Epigenetic Regulation of Neuronal Cell Death

描述(申请人提供)：肌萎缩侧索硬化症(ALS)是一种致命的神经退行性疾病，在全球范围内发生，没有种族、民族或社会经济界限。大多数ALS病例是散发性的，没有已知的遗传成分。流行病学研究表明，一些环境因素和获得性因素参与了ALS的发病机制；然而，以往的遗传学研究一直无法揭示涉及散发性ALS分子机制的基因。我们假设DNA的异常表观遗传调控参与了ALS的神经退行性变。具体地说，我们认为DNA甲基化导致神经元或骨骼肌突触维持基因或细胞生存基因的异常沉默可能是ALS神经退行性变的一个关键因素。我们对ALS患者和年龄匹配的对照组获得的人类基因组DNA的初步研究显示了高甲基化和低甲基化的不同模式。CpG岛微阵列已发现神经连接蛋白-1基因启动子在人类ALS神经组织中高度甲基化。我们还发现，在人类ALS神经组织中，DNA甲基转移酶亚型(DNMT1和3A)和5-甲基胞嘧啶的水平增加。在细胞培养和小鼠模型中，我们发现DNMT的表达和活性增强会导致神经元变性。我们建议进行以下研究。在具体目标1中，我们将对人类肌萎缩侧索硬化症和年龄匹配的对照神经组织和骨骼肌进行额外的CpG微阵列分析，以确定高甲基化或低甲基化的特定基因，并使用甲基化敏感-限制-指纹等替代方法验证基因甲基化模式，还将展示异常DNMT表达和DNA甲基化的细胞类型特异性。在特定的目标2中，我们将检验这一假说，即神经连接素1沉默通过神经元间病变和突触抑制失败参与ALS的机制。通过对ALS中DNA甲基化和基因沉默的研究，我们可以确定ALS发病的基于细胞、基于基因组和基于生理的新机制，从而为ALS的治疗提供新的策略。

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with worldwide incidence and no racial, ethnic, or socioeconomic boundaries. Most ALS cases are sporadic with no known inherited component. Epidemiological studies implicate some environmental and acquired factors in its pathogenesis; however, previous genetic studies have been unable to uncover the genes involved in elucidating the molecular mechanisms of sporadic ALS. We hypothesize the aberrant epigenetic regulation of DNA contributes to the neurodegeneration in ALS. Specifically, we propose that aberrant silencing of neuronal or skeletal muscle synapse maintenance genes or cell survival genes by DNA methylation could be a critical factor in the cause of neurodegeneration in ALS. Our preliminary studies of human genomic DNA obtained from ALS patients and age-matched controls reveal differential patterns of hyper- and hypomethylation. CpG island microarrays have identified the neuroligin-1 gene promoter as being hypermethylated in human ALS nervous tissue. We also find increased levels of DNA methyltransferase isoforms (Dnmt1 and 3A) and 5- methylcytosine in human ALS nervous tissue. In cell culture and mouse models, we find that enforced expression and activity of Dnmts induce neuronal degeneration. We propose to conduct the following studies. In Specific Aim 1 we will perform additional CpG microarray analyses of human ALS and age-matched control nervous tissue and skeletal muscle to determine the specific genes that are hypermethylated or hypomethylated and to validate the patterns of gene methylation using alternative methods such as methylation- sensitive-restriction-fingerprinting and also demonstrate cell-type specificities in aberrant Dnmt expression and DNA methylation. In Specific Aim 2 we will test the hypothesis that neuroligin 1 silencing participates in the mechanisms of ALS through interneuronopathy and failure of synaptic inhibition. By studying the DNA methylation and gene silencing in ALS, we could identify novel cell-based, genome-based, and physiology-based mechanisms of ALS pathogenesis that could lead to novel strategies for treating ALS.