Uncoupling histone H3K4 trimethylation from developmental gene expression via an epigenetic equilibrium of Polycomb, COMPASS and DNA methylation

The COMPASS family catalyzes histone H3 lysine 4 (H3K4) methylation and its members are essential for regulating developmental gene expression. MLL2/COMPASS methylates H3K4 on many genes but only a subset lose expression upon MLL2 loss. To understand MLL2 -dependent transcriptional regulation, we performed a CRISPR screen in mouse embryonic stem cells (mESCs) and found that MLL2 protects developmental genes from repression by repelling PRC2 and DNA methylation machineries from these loci. Repression in the absence of MLL2 is relieved by inhibition of PRC2 and DNA methyltransferases, demonstrating that prevention of active repression and not H3K4me3 underlies their transcriptional state. DNA demethylation on such loci leads to reactivation of MLL2-dependent genes not only by removing DNA methylation but also by opening up previously CpG methylated regions for PRC2 recruitment, diluting PRC2 at Polycomb-repressed genes. These findings reveal how the context and function of these three epigenetic modifiers can orchestrate transcriptional decisions. Overall design: CRISPR screen for regulators of MagohB expression followed ChIP-seq, RNAseq, PRO-seq and bisulfite sequencing analyses of selected candidates and their regulators