Regulation and function of cGMP dependent protein kinase in cardiac hypertrophy

描述(由申请人提供)：心脏肥厚重构是心脏病发病率和死亡率的重要组成部分。鉴于随之而来的高血压和肥厚的高患病率，它影响着世界上近10%的人口。环鸟苷-3‘，5’-单磷酸(CGMP)是细胞内控制多种细胞生理的第二信使。越来越多的证据表明，一氧化氮或利钠肽促进cGMP的合成对心肌肥厚的发生具有负性调节作用。我们最近发现，用磷酸二酯酶5A(PDE5A)抑制剂阻断cGMP的降解具有明显的抗肥厚和抗重构作用。CGMP的一个主要下游调节因子是cGMP依赖的蛋白激酶(CGK)。目前尚不清楚cGMP及其下游CGK如何调节细胞内信号网络，从而对心脏产生这样的生理影响。PDE5A抑制剂目前被广泛用于治疗勃起功能障碍和肺动脉高压。来自美国和其他支持PDE5A抑制剂增强cGMP心脏益处的报告已经引起了人们对这些药物在治疗人类心脏病方面的浓厚兴趣。随着美国国立卫生研究院预计将在心力衰竭患者中启动PDE5A抑制剂的临床试验，澄清这一信号的潜在机制变得更加重要。我们的初步数据表明，cGMP激活的CGK I1(CGKI1)针对两种不同的分子，对应激或疾病心脏发挥有利作用，这取决于它在心脏重构不同阶段的细胞内位置。在肥厚发育的早期，cGKI1通过激活其抑制蛋白G蛋白偶联信号调节因子2(RGS2)来钝化GQ信号，而在肥厚发育的晚期(失败)，它上调PGC-11(过氧化体增殖物-激活受体3共激活因子1-1)，PGC-11是线粒体生物发生和功能的主要调节因子，可能在细胞核内。本研究旨在通过利用缺乏RGS2(常规的和可诱导的条件)的动物和含有突变的cGKI1的动物(其中关键的蛋白质-蛋白质相互作用位点(亮氨酸拉链基序)被破坏)，从两个目的提供关于这一调控机制的关键信息。第一个目的是研究GKI1在早期肥厚发育阶段的心脏保护机制。第二个目的是研究cGKI1对PGC-11的调节作用和机制，以及它在心力衰竭晚期对线粒体生物发生/功能的影响。这些研究的成功完成将极大地扩大我们对cGMP信号在疾病心脏中的作用的理解，并为测试cGMP信号增强药物(如PDE5A抑制剂)治疗心脏病的临床试验提供参考。与公共卫生相关：世界上近10%的人口患有心肌肥大，这增加了他们患心脏病和猝死的风险。我们发现，目前广泛用于治疗勃起功能障碍的药物西地那非(Viagra)可以通过增强固有的cGMP途径来改善心肌肥厚/重构。该项目将阐明心脏中增强的cGMP信号通路对这种有益影响的潜在机制，重点是一个主要的下游分子及其与其他关键调节因子的相互作用。

DESCRIPTION (provided by applicant): Cardiac hypertrophic remodeling underlies a large component of the morbidity and mortality of heart disease. It affects nearly 10% of the world's population given the high prevalence of hypertension and hypertrophy that evolves with it. Cyclic guanosine-3', 5'-monophosphate (cGMP) is an intracellular second messenger which controls diverse cell physiology. Accumulating evidence shows that enhanced synthesis of cGMP by nitric oxide or natriuretic peptides negatively regulate cardiac hypertrophy development. We recently found that blocking cGMP degradation with phospho- diesterase5A (PDE5A) inhibitor have potent anti-hypertrophic and anti-remodeling effects. One major downstream mediator of cGMP is cGMP-dependent protein kinase (cGK). It remains unclear how cGMP and its downstream cGK regulate intra-cellular signaling network, resulting in such physiological impact on the heart. PDE5A inhibitors are currently widely used to treat erectile dysfunction and pulmonary hypertension. Reports from us and others supporting cardiac benefits from cGMP enhancement by PDE5A inhibitors have raised substantial interest for these drugs in treatment of human heart disease. To clarify the underlying mechanism of this signaling takes on more prominence now that the NIH is expected to initiate a clinical trial of PDE5A inhibitor in heart failure patients. Our preliminary data suggest that cGMP- activated cGK type I1 (cGKI1) targets two different molecules to exert beneficial effects over stressed or diseased heart, depending on its intracellular location at different stages of cardiac remodeling. cGKI1 blunts Gq signaling by activating its inhibitory protein named regulator of G-protein coupled signaling 2 (RGS2) at the sarcolemmal membrane in the early phase of hypertrophy development, whereas in the late phase (failure), it up-regulates PGC-11 (peroxisome proliferators-activated receptor 3 coactivator 1-1), a master regulator of mitochondrial biogenesis and function possibly in the nucleus. The research in this proposal aims to provide critical information on the mechanisms of this regulation in two aims by utilizing animals deficient in RGS2 (conventional and inducible conditional) and animals harboring mutant cGKI1 in which critical protein- protein interaction site (leucine zipper motif) is disrupted. The first aim will examine the cardio- protective mechanism of GKI1 in the early hypertrophy development phase. The second aim will examine the role and the mechanism of PGC-11 regulation by cGKI1, and its impact on mitochondrial biogenesis/ function in the late phase of cardiac failure. The successful completion of these studies will greatly expand our understanding of the role for cGMP signaling in diseased hearts, and inform clinical trials testing cGMP signaling-enhancing drugs such as PDE5A inhibitors for treating heart disease. PUBLIC HEALTH RELEVANCE: Nearly 10% of the world's population develops cardiac hypertrophy which increases their risk of suffering from heart disease, and sudden death. We discovered that cardiac hypertrophy/ remodeling is ameliorated by enhancing intrinsic cGMP pathway with sildenafil (Viagra), the drug currently widely used for treating erectile dysfunction. This project will clarify the underlying mechanisms for this beneficial impact from enhanced cGMP signaling pathway in the heart, focusing on a major downstream molecule and its interaction with other key regulators.