Regulation and Function of MEF2 in Cardiomyocytes

Regular formation of the mammalian heart needs precise spatial and temporal transcriptional regulation of gene programs in ardiomyocytes. Cardiac transcription factors are defined, in this context, as essential transcriptional activators that are expressed predominantly in the myocardium which regulate the expression of the cardiac genes encoding structural proteins of cardiomyocytes. Unsurprisingly, disruptions in this elaborate transcriptional machinery can lead to severe cardiac abnormalities including hypertension, cardiomyopathy, and congenital heart disease. In this field, Myocyte Enhancer Factor 2 (MEF2) transcription factor is considered one of only a few core cardiac transcription factors that play important roles in cardiac development, survival, contractility, and in postnatal adaption to a wide array of physiological and pathological signals. MEF2 functions as a transcriptional switch by potently activating or repressing transcription through interaction with a variety of co-factors which serve as positive and negative regulators of transcription. The interaction of MEF2 with its co-factors is controlled by a multitude of signaling pathways that result in post-translational modification of MEF2 and in the subsequent MEF2-dependent repression or activation of target gene transcription. Our project studied regulation and function of MEF2A in cardiomyocytes. We hypothesized that the combinatorial interactions between transcription factors and promoter elements that are required for the regulation of cardiac gene expression may operate in pathological cardiac remodeling and hypertrophy. Therefore, studying and characterizing the regulation of proteins which bind to MEF2A in cardiomyocytes may unravel the underlying dysregulation of the cardiac transcriptome in the pathogenesis of cardiovascular disease and heart failure. In this project, HL-1 cardiomyocytes have been chosen as a model of study. An agonist (Isoproterenol) was used to mimic cardiomyocytes hypertrophy in HL-1 cells. Isoproterenol activates adrenergic signaling which can trigger many mechanisms in the heart contributing to the hypertrophic phenotype. We developed two different methods to capture MEF2A interacting partners (interactome), including immunoprecipitation (IP) of endogenous MEF2A and IP of Flag-MEF2A proteins in normal and hypertrophy conditions. Our optimization will allow characterization of MEF2A interactome partners through state of the art quantitative proteomics approaches. In previous research, transcriptome analysis (RNA-seq) from left ventricular RNA samples and MEF2A depleted cardiomyocytes identified some genes, including kf2, junb, alas2 and rarres2 which may have implications for cardiac hypertrophy. Our ChIP-qPCR data indicated that MEF2A is recruited to the rarres2 promoter in primary cardiomyocytes. Thus, rarres2 is a novel MEF2A target gene and further, it will be interesting to uncover functions of MEF2A interactome partners on rarres2 gene regulation in cardiac diseases. A study has indicated that klf2, junB, alas2, and rarres2 may have a role in promoting cardiomyocyte hypertrophy in cultured HL-1 cells and primary neonatal rat cardiomyocytes. Taken together, this project developed the methods to study the characterization of MEF2A interactome in cardiomyocytes. Additionally, we showed the capacity of some MEF2 target genes, including rarres2 to promote cardiomyocyte hypertrophy