Human genetic polymorphism impact in a mouse model

描述(由申请人提供)： 这个CEBRA应用程序解决了一个高风险、高影响的问题：人类基因变异能否通过转基因小鼠模型来模拟？这是一个迫切需要研究的问题，因为研究表明，微小的遗传变异，如单核苷酸多态(SNP)，是药物成瘾易感性和其他行为变化的主要遗传因素。然而，需要有一种方法来解决遗传变异和表型变化之间的因果关系。这个CEBRA应用程序解决了这个问题，使用转基因小鼠模型复制人类基因多态，允许对由此产生的表型差异进行严格测试，以评估基因变化的影响。重点是一种高度相关的、有充分证据证明的人类遗传变异--人类Mu阿片受体基因中的一种功能性SNP。Mu阿片受体是内源性阿片系统的关键组成部分，介导包括吗啡和海洛因在内的阿片类药物的生理效应。我们和其他人发现的A118G SNP是常见的，导致单一氨基酸的变化，并似乎通过增加细胞中β-内啡肽的敏感性和受体的表达水平来改变受体的功能。最近的人类研究表明，这种基因变异可能在体内具有重要的功能：在人类功能研究中，它与HPA轴的改变有关，并被认为是成瘾的重要因素。因此，A118G SNP是一种理想的遗传多态，可以测试小鼠对人类遗传变异建模的可行性，从而确定基因变化与随后的行为和体内生理功能变化之间的因果关系。这是我们CEBRA提案的目标。目的1.建立人类Mu阿片受体基因多态的小鼠模型。我们将在小鼠身上模拟人类A118G基因的多态。基于同源重组的基因打靶方法将被用来将人类的多态变异“敲入”到小鼠的Mu阿片受体基因中，从而创建这种常见的人类遗传变异的小鼠模型。目的2.研究MU阿片受体基因多态在体内的影响。我们将表征在AIM 1中产生的小鼠模型对体内遗传多态的影响。我们将研究多态对受体表达和配体敏感性的影响。我们还将探索对阿片类药物监管行为的任何潜在影响。由于高风险、高影响的性质，我们选择在CEBRA资助机制下申请。药物滥用研究领域需要知道是否可以在转基因小鼠身上模拟和研究人类遗传易感性因素。我们建议的研究结果将作为药物滥用研究人员未来战略考虑的指导性范例。

DESCRIPTION (provided by applicant): This CEBRA application addresses a high-risk, high-impact question: Can human genetic variations be modeled by transgenic mouse models? This is a question in urgent need of examination, because studies have shown that small genetic variations, such as single nucleotide polymorphism (SNP), are the primary genetic factors for drug addiction vulnerability and other behavioral changes. However, there needs to be an approach to address causal relationship between genetic variations and phenotypic changes. This CEBRA application addresses this question, using genetically modified mouse models to reproduce human genetic polymorphisms, allowing rigorous testing for resulting phenotypic differences to assess the impact of genetic changes. The focus is on a highly relevant, well-documented human genetic variation - a functional SNP in the human mu opioid receptor gene. The mu opioid receptor is a key component of the endogenous opioid system, mediating the physiological effects of opioid drugs including morphine and heroin. The A118G SNP that we and others have identified is common, leads to a single amino acid change, and appears to alter receptor function by increasing beta-endorphin sensitivity and receptor expression level in cells. Recent human studies suggest that this genetic variant may be of functional importance in vivo: it is associated with alterations of the HPA-axis in functional studies in humans, and is implicated as an important contributor to addictions. Thus, the A118G SNP is an ideal genetic polymorphism to test the feasibility of mouse modeling for human genetic variations - so as to determine the causal relationship between a genetic change and subsequent functional alterations in behavior and in vivo physiology. This is the goal of our CEBRA proposal. Aim 1. Generating a mouse model for the human genetic polymorphism in the mu opioid receptor. We will model the human A118G polymorphism in the mouse. The homologous recombination-based gene targeting approach will be used to "knock-in" the human polymorphic variations into the mu opioid receptor gene in mouse, thus creating a mouse model of this common human genetic variation. Aim 2. Characterize impact of the mu opioid receptor polymorphism in vivo. We will characterize the mouse model generated in Aim 1 for impact of genetic polymorphism in vivo. We will examine the effects of the polymorphism on receptor expression and ligand sensitivity. We will also explore any potential impact on opioid-regulated behaviors. Because of the high-risk high-impact nature, we choose to apply under the CEBRA funding mechanism. The field of drug abuse research needs to know whether human genetic vulnerability factors can be modeled and studied in transgenic mice. The outcome from our proposed studies will serve as a guiding example for future strategic considerations by drug abuse researchers.