S-Nitrosylation and Cardioprotection

S-亚硝基化和心脏保护

基本信息

批准号：
9000222
负责人：
Mark Jeffrey Kohr
金额：
$ 24.9万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-08-23 至 2018-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9000222
关键词：
Address Affect Binding Cardiac Myocytes Cause of Death Cell Nucleus Cysteine Cytosol Data Fellowship Goals Healthcare Heart In Situ Intervention Ion Channel Knowledge Lead Mediating Methods Mitochondria Mitochondrial Matrix Modeling Modification Molecular Morbidity - disease rate Mutagenesis Myocardial Myocardial Ischemia Myocardial dysfunction Myocardium Nitric Oxide Nitric Oxide Synthase Nitric Oxide Synthase Type I Pathologic Physiological Play Postdoctoral Fellow Production Protein Isoforms Protein S Protein translocation Proteins Regulation Relative (related person)Reperfusion Injury Research Role Signal Pathway Signal Transduction Site Source Specificity Stimulus Sulfhydryl Compounds Testing Therapeutic Translating United States base extracellular human NOS3 protein insight mortality mutant novel oxidation protective effect protein function protein protein interaction research study therapeutic target

项目摘要

PROJECT SUMMARY/ABSTRACT Ischemic heart disease is one of the most common causes of death in the United States and is responsible for a tremendous healthcare burden. Cardioprotective interventions hold great promise for lessening this burden, but few experimental discoveries have translated successfully into effective therapeutics. This is likely due to a lack of knowledge with regard to the mechanistic details of cardioprotection. Nitric oxide, either produced endogenously or administered exogenously, has been shown to be an important component of cardioprotection. Our recent studies have demonstrated an association between increased levels of nitric oxide-derived protein S-nitrosylation and cardioprotection. S-nitrosylation is a reversible, thiol-based modification that is produced from the covalent attachment of a nitric oxide moiety to the free thiol group of a cysteine residue. The nitric oxide synthase isoforms represent the major source of endogenous nitric oxide production in the cardiac myocyte. S-nitrosylation is thought to provide protective effects by modulating the activity and/or function of target proteins, and by blocking the damaging effects of irreversible cysteine oxidation. Our recent data are consistent with an overall protective role for S-nitrosylation, but the molecular mechanism(s), the relative importance of specific protein targets, and the pathophysiological significance of S- nitrosylation is unknown. Thus, the goal of this proposal is to define the specificity and mechanistic consequences of protein S-nitrosylation in the myocardium by investigating the physiologic and pathologic aspects of S-nitrosylation. To establish the mechanistic details of S-nitrosylation, a novel method to determine the percentage of a given protein that is modified by S-nitrosylation (i.e., S-nitrosylation occupancy) is being developed and this will be used to examine compartmentalized S-nitrosylation signaling. Cysteine mutagenesis will also be used to examine the effects of S-nitrosylation on protein function. We previously identified many S- nitrosylation sites in cardioprotection, thus allowing us to focus on specific cysteine residues that are altered in situ. The specific aims of this proposal are as follows: 1) determine if S-nitrosylation occupancy varies between different cellular compartments of the cardiomyocyte and evaluate how this affects signaling, 2) determine if S- nitrosylation changes protein-protein interaction, alters protein localization, and promotes protein trans-S- nitrosylation, and 3) determine the specific role of S-nitrosylation in the regulation of myocardial ion channel activity. The results of this proposal will define the role of S-nitrosylation in the heart and advance the field by establishing a mechanistic role for S-nitrosylation, thus lending critical insight into potential therapeutic targets.

项目概要/摘要缺血性心脏病是美国最常见的死亡原因之一，导致巨大的医疗负担。心脏保护干预措施有望减轻这种负担，但很少有实验发现能够成功转化为有效的治疗方法。这可能是由于缺乏对心脏保护机制细节的了解。一氧化氮，无论是产生已被证明是内源性或外源性给药的重要组成部分心脏保护。我们最近的研究表明，硝酸盐水平升高与氧化物衍生蛋白S-亚硝基化和心脏保护。 S-亚硝基化是一种可逆的、基于硫醇的由一氧化氮部分与游离硫醇基团共价连接而产生的修饰半胱氨酸残基。一氧化氮合酶异构体代表内源性一氧化氮的主要来源心肌细胞中的产生。 S-亚硝基化被认为通过调节靶蛋白的活性和/或功能，并通过阻断不可逆半胱氨酸的破坏作用氧化。我们最近的数据与 S-亚硝基化的总体保护作用一致，但分子机制、特定蛋白质靶标的相对重要性以及 S-的病理生理学意义亚硝基化未知。因此，该提案的目标是定义特异性和机制通过研究心肌中蛋白质 S-亚硝基化的后果 S-亚硝基化的方面。建立 S-亚硝基化的机制细节，这是一种确定 S-亚硝基化的新方法通过 S-亚硝基化修饰的给定蛋白质的百分比（即 S-亚硝基化占用率）正在计算开发出来，这将用于检查区室化的 S-亚硝基化信号传导。半胱氨酸诱变还将用于检查 S-亚硝基化对蛋白质功能的影响。我们之前发现了许多S- 心脏保护中的亚硝基化位点，因此使我们能够关注在心脏保护中发生改变的特定半胱氨酸残基现场。该提案的具体目标如下： 1) 确定 S-亚硝基化占据率是否存在变化心肌细胞的不同细胞区室并评估其如何影响信号传导，2) 确定 S- 亚硝基化改变蛋白质-蛋白质相互作用，改变蛋白质定位，并促进蛋白质反式-S- 亚硝基化，3)确定S-亚硝基化在调节心肌离子通道中的具体作用活动。该提案的结果将定义 S-亚硝基化在心脏中的作用，并通过以下方式推进该领域的发展：确定 S-亚硝基化的机制作用，从而为潜在的治疗靶点提供重要的见解。