Proteogenomics analysis reveals specific genomic orientations of distal regulatory regions composed by non-canonical histone variants.
Proteogenomics analysis reveals specific genomic orientations of distal regulatory regions composed by non-canonical histone variants.
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DOI：
10.1186/s13072-015-0005-9
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发表时间：
2015
影响因子：
3.9
通讯作者：
Garcia BA
中科院分区：
文献类型：
--
作者：
Won KJ;Choi I;LeRoy G;Zee BM;Sidoli S;Gonzales-Cope M;Garcia BA
组蛋白变体在DNA包装和控制基因表达中起着更重要的作用。然而，我们对它们的组成和功能的了解是有限的。整合蛋白质组学和基因组学方法，我们对包含四种组蛋白变体H3.1，H3.3，H2A.Z和macroH 2A的表观遗传景观进行了全面分析。这些组蛋白在HeLa细胞中被FLAG标记，并使用染色质免疫沉淀（ChIP）纯化。通过采用ChIP随后质谱（ChIP-MS），我们定量组蛋白翻译后修饰（PTM）和组蛋白变体核小体的比例在高度纯化的单核小体。随后的ChIP和下一代测序（ChIP-seq）用于绘制所分析的组蛋白变体的全基因组定位并定义其染色质结构域。最后，我们在我们的研究中包括ENCODE数据库中包含的大型数据集。我们新发现了一组富含H3.1和组蛋白变体的调控区域，这些区域与抑制性标记macroH 2A相关。系统分析确定了对称和不对称模式的组蛋白变异发生在基因间调控区。引人注目的是，这些方向模式与RNA聚合酶II（PolII）有关。这些不对称模式与使用全局连续测序（GR 0-seq）数据测量的增强子活性相关。我们的研究表明，H2A.Z和H3.3描绘了在启动子处观察到的增强子处的转录方向。我们还发现，增强子与倾斜的组蛋白变异模式，以及促进增强子的活性。总的来说，我们的研究表明，组蛋白变体沉积在调控区域，以协助基因调控。本文的在线版本（doi：10.1186/s13072-015-0005-9）包含补充材料，可供授权用户使用。
Histone variants play further important roles in DNA packaging and controlling gene expression. However, our understanding about their composition and their functions is limited. Integrating proteomic and genomic approaches, we performed a comprehensive analysis of the epigenetic landscapes containing the four histone variants H3.1, H3.3, H2A.Z, and macroH2A. These histones were FLAG-tagged in HeLa cells and purified using chromatin immunoprecipitation (ChIP). By adopting ChIP followed by mass spectrometry (ChIP-MS), we quantified histone post-translational modifications (PTMs) and histone variant nucleosomal ratios in highly purified mononucleosomes. Subsequent ChIP followed by next-generation sequencing (ChIP-seq) was used to map the genome-wide localization of the analyzed histone variants and define their chromatin domains. Finally, we included in our study large datasets contained in the ENCODE database. We newly identified a group of regulatory regions enriched in H3.1 and the histone variant associated with repressive marks macroH2A. Systematic analysis identified both symmetric and asymmetric patterns of histone variant occupancies at intergenic regulatory regions. Strikingly, these directional patterns were associated with RNA polymerase II (PolII). These asymmetric patterns correlated with the enhancer activities measured using global run-on sequencing (GRO-seq) data. Our studies show that H2A.Z and H3.3 delineate the orientation of transcription at enhancers as observed at promoters. We also showed that enhancers with skewed histone variant patterns well facilitate enhancer activity. Collectively, our study indicates that histone variants are deposited at regulatory regions to assist gene regulation. The online version of this article (doi:10.1186/s13072-015-0005-9) contains supplementary material, which is available to authorized users.
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