REGULATION AND PROCESSING OF AMYLOID PRECURSOR PROTEIN GENES AND GENE PRODUCTS

淀粉样前体蛋白基因和基因产物的调控和加工

• 批准号: 3745494
• 负责人: J W KUSIAK
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3745494
• 关键词: Alzheimer's disease; PC12 cells; Parkinson's disease; aging; amyloid proteins; cell death; cellular pathology; cyclic AMP; dopamine receptor; genetic promoter element; genetic regulation; genetic regulatory element; laboratory rat; molecular cloning; molecular pathology; neurotrophic factors; phenotype; plasmids; posttranslational modifications; proteolysis; retinoate; tardive dyskinesia; tissue /cell culture; transfection

The major focus of our work is on the amyloid precursor protein (APP) and the role it plays in the neuropathology of Alzheimer's Disease (AD). We also initiated a study on the regulation of the D2 dopamine receptor gene and its role in Parkinson's Disease, tardive dyskinesia and normal aging. We made three independent mutations within the APP gene (mAPPs) which are known to cause early-onset, familial AD or Hereditary Cerebral Hemorrhage with Amyloidoses-Dutch Type. Plasmids containing mAPPs were stably transfected into cell lines in order to examine phenotypic consequences to the cells and proteolytic processing of the expressed APPs. Neuronal and endothelial cells expressing mAPPs showed larger C-terminal fragments. NGF/cAMP treated PC-12 cells expressing mAPPs showed elevated levels of APP and C-terminal fragments. They had severely compacted cell bodies and eventually detached from the plates and died. We have cloned and characterized the 5' upstream regulatory region of the rat APP gene. A 375 bp fragment from this region functioned as a strong promoter in rat and human cell lines and its activity was enhanced by retinoic acid. The basal promoter activity was localized to a 75 bp sequence by deletional analysis. Finally, we isolated several DNA fragments from the functional promoter of the D2 dopamine receptor gene. Nuclear extracts from different tissues of young and old rats are being used with these promoter fragments to identify possible cis-acting transcriptional regulatory sites.

我们工作的主要重点是淀粉样蛋白（APP） 以及它在阿尔茨海默氏病神经病理学中所扮演的作用 （广告）。我们还启动了关于D2多巴胺调节的研究 受体基因及其在帕金森氏病中的作用，迟发性运动障碍 和正常衰老。我们在应用程序中进行了三个独立的突变 已知会引起早发，家族广告或 遗传性脑出血，淀粉样蛋白 - 直属型。质粒 稳定地将含有MAPP转染到细胞系中，以便 检查对细胞和蛋白水解处理的表型后果 表达的应用程序。表达MAPP的神经元和内皮细胞 显示较大的C末端碎片。 NGF/CAMP处理的PC-12细胞 表达MAPP显示APP和C端的水平升高 碎片。他们有严重压实的细胞体，最终 从盘子上脱落并死亡。我们已经克隆并描述了 大鼠应用基因的5'上游调节区域。一个375 bp的片段 该区域起着大鼠和人类细胞中强启动子的作用 视黄酸增强了线条及其活性。基础 通过缺失，启动子活性定位于75 bp序列 分析。最后，我们从 D2多巴胺受体基因的功能启动子。 核提取物 来自不同老鼠的不同组织正在使用这些 启动子片段以识别可能的顺式作用转录 监管地点。