Abstract Body (Do not enter title and authors here): Background: Chromatin structure plays a central role in regulating gene expression and maintaining cellular identity. However, the structural factors driving these processes in cardiomyocytes (CM), particularly in the context of cardiac disease, remain poorly defined.
Hypothesis: The expression of chromatin structure regulators (CSRs) in CM is significantly altered between healthy and diseased individuals, and targeting CSRs affects CM phenotypes.
Approach: To investigate whether the differential expression of these factors can distinguish healthy from diseased CM populations, we generated a comprehensive list of 1,166 CSRs based on an extensive literature review. We re-analyzed a published single-cell RNA sequencing dataset (GSE1838529) from human hearts with and without dilated cardiomyopathy (DCM) using a CSR-specific Autoencoder. CM subpopulations enriched in DCM subjects were then identified. Pathological expression of key CSRs, such as HMGN3, was confirmed in human DCM tissues, mice that underwent transverse aortic constriction (TAC), and pigs that underwent myocardial infarction (MI) using western blotting and immunohistochemistry. The role of HMGN3 in CM was explored by using siRNA transfection in the human cardiomyocyte AC16 cell line. Survival rate was quantified using a CCK8 kit, apoptosis was assessed via cleaved caspase-3, and chromatin accessibility was measured via histone marks western blotting and DNAse digestion protocol.
Results: CSR-specific analysis identified four distinct CM subpopulations (CM1–CM4), with CM1 being exclusive to DCM subjects. This particular subpopulation exhibits an enrichment in DCM markers and a reduction in contractile gene expression. CM1 also shows significant alterations in the expression of 25 CSRs, including HMGN3. Western blotting and immunohistochemistry validated the decrease in HMGN3 expression in human DCM, TAC mouse, and MI pig tissues. In AC16 cells, HMGN3 knockdown led to increased apoptosis and a reduction in active chromatin marks, alongside increased chromatin compaction.
Conclusions: CSR expression can be utilized to effectively stratify CM disease populations, with HMGN3 playing an important role in modulating the CM phenotype through the regulation of chromatin accessibility. These findings further underscore the critical role of chromatin organization in shaping the cardiac phenotype.