Dissertation Research: Pathway Analysis of Ethanol and Acetic Acid Detoxification in Drosophila

乙醇和醋酸耐受性在黑腹果蝇种群和果蝇种群之间呈高度正相关。与乙醇和乙酸解毒途径中不同步骤相关的性状的不独立性强调了对乙醇和乙酸耐受性途径分析的必要性。由于在生物化学和分子水平上只能对ADH和醛氧化酶（ALDOX）进行研究，使得乙酰辅酶a合成酶（AcCoAS）催化的乙醇解毒途径的第三步尚未得到表征，这阻碍了对这一完整途径的分析。从黑腹龙的全基因组序列中鉴定出AcCoAS，以及最近开发的AcCoAS反应分子探针，现在可以对乙醇和醋酸耐受性进行完整的途径分析。我们假设这两种耐受性的选择作用于整个解毒途径，并将形成途径中三种关键酶之间的变异和协变模式。将采用多种技术组合来量化核苷酸序列数据、mRNA表达水平、酶活性以及对乙醇和乙酸的耐受性之间的变异和协变。这项由博士论文改进基金资助的研究采用进化定量遗传方法来表征遗传变异与途径中所有三个步骤的酶活性变异之间的关系，以及这种变异对乙醇和乙酸耐受性的影响。这些发现将在代谢控制理论的背景下进行分析，并将使我们能够对生物性能选择的遗传和生化反应做出预测

0105085ClarkEthanol and acetic acid tolerance are highly positively correlated across populations of D. melanogaster and species of Drosophila. The non-independence of traits associated with different steps in the ethanol and acetic acid detoxification pathway underscores the need for a pathway analysis of ethanol and acetic acid tolerance. Analysis of this complete pathway was hindered by the fact that only ADH and aldehyde oxidase (ALDOX) could be studied at the biochemical and molecular levels, leaving the third step in the ethanol detoxification pathway, catalyzed by acetyl Co-A synthetase (AcCoAS), uncharacterized. Identification of AcCoAS from the complete genome sequence of D. melanogaster and recently developed molecular probes for the AcCoAS reaction now allow for a complete pathway analysis of ethanol and acetic acid tolerance. We hypothesize that selection for both tolerances operates on the entire detoxification pathway and will shape the patterns of variation and covariation between the three key enzymes in the pathway. A combination of techniques will be employed to quantify variation in and covariation between nucleotide sequence data, mRNA expression levels, enzyme activities and tolerance to both ethanol and acetic acid.The research funded by this Doctoral Dissertation Improvement Grant takes an evolutionary quantitative genetic approach to characterize the relationship between genetic variation and enzyme activity variation at all three steps in the pathway and the consequences of this variation on ethanol and acetic acid tolerance. The findings will be analyzed in the context of metabolic control theory and will allow us to make predictions about the genetic and biochemical responses to selection on organismal performance