H4K20me3 co-localizes with activating histone modifications at transcriptionally dynamic regions in embryonic stem cells

Background: Bivalent chromatin domains consisting of the activating histone 3 lysine 4 trimethylation (H3K4me3) and repressive histone 3 lysine 27 trimethylation (H3K27me3) histone modifications are enriched at developmental genes that are repressed in embryonic stem cells but active during differentiation. However, it is unknown whether another repressive histone modification, histone 4 lysine 20 trimethylation (H4K20me3), co-localizes with activating histone marks in ES cells. Results: Here, we describe the previously uncharacterized coupling of the repressive H4K20me3 heterochromatin mark with the activating histone modifications H3K4me3 and histone 3 lysine 36 trimethylation (H3K36me3), and transcriptional machinery (RNA polymerase II; RNAPII), in ES cells. These newly described bivalent domains consisting of H3K4me3/H4K20me3 are predominantly located in intergenic regions and near transcriptional start sites of active genes, while H3K36me3/H4K20me3 are located in intergenic regions and within gene body regions of active genes. Global sequential ChIP, also termed reChlP-Seq, confirmed the simultaneous presence of H3K4me3 and H4K20me3 at the same genomic regions in ES cells. Genes containing H3K4me3/H4K20me3 exhibit decreased RNAPII pausing and are poised for deactivation of RNAPII binding during differentiation relative to H3K4me3 marked genes. An evaluation of transcription factor (TF) binding motif enrichment revealed that DNA sequence may play a role in shaping the landscape of these novel bivalent domains. Moreover, H3K4me3/H4K20me3 and H3K36me3/H4K20me3 bound regions are enriched with repetitive LINE and LTR elements. Conclusions: Overall, these findings highlight a previously undescribed subnetwork of ES cell transcriptional circuitry that utilizes dual marking of the repressive H4K20me3 mark with activating histone modifications.