Regulation of APP Pathway Gene Promoters in Alzheimer's

阿尔茨海默病中 APP 通路基因启动子的调控

基本信息

批准号：
7674686
负责人：
DEBOMOY K LAHIRI
金额：
$ 28.76万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-09-01 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7674686
关键词：
5&apos Flanking Region 5&apos Untranslated Regions Abeta synthesis Accounting Address Affect Affinity Chromatography Aging Alleles Alzheimer disease prevention Alzheimer&apos s Disease Alzheimer&apos s disease risk Amyloid Amyloid Proteins Amyloid beta-Protein Amyloid beta-Protein Precursor Apolipoprotein E Apolipoproteins Applications Grants Biogenesis Brain Brain region C-terminal CREB1 gene Cell Line Cleaved cell DNA-Binding Proteins Deletion Mutagenesis Deposition Diet Disease Drug Delivery Systems Electrophoretic Mobility Shift Assay Enzyme Gene Enzymes Epidemiology Etiology Family Fatty acid glycerol esters Gene Expression Gene Expression Regulation Gene Proteins Generations Genes Genetic Genetic Polymorphism Genetic Transcription Genomics Goals Growth Factor Inflammatory Interleukin-1 alpha Journals Late Onset Alzheimer Disease Length Mediating Memory Metals Molecular Chaperones Nerve Degeneration Neurons Nuclear Nucleic Acid Regulatory Sequences Paper Pathogenesis Pathway interactions Peer Review Peptide Hydrolases Peptides Play Process Production Progress Reports Property Protein Precursors Proteins Proteolytic Processing Publications Publishing Regulation Regulatory Element Regulatory Pathway Research Personnel Resources Risk Role SP1 gene Site Site-Directed Mutagenesis Testing Tissues Transcriptional Regulation Transfection Tumor Necrosis Factor-alpha Up-Regulation Variant Work base brain tissue cell type cognitive function cytokine enzyme activity human TNF protein interest microbial alkaline proteinase inhibitor neuron loss novel programs promoter protein expression research study secretase transcription factor

项目摘要

DESCRIPTION (provided by applicant): Our goal is to study gene regulation in Alzheimer's disease (AD), based on the "amyloid hypothesis" of Alzheimer's disease. Overproduction of the amyloid beta-peptide (Abeta) causes a cascade of neurodegenerative steps resulting in plaque formation and neuronal loss that characterize Alzheimer's disease. The unresolved key question in the field is what factors cause overproduction of Abeta and its large Abeta precursor protein (APP). Increased Abeta production may result from an increase in APP expression, or in its proteolytic processing by the limiting beta-APP cleaving enzyme (BACE). The goals of this proposal are to investigate the transcriptional regulation of i) APP, because APP (and, hence, AP) biogenesis begins at the level of transcription, and ii) BACE gene, as Abeta overproduction may be due to increased BACE level as a result of upregulation in this gene. Specific Aims are: 1) To study the functional domains of the APP promoter and effects of different agents on its activity. We will functionally characterize the 7.9 kb APP promoter and study how intrinsic (cytokines) and extrinsic (metals) factors regulate promoter activity. Promoter will be studied by serial deletions, mutagenesis and transfection experiments in different cell types and primary neuronal cultures. 2) To identify the effects of specific factors and cvtokines common to both APP and BACE gene regulation. We will characterize the role of IL-1alpha, TNF-alpha and CREB transcription factor (TF) on 4.1kb BACE promoter activity. 3) To identify cell type-specific nuclear factors. A 30 bp novel region (-76-47) of the APP promoter contains a regulatory domain that interacts with at least two proteins, PuF and SkiP. We will test i) the candidate TFs that control APP promoter activity and ii) the status of such TF in normal and AD brain tissues using gel shift assay and DNA-affinity chromatography. 4) To characterize APP gene polymorphisms that influences the risk of late-onset Alzheimer's disease. We discovered two polymorphisms at -3829 and -1023 that may be associated with Alzheimer's disease. We will i) do functional and DNA-protein binding studies with promoter variants and ii) correlate promoter studies with levels of APP and Abeta. 5) To study the APP-5'-UTR region. APP expression is also regulated via the 5'-untranslated region (UTR). We will test a dual role for the APP5'-UTR at both transcriptional and post-transcriptional levels, and study its interaction with cytokines. Cell lines from families with characterized FAD will be analyzed for differential expression of the APP and BACE genes. Studying APP and BACE gene regulation is crucial to understand APP production leading to Aa generation. These studies should help developing suitable drug targets for the treatment of Alzheimer's disease.

描述（由申请人提供）：我们的目标是根据阿尔茨海默氏病的“淀粉样假说”研究阿尔茨海默氏病（AD）中的基因调节。淀粉样β-肽（ABETA）的过量生产会导致一系列神经退行性步骤，导致斑块形成和神经元损失，而神经元损失则是阿尔茨海默氏病的特征。该领域未解决的关键问题是哪些因素导致Abeta及其大型Abeta前体蛋白（APP）的过量生产。 ABETA产生的增加可能是由于APP表达的增加或通过限制β-APP切割酶（BACE）而导致其蛋白水解处理。该提案的目标是研究i）应用程序的转录调控，因为APP（并且，因此，AP）生物发生始于转录水平，ii）ii）乳胶基因，因为Abeta过量生产可能是由于该基因上调的乳液水平升高所致。具体目的是：1）研究应用程序启动子的功能域以及不同药物对其活性的影响。我们将在功能上表征7.9 KB应用启动子，并研究内在（细胞因子）和外在（金属）因子如何调节启动子活性。启动子将通过不同细胞类型和原发性神经元培养物中的串行缺失，诱变和转染实验来研究启动子。 2）确定APP和BACE基因调节的特定因素和CVTokines的影响。我们将表征IL-1Alpha，TNF-Alpha和Creb转录因子（TF）在4.1KB BACE启动子活性上的作用。 3）鉴定细胞类型特异性核因子。 APP启动子的30 bp新颖区域（-76-47）包含一个调节域，与至少两个蛋白质PUF和SKIP相互作用。我们将测试i）控制应用程序启动子活性的候选TF和ii）使用凝胶移位测定和DNA亲和力色谱法在正常和AD脑组织中的状态。 4）表征APP基因多态性影响后期发病的阿尔茨海默氏病风险。我们在-3829和-1023上发现了两个与阿尔茨海默氏病有关的多态性。我们将使用启动子变体进行功能性和DNA蛋白结合研究，而II）将启动子研究与APP和ABETA水平相关联。 5）研究App-5'-UTR区域。 APP表达也通过5'-非翻译区域（UTR）调节。我们将在转录和转录后水平上测试APP5'-UTR的双重作用，并研究其与细胞因子的相互作用。将分析具有特征为FAD家族的细胞系，以分析应用程序和贝丝基因的差异表达。研究应用程序和BACE基因调节对于了解导致AA生成的应用程序产生至关重要。这些研究应有助于开发适合治疗阿尔茨海默氏病的药物靶标。