Dystrophin-glycoprotein complex in viral cardiomyopathy

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Dilated cardiomyopathy is a multifactorial disease that includes both hereditary and acquired forms of cardiomyopathy. Recent experiments have shown that hereditary cardiomyopathy in humans can be associated with genetic defects in components of the dystrophin-glycoprotein complex. In addition, we have shown that an acquired form of cardiomyopathy caused by enteroviral infection is associated with viral protease 2A mediated cleavage of dystrophin and disruption of the dystrophin-glycoprotein complex. This raises two important questions regarding the pathogenesis of viral mediated dilated cardiomyopathy. First, how does cleavage of dystrophin affect the kinetics of viral replication and viral propagation; and second, how does cleavage of dystrophin contribute to the cardiomyopathy caused by enteroviral infection? Therefore, the overall goal of this proposal is to precisely identify the protease 2A cleavage site in dystrophin and to determine the effect of protease 2A mediated cleavage of dystrophin in cultured cells and in the intact heart. In order to accomplish this goal, we propose the following Specific Aims: 1. Determine whether cleavage in the hinge 3 region of dystrophin is required for proteolytic cleavage of human and mouse dystrophin by protease 2A in vitro and in cell culture, 2. In cell culture determine the effect of wild type and cleavage resistant dystrophin expression on viral replication kinetics and cell death; in mice determine the impact of CVB3 infection on organization of the dystrophin associated cytoskeletal proteins and other cytoskeletal proteins that are not part of the dystrophin-glycoprotein complex; and in mice, determine the effect of viral replication in CVB3 infected mice that lack either dystrophin or dystrophin-associated sarcoglycans. 3. Determine how cleavage of dystrophin by protease 2A affects the kinetics of viral replication and induction of the full myocarditic/cardiomyopathic phenotype associated with CVB3 infection. 4. Determine whether conditional expression of Coxsackieviral protease 2A in the cardiac myocyte is sufficient to induce cardiomyopathy and disruption of the sarcolemma via cleavage of dystrophin.