Tip60 complex binds to active Pol II promoters and a subset of enhancers and co-regulates the c-Myc network in mouse embryonic stem cells

BackgroundTip60 (KAT5) is the histone acetyltransferase (HAT) of the mammalian Tip60/NuA4 complex. While Tip60 is important for early mouse development and mouse embryonic stem cell (mESC) pluripotency, the function of Tip60 as reflected in a genome-wide context is not yet well understood.ResultsGel filtration of nuclear mESCs extracts indicate incorporation of Tip60 into large molecular complexes and exclude the existence of large quantities of “free” Tip60 within the nuclei of ESCs. Thus, monitoring of Tip60 binding to the genome should reflect the behaviour of Tip60-containing complexes. The genome-wide mapping of Tip60 binding in mESCs by chromatin immunoprecipitation (ChIP) coupled with high-throughput sequencing (ChIP-seq) shows that the Tip60 complex is present at promoter regions of predominantly active genes that are bound by RNA polymerase II (Pol II) and contain the H3K4me3 histone mark. The coactivator HAT complexes, Tip60- and Mof (KAT8)-containing (NSL and MSL), show a global overlap at promoters, whereas distinct binding profiles at enhancers suggest different regulatory functions of each essential HAT complex. Interestingly, Tip60 enrichment peaks at about 200 bp downstream of the transcription start sites suggesting a function for the Tip60 complexes in addition to histone acetylation. The comparison of genome-wide binding profiles of Tip60 and c-Myc, a somatic cell reprogramming factor that binds predominantly to active genes in mESCs, demonstrate that Tip60 and c-Myc co-bind at 50–60 % of their binding sites. We also show that the Tip60 complex binds to a subset of bivalent developmental genes and defines a set of mESC-specific enhancer as well as super-enhancer regions.ConclusionsOur study suggests that the Tip60 complex functions as a global transcriptional co-activator at most active Pol II promoters, co-regulates the ESC-specific c-Myc network, important for ESC self-renewal and cell metabolism and acts at a subset of active distal regulatory elements, or super enhancers, in mESCs.

背景：Tip60（KAT5）是哺乳动物Tip60/NuA4复合物的组蛋白乙酰转移酶（histone acetyltransferase, HAT）。尽管Tip60对小鼠早期发育及小鼠胚胎干细胞（mouse embryonic stem cell, mESC）多能性至关重要，但目前在全基因组层面上对Tip60的功能仍缺乏充分认知。
结果：对小鼠胚胎干细胞细胞核提取物进行凝胶过滤层析实验，结果显示Tip60被整合入高分子量复合物中，未检测到胚胎干细胞细胞核内存在大量“游离”Tip60。因此，通过监测Tip60与基因组的结合情况，可准确反映含Tip60复合物的行为特征。本研究采用染色质免疫共沉淀（chromatin immunoprecipitation, ChIP）结合高通量测序（high-throughput sequencing, ChIP-seq）技术，对小鼠胚胎干细胞中Tip60的结合位点开展全基因组定位分析，结果显示：Tip60复合物主要富集于RNA聚合酶II（RNA polymerase II, Pol II）结合且带有H3K4me3组蛋白修饰的活跃基因启动子区域。含Tip60与Mof（KAT8）的共激活因子型HAT复合物（包括NSL与MSL亚型），在启动子区域呈现全局共定位特征；但二者在增强子区域的结合谱存在显著差异，提示这两种关键HAT复合物具备不同的调控功能。值得注意的是，Tip60的富集峰位于转录起始位点下游约200 bp处，这提示Tip60复合物除介导组蛋白乙酰化外，还具备其他未知功能。将Tip60的全基因组结合谱与c-Myc（一种体细胞重编程因子，在小鼠胚胎干细胞中主要结合活跃基因）的结合谱进行比较后发现，Tip60与c-Myc共同结合的位点占二者各自结合位点的50%~60%。本研究还证实，Tip60复合物可结合一类二价发育基因，并界定了一批小鼠胚胎干细胞特异性的增强子及超级增强子区域。
结论：本研究表明，Tip60复合物可作为多数活跃Pol II启动子区域的全局性转录共激活因子，协同调控与小鼠胚胎干细胞自我更新及细胞代谢密切相关的ESC特异性c-Myc调控网络，并在小鼠胚胎干细胞的一类活跃远端调控元件（即超级增强子）中发挥关键调控作用。