Genome-wide quantitative functional enhancer map of mESCs, in the pluripotent state and after induced differentiation

Precise spatiotemporally regulated gene expression is pivotal for cell homeostasis, cell differentiation and during development. Gene expression networks are tightly regulated by transcription factors (TF) and their targeted regulatory genomic elements (enhancers), which are known to correlate with specific histone modifications. However, the ultimate prerequisites, which determine functionally active enhancers, remain unclear. To elucidate the regulatory landscape of murine ESCs, a massively parallel reporter assay, based on STARR-seq (Self-Transcribing Active Regulatory Region sequencing), assessing genomic fragments prepared from accessible chromatin, (FAIRE-STARR-seq) has been applied. Thus, a genome-wide quantitative map of functional mESC enhancers in naive state and after retinoic acid (RA)-induced differentiation was generated. To investigate sequence features associated with RA-mediated inducibility of enhancers bound by retinoic acid receptor alpha (RARa), the main receptor for RA, genomic binding sites of RARa have been determined. Overall design: mESCs, in the pluripotent state or after induced differentiation, were assessed for genome-wide changes enhancer activity by FAIRE-STARR-seq. Additionally, RARa genomic binding sites after RA-induced differentiation were identified by ChIP-seq.