Transcriptional Coordination and Gene Regulation by MED12 in the Cardiomyocyte

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Heart disease is the leading cause of mortality worldwide and disruption of transcription in cardiomyocytes (CMs) contributes to the progression of heart disease. Transcriptional regulation of some individual genes by transcription factors (TFs) is described. However, much less is known about how functionally related genes, or gene expression programs, are collectively regulated in a coordinated manner in healthy and diseased CMs. The Mediator Complex serves as a bridge to link transcriptional machinery and TFs to control transcription, but the molecular mechanisms of transcriptional regulation by Mediator are not well-understood. Mediator is a large multiprotein complex organized as submodules, with the CDK8 kinase submodule regulating RNA Polymerase II transcriptional activity. MED12 is an essential Mediator component, and within the kinase submodule, is required for CDK8 kinase activity. Although traditionally considered a transcriptional repressor via the CDK8 kinase submodule, our published data demonstrates that increased MED12 is also able to directly activate transcription. Our preliminary studies indicate that MED12 levels are increased in failing hearts from humans and mice, sup- porting the clinical relevance of this novel MED12 function in mis-regulation of transcription in heart disease. Our published studies in mice with CM-specific Med12 deletion demonstrate that MED12 regulates a calcium-handling gene expression program, through direct interaction with the TF MEF2. We identified additional TFs that interact with MED12, and these findings suggest that MED12 has multiple transcriptional functions depending on its interactions, but its functions have not been delineated. To determine the contribution of MED12 to transcriptional mis-regulation in heart disease, it is crucial to define its molecular interactions and functions in CMs and identify its cell-specific roles. Subpopulations of kinase submodules have been reported in other cells, but their functions have not been defined. Additionally, functions of individual kinase submodule proteins have not been investigated for their roles in alternative kinase subcomplexes or for functions independent of the kinase submodule. Our published CM-specific Med12 knockout model (cKO) and newly generated CM Med12 transgenic model (cTg) both develop heart failure, emphasizing the critical contribution of MED12 to transcriptional homeostasis in CMs. We will use our Med12 genetic models to delineate the functions of MED12 in CMs. We will identify MED12 kinase subpopulations, interactions with TFs, and genomic interactions in normal CMs and those with MED12 overexpression. We will also assess the ability of a human MED12 missense mutations to bind TFs and kinase submodule proteins using hi-PSCs. The long-term goal of my lab is to uncover mechanisms that control coordinated transcriptional programs in the heart. In this proposal we aim to 1) determine the mechanisms of MED12-regulated transcription in CMs, and 2) identify the transcriptional mechanisms of MED12- driven heart failure. Consequently, a better understanding of the molecular mechanisms governing cardiac gene expression will provide new insight into therapeutic strategies to restore cardiac function during disease.