RNA Polymerase II Dynamics along the Intestinal Crypt-Villus Axis
The mechanisms underlying intestinal epithelial differentiation are essential for maintaining intestinal health. Gene expression analysis reveals vast changes in the transcriptome as cells transition from crypts to villi, impacting nearly 4000 genes. The regulatory mechanisms driving transcriptome shifts during intestinal differentiation remain incompletely understood. Using ChIP-seq and multi-omic analyses, we have identified differential recruitment of Pol II to gene promoters as the primary driver of transcriptomic shifts during differentiation. Using genetic loss-of-function we show that HNF4, a crucial pro-differentiation transcription factor, is required for recruitment of Pol II to hundreds of genes that are activated during differentiation. Dynamic Pol II recruitment corresponds to dynamic enhancer-promoter looping and chromatin remodeling events, indicating a multi-step mechanism driving differentiation gene expression. Additional regulatory mechanisms, such as differential Pol II pause-release and post-transcriptional processes, may govern specific subsets of differentially expressed genes. In summary, our findings emphasize the central role of Pol II recruitment as the major regulator of intestinal differentiation and highlight the significance of HNF4 in this complex process. Overall design: ChIP-seq of RNA Polymerase II (Pol II) was employed to survey its binding patterns in crypt and villus cells of the intestinal epithelium. Pol II ChIP-seq was also applied to Hnf4DKO and littermate control mice to identify HNF4 dependent recruitment patterns of Pol II in the mouse duodenal epithelium.