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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 信号传导

基本信息

批准号：
10580267
负责人：
ANNIE V BEUVE
金额：
$ 8.66万
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 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10580267
关键词：
Actins Adenoviruses Administrative Supplement Affect Angiotensin II Apoptosis Binding Biochemical Bioinformatics Biological Assay Blood Vessels CRISPR/Cas technology Calcium Cardiac Cardiovascular system Cell Line Cell physiology Cells Co-Immunoprecipitations Complex Consensus Cyclic GMP Cysteine Environment Equilibrium Equipment Functional disorder Funding Guanosine Triphosphate Heart Hypertrophy Heart failure Homeostasis Hypertension Impairment Investigation Knock-in Knock-in Mouse Lead Ligation Mass Spectrum Analysis Measures Mediator of activation protein Metabolic Pathway Modeling Modification Mus Mutation Analysis Nitric Oxide Nitrosation OLFM4 gene Oxidation-Reduction Oxidative Stress Oxides Pathologic Pathway interactions Peptides Physiological Post-Translational Protein Processing Property Proteins Proteomics Regulation Role SKIL gene Signal Pathway Signal Transduction Signaling Molecule Site Skeleton Smooth Muscle Soluble Guanylate Cyclase Specificity Stress Sulfhydryl Compounds System TXN gene Technology Vasodilation Wild Type Mouse angiogenesis blood pressure regulation cGMP production cardioprotection desensitization heme a improved in vivo intravital microscopy mouse model novel oxidation parent grant polymerization pressure protective effect protein function protein protein interaction response

项目摘要

PROJECT SUMMARY (PARENT GRANT) This is an application for an administrative supplemental equipment under our parent grant GM112415. The equipment we are requesting is at the center of our experimental strategy. This equipment is a Pressure Myograph P114 system that will replace and upgrade an intravital microscopy system that is not anymore available to us. We will not be able to complete Aim3 without this type of equipment and because of the upgraded technology we will improve and expand the investigations of Aim1 and Aim2 of the project of our parent grant summarized below: Nitric oxide (NO) is an important signaling molecule that regulates diverse functions relevant to vascular function, apoptosis and angiogenesis. NO is best known for its ability to stimulate soluble guanylyl cyclase (now called GC1) to produce cGMP and stimulate its downstream signaling pathways. However, NO can also covalently modify cysteines (Cys) via S-nitrosation or S-nitrosylation (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, dynamic regulation of protein nitrosation specificity is poorly understood. Our most recent investigations reveal that GC1 has a transnitrosylase activity, i.e. GC1 has the ability to directly transfer SNO to specific targets by protein-protein interaction (transnitrosation). This transnitrosation activity does not require the cGMP forming activity of GC1 and can be accomplished by a single subunit of GC1 (formation of cGMP requires 2 subunits). Furthermore, we showed that one transnitrosation target of GC1 is oxidized thioredoxin 1 (oTrx1), a thiol-redox protein that modulates cellular S-nitrosation. In fact, oxidative/nitrosative conditions appear to favor the GC1-Trx1 complex. Using advanced proteomics approaches, we recently identified the Cys in GC1 and Trx1 that are involved in the SNO transfer in a purified system, and the Cys of proteins targeted by the GC1/Trx1 transnitrosation cascade in smooth muscle and cardiac cells. Our hypothesis is that the function of GC1 transnitrosation activity is an adaptive response to oxidative stress and potentially compensates for the dysfunction of the canonical NO-GC1-cGMP pathway that occurs in oxidative conditions. To explore this provocative hypothesis, we propose to conduct mutational analysis of the Cys we have identified to characterize the mechanism of transnitrosation in smooth muscle and cardiac cells. By comparing the targets of GC1, Trx1 and both we will determine the mechanisms underlying target specificity. We will determine how GC1/Trx1 transnitrosation of specific targets affects their cellular function. For this, we will use cell lines and primary cells isolated from a novel mouse knock-in (KI) of a Cys of GC1 involved in transnitrosation. To determine the physiological relevance of GC1- and GC1/Trx1-transnitrosation in the cardiovascular system and the adaptive response to stress, we will use the Cys KI mouse model and inhibitory peptides that disrupt the GC1/Trx1 transnitrosating complex under Angiotensin II-induced oxidative stress. This project could lead to the discovery of novel cardiovascular protective pathways driven by specific S-nitrosation.

项目摘要（家长资助）