Abstract Body: Background: Cardiovascular diseases remain a leading cause of morbidity and mortality worldwide, largely due to the limited regenerative capacity of cardiomyocytes. Unlike embryonic cardiomyocytes, adult cardiomyocytes are terminally differentiated and unable to re-enter the cell cycle after injury. Epigenetic regulation plays a central role in this developmental transition. DNMT3B, a de novo DNA methyltransferase expressed during early cardiac development, declines sharply during cardiomyocyte maturation, coinciding with cell-cycle exit.
Hypothesis: We hypothesized that reactivation of DNMT3B in adult cardiomyocytes remodels the epigenome to suppress differentiation programs while activating mitotic pathways, enabling dedifferentiation and cell-cycle re-entry.
Approach: DNMT3B was overexpressed in human iPSC-derived cardiomyocytes and in mouse cardiomyocytes following MI. Integrated multi-omics analyses were performed including RNA-seq, DNA methylation profiling, DNMT3B ChIP-seq, and histone modification profiling. Cardiomyocyte proliferation was assessed by Ki67, PHH3 and Aurora-B immunostaining, and cardiac function was evaluated by echocardiography.
Results: DNMT3B overexpression induced extensive epigenetic remodeling characterized by dominant hypermethylation (696 hyper vs 113 hypo). Hypermethylated loci were enriched in differentiation genes and DNA demethylases TET1, TET2, and TET3, indicating suppression of endogenous demethylation pathways. In contrast, cell-cycle regulators MKI67, TOP2A, CCND2, etc. showed hypomethylation and increased expression. The top hypomethylated gene, INCENP, encodes Inner Centromere Protein, a core component of the chromosomal passenger complex that activates Aurora-B kinase. DNMT3B ChIP-seq demonstrated promoter occupancy at proliferative loci including CDK4 and CCND2, accompanied by increased H3K36me3 enrichment. Functionally, DNMT3B increased proliferating cardiomyocytes (p<0.01) and significantly improved cardiac function post MI.
Conclusion: DNMT3B integrates DNA methylation and chromatin remodeling to suppress differentiation programs while activating mitotic regulators, promoting cardiomyocyte plasticity and proliferation.