Loss of Tip60-dependent H2A.Z acetylation reprogramscardiomyocyte identity toward a regenerative state.

Abstract

Approaches to regenerate cardiomyocytes (CMs) after cardiac injury have been insufficient. Toward this end we are targeting the acetyltransferase Tip60, encoded by the Kat5 gene, based on the rationale that its pleiotropic functions block CM proliferation at multiple checkpoints. We previously reported that genetic depletion or pharmacological inhibition of Tip60 in mice after myocardial infarction (post-MI) reduces scarring, restores function, and activates the cell-cycle in CMs, although it remains unresolved whether daughter CMs are generated. For pre-existing adult CMs in the infarcted adult heart to proliferate, they must first undergo dedifferentiation, a process characterized by loss of maturity, epithelial to mesenchymal transitioning (EMT), and a metabolic shift from fatty acid oxidation (FAO) to glycolysis. Recent studies indicate that Tip60 is required to maintain the differentiated state of hematopoietic stem cells and neurons via site-specific acetylation of the histone variant H2A.Z, specifically H2A.ZacK4/K7. Based on these findings we have examined H2A.ZacK4/K7 levels and the expression of dedifferentiation-associated markers in adult hearts following CM-specific knockout of Tip60. In infarcted/Tip60-depleted hearts, H2A.ZacK4/K7 was largely extinguished in CM nuclei, accompanied by the altered expression of genes consistent with EMT induction, extracellular matrix softening, and reduced FAO. Seahorse metabolic analyses of isolated CMs indicated that Tip60 depletion promotes an oxidative-toglycolytic metabolic transition. In parallel, CUT&Tag analysis of nuclei isolated from heart tissue revealed that Tip60 depletion significantly reduced H2A.ZacK4/K7 occupancy within the promoter/transcription start sites of 47 genes that are highly CM-enriched, including cardiac maturity genes that are preceded in development by the expression of embryonic isoforms. RNAseq and RT-qPCR revealed that expression of these genes adult isoforms, as well as [~]50% of maturity genes that are not preceded by embryonic isoforms, were downregulated in Tip60-depleted hearts. These findings are consistent with the hypothesis that the Tip60 H2A.ZacK4/K7 axis maintains the differentiated state of CMs, constituting a major barrier to cardiac regeneration, justifying clinical targeting.