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Stephan, Tabea Leonie

University of British Columbia

Hepatocytes differentiate from definitive endoderm in response to external signaling cues. However, it is unclear how the hepatic lineage is specified and how transcription factors shape the chromatin to allow gene expression. In this study, we used a human stem cell model of liver development, differentiating cells stepwise into endodermal and hepatic progenitors combined with whole-genome epigenomic mapping. To assess the chromatin dynamics during differentiation, we mapped the histone modifications H3K4me1, H3K4me3, H3K27ac, H3K27me3 and H3K36me3 using ChIP-seq at four time points. We used this data to identify 655,208 enhancers and 20,517 promoters and define their activation state across the differentiation. Enhancers were found to exhibit dynamic regulation, with many primed at the definitive endoderm. Tracing the fate of the primed enhancers, we identified hepatic-specific enhancers, which gained active histone modifications, in contrast to non-hepatic enhancers, that subsequently lost H3K4me1 after hepatic specification. Promoters were more stable than enhancers, but also showing dynamic patterns in their regulation. To identify regulating transcription factors, we performed ChIP-seq for two identified predicted regulators, TBX3 and HNF4A, before and after hepatic specification. TBX3, and to lesser extend HNF4A, bound to the hepatic enhancers and formed an interdependent network with known pioneer factors. Mechanistically, TBX3 bound both hepatic enhancers and promoters prior to H3K4 methylation, likely recruiting methyltransferase complexes through interactions with RbBP5. However, TBX3 binding was transient and H3K4 methylation was observed after eviction of TBX3. Interestingly, a subset of promoters that specifically gained H3K4me3 in hepatoblasts and showed immediate upregulation of gene expression was specifically bound by TBX3. This was in contrast to the majority of identified promoters that gained active histone modifications early on but required the activation of associated enhancers to increase gene expression. This suggests that TBX3 plays a role in identifying and priming hepatic enhancers and promoters during specification. These data revealed a very dynamic developmental chromatin landscape, that allowed us to identify TBX3 as a potential driver of hepatic cell identity. Our comprehensive developmental histone modification dataset will serve as a resource for researchers studying the possible reactivation of developmental processes in cancer or regeneration.

Text

2024-03-11

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/87552

Cell and Developmental Biology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/