Genetic basis of clinal variation in ethanol tolerance in Drosophila melanogaster

描述（由申请人提供）：阐明适应的遗传基础是进化生物学的主要目标之一。两个主要未解决的问题是参与适应的基因数量，以及基因表达变化和蛋白质结构变化对表型变化的相对贡献。乙醇是黑腹果蝇（Drosophila melanogaster）孳生地的天然成分，在不同大陆，对乙醇的抗性随纬度的增加而增强，表明该性状是自然选择的结果。然而，温带-热带差异的遗传基础在很大程度上是未知的。强大的遗传工具的可用性和果蝇-乙醇系统的丰富背景信息使其成为研究自然适应遗传基础的绝佳选择。在提出的工作中要测试的主要假设是，温带种群对高乙醇水平的适应部分是由顺式调节进化带来的。将使用微阵列和实时PCR测量温带和热带种群之间基因表达的差异，并将其与人工选择增加抗性所产生的差异进行比较。在一定程度上，表达差异促成了乙醇抗性的自然地理差异，一个或多个基因在温带种群中表现出始终高于（低于）热带种群的表达，作为抗性选择的结果，其表达将始终增加（减少）。对于符合这一标准的基因，我们将使用等位基因特异性表达测定来测量顺式调控变异对表达差异的贡献，并将在数量性状位点（QTL）定位实验中估计基因对抗性的影响。如果某一基因在温带/热带和选择/对照中的表达差异都具有顺式调控成分，且该基因接近抗性QTL，则该结果将有力地证明该基因是乙醇选择的靶标。对于这些基因，将使用敲除突变和GAL4-UAS系统来操纵表达，以确认它们在耐药性中的作用。酗酒是一个重大的公共卫生问题。这项研究的结果可能会对酒精在神经和生理上的影响有深入的了解，可能会确定乙醇在神经系统中的新靶点。这些发现可能被用于开发酒精中毒的药物或其他治疗方法。

DESCRIPTION (provided by applicant): Elucidating the genetic basis of adaptation is one of the primary goals of evolutionary biology. Two major unresolved questions are the number of genes involved in adaptations, and the relative contributions of changes in gene expression and changes in protein structure to phenotypic change. Ethanol is a natural constituent of breeding sites of Drosophila melanogaster, and resistance to ethanol increases strongly with latitude on different continents, indicating that the trait is under natural selection. The genetic basis of the temperate-tropical difference is largely unknown, however. The availability of powerful genetic tools for D. melanogaster and the wealth of background information on the Drosophila-ethanol system make it an excellent choice for investigating the genetic basis of a natural adaptation. The principal hypothesis to be tested in the proposed work is that adaptation to high ethanol levels in temperate populations has been brought about in part by cis-regulatory evolution. Differences in gene expression between temperate and tropical populations will be measured using microarrays and real-time PCR, and compared to differences resulting from artificial selection for increased resistance. To the extent that expression differences contribute to the natural geographic variation in ethanol resistance, one or more genes showing consistently higher (lower) expression in temperate than tropical populations will consistently increase (decrease) in expression as a result of selection for resistance. For genes meeting this criterion, we will use allele-specific expression assays to measure the contribution of cis-regulatory variation to the expression differences, and will estimate the effects of the genes on resistance in a quantitative trait locus (QTL) mapping experiment. If, for a given gene, the temperate/tropical and selected/control expression differences both have cis-regulatory components, and the gene is close to a QTL for resistance, the results will give strong evidence that the gene is a target of selection by ethanol. For such genes, expression will be manipulated using knockout mutations and the GAL4-UAS system to confirm their role in resistance. Alcohol abuse is a major public health problem. The results of this study are likely to give insights into alcohol's neurological and physiological effects, possibly identifying new targets for ethanol in the nervous system. These insights could potentially be used to develop drug or other treatments for alcoholism.