Identifying Brain Epitranscriptomic Changes Associated with Alcohol Use Disorder

项目摘要/摘要 酒精使用障碍(AUD)每年影响约4.2%(或1410万)的美国成年人，并导致 相当高的发病率和死亡率。虽然基因变异会影响个体对AUD的易感性， 长期饮酒也会导致对酒精的耐受和依赖，但其潜在机制 目前还不清楚。有证据表明，长期饮酒会改变特定基因的DNA甲基化，导致 基因表达发生改变，并可能增加患AUD的风险。RNA甲基化是一种常见的后- 转录修饰，它对各种刺激快速反应，并将刺激信号翻译成 细胞活动。我们假设饮酒会改变个体的RNA甲基组(或表位转录组)， 导致参与AUD相关通路的基因表达发生变化。这项研究的目的是 探讨AUD患者脑内信使RNA(MRNA)甲基组的变化并分析其影响 信使核糖核酸甲基化对信使核糖核酸表达和神经元活性的影响，从而为寻找新基因提供证据 AUD的表达调控机制。为了实现这一目标，我们提出了三个具体目标。首先，我们将 八个区域(杏仁核、尾状核、小脑、 海马体、伏隔核、前额叶皮质、壳核和腹侧被盖区) AUD受试者大脑。其次，我们将使用慢性间歇性乙醇(CIE)暴露的小鼠作为模型来验证 AUD相关脑内基因甲基化、表达变化及与酒精的相关性 暴露诱导的mRNA甲基化和表达随乙醇自我给药的升级而变化 在老鼠身上。第三，我们将使用一种新的表位转录组学编辑方法来研究AUD- 相关的mRNA甲基化改变对mRNA表达和神经元活性的影响。我们期望(1)观察 AUD相关的mRNA甲基化和特定(或跨)脑区表达的变化；(2)验证AUD- 通过小鼠模型研究相关基因甲基化和表达的变化；(3)确认其功能作用 AUD相关信使核糖核酸甲基化在调节信使核糖核酸表达和神经元活性中的作用。我们的书房设计 是创新的。AUD相关的mRNA甲基化变化将在单碱基分辨率下进行检测。一个 综合研究方法(人类死后脑研究、小鼠建模和外显文组编辑) 将会被雇佣。这项拟议的研究将为大脑表位转录的影响提供证据。 澳元风险的变化。鉴于mRNA甲基化是动态的和可逆的，具有差异的mRNAs AUD患者脑内甲基化可能成为AUD药物治疗的潜在靶点 改变信使核糖核酸甲基化状态。

PROJECT SUMMARY/ABSTRACT Alcohol use disorder (AUD) affects about 4.2% (or 14.1 million) of American adults each year and causes substantial morbidity and mortality. While genetic variation can influence an individual's vulnerability to AUD, chronic alcohol consumption can also lead to alcohol tolerance and dependence, but the underlying mechanism is unclear. There is evidence that that chronic alcohol use alters DNA methylation of specific genes, leading to gene expression changes and possibly an increased risk of AUD. RNA methylation is a common post- transcriptional modification, and it responds rapidly to a variety of stimuli and translates stimulatory signals into cellular activity. We hypothesize that alcohol use alters an individual's RNA methylome (or epitranscriptome), resulting in altered expression of genes involved in AUD-related pathways. The objective of this research is to explore messenger RNA (mRNA) methylomic changes in the brain of AUD subjects and analyze the effect of mRNA methylation on mRNA expression and neuronal activity, thus providing evidence for a new gene expression regulatory mechanism of AUD. To achieve this goal, we propose three specific aims. First, we will profile mRNA methylomic and transcriptomic changes in eight regions (amygdala, caudate, cerebellum, hippocampus, nucleus accumbens, prefrontal cortex, putamen, and ventral tegmental area) of postmortem brains of AUD subjects. Second, we will use chronic intermittent ethanol (CIE)-exposed mice as models to verify AUD-associated brain mRNA methylation and expression changes as well as the correlation of ethanol exposure-induced mRNA methylation and expression changes with the escalation of ethanol self-administration in mice. Third, we will employ a novel epitranscriptome editing approach to investigate the effect of AUD- associated mRNA methylation changes on mRNA expression and neuronal activity. We expect to (1) observe AUD-associated mRNA methylation and expression changes in specific (or across) brain regions; (2) verify AUD- associated mRNA methylation and expression changes by mouse modeling; and (3) confirm the functional role of AUD-associated mRNA methylation in regulating mRNA expression and neuronal activity. Our study design is innovative. AUD-associated mRNA methylation changes will be examined at single-base resolution. An integrative study approach (human postmortem brain studies, mouse modeling, and epitranscriptome editing) will be employed. The proposed research will provide evidence about the influence of brain epitranscriptomic changes on AUD risk. Given that mRNA methylation is dynamic and reversible, mRNAs with differential methylation in the brain of AUD subjects could be potential targets for AUD treatment by pharmacologically altering mRNA methylation status.