Two enhancers synergistically regulate HoxB genes expressions and retinoic acid-induced early ESCs differentiation through 3D chromatin structure

Homeobox B cluster (HoxB) genes play important roles in RA-induced early ESCs differentiation. In this study, we identified two enhancers synergistically regulate HoxB genes expressions through 3D chromatin structure and thus regulate RA-induced early ESCs differentiation. 4C data showed interactions between HoxB genes and the two enhancers. CRISPR/Cas9-mediated individual or compound deletion of the two enhancers significantly inhibits HoxB genes expressions while transcriptome analysis revealed that RA induced early ESCs differentiation was blocked in the enhancer KO cells. Our study suggests new mechanism by which two enhancers regulate HoxB genes expressions and retinoic acid-induced early ESCs differentiation through 3D chromatin structure. 3 biological replicates were analyzed for WT, E1-KO, E2-KO and D-KO by RNA-seq in ESC；2 biological replicates were analyzed for 1-1 WT, 2-1 WT and 2-3 WT by 4C-seq in ESC; 2 biological replicates were analyzed for WT-1 and WT-2 by ChIP-seq in ESC.