Physiologic Mechanisms of Action of APP and APLP2 in Axon Targeting

APP 和 APLP2 在轴突靶向中作用的生理机制

• 批准号: 8729518
• 负责人: MARK W ALBERS
• 金额: $ 20.56万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-15 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8729518
• 关键词: Adverse effects; Afferent Neurons; Alzheimer' s Disease; Amyloid; Amyloid beta-Protein Precursor; Anatomy; Axon; Binding; Biochemical; Biological Models; Brain; Cell Adhesion Molecules; Chondroitin Sulfates; Clinical; Cues; Data; Defect; Development; Exhibits; Extracellular Domain; Genes; Genetic; Growth Cones; Heparan Sulfate Proteoglycan; Heparitin Sulfate; Homologous Gene; Homologous Protein; Human; Immunoprecipitation; In Vitro; Intervention; Investigation; Knock-out; Knowledge; Maintenance; Mammals; Maps; Mus; Neuromuscular Junction; Olfactory Epithelium; Orthologous Gene; Pathogenesis; Patients; Peptide Hydrolases; Peripheral; Physiological; Play; Process; Protein Family; Protein Isoforms; Proteins; Proteoglycan; Proteomics; Publishing; RNA Splicing; Relative (related person); Reporting; Role; Series; Sorting - Cell Movement; System; Testing; Tissue Embedding; Transcript; Yeasts; adverse outcome; axon guidance; beta-site APP cleaving enzyme 1; contactin; in vivo; insight; intercellular communication; loss of function; member; neural circuit; next generation sequencing; olfactory bulb; pre-clinical; protein expression; public health relevance; research study; tool; transcriptome sequencing

DESCRIPTION (provided by applicant): The amyloid precursor protein (APP) is thought to play a central role in the pathogenesis of Alzheimer's disease. In mammals, APP has two amyloid precursor like protein (APLP) homologs, and the physiological functions of this family of proteins remain poorly understood. Orthologs of APP are not present in yeast, but are conserved from worms to humans, suggesting they provide a fundamental role in intercellular communication. In preliminary data, we demonstrate that mouse lines with genetic deletion of either APP or APLP2 exhibit a loss of fidelity of axon projections of olfactory sensory neurons (OSNs), indicating an essential role for these proteins in the establishment and/or maintenance of the precise wiring diagram of the olfactory neural circuit. Next generation sequencing data derived from isolated OSNs indicates that APLP2 and APP are highly expressed in OSNs (ranked #26 and #29, respectively, of 27,390 annotated transcripts). We recently published that loss of function of the BACE1 protease, a key protease that processes proteins of the APP family as well as many other axon guidance molecules, also leads to connectivity errors in the mouse olfactory neural circuit. Here, we propose that APP and APLP2 function physiologically in the formation and maintenance of the precise axon projection map of the mouse olfactory neural circuit. This hypothesis is supported by several observations from others. APP associates with numerous adhesion molecules as well as proteins with established roles in axon guidance, such as netrin and contactin 4. Indeed, many of these associated proteins are expressed in OSNs, too, as evidenced by our next generation sequencing data. Alternatively spliced extracellular domains of APP and APLP2 can be modified posttranslationally by chondroitin sulfate. This alternatively spliced APLP2 isoform is expressed at high levels in OSNs. In addition, the extracellular domains of APP and APLP2 bind heparan sulfate in vitro. Both of these proteoglycans contribute to the environmental cues that repel or attract growth cones and are present in tissue embedding the olfactory neural circuit. To test the role of APP and APLP2 to precisely map axon projections, we deploy a combination of genetic tools developed to manipulate the mouse olfactory circuit to examine the consequences of selective deletion of APP and/or APLP2 exclusively in OSNs (Aim 1). Furthermore, we take advantage of the high levels of expression of APP and APLP2 to purify and identify proteins associated with APP and/or APLP2 derived from mouse OSNs (Aim 2). Together, the knowledge gained from these studies will advance our understanding of the physiologic function of this important protein family and may provide insight into the pathogenesis of Alzheimer's disease as well as into adverse consequences of the therapies under development for Alzheimer's disease to alter the levels of APP and its cleavage products.

描述（由申请人提供）：淀粉样前体蛋白（APP）被认为在阿尔茨海默病的发病机制中发挥核心作用。在哺乳动物中，APP 有两种淀粉样前体样蛋白 (APLP) 同源物，但对该蛋白家族的生理功能仍知之甚少。 APP 的直系同源物不存在于酵母中，但从蠕虫到人类都是保守的，这表明它们在细胞间通讯中发挥着重要作用。在初步数据中，我们证明具有 APP 或 APLP2 基因缺失的小鼠品系表现出嗅觉感觉神经元 (OSN) 轴突投射保真度的丧失，表明这些蛋白质在建立和/或维持精确嗅觉的过程中发挥着重要作用。嗅觉神经回路接线图。来自分离的 OSN 的下一代测序数据表明 APLP2 和 APP 在 OSN 中高度表达（在 27,390 个带注释的转录本中分别排名第 26 和第 29）。我们最近发表了 BACE1 蛋白酶（一种处理 APP 家族蛋白质以及许多其他轴突引导分子的关键蛋白酶）功能丧失也会导致小鼠嗅觉神经回路的连接错误。在这里，我们提出 APP 和 APLP2 在小鼠嗅觉神经回路精确轴突投影图的形成和维持中发挥生理功能。这一假设得到了其他人的一些观察结果的支持。 APP 与许多粘附分子以及在轴突引导中具有既定作用的蛋白质相关联，例如 netrin 和 contactin 4。事实上，正如我们的下一代测序数据所证明的那样，许多这些相关蛋白也在 OSN 中表达。 APP 和 APLP2 的选择性剪接胞外结构域可以通过硫酸软骨素进行翻译后修饰。这种选择性剪接的 APLP2 同工型在 OSN 中高水平表达。此外，APP 和 APLP2 的胞外结构域在体外结合硫酸乙酰肝素。这两种蛋白聚糖都有助于排斥或吸引生长锥的环境信号，并且存在于嵌入嗅觉神经回路的组织中。为了测试 APP 和 APLP2 在精确绘制轴突投影方面的作用，我们部署了一系列用于操纵小鼠嗅觉回路的遗传工具，以检查仅在 OSN 中选择性删除 APP 和/或 APLP2 的后果（目标 1）。此外，我们利用 APP 和 APLP2 的高水平表达来纯化和鉴定与源自小鼠 OSN 的 APP 和/或 APLP2 相关的蛋白质（目标 2）。总之，从这些研究中获得的知识将增进我们对这一重要蛋白质家族的生理功能的理解，并可能深入了解阿尔茨海默病的发病机制，以及正在开发的阿尔茨海默病疗法的不良后果，以改变阿尔茨海默病的水平APP及其裂解产物。