lncRNA Gas5 dysregulation alters ethanol drinking behavior and ethanol-related phenotypes

项目摘要 酒精使用障碍(AUD)是一种慢性、衰弱和复发的脑部疾病。压倒性的数量 每年都有人死于酒精相关的原因；然而，其作用机制(S)很难破译 而且在很大程度上仍然难以捉摸。 长非编码RNA(LncRNAs)是基因组的关键调节因子；持续的转录变化诱导 慢性酒精暴露是AUD发生、戒断和复发的一种假想机制。我们的 实验室和其他研究人员已经表明，对乙醇反应的lncRNA具有直接调节乙醇的能力。 在体内发生突变时的相关行为，但到目前为止只有少数被表征。在这项建议中，我 我将学习和应用最先进的CRISPR/CAS9技术来从功能上研究乙醇响应 LncRNA Gas5对饮酒和与酒精相关的行为的调节。 随着乙醇改变基因表达和调节神经炎症的分子途径，我想针对一个 乙醇反应和高度互联的竞争内源RNA(Cerna)lncRNA，可能 协调与AUD相关的大型内源性免疫网络。 我将专注于体内CRISPR/Cas9对感兴趣的特定“Hub”lncRNA的调节，生长停滞特异性5 (Gas5)。Gas5作为Cerna调节免疫信号，显著且持续下调 在慢性间歇性乙醇蒸气暴露后，并已被证明可减少可卡因的摄入量， 对药物滥用的5号气体。因此，检验Gas5是一个关键的假设是很有意义的 乙醇作用的决定因素。Gas5将以特定于时间、细胞和大脑区域的方式被敲除 研究其在酒精饮酒中的作用。 在这个项目完成后，我将推进我们对乙醇分子影响的理解- 反应性lncRNA Gas5及其与成瘾行为的关系。该项目将提供大量的培训 机会，适用于各种科学领域的强大研究基础，以及合作 环境与该领域的其他研究人员，包括关于酒精中毒的综合神经科学倡议- 神经免疫联盟。丰富的技术技能，从板凳工作到复杂的外科技术， 对项目设计的理解和发展，以及几个科学写作手稿的展望 在每个目标中都有，这对我继续科学训练将是有价值的。

Project Summary Alcohol use disorder (AUD) is a chronic, debilitating, and relapsing brain disease. An overwhelming number of people die yearly due to alcohol-related causes; however, the mechanism(s) of action are challenging to decipher and remain largely elusive. Long noncoding RNAs (lncRNAs) are key regulators of the genome; persistent transcriptional changes induced by chronic ethanol exposure is a hypothesized mechanism for AUD development, withdrawal, and relapse. Our laboratory and others have shown that ethanol-responsive lncRNAs have the ability to directly regulate ethanol- related behavior when mutated in vivo, but only a handful have been characterized to date. In this proposal, I will learn and apply state-of-the-art CRISPR/Cas9 techniques to functionally investigate ethanol-responsive lncRNA Gas5 for its regulation of ethanol drinking and ethanol-related behaviors. As ethanol alters gene expression and molecular pathways that regulate neuroinflammation, I want to target an ethanol-responsive and highly-interconnected competing endogenous RNA (ceRNA) lncRNA that may coordinate large endogenous immune networks related to AUD. I will focus on in vivo CRISPR/Cas9 modulation of a specific ‘hub’ lncRNA of interest, growth arrest-specific 5 (Gas5). Gas5 acts as a ceRNA to regulate immune signaling, is significantly and persistently downregulated following chronic intermittent ethanol vapor exposure and has been shown to reduce cocaine-intake, linking Gas5 to substance abuse. It is therefore of substantial interest to test the hypothesis that Gas5 is a key determinant of ethanol action. Gas5 will be knocked out in a temporal, cellular, and brain region-specific manner to investigate its role on ethanol drinking. Upon completion of this project, I will have advanced our understanding for the molecular impact of ethanol- responsive lncRNA Gas5 and how it relates to addictive behavior. This project will provide substantial training opportunities, a strong research foundation applicable to a variety of scientific fields, and a collaborative environment with other researchers in the field including the Integrative Neuroscience Initiative on Alcoholism – Neuroimmune consortium. A wealth of technical skills ranging from bench work to complex surgical techniques, the understanding and development of project design, and several scientific writing manuscript prospects are available within each aim that will be valuable for my continued scientific training.