ACTIVITY OF THE ALZHEIMERS DISEASE PRESENIL IN PROTEIN

描述(摘自申请人的摘要)：本报告中提出的研究 申请旨在继续申请人的遗传和分子研究 早老素功能及其在细胞内转运和转运中的作用 其底物蛋白质的蛋白水解性加工。突变型人早老素 影响淀粉样前体蛋白(APP)的蛋白分解，导致 神经毒性淀粉样多肽在阿尔茨海默病中的加速积累 疾病。在模式生物线虫和果蝇中，早老素是 Notch/LIN-12发育信号所必需的。老年前期患者最近 已被证明调节Notch受体过程中的蛋白分解处理事件 可能类似于依赖早老素的成熟和信号转导 APP在阿尔茨海默病中的裂解。最后，早衰者也被 与两种哺乳动物对凋亡刺激的细胞反应有关 细胞和果蝇。 将使用新分离的早老素在体内进行马赛克组织研究 基因变种人。初步实验已经揭示了整合素样表型 突变组织克隆，促使申请者分析 用遗传学和生化方法研究早老素对整合素的切割作用 以前已经被用来演示早老素对 凹槽加工。这些研究可能揭示早衰的共同特征 底物，并导致更好地理解特定的途径 蛋白质加工受早老素控制。这项提议的一个核心目标是 开发一系列新的分子探针来解剖Notch 以比目前可能的分辨率高得多的分辨率进行处理。这些 试剂，包括新的抗体和表位标记的构建体，可以 区分Notch裂解产物，将与突变相结合 和蛋白质降解抑制研究以确定Notch的生化步骤 涉及早老素的加工。基因和分子筛查 还将执行早老素相互作用的因素，利用 申请人最近发现，早老素保守的C-末端是一个 关键功能领域。最后，详细的平行研究 Notch和其他蛋白质的运输将在缺乏的组织中进行 要么是早老素，要么是另一种具有亚细胞效应的已知蛋白质 贩卖，SERCA类型的钙-ATPase。这些实验成为可能 通过申请人最近分离的钙-ATPase突变体，他们将 解决蛋白质是否需要早老素这一悬而未决的问题 贩卖或仅仅是蛋白质分解。这里提出的研究将澄清 早老素在Notch、APP等食品加工中的生化活性 蛋白质，并可能最终增加我们对分子原因的理解 阿尔茨海默氏症。

DESCRIPTION (From the applicant's abstract): Research proposed in this application seeks to continue the applicant's genetic and molecular studies on Presenilin function and its role in the intracellular trafficking and proteolytic processing of its substrate proteins. Mutant human Presenilins influence the proteolysis of amyloid precursor protein (APP), resulting in an accelerated accumulation of the neurotoxic amyloid peptides during Alzheimer's disease. In the model organisms Caenorhabditis and Drosophila, Presenilins are required for Notch/Lin-12 developmental signaling. Presenilins have recently been shown to regulate proteolytic processing events during Notch receptor maturation and signaling that may be analogous to the Presenilin-dependent cleavages of APP in Alzheimer's disease. Finally, Presenilins have also been implicated in the cellular response to apoptotic stimuli in both mammalian cells and Drosophila. Mosaic tissue studies will be performed in vivo using newly isolated Presenilin gene mutants. Preliminary experiments have revealed integrin-like phenotypes in the mutant tissue clones, prompting the applicants to analyze the role of Presenilin in integrin cleavage using the genetic and biochemical approaches that have been used previously to demonstrate the effects of Presenilin on Notch processing. These studies may reveal shared feature of Presenilin substrates and lead to a better understanding of the specific pathway of protein processing controlled by Presenilin. A central goal of this proposal is to develop an extensive collection of new molecular probes to dissect Notch processing at much higher resolution than is currently possible. These reagents, including new antibodies and epitope-tagged constructs that can discriminate among Notch cleavage products, will be combined with mutational and proteolysis inhibition studies to identify the biochemical steps of Notch processing that involve Presenilin. Genetic and molecular screens for Presenilin-interacting factors will also be performed, taking advantage of the applicant's recent finding that the conserved C-terminus of Presenilin is a crucial functional domain. Finally, detailed parallel studies on the trafficking of Notch and other proteins will be undertaken in tissues lacking either Presenilin or another protein with known effect of subcellular trafficking, the SERCA-type Calcium-ATPase. These experiments are made possible by the applicant's recent isolation of Calcium-ATPase mutants, and they will address the unresolved issue of whether Presenilin is required for protein trafficking or only proteolysis. The studies proposed here will clarify the biochemical activity of Presenilin in the processing of Notch, APP and other proteins, and may ultimately increase our understanding of the molecular causes of Alzheimer's disease.