3D chromatin organisation contributes to the regulatory logic in differentiating lymphatic endothelium

Abstract

Gene activation and repression is an integral part of embryonic development and tissue formation. Changes in chromatin organisation dictate accessibility to gene regulatory elements, controlling gene expression. Although several molecular regulators of lymphatic endothelial cell (LEC) development have been identified, the role of chromatin organisation in the acquisition of LEC identity remains unclear. In this study, we combine HiC and ATAC-sequencing to map 3D chromatin architecture and accessibility in LECs and blood endothelial cells (BECs). We identify cell type-specific topologically associating domains (TADs), and discovered TAD boundaries changes and differentially segregating enhancers in lymphatic-associated loci, such as prox1a and tbx1. Our multi-omic approach also defines the regulatory logic of nine LEC-enriched genes. In vivo validation of ATAC- and HiC-based enhancers confirms their activity in LECs. Leveraging these datasets, we reconstructed mafba tissue-specific regulatory networks identifying a genetic interaction with tfe3a in vivo limiting ectopic vessel formation. Overall, our work provides a powerful resource of multi-omic datasets that can be used to systematically determine the regulatory networks governing LEC identity and genes linked to lymphatic disease. The topologically associating domains (TADs) and chromatin accessibility of lymphatic endothelial cells diverge from those of blood endothelial cells. These differences underlie the activation of LEC-specific regulatory networks governing cell identity and involving genes linked to lymphatic disease. The LEC and BEC populations present lineage-specific chromatin organisation at the level of both TADs and chromatin accessibility. Tissue-specific chromatin structural changes are association with key genes involved in LEC differentiation. Novel local enhancers of lymphatic enriched genes are marked by differentially accessible chromatin. Tissue-specific chromatin loops connect the promoters of known vascular and lymphatic regulators, such as mafba and tbx1, with long-range enhancers active in the lymphatic endothelium. Transcription factor footprinting revealed an interaction between mafba and tfe3, which limits ectopic vessel formation and ensures a correct vascular differentiation. The LEC and BEC populations present lineage-specific chromatin organisation at the level of both TADs and chromatin accessibility. Tissue-specific chromatin structural changes are association with key genes involved in LEC differentiation. Novel local enhancers of lymphatic enriched genes are marked by differentially accessible chromatin. Tissue-specific chromatin loops connect the promoters of known vascular and lymphatic regulators, such as mafba and tbx1, with long-range enhancers active in the lymphatic endothelium. Transcription factor footprinting revealed an interaction between mafba and tfe3, which limits ectopic vessel formation and ensures a correct vascular differentiation. The topologically associating domains (TADs) and chromatin accessibility of lymphatic endothelial cells diverge from those of blood endothelial cells. These differences underlie the activation of LEC-specific regulatory networks governing cell identity and involving genes linked to lymphatic disease.