Translational Control of Cardiac Excitability

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Project Summary/Abstract The expression of ion channels underlying the rhythmic beating of the heart must be precisely coordinated to fulfill their physiological roles and protect the heart from arrhythmia. Many aspects of how this task is accomplished remain poorly understood. Our preliminary findings suggest a novel way in which ion channel expression is coordinated. The central hypothesis of this proposal is that a “micro- translatome” of interacting mRNA species encodes functionally related proteins, such as those encoding the ventricular action potential. These ion channels assemble co-translationally into macromolecular complexes that govern higher-order cardiac excitability. We will test this hypothesis using a range of experimental preparations including human ventricular myocardium, cardiomyocytes derived from human induced pluripotent stem cells (iPSC-CM's), animal models and HEK293 cells. We will probe the co- regulation and interaction of transcripts encoding ventricular ion channels, and determine whether such assemblies predict stable macromolecular protein complexes within cardiomyocytes using RNA immunoprecipitation experiments, protein co-immunoprecipitation, patch- clamp electrophysiology and super-resolution microscopy. Using RNA-seq, we will identify other transcripts in the micro-translatome, including those encoding RNA binding proteins that tether the transcripts together, and test their roles using RNAi. We will test the hypothesis that mechanisms of mRNA processing, such as nonsense- mediated decay and miRNA regulation, coordinately control the components of the action potential micro-translatome to modify the disease state and fulfill normal, physiological roles. These experiments will uncover mechanisms that quantitatively regulate the critical balance of cardiac excitability, the perturbation of which triggers catastrophic ventricular arrhythmias.