GENOME RESPONSES TO DROSOPHILA ADDICTION USING DNA CHIPS

描述：(申请者摘要)吸毒成瘾的机制似乎是 非常保守。因此，在动物模型系统中的工作可以极大地帮助 我们对人类成瘾的理解。规模经济和 果蝇可用的分子遗传学工具将被用来识别基因 以及成瘾的遗传途径。最近对果蝇的研究已经 建立了可卡因和乙醇的行为敏感性模型。在 后者是健忘症的一个等位基因，是一种学习和记忆基因，有一些 与一种刺激cAMP信号的脊椎动物神经肽的同源性是 已确认身份。健忘症突变体对乙醇的耐受性降低，这是一个缺陷 可通过药物刺激cAMP水平逆转。在前者中， 几个单基因突变体被证明影响可卡因的敏化，这 也是由多巴胺能信号介导的。这些研究证实了果蝇 作为可卡因和酒精成瘾的遗传模型系统 CAMP信号通路在对两者的反应中。 这种果蝇成瘾模型系统将通过DNA的使用而得到加强 用于识别药物相关基因表达变化的微阵列芯片 全基因组的规模。在初步实验中，一个部分基因组原型 利用果蝇芯片对176个候选基因进行转录鉴定 在健忘症突变体中发生改变，75个在转录上发生改变 对cAMP水平的药理学操纵。我们已经确认了一些 来自芯片的统计候选数据使用Northern杂交分析或 定量聚合酶链式反应。从对100多个芯片的初步分析来看，我们还拥有 开发了一种新的通用统计方法来分析DNA芯片数据。 这些初步结果现在将与暴露于 可卡因和乙醇。我们方法的核心是三个基因组的比较 回答：(1)药物暴露与对照，(2)药物操作与 对照；(3)突变体与野生型。首选用于进一步的体内研究 将是那些在所有三种情况下都有转录反应的基因 -以及可卡因和酒精暴露。给出了“函数的同调” 已经在苍蝇和脊椎动物身上观察到了药物反应，这些基因 在果蝇中发现的这种病毒很可能在脊椎动物中也有同源物。因此是这样的 这项工作将直接导致对哺乳动物成瘾模型的研究，并最终 将对人类成瘾的遗传基础有更深入的了解。

DESCRIPTION: (Applicant's Abstract) Mechanisms of drug addiction appear to be remarkably conserved. Work in animal model systems can therefore greatly inform our understanding of human addiction. The economy-of-scale and molecular-genetic tools available in fruit flies will be used to identify genes and genetic pathways of addiction. Recent studies in Drosophila have established behavioral sensitivity models for both cocaine and ethanol. In the latter, an allele of amnesiac, a learning and memory gene that has some homology to a vertebrate neuropeptide that stimulates cAMP signaling, was identified. Amnesiac mutants show decreased tolerance to ethanol, a defect that can be reversed with pharmacological stimulation of cAMP levels. In the former, several single-gene mutants were shown to affect cocaine sensitization, which also was mediated by dopaminergic signaling. These studies validate Drosophila as a genetic model system for both cocaine and alcohol addiction and implicate the cAMP signaling pathway in the response to both. This Drosophila model system of addiction will be enhanced with the use of DNA microarray chips to identify drug-related changes in gene expression on a genomic-wide scale. In preliminary experiments, a partial genome prototype Drosophila chip has been used to identify 176 candidate genes transcriptionally altered in amnesiac mutants and 75 that are transcriptionally altered by pharmacological manipulation of cAMP levels. We have confirmed some of the statistical candidates from the chip using either Northern blot analysis or quantitative PCR. From initial analyses of more than 100 chips, we also have developed a novel and general statistical method to analyze DNA chip data. These initial results now will be integrated with data from exposures to cocaine and ethanol. Central to our approach is the comparison of three genomic responses: (1) drug-exposed vs. control, (2) pharmacological manipulation vs. control and (3) mutant vs. wild-type. Chosen first for further in vivo study will be those genes that are transcriptionally responsive in all three contexts - and to both cocaine and ethanol exposures. Given the "homology of function" already observed for drug responses in flies and vertebrates, the genes identified in flies most likely will have homologues in vertebrates. Hence this work will lead directly to work in mammalian models of addiction and ultimately will yield a greater understanding of the genetic basis of addiction in humans.