Combinatorial Actions of Genetic Variants and Gender Bias of Alzherimer's Disease

项目摘要 阿尔茨海默病(AD)通常以特定的神经病理特征为特征，包括 出现细胞外淀粉样物沉积和细胞内神经原纤维缠结堆积。而当 几个基因突变显然与早发性阿尔茨海默病有关，大量的 表现出延迟发病、与衰老相关的AD的个体可能在关联修饰物中存在许多变化 易患阿尔茨海默病的基因。遗传和全基因组关联研究(GWAS)有 发现了大量表明细胞自主和非细胞自主的基因和风险等位基因 导致神经元丧失和认知衰退的机制。在这方面，大多数风险变种 Gwas鉴定的基因位于基因组的非编码区，这意味着它们在一定程度上改变了基因 表情。这项提议回应了RFA指出的特别需要旨在描绘的方法 风险等位基因中SNPs组合产生新细胞的转录和细胞后果 帮助解开特定神经元中致病SNPs及其靶基因问题的系列试剂 来源于AD患者的iPS细胞。散发性AD有两个特征需要分子 解释--衰老在AD易感性中的潜在作用，以及显著的性别差异，随着发病率的增加 AD在女性身上被夸大的可能性。这些问题现在只能基于新技术和 患者来源样本的可用性。我们提出的研究计划利用了无价的 样本存储在加州大学圣迭戈分校Shiley-Marcos阿尔茨海默病研究中心(ADRC)的脑库中， 和IPSC来源的神经元(SOLK)。这一方法将询问不同基因变异的影响 其他风险因素(如年龄、性别)，并评估它们对特定细胞类型增强子环境的影响。通过合并 这些数据，我们可以开始识别导致特定细胞类型功能改变的潜在致病SNPs 增强剂。我们建议使用高通量4C筛选方法(UMI-4C)和Hi-Chip来识别 最有可能的是与功能相关的编码目标基因的致病、增强子相关的SNPs。利用 当代基因编辑方法在对照或患者来源的iPS细胞中对特定神经细胞的能力 对于星形胶质细胞，我们可以评估神经元的转录表型和功能行为 存在不同的风险等位基因组合，无论是单独在分离的细胞系中还是在与 星形胶质细胞和小胶质细胞共培养实验，因为这些SNP的影响可能只在 星形胶质细胞：神经元之间的相互作用。这些研究将共同使用强大的当代全球基因组 确定增强子中几个最重要的SNP的编码转录靶标的方法， 以及雌激素受体在AD性别差异中的作用。

PROJECT ABSTRACT Alzheimer’s disease (AD) is conventionally characterized by specific neuropathological features, including the appearance of extracellular amyloid deposits and the accumulation of intracellular neurofibrillary tangles. While several gene mutations are clearly associated with early onset Alzheimer’s disease, the large number of individuals exhibiting delayed onset, aging-associated AD, are likely to harbor many alterations in linked modifier genes that predispose to AD susceptibility. Genetic and genome wide association studies (GWAS) have identified numerous genes and risk alleles that indicate both cell autonomous and non-cell autonomous mechanisms contributing to loss of neurons and cognitive decline. In this regard, the majority of risk variants identified by GWAS reside in non-coding regions of the genome, implying that they act in part to alter gene expression. This proposal responds to the RFA indicating a particular need for approaches designed to delineate the transcriptional and cellular consequences of combinations of SNPs in the risk alleles by generating new cell line reagents to help unravel the question of the causative SNPs and their target genes in specific neurons derived from iPS cells of AD individuals. There are two features of sporadic AD that require molecular explanation- the potential role of aging in AD susceptibility, and the striking gender disparity, with the incidence of AD being exaggerated in females. These issues can only now be addressed based on new technologies and the availability of patient-derived samples. Our proposed research plan takes advantage of the invaluable samples stored at the brain bank of the Shiley-Marcos Alzheimer's Disease Research Center (ADRC) at UCSD, and the iPSC-derived neurons (Salk). This approach will interrogate the effects of different genetic variants with other risk factors (e.g. age, sex), and assess their effects on cell type-specific enhancer landscapes. By merging these data, we can begin to identify the potential causative SNPs that result in altered function of cell-type specific enhancers. We propose using a high throughput 4C screening approach (UMI-4C), and Hi-ChIP, to identify the most likely causative, enhancer-associated SNPs for functionally-implicated coding target genes. Exploiting the power of contemporary gene editing approaches in control or patient-derived iPS cells to specific neuronal cell types, and to astroglia, we can assess the transcriptional phenotypes and functional behaviors of neurons harboring different combinations of risk alleles, both in the isolated cell lines alone and in combination with coculture experiments with astroglia and microglia, as effects of these SNPs may be manifest only with astroglial:neuronal interactions. Together these studies will use powerful contemporary global genomic approaches to determine the coding transcriptional targets of several of the most significant SNPs in enhancers, and the link to roles of estrogen receptor in the gender disparity for AD.