Genome-wide maps of Dot1l associated epigenetic modifications during pluripotency acquistion

Two independent, scaled, chromatin immunoprecipitation experiments were performed to uncover how Dot1l affects epigenetic modification (1) and RNA polymerase II (RNAPII) (2) localization. Mouse embryonic fibroblasts (MEFs), embryonic stem cells (ESCs), and reprogramming MEFs on day 4 treated with control (DMSO) or Dot1l inhibitor (Dot1li -SGC0946) were profiled. 1) Although ESCs have less global enrichment of H3K79me2 by mass spectrometry than MEFs, H3K79me2 is largely localized on the same shared genes in both cells, yet with reduced levels in pluripotent cells. All degrees of H3K79me (1/2/3) are predominately enriched on the same genes across cell types. Treatment with Dot1li evicts H3K79me1/2/3 from MEF specific, ESC specific, and shared genes with H3K79me2 peaks and enables gain of H3K9ac during reprogramming. 2) To understand how genebody epigenetic modifications affect transcription, RNAPII was analyzed. ESCs have less RNAPII enrichment at the transcriptional start site (TSS), but have similar genebody enrichment as somatic cells indicative of more relative elongation. Dot1li treatment during reprogramming reduces RNAPII at the TSS compared to control. H3K79me1, H3K79me2, H3K79me3, H3K9ac, and RNA polymerase II ChIP-Seq in MEFs, ESCs, and reprogramming cells