CORE--SCIENTIFIC

淀粉样前体蛋白（APP）的代谢命运受到调节 蛋白质磷酸化。 蛋白激酶C（PKC）激活，或 抑制蛋白磷酸酶-1（PP-1）刺激APP加工 通过非淀粉样蛋白生成的α-分泌酶途径。 更大 了解控制信号转导通路的 APP的细胞内运输和加工，并控制 A-β淀粉样蛋白（A/β）的产生和分泌可能导致 用于治疗阿尔茨海默病的新的治疗靶点。 本计划项目提出了一种多学科方法， 定义APP的细胞作用和 APP代谢的磷酸化依赖性调节。 研究将 在组织复杂性的几个层次上执行，包括 用纯化分子进行体外生物化学研究，细胞生物学 探索细胞内运输事件的系统， 完整动物分析APP代谢的变化， 特异性PP-1亚型和两种内源性抑制剂的靶向缺失 的PP-1 核心B旨在提供一系列技术支持服务， 项目的其他成员。 中央集权组织 支持功能将促进可靠，高效和成本- 确保物资供应充足的有效手段。 芯会 负责准备和维护充足的供应品 关键试剂（特异性目标1），包括纯化的蛋白激酶， 蛋白磷酸酶和这些酶的蛋白和肽抑制剂。 核心小组将提供咨询意见，并协调利用 蛋白质化学支持功能，如肽合成，蛋白质 测序和质谱。 APP的结构域特异性抗体 及其代谢产物已成为研究APP的重要工具 处理. 核心将保持目前使用的抗体库存， 并将产生额外的多克隆和磷酸化状态特异性 抗体，并协助开发其使用的测定方法 （具体目标2）。 核心将维持基因敲除小鼠的群体 并提供拟用于研究的所需动物数量 项目3（具体目标3）。 这些将包括基因敲除小鼠， 磷酸酶抑制剂-1（已生成）、抑制剂-2（待生成 在项目3中）、PP-1 α和PP-1 γ（由其他机构提供 合作者）。 l-1/l-2和PP-1 α/γ的“双重”敲除将 通过杂交繁殖产生。

The metabolic fate of the amyloid precursor protein (APP) is regulated by protein phosphorylation. Activation of protein kinase C (PKC), or inhibition of protein phosphatase-1 (PP-1) stimulates processing of APP through the non-amyloidogenic, alpha-secretase pathway. A greater understanding of the signal transduction pathways that control the intracellular trafficking and processing of APP, and govern the production and secretion of the A-beta amyloid protein (A/beta), may lead to novel therapeutic targets for the treatment of Alzheimer's disease. A multi-disciplinary approach is proposed in this Program Project to define the cellular role of APP and the molecular basis of the phosphorylation-dependent regulation of APP metabolism. Studies will be performed at several levels of organizational complexity, encompassing in vitro biochemical studies with purified molecules, cell biological systems to explore intracellular trafficking events, and studies in intact animals analyzing changes in APP metabolism resulting from targeted deletion of specific PP-1 isoforms and two endogenous inhibitors of PP-1 Core B is designed to provide a range of technical support services to the other members of the Program Project. The centralized organization of support functions will facilitate a reliable, efficient, and cost- effective means to ensure an adequate supply of materials. The Core will be responsible for the preparation and maintenance of adequate supplies of key reagents (Specific Aim 1), including purified protein kinases, protein phosphatases and protein and peptide inhibitors of these enzymes. The Core will provide advice and coordinate the utilization of other protein chemical support functions, such as peptide synthesis, protein sequencing, and mass spectroscopy. Domain-specific antibodies for APP and its metabolites have been essential tools for the study of APP processing. The Core will maintain stocks of antibodies in current use, and will produce additional polyclonal and phosphorylation state-specific antibodies and assist in the development of assays for their use (Specific Aim 2). The Core will maintain the colonies of knock-out mice and provide the required number of animals that are proposed for study in Project 3 (Specific Aim 3). These will include the knock-out mice for phosphatase inhibitor-1 (already created), inhibitor-2 (to be produced in project 3), PP-1alpha, and PP-1gamma (to be provided by other collaborators). "Double" knock-outs of l-1/l-2 and PP-1alpha/gamma will be produced by cross-breeding.