EPIGENETIC MODIFICATIONS OF GABA NEURONS IN PSYCHOSIS

精神病中 GABA 神经元的表观遗传修饰

基本信息

批准号：
7619304
负责人：
Rajiv Pandit Sharma
金额：
$ 17.72万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-04-01 至 2011-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7619304
关键词：
Antisense Oligonucleotides Binding Binding Proteins Biological Assay Brain Brain-Derived Neurotrophic Factor Cell Line Characteristics Chromatin Chromatin Structure Co-Immunoprecipitations CpG Islands DNA DNA Binding DNA Methylation DNA Sequence DNA-Binding Proteins DNA-Protein Interaction Data Development Disease Dose Down-Regulation Enzymes Epigenetic Process Exons Gene Expression Gene Expression Regulation Genes Genetic Transcription Genome Histone Deacetylase Inhibitor Histones Hypermethylation Interneurons Intervention K-Series Research Career Programs Laboratories Location Measures Membrane Messenger RNA Methionine Methods Methyl-CpG-Binding Protein 2 Methylation Mitotic Modification Molecular Mus Mutation Neocortex Neuron-Specific Enolase Neurons Output Patients Pharmaceutical Preparations Pharmacological Treatment Phenotype Play Proteins Psychotic Disorders Recording of previous events Reporting Repressor Proteins Rett Syndrome Reverse Transcriptase Polymerase Chain Reaction Role Schizophrenia Site Stimulus Training Valproic Acid Variant Western Blotting Work bisulfite chromatin remodeling gamma-Aminobutyric Acid inhibitor/antagonist mRNA Expression molecular pathology novel therapeutics promoter response therapeutic target

项目摘要

DESCRIPTION (provided by applicant): The GABA intemeuron plays a central role in integrating cortical output and is emerging as a major locus of schizophrenia molecular pathology. Reelin (RELN), GAD67 and DNA methylating enzyme (DNMT1) are all expressed in GABA interneurons, and are each abnormally regulated in schizophrenic brain. Further, DNMT1 is a critical component of the epigenetic regulatory cascade, now shown to play an essential role in neuronal gene expression. We have earlier reported on the effects of epigenetic modifiers (DNA methylation, chromatin structure, and methylated-DNA binding proteins) on the levels of RELN and GAD67 expression. This career development award will extend these observations using primary neuronal cultures (PNC) to describe variations in epigenetic modifications of the RELN and GAD67 promoters resulting from pharmacological treatment and membrane depolarization. Supporting the use of PNCs, we and others have observed El4 mouse neuron cultures to be predominantly GABAergic in phenotype (co-expressing RELN, GAD and DNMT1). Our first objective, using these homogenous neuronal cultures, will be to analyze DNA methylation, local chromatin structure and promoter DNA-protein interactions in response to pharmacological treatments with known epigenetic effects, specifically L-methionine, a hypermethylating stimulus, and histone deactylase (HDAC) inhibitors such as valproic acid. Supporting this first objective is our earlier demonstration that L-methionine down regulates RELN and GAD67 mRNA expression as a consequence of epigenetic modifications along their promoters, and this is reversed by the application of valproic acid. Our second objective will be to examine the effects of membrane depolarization on epigenetic modification of depolarization inducible promoters within these GABAergic neurons. We will apply antisense oligonucleotide strategies to down regulate epigenetic repressor proteins such as DNMT1 and MeCP2 to dissect their contribution to depolarization induced gene transcription. Supporting this second objective, we have now shown that antisense induced down regulation of DNMT1 mRNA and protein will result in significant increases in RELN mRNA expression. Also, we have preliminary data indicating robust changes in GAD67 and DNMT1 mRNA in response to depolarization. With this KO1 career development award, the candidate seeks training in focused methods in the molecular study of epigenetic gene regulation. Because epigenetic mechanisms are modifiable by drug interventions, this work has immediate relevance to the development of novel therapeutics.

描述（由申请人提供）：GABA 中间神经元在整合皮质输出中发挥着核心作用，并且正在成为精神分裂症分子病理学的主要基因座。 Reelin (RELN)、GAD67 和 DNA 甲基化酶 (DNMT1) 均在 GABA 中间神经元中表达，并且在精神分裂症患者的大脑中均受到异常调节。此外，DNMT1 是表观遗传调控级联的关键组成部分，现已证明在神经元基因表达中发挥重要作用。我们之前报道过表观遗传修饰剂（DNA 甲基化、染色质结构和甲基化 DNA 结合蛋白）对 RELN 和 GAD67 表达水平的影响。该职业发展奖将使用原代神经元培养物 (PNC) 扩展这些观察结果，以描述药物治疗和膜去极化导致的 RELN 和 GAD67 启动子表观遗传修饰的变化。为了支持 PNC 的使用，我们和其他人观察到 El4 小鼠神经元培养物的表型主要是 GABA 能（共表达 RELN、GAD 和 DNMT1）。我们的首要目标是使用这些同质神经元培养物来分析 DNA 甲基化、局部染色质结构和启动子 DNA-蛋白质相互作用，以响应具有已知表观遗传效应的药物治疗，特别是 L-蛋氨酸（一种超甲基化刺激物）和组蛋白脱乙酰酶 (HDAC) ) 抑制剂，例如丙戊酸。支持第一个目标的是我们之前的证明，即 L-蛋氨酸下调 RELN 和 GAD67 mRNA 表达是其启动子表观遗传修饰的结果，并且通过应用丙戊酸可以逆转这种情况。我们的第二个目标是检查膜去极化对这些 GABA 能神经元内去极化诱导启动子的表观遗传修饰的影响。我们将应用反义寡核苷酸策略来下调表观遗传阻遏蛋白，例如 DNMT1 和 MeCP2，以剖析它们对去极化诱导的基因转录的贡献。为了支持第二个目标，我们现已证明反义诱导的 DNMT1 mRNA 和蛋白下调将导致 RELN mRNA 表达显着增加。此外，我们有初步数据表明 GAD67 和 DNMT1 mRNA 响应去极化而发生剧烈变化。凭借 KO1 职业发展奖，候选人将寻求表观遗传基因调控分子研究重点方法的培训。由于表观遗传机制可以通过药物干预来改变，因此这项工作与新型疗法的开发具有直接相关性。