PKG Redox Modulation of Cardiac Function and Disease

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): In the hypertrophied or failing heart, signaling cascades that both induce and ameliorate maladaptive changes in cell and organ function are co-activated. Current clinical therapy largely aims at blocking pathological signaling, but efforts to enhance intrinsic adaptive pathways are also gaining interest. The cyclic GMP-protein kinase G1a (PKG1a) pathway is a prime example of the latter. Recent studies, many from my laboratory, have revealed how inhibiting phosphodiesterase type 5 to stimulate PKG blocks the progression and even reverses established heart disease. This has led to two major NIH- sponsored multicenter trials of PDE5 inhibitors in heart failure with either reduced or preserved ejection fraction. At the core of this translation is the functionality of PKG1a in the myocardium, and in new data we show for the first time that the enzyme is oxidized in diseased left ventricles, reducing its capacity to offset pathology. This involves a di-sulfide between cysteine-42 (C42) in each PKG1a monomer, a residue lying just within an N-terminus docking domain critical for protein interactions. Mice with a knock-in mutation (C42S) that precludes C42 oxidation show improved pathophysiology to sustained pressure-overload. However, they also show that PKG can no longer be activated by PDE5-inhibition. This indicates that therapies leveraging PKG activation, such as natriuretic peptides, nitric oxide donors, or PDE5-inhibitors, may themselves critically depend on PKG redox state. In this project, we determine the impact of PKG1a C42 oxidation/dimerization on the diseased heart, testing the hypothesis that redox modification alters its protein-interactome and thus kinase signaling, as well as activation by intracellular cGMP pools. This is accomplished in three Aims. In aim 1, we determine how prevention of PKG1a C42-dimer impacts hearts subjected to pressure overload or infarction versus physiological (exercise) stress, identifying differences in kinase targeting and patho- physiological regulation. We also test if PKG1a-redox impacts the capacity of clinical methods to stimulate the kinase, and identify mechanisms for such changes. In Aim 2, we assess how PKG1a- redox impacts kinase modification of myocyte sarcomere function, identifying changes in the myofibrillar enriched and depleted phospho-proteome. In Aim 3, we examine the impact of PKG1a- redox on intracellular kinase localization and protein interactions. The interactome is identified, and mechanisms by which the C42-dimer impacts the protein docking-domain determined. Collectively, these studies will determine the role, mechanisms, and translational implications for PKG redox modulation, information central to optimizing therapies designed to leverage its signaling for the treatment of treat heart disease.