Guermah Mohamed, Palhan Vikas B, Tackett Alan J, Chait Brian T, Roeder Robert G
Laboratory of Biochemistry and Molecular Biology, The Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.
Cell. 2006 Apr 21;125(2):275-86. doi: 10.1016/j.cell.2006.01.055.
We have reconstituted a highly purified RNA polymerase II transcription system containing chromatin templates assembled with purified histones and assembly factors, the histone acetyltransferase p300, and components of the general transcription machinery that, by themselves, suffice for activated transcription (initiation and elongation) on DNA templates. We show that this system mediates activator-dependent initiation, but not productive elongation, on chromatin templates. We further report the purification of a chromatin transcription-enabling activity (CTEA) that, in a manner dependent upon p300 and acetyl-CoA, strongly potentiates transcription elongation through several contiguous nucleosomes as must occur in vivo. The transcription elongation factor SII is a major component of CTEA and strongly synergizes with p300 (histone acetylation) at a step subsequent to preinitiation complex formation. The purification of CTEA also identified HMGB2 as a coactivator that, while inactive on its own, enhances SII and p300 functions.
我们重新构建了一个高度纯化的RNA聚合酶II转录系统，该系统包含由纯化的组蛋白和组装因子组装而成的染色质模板、组蛋白乙酰转移酶p300以及通用转录机制的组分，这些组分本身足以在DNA模板上进行激活转录（起始和延伸）。我们发现该系统可介导染色质模板上依赖激活因子的起始，但不能进行有效的延伸。我们进一步报道了一种染色质转录促进活性（CTEA）的纯化，该活性以依赖p300和乙酰辅酶A的方式，强烈增强转录通过几个连续核小体的延伸，这在体内必然会发生。转录延伸因子SII是CTEA的主要成分，并且在起始前复合物形成后的一个步骤中与p300（组蛋白乙酰化）强烈协同作用。CTEA的纯化还鉴定出HMGB2作为一种共激活因子，它虽然自身无活性，但可增强SII和p300的功能。
