Cardiac effects of histone 3 modifications by CaMKII-EZH2 crosstalk

Histone H3 tail modifications are pivotal in epigenetic regulation of the chromatin state. CaMKII - a catecholamine-sensitive kinase plays both adaptive and maladaptive roles in different forms of heart failure - has been reported to phosphorylate Histone 3 at Serine 28 residue (H3S28p). However, the contribution of this mechanism to heart failure development has not yet been determined. Our initial findings in neonatal rat ventricular myocytes (NRVMs) indicate EZH2 and CaMKII may interact epigenetically at Histone 3 tail in cardiac tissues. An increased CaMKII-dependent H3S28p mark inversely correlates with EZH2-mediated histone H3 lysine 27 trimethylation levels (H3K27me3) as a transcription silencing mark. This in turn correlates with a time dependent increased levels of the open chromatin mark H3K27ac. While pressure overload-induced heart failure in wild-type mice resulted in increased H3S28p levels, this histone modification was unaffected in cardiac-specific CaMKII-deficient mice upon pressure overload. Using isolated cardiomyocyte nuclei, ChIP-seq analysis reveals a genome-wide association between H3S28p and H3K27ac PTMs, strongly indicative of CaMKII mediated H3S28p mark as an open chromatin mark enhancing transcription events at the target sites. The main target regions impacted by these PTMs are gene promoter and transcription regulatory sites. Several genes associated with cardiac metabolism, transcription regulation and sarcomere and cytoskeleton organization were detected to be associated with H3S28p and H3K27ac enrichment, using gene-ontology (GO) analysis. We were able to identify two predominant cardiac-specific H3 genes with the aid of RNA-seq analysis using cardiomyocyte nuclei. AAV9-mediated, cardiomyocyte-specific expression of CaMKII and PKA enhanced H3S28p levels. CaMKII expression resulted in severe systolic dysfunction, whereas PKA expression left mice unaffected. These findings indicate a locus-specific role for the H3S28p mark, mediated by distinct kinases. A novel mouse model with an alanine substitution at serine-28 of H3 has been generated and will shed light on the significance of this H3 PTM under cardiac stress. This work allows to investigate chromatin remodeling downstream of CaMKII and its contribution to cardioprotection versus -damage. Furthermore, this study sheds light on exact modifications at the chromatin landscape mediated by CaMKII/PKA activity during cardiac stress and homeostasis.