Gonzales-Cope Michelle, Sidoli Simone, Bhanu Natarajan V, Won Kyoung-Jae, Garcia Benjamin A
Epigenetics Program, Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA.
The Institute for Diabetes, Obesity, and Metabolism, Philadelphia, PA, 19104, USA.
BMC Genomics. 2016 Feb 4;17:95. doi: 10.1186/s12864-016-2414-y.
Pluripotent cells can be differentiated into many different cell types in vitro. Successful differentiation is guided in large part by epigenetic reprogramming and regulation of critical gene expression patterns. Recent genome-wide studies have identified the distribution of different histone-post-translational modifications (PTMs) in various conditions and during cellular differentiation. However, our understanding of the abundance of histone PTMs and their regulatory mechanisms still remain unknown.
Here, we present a quantitative and comprehensive study of the abundance levels of histone PTMs during the differentiation of mouse embryonic stem cells (ESCs) using mass spectrometry (MS). We observed dynamic changes of histone PTMs including increased H3K9 methylation levels in agreement with previously reported results. More importantly, we found a global decrease of multiply acetylated histone H4 peptides. Brd4 targets acetylated H4 with a strong affinity to multiply modified H4 acetylation sites. We observed that the protein levels of Brd4 decreased upon differentiation together with global histone H4 acetylation. Inhibition of Brd4:histone H4 interaction by the BET domain inhibitor (+)-JQ1 in ESCs results in enhanced differentiation to the endodermal lineage, by disrupting the protein abundance dynamics. Genome-wide ChIP-seq mapping showed that Brd4 and H4 acetylation are co-occupied in the genome, upstream of core pluripotency genes such as Oct4 and Nanog in ESCs and lineage-specific genes in embryoid bodies (EBs).
Together, our data demonstrate the fundamental role of Brd4 in monitoring cell differentiation through its interaction with acetylated histone marks and disruption of Brd4 may cause aberrant differentiation.
多能细胞可在体外分化为多种不同的细胞类型。成功的分化在很大程度上由表观遗传重编程和关键基因表达模式的调控所引导。最近的全基因组研究已经确定了不同组蛋白翻译后修饰（PTM）在各种条件下以及细胞分化过程中的分布。然而，我们对组蛋白PTM丰度及其调控机制的理解仍然未知。
在此，我们使用质谱（MS）对小鼠胚胎干细胞（ESC）分化过程中组蛋白PTM的丰度水平进行了定量和全面的研究。我们观察到组蛋白PTM的动态变化，包括H3K9甲基化水平升高，这与先前报道的结果一致。更重要的是，我们发现多重乙酰化组蛋白H4肽的整体水平下降。Brd4以高亲和力靶向乙酰化H4，作用于多重修饰的H4乙酰化位点。我们观察到，分化时Brd4的蛋白水平与整体组蛋白H4乙酰化水平一起下降。在ESC中，BET结构域抑制剂（+）-JQ1抑制Brd4与组蛋白H4的相互作用，通过破坏蛋白质丰度动态变化，导致向内胚层谱系的分化增强。全基因组ChIP-seq图谱显示，Brd4和H4乙酰化在基因组中共定位，位于ESC中核心多能性基因（如Oct4和Nanog）以及胚状体（EB）中谱系特异性基因的上游。
总之，我们的数据证明了Brd4通过与乙酰化组蛋白标记相互作用在监测细胞分化中的基本作用，破坏Brd4可能导致异常分化。
