Whole-genome analysis of DNA methylation in alcohol dependence

描述（由申请人提供）：酒精中毒是一种慢性复发性疾病，其特征是强迫性使用酒精，并损害个人的社会和职业功能。越来越多的证据表明，表观遗传机制可能有助于参与酒精依赖和耐受性的神经适应。然而，需要对酒精依赖和戒断引起的表观遗传改变进行更彻底的调查，以了解表观遗传机制在酒精持久作用中的作用。作为我们正在进行的识别酒精诱导适应基因的努力的一部分，我们已经观察到酒精暴露模型中DNA甲基化和基因表达的协调变化，无论是一次一个基因还是高通量方法。这些初步结果表明，DNA甲基化的变化可能是酒精影响的关键机制，并激发了对酒精依赖和戒断中甲基化模式的全基因组调查。新策略和工具的引入，如高密度DNA阵列，旨在研究全基因组范围内的染色质调控，现在可以系统地分析表观遗传改变。虽然DNA甲基化不是唯一的表观遗传修饰，但它是高等真核生物中DNA的唯一共价修饰，并且大量证据已经确立了其在神经和外周器官疾病中的作用。由于Affytechnology不生产也不计划生产大鼠基因组的平铺阵列，因此在本申请中，我们将限于小鼠。该提案的目的是在酒精依赖、戒断和依赖诱导的饮酒的小鼠模型中，在全球范围内分析DNA甲基化的变化。为了这个目的，我们将使用最近发布的称为“平铺”的新一代Affychip GeneChip（r）微阵列，因为它们的探针被设计成用单个微阵列覆盖基因组中所有已知和推定基因的整个启动子区域，或者用一组微阵列以无偏的方式覆盖整个基因组。我们将使用甲基胞嘧啶免疫沉淀（mCIP）作为检测差异DNA甲基化的主要手段。预计系统阐明DNA甲基化改变在酒精诱导的基因表达变化中的作用，将有助于确定新的致病假设和预防和治疗酒精滥用的治疗靶点。

DESCRIPTION (provided by applicant): Alcoholism is a chronic relapsing disorder characterized by compulsive use of alcohol and impairment in an individual's social and occupational functioning. Mounting evidence suggests that epigenetic mechanisms may contribute to the neuroadaptation that are involved in alcohol dependence and tolerance. However, a more thorough investigation of epigenetic alterations induced by alcohol dependence and withdrawal is needed to understand the role of epigenetic mechanisms in alcohol's long-lasting actions. As part of our ongoing effort to identify genes involved in alcohol-induced adaptations, we have observed coordinated changes in DNA methylation and gene expression in models of alcohol exposure both with one-gene-at-a-time and high-throughput approaches. These preliminary results suggest that changes in DNA methylation are likely to be a key mechanism of alcohol's effects and motivate the proposed genome-wide survey of patterns of methylation in alcohol dependence and withdrawal. The introduction of new strategies and tools such as high-density DNA arrays designed to investigate chromatin regulation on a genome-wide scale now makes it possible to systematically analyze epigenetic alterations. While DNA methylation is not the only epigenetic modification, it is the only covalent modification of DNA in higher eukaryotes and a large body of evidence has established its role in neurological and peripheral organ disease. Since Affymetrix does not produce and is not planning to produce tiling arrays for the rat genome, in the present application we will be limited to mice. This proposal is aimed at profiling changes in DNA methylation on a global basis in a mouse model of alcohol dependence, withdrawal and dependence-induced drinking. To this aim, we will use the recently released generation of Affymetrix GeneChip(r) microarrays denominated 'tiling' because their probes are designed to cover either the entire promoter region of all known and putative genes in the genome with a single microarray or the entire genome in an unbiased way with a set of microarrays. We will use methylcytosine-immunoprecipitation (mCIP) as the primary means to detect differential DNA methylation. It is expected that a systematic elucidation of the role of altered DNA methylation in alcohol-induced gene expression changes will foster the identification of new pathogenic hypotheses and therapeutic targets for the prevention and treatment of alcohol abuse.