Impact of ethanol-induced protein nitration on the histone modification code

乙醇诱导的蛋白质硝化对组蛋白修饰代码的影响

• 批准号: 8805809
• 负责人: Stanley M Stevens
• 金额: $ 20.55万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-02-15 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8805809
• 关键词: Acetylation; Active Sites; Acute; Affect; Alcohol consumption; Alcoholic Liver Diseases; Alcohols; Animal Model; Bioinformatics; Biological Assay; Cardiovascular Diseases; Cell Culture Techniques; Cells; Charge; Chronic; Code; Custom; Data; Dependency; Development; Disease; Disease Outcome; Dose; Drug Targeting; Environment; Enzymes; Epigenetic Process; Ethanol; Etiology; Exposure to; Gene Expression Process; Generations; Genetic Transcription; Goals; Health; Hepatic; Hepatitis; Hepatocyte; Histone Deacetylase; Histone H2A; Histone H3; Histones; Human; In Vitro; Incubators; Injury; Laboratories; Lead; Light; Link; Liver; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Modeling; Modification; Molecular; Molecular Weight; Mus; Neurodegenerative Disorders; Nitrates; Nitric Oxide; Nuclear; Nuclear Protein; Nuclear Proteins; Oxidants; Oxidative Stress; Peptides; Peroxonitrite; Pharmacotherapy; Phosphorylation; Phosphotyrosine; Post-Translational Protein Processing; Prevention; Process; Protein Isoforms; Proteins; Proteomics; Publishing; Rattus; Relative (related person); Research; Role; Severities; Site; Specificity; Stable Isotope Labeling; Stress; System; Testing; Time; Tissues; Transcription Process; Tyrosine; Tyrosine Phosphorylation; Uric Acid; Validation; Western Blotting; alcohol exposure; base; cell type; epigenetic marker; histone modification; human NOS2A protein; in vivo; inhibitor/antagonist; link protein; liver injury; mortality; multicatalytic endopeptidase complex; multiple reaction monitoring; nitration; novel; tool

DESCRIPTION (provided by applicant): Increase in protein tyrosine nitration (PTN) has been associated with numerous diseases and mounting evidence suggests this modification could be a causative factor rather than just a general outcome of disease. The generation of PTN has been implicated in early alcohol-induced hepatitis in mice where inducible nitric oxide synthase (iNOS) and the nitric oxide-derived product, peroxynitrite (a potent agent that causes PTN in the cell), have been identified as major players in the development of this pathophysiological condition of the liver. In an attempt to identify potential hepatic PTN targets, preliminary proteomic studies were performed in our laboratory and the results suggest that histone proteins undergo selective nitration in primary human hepatocyte culture after acute ethanol exposure. Accordingly, the goal of this R21 project is to examine the impact of ethanol-induced PTN on the histone modification code by testing the hypotheses that 1) protein nitration serves as a novel histone modification that could result in subsequent alteration of gene transcription processes and 2) oxidative/nitrative modifications of enzymes that directly modulate histone modification levels could have an impact on various ethanol-induced epigenetic markers (for example, acetylation of histone H3 at Lys9). In Specific Aim 1 of this proposal, we will characterize the process of ethanol-induced tyrosine nitration in histone H2A, H2B, H3, and H4 isoforms. In Specific Aim 2, we will characterize the nuclear nitroproteome after ethanol exposure to identify nitrated proteins associated with altering the histone modification code (for example, HAT or HDAC enzymes). Additionally, our findings from in vitro studies will be validated in an in vivo animal model of chronic ethanol exposure. The results from this project will help shed light on the role of oxidative/nitrative stress in ethanol-induced alterations of the histone modification code and provide further evidence to the link of PTN and disease causation. Identification of nuclear nitration targets and enzyme processes that affect PTN levels will likely provide new information on potential targets for drug therapy in ethanol-induced tissue injury where PTN is linked to injury onset or severity.

描述（由申请人提供）：蛋白质酪氨酸硝化（PTN）的增加与多种疾病有关，并且越来越多的证据表明这种修饰可能是导致因素，而不仅仅是疾病的一般结果。 PTN的产生与早期酒精诱导的肝炎有关小鼠的早期肝炎，其中可诱导的一氧化氮合酶（INOS）和一氧化氧化物衍生的产物（一种导致细胞中PTN的有效药物）被确定为这种病理学条件的主要参与者。为了鉴定潜在的肝PTN靶标，在我们的实验室中进行了初步蛋白质组学研究，结果表明，急性乙醇暴露后，组蛋白蛋白在原发性人肝细胞培养中经历选择性硝化。因此，该R21项目的目的是通过测试1）蛋白硝化的假设来检查乙醇诱导的PTN对组蛋白修饰代码的影响，作为一种新型组蛋白的修饰，可能导致基因转录的随后变化，并直接对氧化含量的改性进行改性 - 可能对enzy含量的改性 - 可能对enzy的含量进行改性 - 可能会导致氧化含量的含量，以使氧化含量适用于enzy级别，以实现氧化含量的含量。表观遗传标记（例如，在lys9处组蛋白H3的乙酰化）。在该提案的特定目的1中，我们将表征组蛋白H2A，H2B，H3和H4同工型中乙醇诱导的酪氨酸硝化的过程。在特定的目标2中，我们将表征乙醇暴露后的核硝化蛋白体，以鉴定与改变组蛋白修饰代码相关的硝化蛋白（例如，HAT或HDAC酶）。此外，我们从体外研究中的发现将在慢性乙醇暴露的体内动物模型中得到验证。该项目的结果将有助于阐明氧化/硝化应激在乙醇诱导的组蛋白修饰代码改变中的作用，并为PTN和疾病因果关系的联系提供进一步的证据。鉴定影响PTN水平的核硝化靶标和酶过程可能会提供有关乙醇诱导的组织损伤药物治疗靶标的新信息，而PTN与损伤发作或严重程度有关。

项目成果

Evaluation of a method for nitrotyrosine site identification and relative quantitation using a stable isotope-labeled nitrated spike-in standard and high resolution fourier transform MS and MS/MS analysis.

使用稳定同位素标记的硝化掺入标准品和高分辨率傅立叶变换 MS 和 MS/MS 分析来评估硝基酪氨酸位点鉴定和相对定量的方法。

• DOI: 10.3390/ijms15046265
• 发表时间: 2014
• 期刊: International journal of molecular sciences
• 影响因子: 5.6
• 作者: Seeley,KentW;Fertig,AlisonR;Dufresne,CraigP;Pinho,JoaoPC;StevensJr,StanleyM
• 通讯作者: StevensJr,StanleyM

Site-specific identification and validation of hepatic histone nitration in vivo: Implications for alcohol-induced liver injury.

体内肝组蛋白硝化的位点特异性鉴定和验证：对酒精性肝损伤的影响。

• DOI: 10.1002/jms.4713
• 发表时间: 2021
• 期刊: Journal of mass spectrometry : JMS
• 作者: Kriss,CrystinaL;Duro,Nalvi;Nadeau,OwenW;Guergues,Jennifer;Chavez-Chiang,Omar;Culver-Cochran,AshleyE;Chaput,Dale;Varma,Sameer;StevensJr,StanleyM
• 通讯作者: StevensJr,StanleyM