Brd4 binds to active enhancers to control cell differentiation and tissue development

Authors

• JE Lee
• YK Park
• A Dey
• K Ozato
• W Peng
• G Kai

Abstract

The epigenomic reader Brd4 is an important drug target for cancers. To investigate its role in cell differentiation and tissue development, we used two independently developed Brd4 conditional knockout (cKO) mouse strains, Brd4 f/f (flanking exon 3) and Brd4 f/f #2 (flanking exon 5). By crossing cKO mice with Myf5-Cre mice to delete Brd4 gene in progenitor cells of brown adipose tissue and muscle, we demonstrate that Brd4 is essential for adipose tissue and muscle development. Consistently, Brd4 is essential for adipogenesis and myogenesis in culture. To map the genomic binding of Brd4, we performed chromatin immunoprecipitation assays with sequencing (ChIP-Seq). Brd4 co-localizes with lineage-determining transcription factors (LDTFs) on active enhancers during cell differentiation. LDTFs coordinate with H3K4 mono-methyltransferases MLL3/MLL4 (KMT2C/KMT2D) and H3K27 acetyltransferases CBP/p300 to recruit Brd4 to enhancers activated during differentiation. Brd4 deletion prevents the enrichment of Mediator and RNA polymerase II transcription machinery, but not that of LDTFs, MLL3/MLL4-mediated H3K4me1, and CBP/p300-mediated H3K27ac, on enhancers. Consequently, Brd4 deletion prevents enhancer RNA production, cell identity gene induction and cell differentiation. Interestingly, Brd4 is dispensable for maintaining cell identity genes in differentiated cells. Our data suggest a model of sequential actions of epigenomic regulators on enhancers: 1) pioneer TFs and LDTFs recruit MLL3/MLL4 to prime enhancer regions and label them with H3K4me1; 2) MLL3/MLL4 facilitate the binding of CBP/p300, which activate enhancers and label them with H3K27ac; 3) H3K27ac and acetylated TFs are recognized by the epigenomic reader Brd4, which recruits Mediator and RNA Polymerase II to establish enhancer-promoter interactions and activate cell type-specific gene expression.

Scientific Focus Area: Developmental Biology

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