Determining the role of Rac1 palmitoylation in cardiac hypertrophy and oxidative stress

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PROJECT SUMMARY Cardiovascular disease, including heart failure, is the leading cause of mortality in the USA and new therapeutics are needed to better treat these diseases. Cardiac hypertrophy is the process by which the heart becomes larger in response to an increased workload and although initially adaptive, this often leads to maladaptive cardiac remodeling and is involved in the pathogenesis of several cardiac diseases. Rac1, a Rho family small GTPase, is required for the development of cardiac hypertrophy and relatedly plays an important role in mediating oxidative stress in the heart as a regulatory subunit of the NAPDH oxidase 2 (Nox2) enzyme complex. Both hypertrophy and oxidative stress promote the progression of heart failure and therefore determining novel regulatory mechanisms controlling these processes is of great interest to the pharmaceutical industry. Rac1 is palmitoylated at Cys-178 which promotes its activation and membrane targeting. Palmitoylation is a reversible post-translational lipid modification that dynamically regulates protein signaling and has been shown to be required for pathologic signaling in several disease states. Importantly, both genetic and small molecule strategies have been effectively utilized to target palmitoylation of specific proteins in pre-clinical models of diseases such as cancer and inflammation. However, the role of palmitoylation in regulating pathologic signaling in the heart remains untested. Excitingly, our preliminary data demonstrate that genetic inhibition of Rac1 palmitoylation protects neonatal rat cardiomyocytes, a common in vitro model of cardiac disease, from both hypertrophy and oxidative stress induced by hyperactive Rac1 signaling. Therefore, we hypothesize that palmitoylation of Rac1 at Cys-178 targets Rac1 to the membrane where it is activated and induces pathogenic signaling in part through regulation of the Nox2 complex. This proposal seeks to determine the function of Rac1 palmitoylation in regulating cardiac hypertrophy and oxidative stress in vivo using two relevant models of cardiac hypertrophy. We will use a chronic AngII infusion model of cardiac hypertrophy as well as a model using AAV- mediated overexpression of constitutively active Rac1, which causes dilated cardiomyopathy. In these models, we will test the effect of mutating Cys-178 to serine (C178S) which cannot be palmitoylated to rigorously determine the necessity of palmitoylation for the development of cardiac hypertrophy and myocardial oxidative stress. The results of this work will test, for the first time, a role for palmitoylation-dependent signaling in regulating the progression to heart failure. This fellowship will provide me with the training necessary to succeed as an independent scientist in the pharmaceutical industry as well as make significant contributions to our shared scientific knowledge that have the potential to improve human health.