Department of Urology, USC Norris Comprehensive Cancer Center, Keck School of Medicine, University of Southern California, Los Angeles, CA 90089, USA.
Proc Natl Acad Sci U S A. 2011 Aug 30;108(35):14497-502. doi: 10.1073/pnas.1111309108. Epub 2011 Aug 15.
Recent epigenome-wide mapping studies describe nucleosome-depleted regions (NDRs) at transcription start sites and enhancers. However, these static maps do not address causality or the roles of NDRs in gene control, and their relationship to transcription factors and DNA methylation is not well understood. Using a high-resolution single-molecule mapping approach to simultaneously investigate endogenous DNA methylation and nucleosome occupancies on individual DNA molecules, we show that the unmethylated OCT4 distal enhancer has an NDR, whereas NANOG has a clear NDR at its proximal promoter. These NDRs are maintained by binding of OCT4 and are required for OCT4 and NANOG expression. Differentiation causes a rapid loss of both NDRs accompanied by nucleosome occupancy, which precedes de novo DNA methylation. NDRs can be restored by forced expression of OCT4 in somatic cells but only when there is no cytosine methylation. These data show the central role of the NDRs, established by OCT4, in ensuring the autoregulatory loop of pluripotency and, furthermore, that de novo methylation follows the loss of NDRs and stabilizes the suppressed state.
最近的全基因组图谱研究描述了转录起始位点和增强子处的核小体缺失区域（NDR）。然而，这些静态图谱并不能解决因果关系或 NDR 在基因调控中的作用，并且人们对其与转录因子和 DNA 甲基化的关系也了解甚少。我们使用高分辨率单分子作图方法，同时研究单个 DNA 分子上的内源性 DNA 甲基化和核小体占有率，结果表明，未甲基化的 OCT4 远端增强子具有 NDR，而 NANOG 在其近端启动子处则具有明显的 NDR。这些 NDR 通过 OCT4 的结合得以维持，并且对于 OCT4 和 NANOG 的表达是必需的。分化导致这两个 NDR 迅速丢失，伴随着核小体占有率的增加，这发生在新的 DNA 甲基化之前。在体细胞中强制表达 OCT4 可以恢复 NDR，但只有在没有胞嘧啶甲基化的情况下才可以。这些数据表明，OCT4 建立的 NDR 在确保多能性的自我调节环中起着核心作用，此外，新的甲基化紧随 NDR 的丢失，并稳定抑制状态。
