Ethanol Withdwawal QTL and Candidate Genes

乙醇戒断QTL和候选基因

• 批准号: 6785931
• 负责人: KARI J BUCK
• 金额: $ 26.43万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2006-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6785931
• 关键词: GABA receptor; brain electrical activity; chromosomes; disease /disorder proneness /risk; drug withdrawal; ethanol; gene expression; genetically modified animals; genotype; immunocytochemistry; laboratory mouse; neural transmission; neurogenetics; polymerase chain reaction; protein structure function; quantitative trait loci; synapses; western blottings

DESCRIPTION (provided by applicant): Susceptibility to physiological dependence on ethanol (EtOH), which is manifested as a withdrawal syndrome, is influenced by multiple genes and environmental factors. We have identified three chromosomal regions that contain genes that influence risk for physiological dependence on EtOH in mice. These chromosomal regions are referred to as quantitative trait loci (QTL). This application focuses on the Chr 4 QTL (LOD>8, p<10[-9]). We have fine mapped this QTL to a 1.5 Mb (0.7 cM) interval that contains 3 known genes and at least 2 novel genes. This proposal will use novel genetic animal models for rigorous, unbiased evaluations of the known and predicted genes in this QTL interval as potential candidate genes. In addition, we will evaluate the influence of the QTL (gene) on neuronal activation related to physiological dependence on EtOH and associated withdrawal. Using a novel congenic strain that isolates the QTL on a uniform (inbred) genetic background, we propose the following. (1) Evaluate the predicted genes in the QTL interval for expression to identify novel expressed genes. Expression will be assessed in the brain, peripheral tissues, and in embryonic tissue. (2) Evaluate the expressed genes to identify promising candidates that show genotype-dependent (i.e., congenic vs. background strain mice) differences in expression. Expressed genes will also be assessed for differential regulatory and coding sequence (structural differences) between these strains. Whenever feasible, their protein products will be assessed immunohistochemically to determine the anatomical distribution of their expression. Genotype-dependent differences in protein abundance will be assessed using Western blot analysis. (3) Evaluate neuronal activation to identify the circuit(s) that show genotype dependent activation in EtOH withdrawn mice. (4) Develop an innovative, transgenic model to rigorously test the hypothesis that a promising candidate gene (Mpdz) underlies the QTL. An innovative feature of this proposal is to combine a robust behavioral model of physiological dependence on EtOH with sophisticated molecular techniques to identify candidate genes of high quality as well as the circuit(s) involved in their influence on EtOH withdrawal, and novel transgenic models that can establish with certainty that a promising candidate gene in fact underlies the QTL.

描述（申请人提供）：对乙醇（EtOH）的生理依赖性易感性，表现为戒断综合征，受到多种基因和环境因素的影响。我们已经确定了三个染色体区域，其中包含影响小鼠对乙醇生理依赖风险的基因。这些染色体区域称为数量性状基因座 (QTL)。该应用重点关注 Chr 4 QTL (LOD>8, p<10[-9])。我们已将此 QTL 精细映射到 1.5 Mb (0.7 cM) 区间，其中包含 3 个已知基因和至少 2 个新基因。该提案将使用新型遗传动物模型对该 QTL 区间中的已知和预测基因作为潜在候选基因进行严格、公正的评估。此外，我们将评估 QTL（基因）对与乙醇生理依赖性和相关戒断相关的神经元激活的影响。使用一种在统一（近交）遗传背景上分离 QTL 的新型同源菌株，我们提出以下建议。 (1)评估QTL区间内的预测基因的表达，以鉴定新的表达基因。将评估大脑、外周组织和胚胎组织中的表达。 (2) 评估表达的基因，以确定有希望的候选基因，这些候选基因表现出基因型依赖性（即同类小鼠与背景品系小鼠）的表达差异。还将评估表达基因的差异调节和编码序列（结构差异）。只要可行，就会对它们的蛋白质产物进行免疫组织化学评估，以确定其表达的解剖分布。将使用蛋白质印迹分析来评估蛋白质丰度的基因型依赖性差异。 (3) 评估神经元激活，以确定在 EtOH 戒断小鼠中表现出基因型依赖性激活的回路。 (4) 开发一种创新的转基因模型，以严格检验 QTL 背后有一个有前途的候选基因 (Mpdz) 的假设。该提案的一个创新特征是将乙醇生理依赖性的稳健行为模型与复杂的分子技术相结合，以确定高质量的候选基因以及影响乙醇戒断的电路，以及新颖的转基因模型可以肯定地确定一个有希望的候选基因实际上是 QTL 的基础。