Function and polymorphisms of complement receptor 1 (CR1) in Alzheimer's disease

补体受体1（CR1）在阿尔茨海默病中的功能和多态性

• 批准号: 8286201
• 负责人: JOSEPH B ROGERS
• 金额: $ 54.01万
• 依托单位: SRI INTERNATIONAL
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8286201
• 关键词: Accounting; Address; Adopted; Alzheimer' s Disease; Alzheimer' s disease risk; Antigen-Antibody Complex; Apolipoprotein E; Back; Bar Codes; Binding; Biological Assay; Blood Circulation; Blood specimen; Brain; Collaborations; Complement; Complement Activation; Complement Receptor; Complement component C1; Coupled; Data; Defect; Disease; Elderly; Engineering; Enzyme-Linked Immunosorbent Assay; Erythrocytes; Exhibits; Functional RNA; Genes; Genetic; Genetic Polymorphism; Genetic Risk; Genetic Variation; Genomics; Goals; Haplotypes; Left; Link; Linkage Disequilibrium; Mediating; Membrane; Molecular Weight; Nature; Outcome Measure; Peripheral; Personal Communication; Physiological; Play; Population; Primates; Property; Proteins; Publishing; Regulation; Reporting; Research; Research Personnel; Risk; Risk Assessment; Role; Sampling; Single Nucleotide Polymorphism; Site-Directed Mutagenesis; Structure; Techniques; Testing; Validation; Variant; Western Blotting; base; disorder risk; genetic risk factor; genome wide association study; instrument; next generation; novel; pathogen; protein structure

DESCRIPTION (provided by applicant): A recent genome-wide association study (GWAS) found that single nucleotide polymorphisms (SNPs) in CR1 exhibited the third highest association with genetic risk for Alzheimer's disease (AD), following only ApoE and ApoJ. Three non-overlapping GWAS efforts have confirmed a highly significant CR1 SNP association with AD risk. The overall goal of the present project is to fill in two major gaps left by the GWAS reports. 1) How is CR1 functionally altered in AD? and 2) To what extent do CR1 polymorphisms account for the functional alterations? These two issues are linked in our proposal by a reverse-engineering strategy that will first assess functional CR1 changes in AD, then use that information to identify the responsible CR1 SNPs. Specific Aim 1. Identify abnormalities in CR1 protein molecular weight, expression, binding, or function in AD and ND subjects. Despite its association with AD risk, CR1 is actually very poorly expressed in brain. Instead, its primary roles are regulation of spontaneous complement activation and clearance of pathogens and immune complexes (ICs)-both of which occur almost exclusively in the peripheral circulation. Aim 1 will therefore evaluate CR1 structure, expression, binding, complement regulation, and pathogen/IC clearance in blood samples from 300 AD and 300 ND subjects using ELISA, Western blot, complement activation and other techniques with which the investigators have documented expertise. SNPs themselves do not cause disease risk; functional changes induced by SNPs do. Testing mechanisms of CR1 function in AD is therefore an essential first step in understanding the full significance of AD-related CR1 polymorphisms. Specific Aim 2. Test the extent to which the CR1 protein abnormality or abnormalities identified in Aim 1 are linked to specific CR1 gene polymorphisms. Previous GWAS analyses were able to sample only a small fraction of the 476 known SNPs in CR1, and the CR1 SNPs they identified turn out to be in non- coding regions. The elucidation of exact, risk-enhancing SNPs, however, entails many well known challenges and thousands of samples. We have therefore chosen the more modest goal of seeking functional correlates. Using the Aim 1 samples, a RainDance enrichment approach will be employed to select for the CR1 genomic region, followed by bar-coded next generation sequencing on the Illumina GA HiSeq instrument. CR1 SNPs will then be tested, within subjects, for association with any CR1 protein alterations found in Aim 1. A potential shortcut lies in the fact that nearly a dozen CR1 SNPs have already been linked to specific CR1 protein abnormalities in other disease contexts. Candidate SNPs will be further evaluated by site-directed mutagenesis to determine which of them recapitulates Aim 1 CR1 protein defects. An ultimate goal will be to perform validation and true risk assessment in a much larger sample population, an undertaking that would be warranted and facilitated by our prior identification of CR1 SNPs that induce specific CR1 functional alterations.

描述（由申请人提供）：最近的一项全基因组关联研究（GWAS）发现，CR 1中的单核苷酸多态性（SNP）与阿尔茨海默病（AD）遗传风险的关联性排名第三高，仅次于ApoE和ApoJ。三个非重叠的GWAS工作已经证实了CR 1 SNP与AD风险的高度显著相关性。本项目的总体目标是填补全球WAS报告留下的两个主要空白。1)CR 1在AD中的功能是如何改变的？2）CR 1基因多态性在多大程度上解释了功能改变？在我们的提案中，这两个问题通过逆向工程策略联系在一起，该策略将首先评估AD中的功能性CR 1变化，然后使用该信息来识别负责的CR 1 SNP。 具体目标1。确定AD和ND受试者中CR 1蛋白分子量、表达、结合或功能的异常。尽管CR 1与AD风险相关，但它在大脑中的表达实际上非常差。相反，它的主要作用是调节自发补体激活和病原体和免疫复合物（IC）的清除-这两者几乎完全发生在外周循环中。因此，目标1将使用ELISA、蛋白质印迹、补体激活和研究者已记录专业知识的其他技术，评价来自300名AD和300名ND受试者的血液样本中的CR 1结构、表达、结合、补体调节和病原体/IC清除。SNPs本身并不引起疾病风险; SNPs引起的功能变化会引起疾病风险。因此，在AD中检测CR 1功能的机制是理解AD相关CR 1多态性的全部意义的重要的第一步。 具体目标2。检测目标1中确定的CR 1蛋白异常或异常与特定CR 1基因多态性相关的程度。以前的GWAS分析只能对CR 1中476个已知SNPs中的一小部分进行采样，他们发现的CR 1 SNPs原来是在非编码区。然而，精确的、风险增强的SNP的阐明需要许多众所周知的挑战和数千个样本。因此，我们选择了寻求功能相关性的更温和的目标。使用Aim 1样本，将采用RainDance富集方法选择CR 1基因组区域，然后在Illumina GA HiSeq仪器上进行条形码下一代测序。然后将在受试者中测试CR 1 SNP与Aim 1中发现的任何CR 1蛋白质改变的关联。一个潜在的捷径在于，近12个CR 1 SNP已经与其他疾病背景下的特定CR 1蛋白异常相关。将通过定点诱变进一步评价候选SNP，以确定它们中的哪一个重现Aim 1 CR 1蛋白缺陷。最终目标将是在更大的样本人群中进行验证和真正的风险评估，这一任务将通过我们先前鉴定诱导特定CR 1功能改变的CR 1 SNP来保证和促进。