Analysis of bipolar and schizophrenia candidate alleles

双相情感障碍和精神分裂症候选等位基因分析

• 批准号: 7148929
• 负责人: HERBERT M LACHMAN
• 金额: $ 26.15万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7148929
• 关键词: DNA binding protein; alleles; biotechnology; bipolar depression; clinical research; developmental genetics; developmental neurobiology; gel mobility shift assay; gene expression; gene mutation; genetic enhancer element; genetic mapping; genetic markers; genetic polymorphism; genetic promoter element; genetic regulatory element; genetic screening; genetic susceptibility; human genetic material tag; human tissue; laboratory mouse; neuregulins; neurogenetics; schizophrenia; single nucleotide polymorphism

DESCRIPTION (provided by investigator): Several candidate genes for schizophrenia (SZ) have been identified as a result of genetic linkage and association studies, including NRG1 and DTNBP1, among others. However, despite intensive analysis of exons, intron-exon borders, and promoter regions immediately upstream from transcription start sites, specific disease-causing alleles have not yet been identified. One possibility is that the expression of these candidate genes is being affected by variant alleles located in regulatory elements that have not yet been identified, that can function many kilobases away from transcription start sites, such as enhancers and locus control regions. The primary aims of this proposal are to identify distal regulatory elements of several SZ candidate genes and determine if potential functional alleles, which would be candidates for SZ susceptibility, exist within. We will screen for distal regulatory elements by mapping DNAse 1 hypersensitive sites in the NRG1 and DTNB1 loci by quantitative real-time PCR, and testing for promoter and enhancer function in the DNA near these sites using the pGL2-luciferase reporter system. Variant alleles will be tested in the same system to determine if promoter/enhancer function is impaired. They will also be tested for protein binding affinity using electromobility gel shift assays (EMSA) to determine whether or not allele-specific differences in DNA-protein complexes exist. Functional mutations will be analyzed in a case control association study to determine if the allele and genotype distributions differ between controls and patients with SZ. For the most promising variants, we will identify the DNA-binding protein by EMSA supershift using antibodies to the most likely candidates, or by purifying them using DNA-binding proteins using heparin Sepharose and DNA Sepharose chromatography. By identifying the proteins that bind to a gene's regulatory regions, we will gain new insight into the pathogenesis of SZ, in particular, gene-environment interactions during development that may play a role in some cases. It is through alterations in gene expression that environmental stressors interact with the brain.

描述（研究人员提供）：由于遗传联系和关联研究，包括NRG1和DTNBP1等，已经确定了精神分裂症（SZ）的几个候选基因（SZ）。 然而，尽管对外显子，内含子 - 外观边界和启动子区域进行了深入分析，但尚未确定引起疾病的特定等位基因的转录起始位点上游，但尚未确定。 一种可能性是，这些候选基因的表达受到尚未鉴定的调节元素中的变异等位基因的影响，这些等位基因尚未鉴定出来，这些等位基因可以运行许多千叠酶，从而从转录起始位点（例如增强子和基因座控制区域）发挥作用。 该提案的主要目的是确定几个SZ候选基因的远端调节元素，并确定内部是否存在潜在的功能等位基因，这可能是SZ敏感性的候选者。 我们将通过绘制NRG1和DTNB1基因座中的DNase 1超敏位点的远端调节元件，并通过定量实时PCR进行测试，并使用PGL2-荧光酶报告基因系统在这些位点附近的DNA中测试启动子和增强子功能。 变体等位基因将在同一系统中测试，以确定启动子/增强子功能是否受损。 它们还将使用电动性凝胶移位测定（EMSA）来测试它们的蛋白质结合亲和力，以确定是否存在DNA-蛋白复合物的等位基因特异性差异。 在案例控制关联研究中将分析功能突变，以确定对照和SZ患者之间的等位基因和基因型分布是否不同。 对于最有前途的变体，我们将使用抗体最有可能候选的抗体来鉴定EMSA Supershift的DNA结合蛋白，或者使用肝素sepharose和DNA Sepharose色谱使用DNA结合蛋白纯化它们。 通过鉴定与基因调节区域结合的蛋白质，我们将获得对SZ发病机理的新见解，尤其是在发育过程中可能在某些情况下起作用的基因 - 环境相互作用。 环境压力源与大脑相互作用是通过基因表达的改变。