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）：淀粉样蛋白前体蛋白（APP）被认为在阿尔茨海默氏病的发病机理中起着核心作用。在哺乳动物中，APP具有两个淀粉样蛋白前体，例如蛋白（APLP）同源物，并且该蛋白质家族的生理功能仍然很熟悉。 APP的直系同源物不存在于酵母中，而是从蠕虫到人的保守，这表明它们在细胞间交流中提供了基本作用。在初步数据中，我们证明了具有遗传缺失或APLP2的遗传缺失的小鼠线表现出嗅觉感觉神经元（OSN）的轴突投影的遗迹，表明这些蛋白质在建立和/或维持嗅觉神经循环的精确接线图中的重要作用。从孤立的OSN得出的下一代测序数据表明APLP2和APP在OSN中高度表达（分别排名第26和＃29，分别为27,390个带注释的转录本）。我们最近发表了BACE1蛋白酶的功能丧失，Bace1蛋白酶是一种关键蛋白酶，该蛋白酶处理APP家族的蛋白质以及许多其他轴突引导分子，也导致小鼠嗅觉神经回路中的连通性误差。在这里，我们在生理上提出了该应用程序和APLP2在生理上的函数，以形成和维护小鼠嗅觉神经回路的精确轴突投影图。该假设得到了其他观察结果的支持。 APP与许多粘附分子以及在轴突引导中具有确定作用的蛋白质相关联，例如Netrin和Contactin 4。的确，这些相关蛋白中的许多相关蛋白也在OSN中表达，我们的下一代测序数据也证明了这些相关的蛋白质。或者，APP和APLP2的剪接细胞外结构域可以通过硫酸软骨素后翻译后修饰。另一种剪接的APLP2同工型在OSN中高水平表达。另外，APP和APLP2的细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驱除或吸引生长锥的环境线索，并存在于嵌入嗅觉神经回路的组织中。为了测试APP和APLP2对轴突投影的作用，我们部署了开发的遗传工具的组合，以操纵鼠标嗅觉电路，以检查OSN中专用APP和/或APLP2的选择性缺失的后果（AIM 1）。此外，我们利用APP和APLP2的高度表达来纯化和识别与App和/或APLP2相关的蛋白质（AIM 2）。总之，从这些研究中获得的知识将提高我们对这一重要蛋白质家族的生理功能的理解，并可以深入了解阿尔茨海默氏病的发病机理，并带来对阿尔茨海默氏病正在开发的疗法的不良后果，以改变APP的水平及其裂解产物。