NO signaling by a Soluble Guanylyl Cyclase-Thioredoxin transnitrosation complex

可溶性鸟苷酸环化酶-硫氧还蛋白转亚硝基复合物的 NO 信号转导

基本信息

批准号：
8894270
负责人：
ANNIE V BEUVE
金额：
$ 41.16万
依托单位：
RBHS-NEW JERSEY MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2019-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8894270
关键词：
Angiotensin II Apoptosis Apoptosis Regulator Apoptotic Binding Biochemical Bioinformatics Biological Biological Assay Cardiac Cardiac Myocytes Cardiovascular Physiology Cardiovascular system Cell Line Cell Nucleus Cell Survival Cells Cessation of life Complex Consensus Sequence Cyclic GMP Cysteine Data Environment Functional disorder Guanosine Triphosphate Heart Heart Hypertrophy Heart failure In Vitro Investigation Lead Mass Spectrum Analysis Mediating Mediator of activation protein Modeling Modification Mus Mutagenesis Myocardium Nitric Oxide Nitrosation Nuclear Translocation Oxidation-Reduction Pathway interactions Play Post-Translational Protein Processing Process Property Proteins Proteome Proteomics Regulation Role SKIL gene Signal Pathway Signal Transduction Signaling Molecule Site Smooth Muscle Myocytes Soluble Guanylate Cyclase Specificity Stress Sulfhydryl Compounds Thioredoxin Transgenic Mice angiogenesis base cGMP production heme a human CLIC4 protein mutant nitrosative stress novel oxidation prevent protein function protein protein interaction public health relevance research study response small hairpin RNA

项目摘要

DESCRIPTION (provided by applicant): Nitric oxide (NO) is an important signaling molecule that regulates diverse functions relevant to cardiovascular function, apoptosis and angiogenesis. NO is best known for its ability to stimulate soluble guanylyl cyclase (sGC) to produce cGMP and stimulate its downstream signaling pathways. However, NO can also covalently modify cysteines via S-nitrosation (addition of a NO moiety to the cysteine of a protein, SNO). Although this reversible post-translational modification is increasingly recognized as an important regulatory mechanism of protein function, and to play a role in cardiac protection, dynamic regulation of protein nitrosation specificity is poorly understood. Our collaborative team has made the exciting observation that sGC, the key NO receptor, modulates the level of nitrosation of specific proteins in cardiomyocytes and smooth muscle cells. Preliminary data showed that sGC increases nitrosation by a protein-protein interaction-driven SNO transfer (transnitrosation). Moreover, this increased nitrosation is due, for a specific subset of proteins, to the association of sGC with thioredoxin 1 (Trx1), a cardiac protective thiol-redox protein with both transnitrosation and denitrosation activities. Initial mass spectrometry and biochemical analyses showed that sGC transnitrosates Trx1, which in turn nitrosates a specific subset of targets, a finding supported by shTrx1 knockdown experiments in cardiomyocytes. These novel observations lead to the provocative idea that sGC modulates S-nitrosation specificity via a transnitrosation cascade that includes an S-nitrosated Trx1 intermediate. This study aims to answer three critical questions based on this hypothesis. Aim1: What is the mechanism of sGC transnitrosation of Trx1? We will identify key cysteines (Cys) responsible for sGC transfer of SNO to Trx1 and for interaction via mutagenesis and biochemical analyses. Aim2: What are the specific targets of the sGC/Trx1 transnitrosation cascade and the mechanisms underlying target specificity? Using novel and highly specific proteomics approaches, we will quantify the SNO-proteomes modulated by the sGC/Trx1 transnitrosation cascade under nitrosative and oxidative conditions and determine consensus sequence motifs among the target proteins. Aim3: Is sGC-mediated transnitrosation an anti-apoptotic mechanism? NO signaling and Trx1 are crucial components of the anti-apoptotic response to stress. Among the sGC/Trx1 transnitrosation targets identified is the chloride intracellular channel 4 (CLIC4), a regulator of apoptosis, whose nuclear translocation is modulated via specific nitrosation. We will determine whether sGC/Trx1 transnitrosation of CLIC4 is an important inhibitory mechanism of angiotensin II-induced apoptosis of cardiomyocytes, underlying the potential role of this newly discovered transnitrosation cascade in cardiac remodeling following heart failure. This multi-PI project could lead to the discovery of novel cardioprotective pathway driven by specific S-nitrosation.

描述（由适用提供）：一氧化氮（NO）是一个重要的信号分子，可调节与心血管功能，凋亡和血管生成相关的分流功能。 NO以其刺激固体Guanylyl Cyclase（SGC）产生CGMP并刺激其下游信号通路的能力而闻名。但是，不能通过S-硝化来共价修饰半胱氨酸（在半胱氨酸中添加无部分的部分，尽管这种可逆的翻译后修饰越来越多地被认为是蛋白质功能的重要调节机制，并且在心脏保护中起作用，并且在心脏保护中起作用，对蛋白质硝化特异性的动态调节使我们的蛋白硝化特异性变得不足。调节心肌细胞和平滑肌细胞中特定蛋白的硝化水平。具有跨硝化和反硝化活性的受保护的硫醇 - 雷多斯蛋白。这些新颖的观察结果导致了一个挑衅的观念，即SGC通过跨硝化级联反应调节S-硝化特异性，该级别包括S-硝化TRX1中间体。这项研究旨在基于这一假设回答三个关键问题。 AIM1：SGC TRX1的SGC转硝化机理是什么？我们将确定负责SNO转移到TRX1的SGC转移以及通过诱变和生化分析的相互作用的关键半胱氨酸（CYS）。 AIM2：SGC/TRX1翻译级联的特定目标是什么以及目标特异性的基础机制？使用新型和高度特定的蛋白质组学方法，我们将在硝化和氧化条件下量化由SGC/TRX1翻译级联反应的SNO蛋白质体，并确定靶蛋白之间的共识序列基序。 AIM3：SGC介导的跨硝化是一种抗凋亡机制吗？没有信号传导和TRX1是对应激反应的抗凋亡反应的关键组成部分。在确定的SGC/TRX1转硝化靶标中，是氯化物内通道4（CLIC4），一种凋亡的调节剂核易位是通过特异性硝化调节的。我们将确定CLIC4的SGC/TRX1跨硝化是否是血管紧张素II诱导的心肌细胞凋亡的重要抑制作用机制，这是这种新发现的转硝化级联反应在心脏失败后心脏重塑中的潜在作用。这个多PI项目可以导致发现由特定S-硝化驱动的新型心脏保护途径。