Genome-wide profiling of transcription, H3K27me3 and RING1B occupancy in mouse ESCs following EZH1/2 inhibition

Mouse embryonic stem cells (mESCs) were treated with DMSO (control) or EPZ-6438 (EPZ; a potent inhibitor of the EZH1/2) for 24 h. ChIP-seq was used to determine the impact of this treatment on the level and distribution of H3K27me3 and RING1B genome-wide. In parallel, gene expression was assessed by deep sequencing of both total RNA (RNA-seq) and 4-thiouridine metabolically labelled RNA (4sU-seq). Calibrated ChIP-seq, utilizing Drosophila chromatin as an exogenous control, demonstrated a marked global depletion of H3K27me3 following 24 h of EPZ treatment concomitant with a substantial reduction in RING1B occupancy. A rather modest change in the levels of total and nascent transcripts allowed us to identify a direct contribution of the polycomb system in controlling chromatin architecture and 3D nuclear organization. Overall design: Following 24 h of treatment with either DMSO or EPZ-6438, mESCs were processed for ChIP-seq (calibrated H3K27me3 and non-calibrated RING1B), RNA-seq and 4sU-seq. Two independent replicate experiments were prepared for each sample.