The role of histone demethylase KDM5B in ethanol-induced microglial activation

组蛋白去甲基化酶 KDM5B 在乙醇诱导的小胶质细胞激活中的作用

• 批准号: 10247833
• 负责人: Stanley M Stevens
• 金额: $ 34.11万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-10 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10247833
• 关键词: Acute; Affect; Alcohol abuse; Alcoholism; Alcohols; Amino Acids; Bioinformatics; Biological; Brain; Brain Injuries; Carbon; Cell Culture Techniques; Cells; ChIP-seq; Chromatin; Chromatin Structure; Chronic; Classification; Coculture Techniques; Code; Data; Development; Dose; Drug Targeting; Epigenetic Process; Ethanol; Excision; Exhibits; Functional disorder; Future; Genetic; Genetic Transcription; Histone H3; Histones; Immune; Immunoprecipitation; In Vitro; Inflammatory; Inflammatory Response; KDM5B gene; Mass Spectrum Analysis; Mediating; Metabolism; Methylation; Methyltransferase; Microglia; Modification; Mus; Neuraxis; Neurons; Outcome; Pathway interactions; Pharmacology; Phenotype; Process; Proteins; Proteome; Proteomics; Publishing; Role; Site; Small Interfering RNA; Stable Isotope Labeling; Structure; Testing; Time; Validation; activity-based protein profiling; alcohol exposure; base; brain cell; chromatin immunoprecipitation; epigenetic therapy; glial activation; histone demethylase; histone methylation; histone methyltransferase; improved; in vitro activity; in vivo; innovation; knock-down; molecular dynamics; multidisciplinary; neuroinflammation; next generation sequencing; novel; overexpression; response; small molecule inhibitor; treatment strategy; validation studies

Ethanol has profound effects on the central nervous system (CNS) including pathophysiological sequelae resulting from glial cell activation. Microglia, as the resident immune cells of the brain, have been implicated in neuroinflammatory processes that occur from chronic ethanol exposure. Emerging evidence now suggests, however, that microglia can exhibit activation phenotypes other than a pro-inflammatory state depending on dose and time of ethanol exposure. Based on recent proteomic analyses of ethanol-treated microglia, we demonstrate that a significant portion of the ethanol-induced proteome response in microglia can be attributed to changes in the activity of KDM5B, a histone demethylase that catalyzes the removal of tri-methylation on Lys 4 of histone H3 (H3K4me3). Moreover, we have strong preliminary data that show ethanol induces histone methylation changes both in vitro and in vivo and that experimental modulation of KDM5B activity affects histone methylation status and subsequent pro-inflammatory response of microglia. Therefore, we hypothesize that (a) methylation of H3K4me3 and its potential impact on the overall histone methylation code is an important epigenetic mechanism that influences ethanol-induced activation of microglia and (b) changes in KDM5B-mediated histone methylation promotes the exposure time-dependent transition of microglia to a pro- inflammatory phenotype. In order to test our hypothesis, we will 1) determine ethanol-induced changes in the histone methylation code and related impact on microglial activation phenotype upon genetic and pharmacological modulation of KDM5B activity in vitro and 2) characterize the ethanol dose- and time- dependent role of KDM5B on chromatin structural and functional changes related to microglial activation phenotype in vivo. In Aim 1, unbiased, mass spectrometry-based approaches will be utilized in order to determine the impact of KDM5B activity on the histone methylation code and on the activity of epigenetic writers such as methyltransferases. Additionally, we will accurately define activation phenotype driven by histone methylation through a novel bioinformatics approach developed by our lab. In Aim 2, we will employ mass spectrometry, ChIP-Seq and computational approaches to determine chromatin functional and structural consequences related to ethanol-induced histone methylation changes in vivo. This project will be the first of its kind to accurately classify activation phenotype of microglia through novel proteomics and bioinformatics- based approaches in order to better understand microglial functional changes that occur during chronic ethanol exposure. Moreover, this project will clarify the role of the histone demethylase, KDM5B, in histone methylation changes that regulate ethanol-induced microglial phenotype, potentially allowing for the development of novel epigenetic therapies for the treatment of CNS dysfunction resulting from alcohol abuse.

乙醇对中枢神经系统（CNS）有深远的影响，包括病理生理学后遗症 由神经胶质细胞激活引起。小胶质细胞作为大脑的常驻免疫细胞， 慢性乙醇暴露引起的神经炎症过程。现在新出现的证据表明， 然而，小神经胶质细胞可以表现出除促炎状态以外活化表型，这取决于 乙醇暴露的剂量和时间。基于最近对乙醇处理的小胶质细胞的蛋白质组学分析，我们 表明乙醇诱导的小胶质细胞蛋白质组反应的重要部分可以归因于 KDM 5 B是一种组蛋白去甲基化酶，催化细胞表面三甲基化的去除， 组蛋白H3的赖氨酸4（H3 K4 me 3）。此外，我们有强有力的初步数据表明，乙醇诱导组蛋白 甲基化在体外和体内都发生了变化，KDM 5 B活性的实验性调节影响了 组蛋白甲基化状态和随后小胶质细胞促炎反应。因此，我们假设 （a）H3 K4 me 3的甲基化及其对整个组蛋白甲基化密码的潜在影响是一个重要的因素。 影响乙醇诱导的小胶质细胞激活的重要表观遗传机制和（B）小胶质细胞的变化 KDM 5 B介导的组蛋白甲基化促进暴露时间依赖性的小胶质细胞向前- 炎症表型为了检验我们的假设，我们将1）确定乙醇诱导的变化， 组蛋白甲基化密码及其对遗传和小胶质细胞活化表型的相关影响 体外KDM 5 B活性的药理学调节和2）表征乙醇剂量和时间 KDM 5 B对小胶质细胞活化相关染色质结构和功能变化的依赖性作用 体内表型。在目标1中，将利用无偏的、基于质谱的方法， 确定KDM 5 B活性对组蛋白甲基化密码和表观遗传修饰活性的影响。 例如甲基转移酶。此外，我们将准确地定义由以下因素驱动的激活表型： 通过我们实验室开发的一种新的生物信息学方法进行组蛋白甲基化。在目标2中，我们将采用 质谱、ChIP-Seq和计算方法来确定染色质的功能和结构 结果与乙醇诱导的组蛋白甲基化变化在体内。该项目将是其第一个 一种通过新的蛋白质组学和生物信息学准确分类小胶质细胞活化表型的方法- 为了更好地了解慢性乙醇中毒过程中发生的小胶质细胞功能变化， exposure.此外，本项目将阐明组蛋白去甲基化酶KDM 5 B在组蛋白甲基化中的作用 调节乙醇诱导的小胶质细胞表型的变化，可能允许开发新的 表观遗传疗法用于治疗由酒精滥用引起的CNS功能障碍。

项目成果

Methyl-Induced Polarization Destabilizes the Noncovalent Interactions of N-Methylated Lysines.

• DOI: 10.1002/chem.202100644
• 发表时间: 2021-07-26
• 期刊: CHEMISTRY-A EUROPEAN JOURNAL
• 影响因子: 4.3
• 作者: Rahman, Sanim;Wineman-Fisher, Vered;Nagy, Peter R.;Al-Hamdani, Yasmine;Tkatchenko, Alexandre;Varma, Sameer
• 通讯作者: Varma, Sameer

Enhancement of Proteome Coverage by Ion Mobility Fractionation Coupled to PASEF on a TIMS-QTOF Instrument.

• DOI: 10.1021/acs.jproteome.2c00336
• 发表时间: 2022-08-05
• 期刊: JOURNAL OF PROTEOME RESEARCH
• 影响因子: 4.4
• 作者: Guergues, Jennifer;Wohlfahrt, Jessica;Stevens, Stanley M., Jr.
• 通讯作者: Stevens, Stanley M., Jr.

Predictive QM/MM Modeling of Modulations in Protein-Protein Binding by Lysine Methylation.

• DOI: 10.1016/j.jmb.2020.166745
• 发表时间: 2021-02-05
• 期刊: JOURNAL OF MOLECULAR BIOLOGY
• 影响因子: 5.6
• 作者: Rahman, Sanim;Wineman-Fisher, Vered;Al-Hamdani, Yasmine;Tkatchenko, Alexandre;Varma, Sameer
• 通讯作者: Varma, Sameer