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.

乙醇对包括病理生理后遗症的中枢神经系统（CNS）具有深远的影响 由神经胶质细胞激活产生。小胶质细胞作为大脑的驻留免疫细胞已与 慢性乙醇暴露发生的神经炎症过程。现在有新兴的证据表明， 但是，小胶质细胞可以表现出激活表型，而不是促炎性状态 乙醇暴露的剂量和时间。基于乙醇处理的小胶质细胞的最新蛋白质组学分析，我们 证明可以归因于小胶质细胞中乙醇诱导的蛋白质组反应的很大一部分可以归因于 对于KDM5B活性的变化，KDM5B的活性是一种组蛋白脱甲基酶，可催化三甲基化的去除 组蛋白H3的LYS 4（H3K4ME3）。此外，我们有强大的初步数据，显示乙醇诱导组蛋白 甲基化在体外和体内都会改变，并且KDM5B活性的实验调节会影响 小胶质细胞的组蛋白甲基化状态和随后的促炎反应。因此，我们假设 （a）H3K4me3的甲基化及其对整个组蛋白甲基化代码的潜在影响是一种 影响乙醇诱导的小胶质细胞激活的重要表观遗传机制，并且（b） KDM5B介导的组蛋白甲基化促进了小胶质细胞的暴露时间依赖性过渡 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 组蛋白甲基化代码及其对小胶质激活表型对遗传和 在体外KDM5B活性的药理调节和2）表征乙醇剂量和时间 - KDM5B在与小胶质细胞激活有关的染色质结构和功能变化上的依赖性作用 体内表型。在AIM 1中，将利用公正的基于质谱的方法来 确定KDM5B活性对组蛋白甲基化代码和表观遗传学活性的影响 作家，例如甲基转移酶。此外，我们将准确定义由 通过我们实验室开发的新型生物信息学方法，组蛋白甲基化。在AIM 2中，我们将雇用 质谱，芯片序列和计算方法，以确定染色质功能和结构 与乙醇诱导的组蛋白甲基化变化有关的后果。这个项目将是其第一个项目 通过新颖的蛋白质组学和生物信息学来准确地对小胶质细胞的激活表型进行分类 为了更好地了解慢性乙醇期间发生的小胶质细胞功能变化 接触。此外，该项目将阐明组蛋白脱甲基酶KDM5B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，有可能允许新的 酗酒引起的中枢神经系统功能障碍的表观遗传疗法。

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

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

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.