Delineating the genetic basis of amphetamine sensitivity using a Drosophila behavioral model

项目摘要 滥用精神刺激剂，包括苯丙胺(AMPHs)，是一个严重的公共卫生问题，具有深远的 精神、医学和社会心理方面的并发症。遗传因素在很大程度上决定了个人的 对成瘾的易感性；然而，寻找危险等位基因取得的成功有限。这个 最初对精神刺激剂的敏感性差异很大，并与持续使用和 虐待。这一特征可以在动物模型中进行研究，动物模型已经成为研究 有节制的、系统的对药物的行为反应。我们的方法组合 在这一应用中的提议将使我们能够利用果蝇遗传学的力量来发现新的基因 以及对Amph敏感的基因变异体。凭借其快速的生命周期和对遗传、细胞 分子分析，苍蝇使在更难调查的领域进行行为研究成为可能 在脊椎动物模型中。本申请中提供的初步数据表明，苍蝇对 通过增加他们的运动活动和减少他们的睡眠来缓解疼痛。基因突变破坏了 多巴胺(DA)合成或多巴胺转运体基因(DAT)功能抑制这些行为反应， 证明我们已经开发了一种强大的行为工具来将遗传变异与表型联系起来 改变。我们已经制定了将此行为分析与下一代(Next-Gen)相结合的策略 用测序技术和系统遗传学方法研究Amph的遗传结构 并找出影响这一特性的新的基因变异。这种综合方法通过以下方式得以实现 我们与大卫·戈德曼博士和科林·霍奇金森博士在神经遗传学实验室的积极合作 在NIAAA，最先进的下一代技术、遗传连锁研究和功能基因组学方面的专家 研究行为特征的方法。我们提出了以下具体目标：1)鉴定基因 野生亚株内和亚株之间Amph敏感度的巨大差异背后的变异- 模式，非等基因果蝇株广州S(CS)。我们将(A)使用选择性育种，结合 基因组学方法，如全基因组测序和缺陷图，以识别 改变不同CS亚株的Amph敏感性和b)使用RNA测序(RNA-seq)来分析基因 在不同亚株中与Amph敏感性改变相关的表达变化和2)筛选 大量近交系菌株，以确定与Amph敏感性改变相关的遗传位点。 我们将(A)筛选果蝇遗传参考小组，该小组由203个基因分型的近交系组成，用于 (B)使用系统遗传学方法，包括数量性状基因座(QTL)分析 以及极端QTL定位，将Amph敏感性的表型变异与DNA多态联系起来。

Project Summary Abuse of psychostimulants, including amphetamines (AMPHs), is a major public health problem with profound psychiatric, medical and psychosocial complications. Genetic factors contribute substantially to an individual's susceptibility to developing addiction; however, the search for risk alleles has yielded limited success. The initial sensitivity to psychostimulants varies significantly, and has been associated with continued use and abuse. This trait can be studied in animal models, which have emerged as powerful tools to investigate the behavioral response to drugs in a controlled and systematic manner. The combination of approaches we propose in this application will allow us to harness the power of Drosophila genetics to uncover novel genes and gene variants that confer sensitivity to AMPH. With its rapid life cycle and accessibility to genetic, cellular and molecular analyses, the fly has enabled behavioral studies in areas that are far more difficult to investigate in vertebrate animal models. The preliminary data presented in this application show that that flies respond to AMPH by increasing their locomotor activity and decreasing their sleep. Genetic mutations that disrupt dopamine (DA) synthesis or dopamine transporter gene (DAT) function inhibit these behavioral responses, demonstrating that we have developed a robust behavioral tool to associate genetic variations with phenotypic changes. We have developed a strategy combining this behavioral analysis with next-generation (Next-Gen) sequencing technology and systems genetics approaches to investigate the genetic architecture of AMPH sensitivity and identify new gene variants that influence this trait. This integrated approach is made possible by our active collaboration with Dr. David Goldman and Dr. Colin Hodgkinson at the Laboratory of Neurogenetics at NIAAA, experts in state-of-the-art Next-Gen technologies, genetic linkage studies and functional genomics approaches to the study of behavioral traits. We propose the following specific aims: 1) To identify gene variants that underlie wide variation in AMPH sensitivity within and between substrains of the wild- type, non-isogenic Drosophila strain Canton S (CS). We will (a) use selective breeding, combined with genomic approaches such as whole genome sequencing and deficiency mapping, to identify gene variants that alter AMPH sensitivity in different CS substrains and b) use RNA-sequencing (RNA-seq) to profile gene expression changes associated with altered sensitivity to AMPH in the different substrains and 2) To screen a large population of inbred strains to identify genetic loci associated with altered sensitivity to AMPH. We will (a) screen the Drosophila Genetic Reference Panel, which consists of 203 genotyped inbred lines, for response to AMPH and (b) use systems genetics approaches, including quantitative trait locus (QTL) analysis and extreme QTL mapping, to associate phenotypic variation in sensitivity to AMPH with DNA polymorphisms.