Identification of downstream effectors of retinoic acid specifying the zebrafish pancreas by integrative genomics

Retinoic acid (RA) is a key signal for the specification of the pancreas. Still, the gene regulatory cascade triggered by RA in the endoderm remains poorly characterized. In this study, we investigated this regulatory network in zebrafish by combining RNA-seq, RAR ChIP-seq and ATAC-seq assays. By analysing the effect of RA and of the RA receptor (RAR) inverse-agonist BMS493 on the transcriptome and on the chromatin accessibility of endodermal cells, we identified a large set of genes and regulatory regions regulated by RA signalling. RAR ChIP-seq further defined the direct RAR target genes in zebrafish, including hox genes as well as several pancreatic regulators like mnx1, insm1b, hnf1ba and gata6. Comparison of zebrafish and murine RAR ChIP-seq data highlighted the conserved direct target genes and revealed that some RAR sites are under strong evolutionary constraints. Among them, a novel highly conserved RAR-induced enhancer was identified downstream of the HoxB locus and driving expression in the nervous system and in the gut in a RA-dependent manner. Finally, ATAC-seq data unveiled the role of the RAR-direct targets Hnf1ba and Gata6 in opening chromatin at many regulatory loci upon RA treatment.