Histone methylation and transcription factor binding during intestinal cell differentation

Cell differentiation requires epigenetic modulation of tissue-specific genes and activities of master transcriptional regulators, which are recognized for their dominant control over cellular programs. Using novel epigenomic methods, we characterized enhancer elements specifically modified in differentiating intestinal epithelial cells and found enrichment of transcription factor-binding motifs corresponding to CDX2, a master regulator of the intestine. Directed investigation revealed surprising lability in CDX2 occupancy of the genome, with redistribution from hundreds of sites occupied only in progenitors to thousands of new sites in mature cells. Knockout mice confirmed distinct Cdx2 requirements in dividing and differentiated adult intestinal cells, including responsibility for the active enhancer configuration associated with maturity. Dynamic CDX2 occupancy corresponds with condition-specific gene expression and, importantly, to differential co-occupancy with other tissue-restricted transcription factors: HNF4A in mature cells and GATA6 in progenitors. These results reveal dynamic, context-specific functions and mechanisms of a master transcription factor within a cell lineage. H3K4me2, CDX2, GATA6, and HNF4A ChIP-seq patterns were mapped in proliferating and 26 day post-confluent (differentatied) Caco-2 intestinal cell lines and compared to published expression data or newly generated Cdx2 knockout mouse intestine expression data细胞分化需要对组织特异性基因进行表观遗传调控（epigenetic modulation），以及主控转录调节因子（master transcriptional regulators）的活性调控——这类因子因对细胞程序的显性调控作用而被学界熟知。本研究采用新型表观基因组学方法，对分化过程中肠道上皮细胞内发生特异性修饰的增强子元件（enhancer elements）进行了系统表征，发现与肠道主控调节因子CDX2对应的转录因子结合基序（transcription factor-binding motifs）存在显著富集。定向追踪研究揭示了CDX2在基因组上的结合具有出人意料的不稳定性：其结合位点从仅在肠道祖细胞中占据的数百个位点发生重分布，转而在成熟细胞中形成数千个全新结合位点。基因敲除小鼠（knockout mice）实验证实，Cdx2在增殖型与成熟型成年肠道细胞中具有截然不同的功能需求，其中包括负责维持与细胞成熟相关的活性增强子构型（active enhancer configuration）。CDX2的动态结合与细胞状态特异性的基因表达模式高度相关，尤为重要的是，其与其他组织限制性转录因子（tissue-restricted transcription factors）的共结合模式存在显著差异：在成熟细胞中为HNF4A，在祖细胞中则为GATA6。上述研究结果揭示了细胞谱系内主控转录因子的动态、环境依赖性功能与作用机制。本研究对增殖型及汇合后培养26天（分化成熟型）的Caco-2肠道细胞系中H3K4me2、CDX2、GATA6与HNF4A的染色质免疫共沉淀测序（ChIP-seq）图谱进行了定位，并将其与已发表的基因表达数据，或新生成的Cdx2基因敲除小鼠肠道组织基因表达数据进行了比对。