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

在黑腹果蝇和果蝇的种群中，对乙醇和醋酸的耐受性呈高度正相关。乙醇和醋酸解毒途径中与不同步骤相关的性状的非独立性突出了乙醇和醋酸耐受性的途径分析的必要性。由于在生物化学和分子水平上只能研究ADH和醛氧化酶(ALDOX)，而乙酰辅酶A合成酶(AcCoAS)催化的乙醇解毒途径的第三步尚未确定，这一事实阻碍了对这一完整途径的分析。从黑腹果蝇全基因组序列中鉴定AcCoAS，以及最近开发的用于AcCoAS反应的分子探针，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，对这两种耐受性的选择都在整个解毒途径上进行，并将塑造该途径中三种关键酶之间的变异和协变模式。我们将使用多种技术的组合来量化核苷酸序列数据、mRNA表达水平、酶活性和对乙醇和醋酸的耐受性之间的差异和协变。由这篇博士论文Improval Grant资助的研究采用进化定量遗传学的方法来表征基因变异和酶活性变化在途径的所有三个步骤中的关系以及这种变异对乙醇和醋酸耐受性的影响。这些发现将在代谢控制理论的背景下进行分析，并将使我们能够预测对生物性能选择的遗传和生化反应。

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