YTHDF1 impacts cardiomyocyte differentiation by regulating the TET2 mRNA

Abstract

Cardiac diseases frequently arise from compromised cardiomyocyte differentiation and function. Although studies have demonstrated that loss of the m 6 A modification reader YTHDF1 impairs cardiomyocyte differentiation, the specific mRNA transcripts it directly regulates remain to be identified. Comprehensive analysis of public databases was conducted to examine the correlation between YTHDF1 expression and cardiac differentiation processes as well as specific cardiac pathology. Stable YTHDF1-knockdown cell lines were generated in the rat H9C2 cardiomyoblasts. After retinoic acid (RA) induction, cardiomyocyte differentiation was assessed. RNA immunoprecipitation sequencing (RIP-seq) was performed in H9C2 cells, and the resulting data were integrated with known cardiomyocyte differentiation regulators to identify direct YTHDF1 mRNA targets. Here we found that YTHDF1 expression increases progressively during cardiomyocyte differentiation but gradually declines upon cellular/organ maturation in mice. Human left ventricle (LV) exhibited higher YTHDF1 expression than right ventricle (RV), while LV from dilated cardiomyopathy (DCM) patients showed modestly reduced YTHDF1 levels compared to healthy controls. In differentiating H9C2 cardiomyoblasts, YTHDF1 expression progressively increased. YTHDF1 knockdown impaired differentiation, reducing maturation markers cTnT/cTnI, which was aligned with RNA-seq analysis. RIP-seq identified significant TET2 mRNA enrichment in YTHDF1 complexes. YTHDF1 knockdown selectively reduced TET2 protein level without affecting its mRNA level, while YTHDF1 overexpression enhanced TET2 translation. Additionally, we identified a novel rat tet2 variant. Complementation with this variant in YTHDF1-knockdown H9C2 cells rescued the differentiation defect. Collectively, YTHDF1 promotes cardiomyocyte differentiation by regulating TET2 mRNA during cardiac development.