激活因子Pho 4的两个结合位点，其在阻遏的启动子处分别位于核小体内和两个核小体之间的接头区域中。早期的研究表明，在没有染色质重塑的情况下，核小体结合位点无法与Pho 4结合。然而，最近的几份报告对这一概念提出了质疑。因此，我们重新分析了转录因子结合到体内的PHO 5启动子，使用“染色质内源性切割”（ChEC）。我们的研究结果明确表明，核小体有效地干扰PHO 4和其他关键转录因子的PHO 5启动子的调控序列的结合。我们的数据还表明，PHO 4招募TATA盒结合蛋白的PHO 5启动子。
Nucleosomes are believed to inhibit DNA binding by transcription factors. Theoretical attempts to understand the significance of nucleosomes in gene expression and regulation are based upon this assumption. However, nucleosomal inhibition of transcription factor binding to DNA is not complete. Rather, access to nucleosomal DNA depends on a number of factors, including the stereochemistry of transcription factor- DNA interaction, the in vivo kinetics of thermal fluctuations in nucleosome structure, and the intracellular concentration of the transcription factor. In vitro binding studies must therefore be complemented with in vivo measurements. The inducible PHO5 promoter of yeast has played a prominent role in this discussion. It bears two binding sites for the transcriptional activator Pho4, which at the repressed promoter are positioned within a nucleosome and in the linker region between two nucleosomes, respectively. Earlier studies suggested that the nucleosomal binding site is inaccessible to Pho4 binding in the absence of chromatin remodeling. However, this notion has been challenged by several recent reports. We therefore have reanalyzed transcription factor binding to the PHO5 promoter in vivo, using 'chromatin endogenous cleavage' (ChEC). Our results unambiguously demonstrate that nucleosomes effectively interfere with the binding of Pho4 and other critical transcription factors to regulatory sequences of the PHO5 promoter. Our data furthermore suggest that Pho4 recruits the TATA box binding protein to the PHO5 promoter.