MECHANISMS OF ETHANOL TOXICITY IN DICTYOSTELIUM

为了了解乙醇对人类的影响，我们必须首先确定乙醇毒性的分子靶点。这些分子将是诊断酒精中毒遗传易感性和研究酒精相关疾病治疗的主要候选者。这项建议的广泛目标是通过模式生物盘基网柄菌的遗传筛选来确定乙醇毒性的分子靶标。CAMP信号转导是哺乳动物乙醇中毒的可能靶点之一，而介导cAMP信号转导的分子机制在进化差异的生物中是高度保守的。在网柄苔藓中，就像在人类中一样，cAMP信号对乙醇的影响很敏感。然而，对网柄苔藓的遗传学研究远没有在其他生物中那么复杂，部分原因是cAMP信号转导机制对于生长是必不可少的。这提供了一个强大的遗传模型，其中cAMP信号基因可以通过缺失、过度表达和其他突变来操纵。遗传操作的后果可以通过多细胞发育和孢子萌发的变化进行监测，这些过程高度依赖于cAMP信号转导。这一建议的实验目标是确定DictyostelialcAMP信号中的已知基因是否为乙醇毒性的相关靶标，并筛选具有改变乙醇敏感性的新突变体。在对相关基因进行鉴定和鉴定后，将克隆人的同源物，并在突变的Dictyostelius细胞中表达。与乙醇相关的突变互补的人类基因很可能是乙醇毒性的分子靶标。表达人类基因的网柄菌菌株可能会提供一种方便的实验模型，这将有助于未来对人类的研究。

In order to understand the effects of ethanol on humans we must first identify the molecular targets for ethanol toxicity. These molecules will be prime candidates for the diagnosis of genetic predisposition to alcoholism and for investigations into therapy for alcohol-related disorders. The broad objective of this proposal is to identify molecular targets for ethanol toxicity by means of genetic screens in the model organism Dictyostelium discoideum. cAMP signal transduction is one of the likely targets for ethanol toxicity in mammals, and the molecular mechanisms that mediate cAMP signaling are highly conserved between evolutionary divergent organisms. In Dictyostelium, as in humans, cAMP signaling is sensitive to the effects of ethanol. However, genetic studies in Dictyostelium are far less complicated then in other organisms, partly because the cAMP signal transduction mechanism is dispensable for growth. This provides a powerful genetic model in which cAMP signaling genes can be manipulated by deletion, over expression, and other mutations. Consequences of the genetic manipulations can be monitored as changes in multicellular development and in spore germination, processes that are highly dependent on cAMP signal transduction. The experimental goals of this proposal are to determine whether known genes in Dictyostelium cAMP signaling are relevant targets for ethanol toxicity, and to select for new mutants with altered ethanol sensitivity. Following identification and characterization of the relevant genes, human homologs will be cloned and expressed in mutants Dictyostelium cells. Humans genes that complement ethanol related mutations in Dictyostelium are likely to be molecular targets for ethanol toxicity. Dictyostelium strains that express the human genes may provide a convenient experimental model that will facilitate future studies in humans.