Transcription Factor Nfya Regulates Mitochondrial Metabolism And Cardiomyocyte Proliferation In The Fetal Heart

We previously identified a neonatal cardiomyocyte population that enters the cell-cycle following injury and disappears as the heart loses the ability to regenerate. These regenerative neonatal cardiomyocytes display a unique transcriptional program regulated by NFYA, a subunit of the trimeric complex Nuclear Transcription Factor Y (NFY) that functions as a pioneer factor to promote cell proliferation. To study whether NFYA is required for cardiomyocyte proliferation, we generated mice with cardiomyocyte-specific deletion of NFYA using alpha-MHC-Cre. NFYA KO mice died before birth; their hearts were smaller in ventricular volumes and developed a distinct cardiac noncompaction phenotype, with hyper trabeculation and reduced cardiomyocyte proliferation. Integration of single-nuclear RNA and ATAC sequencing with spatial transcriptomics identified different embryonic cardiomyocyte subtypes in wildtype and NFYA KO hearts, including those from atria, ventricles, trabeculae, and the conduction system. We found that NFYA deletion reduced the reservoir of immature regenerative cardiomyocytes in the fetal heart and increased the number of trabecular cardiomyocytes. Mechanistically, NFYA deletion reduced the expression of genes regulating oxidative phosphorylation metabolism. The defect in mitochondrial metabolism was further supported by decreased activity of NADH oxidation at mitochondrial membranes and reduced levels of TCA cycle metabolites in NFYA KO hearts. Using chromatin immunoprecipitation and reporter assays, we show that NFYA directly regulates the transcription of those mitochondrial metabolic genes as well as cell-cycle genes in cardiomyocytes, thus providing a transcriptional coupling between mitochondrial metabolism and cardiomyocyte proliferation. Our study identified a previously unrecognized transcriptional control mechanism of metabolic genes controlled by NFYA and highlights the importance of oxidative metabolism during fetal heart development and regeneration.