Probing the role of cysteine sulfenylation in cell signaling

探讨半胱氨酸磺酰化在细胞信号传导中的作用

基本信息

批准号：
8342423
负责人：
Kate Suzanne Carroll
金额：
$ 39.83万
依托单位：
SCRIPPS FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-01 至 2016-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8342423
关键词：
Active Sites Address Area Binding Binding Sites Biochemical Biological Biological Markers Cancer Biology Cell physiology Cells Chemicals Chemistry Clinical Medicine Code Complex Cysteine Development Disease Environment Enzymes Epidermal Growth Factor Receptor Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Fostering Functional disorder Genetic Growth Growth Factor Health Human Human Pathology Hydrogen Peroxide Isotopes Kinetics Knowledge Lesion Link Malignant Neoplasms Mass Spectrum Analysis Measures Mediating Methods Mission Modification Molecular Monitor Neurodegenerative Disorders Normal Cell Oxidation-Reduction Oxidative Stress Oxidoreductase Pathology Pathway interactions Patients Phenotype Phosphoric Monoester Hydrolases Phosphotransferases Physiological Physiology Post-Translational Protein Processing Protein Tyrosine Kinase Protein Tyrosine Phosphatase Proteins Proteome Proteomics Public Health Receptor Protein-Tyrosine Kinases Recycling Regulation Reporter Research Role Side Signal Pathway Signal Transduction Source Specificity Stratification Structure Sulfhydryl Compounds System Techniques Technology Testing Therapeutic analytical tool base cell growth disease diagnosis enzyme activity improved inhibitor/antagonist innovation migration mutant novel oxidation protein profiling receptor response therapeutic target tool tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): Protein sulfenylation, the redox-based modification of cysteine thiol side chains by hydrogen peroxide (H2O2), is an important mechanism in signal transduction. Dysregulated protein sulfenylation contributes to a range of human pathologies, including cancer. However, efforts to elucidate the diverse roles of protein sulfenylation in physiology and disease have, to date, suffered from a lack of techniques to probe these modifications in native environments. To address this problem, we have recently introduced a new chemical proteomic strategy to detect changes in protein sulfenylation directly in cells. To date, our preliminary studies have identified several novel intracellular protein targets of H2O2 during growth factor signaling, including the epidermal growth factor receptor (EGFR). Specifically, we have discovered that H2O2 directly modifies a cysteine residue within the ATP-binding site of EGFR, and that oxidation stimulates its tyrosine kinase activity, though the biochemical mechanism for this effect remains to be fully elucidated. In this proposal, we will apply our suite of chemical probes and analytical tools to address four major questions of high significance to the fields of redox signaling, chemical biology, and cancer. Aim 1 of the proposal will define the molecular mechanism by which sulfenylation of EGFR regulates its kinase activity. To identify features that dictate selectivity in H2O2-mediated signaling, we will examine sulfenylation, localization, and enzyme activity of EGFR-targeted protein tyrosine phosphatases (PTPs) in Aim 2. We will evaluate additional targets of intracellular H2O2 generated in response to growth factors using target-based and chemical reporter/proteomic methods in Aim 3. While sulfenylation is a reversible modification in cells, the factors that recycle sulfenylated proteinsto their reduced thiol form (RSH) are largely ill defined. We will test candidate reductases responsible for reversible sulfenylation in Aim 4. The development and application of our chemical tools in cells provides an unprecedented opportunity to elucidate mechanisms that govern sulfenylation of proteins. Given that aberrant sulfenylation of proteins has been linked to aggressive cancer phenotypes and that genetic lesions in H2O2-metabolizing enzymes can contribute to tumorigenesis, defining the mechanisms that control reversible protein sulfenylation is vital for understanding human physiology and disease. We anticipate that these studies will define how sulfenylation of proteins regulates signaling networks that underlie cell growth and identify key enzymes that controls desulfenylation. Ultimately, this will facilitate the identification of new biomarkers and therapeutic targets for cancer, as well as produce methodological advances that expand the scope and utility of proteomic technologies for biological and biomedical discoveries. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because the discovery of cellular mechanisms that regulate physiological protein sulfenylation is ultimately expected to increase our understanding of the pathophysiology associated with oxidative stress and abnormal H2O2-based signal transduction, with translational potential for clinical medicine in the key areas of disease diagnosis, patient stratification, and monitoring efficacy in the new era of redox-based therapeutics. Thus, the proposed research is relevant to the part of NIH's mission that pertains to developing fundamental knowledge that will help to foster fundamental discoveries, innovative research strategies, and their applications as a basis for ultimately protecting and improving health.

描述（由申请人提供）：蛋白质磺酰化，基于氧化氢的基于氧化还原的修饰，通过过氧化氢（H2O2）是信号转导的重要机制。失调的蛋白质磺酰化有助于包括癌症在内的一系列人类病理。然而，迄今为止，缺乏在本机环境中探究这些修饰的技术，阐明蛋白质磺苯二苯基化在生理和疾病中的各种作用的努力。为了解决这个问题，我们最近引入了一种新的化学蛋白质组学策略，以检测细胞中蛋白质磺苯基的变化。迄今为止，我们的初步研究已经确定了生长因子信号传导期间H2O2的几个新型细胞内蛋白靶标，包括表皮生长因子受体（EGFR）。具体而言，我们发现H2O2直接修饰了EGFR ATP结合位点内的半胱氨酸残基，并且氧化刺激其酪氨酸激酶活性，尽管这种作用的生化机制尚待充分阐明。在此提案中，我们将应用我们的化学探针和分析工具套件，以解决对氧化还原信号，化学生物学和癌症领域具有很高意义的四个主要问题。该提案的目标1将定义EGFR磺酰化调节其激酶活性的分子机制。为了确定在H2O2介导的信号传导中决定选择性的功能，我们将检查 sulfenylation, localization, and enzyme activity of EGFR-targeted protein tyrosine phosphatases (PTPs) in Aim 2. We will evaluate additional targets of intracellular H2O2 generated in response to growth factors using target-based and chemical reporter/proteomic methods in Aim 3. While sulfenylation is a reversible modification in cells, the factors that recycle sulfenylated proteinsto their reduced硫醇形式（RSH）在很大程度上定义。我们将测试负责在AIM 4中可逆磺酰化的候选还原酶。我们的化学工具在细胞中的开发和应用提供了前所未有的机会，以阐明控制蛋白质磺酰化的机制。鉴于蛋白质的异常磺酰化与侵袭性癌症表型有关，并且H2O2-替代酶中的遗传病变可以导致肿瘤发生，从而定义控制可逆蛋白质硫苯基化的机制，对理解人类物理学和疾病至关重要。我们预计这些研究将定义蛋白质的磺苯基如何调节细胞生长的信号网络并鉴定控制脱硫硝基化的关键酶。最终，这将有助于鉴定新的生物标志物和癌症治疗靶标，并产生方法论进步，以扩大生物学和生物医学发现的蛋白质组学技术的范围和实用性。公共卫生相关性：拟议的研究与公共卫生有关，因为发现调节生理蛋白质磺酰基化的细胞机制最终会增加我们对与基于H2O2信号的氧化应激和基于H2O2信号转导相关的病理生理学的理解，并在疾病诊断和监测良好的诊断性疗法中，具有良好的临床药物的临床潜力。因此，拟议的研究与NIH使命的一部分有关，该任务与开发基本知识有关，这些知识将有助于促进基本发现，创新的研究策略及其作为最终保护和改善健康的基础。