Genome-wide distribution of KAT6B and H3K23ac in mouse embryonic stem cells
Promoters of developmental genes in embryonic stem cells (ESCs) are marked by histone H3 lysine 4 trimethylation (H3K4me3) and H3K27me3 in an asymmetric nucleosomal conformation, with each sister histone H3 carrying one of the two marks. These so-called bivalent domains are thought to poise genes for timely activation upon differentiation. Here we show that asymmetric bivalent nucleosomes recruit repressive H3K27me3 binders but fail to enrich activating H3K4me3 binders, despite presence of H3K4me3, thereby promoting a poised state. Strikingly, the bivalent mark combination further promotes binding of chromatin proteins that are not recruited by each mark individually, including the histone acetyltransferase complex KAT6B (MORF). Knockout of KAT6B blocks neuronal differentiation, demonstrating that bivalency-specific readers are critical for proper ESC differentiation. These findings reveal how histone mark bivalency directly promotes establishment of a poised state at developmental genes, while highlighting how nucleosomal asymmetry is critical for histone mark readout and function. Overall design: E14 mouse embryonic stem cells were engineered to express HA-tagged KAT6B from its endogenous locus. In addition, mutations were introduced to render MLL2 catalytically inactive (Y2688A mutation) or to abrogate PRC2 function (EZH2-/-; EZH1-/-). KAT6B distribution was profiled via the added HA tag, while antibodies against H3K23ac, H3K4me3, and H3K27me3 were used to profile the corresponding histone modifications. Further, mutations were introduced in subunits of the KAT6B complex (ING2 Y188A, KAT6B D290A, BRPF1 D270/272A, BRPF2 D212/214A) and KAT6B (via HA tag) and H3K23ac were profiled. Furthermore, E14 mouse embryonic stem cells both wild-type and deficient for KAT6A and KAT6B (KAT6A-/-; KAT6B-/-) were profiled for distribution of H3K23ac, H3K4me3, and H3K27me3. Drosophila spike-in calibration was applied to enable quantitative comparisons within the following sets of samples: HA-KAT6B line with and without catalytically inactive MLL2 and EZH1/2 double knockout; HA-KAT6B line with and without point mutations in histone mark binding domains; E14 cells with and without KAT6A/B knockouts.