Genetic basis of clinal variation in ethanol tolerance in Drosophila melanogaster

描述(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。两个尚未解决的主要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。乙醇是果蝇繁殖地的天然成分，在不同的大陆上，对乙醇的抗性随着纬度的增加而强烈增加，表明这一特征是自然选择的结果。然而，温带-热带差异的遗传基础在很大程度上是未知的。对于黑腹果蝇来说，强大的遗传工具的可用性和关于果蝇-乙醇系统的丰富的背景信息使其成为研究自然适应的遗传基础的极佳选择。在拟议的工作中要检验的主要假设是，温带种群对高酒精水平的适应部分是由顺式调控进化带来的。温带和热带种群之间的基因表达差异将使用微阵列和实时聚合酶链式反应进行测量，并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，表达差异有助于乙醇抗性的自然地理差异，一个或多个基因在温带种群中的表达持续高于(低于)热带种群，将持续增加(降低)表达作为抗性选择的结果。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分析来衡量顺式调控变异对表达差异的贡献，并将在数量性状基因座(QTL)定位实验中估计基因对抗性的影响。如果对于一个给定的基因，温带/热带和选择/对照的表达差异都具有顺式调控成分，并且该基因接近抗病的QTL，则结果将提供强有力的证据，表明该基因是乙醇选择的目标。对于这类基因，将使用敲除突变和GAL4-UAS系统来操纵表达，以确认它们在抗药性中的作用。酗酒是一个主要的公共卫生问题。这项研究的结果可能会对酒精的神经和生理影响提供深入的见解，可能会确定酒精在神经系统中的新靶点。这些见解可能会被用来开发治疗酒精中毒的药物或其他疗法。

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.