Roles of Recycling Endosomes in Neuronal Extracellular Vesicle Cargo Traffic

回收内体在神经元细胞外囊泡货物运输中的作用

• 批准号: 10584339
• 负责人: Avital Adah Rodal
• 金额: $ 47.48万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-12-01 至 2027-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10584339
• 关键词: Acute; Alzheimer' s Disease; Behavior; Binding; Biochemistry; Biogenesis; Brain; Carrier Proteins; Cell membrane; Cells; Cellular Membrane; Chemicals; Clathrin; Communication; Complex; Data; Defect; Disease; Drosophila genus; Endocytosis; Endosomes; Event; Foundations; Genetic; Goals; Human; Knowledge; Link; Lipids; Mediating; Membrane; Membrane Protein Traffic; Microscopy; Monitor; Morphogenesis; Morphology; Nature; Nerve; Nervous System; Nervous System Physiology; Neurodegenerative Disorders; Neuroglia; Neurons; Nucleic Acids; Parkinson Disease; Pathologic; Pathology; Pathway interactions; Phosphotransferases; Physiological; Plasma; Play; Population; Presynaptic Terminals; Process; Proteins; Receptor Signaling; Recycling; Regulation; Research; Resolution; Role; Route; SYNJ1 gene; Signal Transduction; Sorting; Synapses; Synaptic Membranes; Synaptic Transmission; Testing; Therapeutic; Vesicle; Work; cell type; experimental study; extracellular vesicles; genetic approach; imaging approach; in vivo; insight; intercellular communication; live cell imaging; mutant; nervous system disorder; novel; optogenetics; presynaptic; rab11 protein; response; retrograde transport; scaffold; tool; trafficking; vesicular release

PROJECT SUMMARY The goal of this proposal is to understand how cellular membrane trafficking machinery controls the packaging and release of extracellular vesicle (EV) cargoes from synapses in vivo. EVs are small membrane-bound vesicles released by numerous cell types, including neurons, carrying cargoes critical for signaling and disease. However, we understand very little about how EV cargo traffic is spatially and temporally regulated within the polarized and complex morphology of neurons. We have developed tools to track and manipulate EV traffic at Drosophila presynaptic terminals in vivo, and discovered that flux of cargoes through a plasma membrane-recycling endosome route determines whether they are locally sorted for packaging and release in EVs, rather than depleted from synapses by retrograde transport. Recycling endosomes have primarily been studied in non-neuronal cells, and very little is known about their lifetime, functions, or dynamics at presynaptic terminals. We do know that recycling endosomes play critical roles in signaling, neuronal morphogenesis, EV traffic, and synaptic transmission. Understanding and therapeutically intervening in these important processes will require a deeper knowledge of the mechanisms of neuronal recycling endosome function. In this proposal, we will elucidate the mechanisms of synaptic EV cargo and recycling endosome traffic in vivo. To achieve these goals, we will use Drosophila genetics, biochemistry, high-resolution microscopy, and live cell imaging. 1) We will determine the functions, dynamics, and regulation of different types of synaptic recycling endosomes. To this end, we will develop new tools and approaches to define and control functionally distinct recycling compartments at synapses. Using these tools, we will test novel mechanistic hypotheses for how membrane traffic machinery sorts cargoes at synaptic recycling compartments. 2) We will determine how EV cargo traffic depends on distinct modes of synaptic endocytosis: clathrin-mediated endocytosis, which operates under low neuronal activity and activity-dependent bulk endocytosis, which operates during intense neuronal activity. These experiments will ascertain if EV fate is determined by different modes of internalization, how recycling endosomes contribute to these functions, and provide new mechanisms to link activity, endosomal traffic, and EV release. Given the conserved nature of synaptic membrane trafficking machinery, our findings and tools will lay the foundation for new insights into EV traffic in many aspects of nervous system function, including in human neurological disease.

项目摘要 该提案的目的是了解细胞膜贩运机械如何控制包装 并释放体内突触中的细胞外囊泡（EV）货物。电动汽车是小膜结合的 由许多细胞类型释放的囊泡，包括神经元，载有对信号传导至关重要的货物 疾病。但是，我们对电动汽车货物流量如何在空间和时间上进行监管一无所知 在神经元的两极化和复杂的形态中。我们开发了跟踪和操纵电动汽车的工具 果蝇的交通在体内突触前终端，发现货物的通量通过等离子体 膜回收的内体路线确定它们是否本地分类以包装并释放 电动汽车，而不是通过逆行运输从突触中耗尽。回收内体主要是 在非神经元细胞中研究，对它们的寿命，功能或动力学在突触前的动力学知之甚少 终端。我们确实知道，回收内体在信号传导，神经元形态发生，EV中起关键作用 流量和突触传输。理解并在这些重要过程中介入 将需要更深入地了解神经元回收内体功能的机制。在此提案中， 我们将阐明体内突触EV货物和回收内体流量的机制。实现 这些目标，我们将使用果蝇遗传学，生物化学，高分辨率显微镜和活细胞成像。 1）我们将确定不同类型的突触回收的功能，动力学和调节 内体。为此，我们将开发新的工具和方法来定义和控制功能不同 突触的回收室。使用这些工具，我们将测试新的机械假设 膜交通机械在突触回收室中分类货物。 2）我们将确定EV 货物交通取决于突触内吞作用的不同模式：网格蛋白介导的内吞作用，可运作 在较低的神经元活性和活性依赖性体积内吞作用下，在强烈的神经元中起作用 活动。这些实验将确定ev命运是否由不同的内在化模式确定 回收内体有助于这些功能，并提供新的机制来连接活动，内体 流量和电动汽车释放。鉴于突触膜贩运机械的保守性质，我们的发现 工具将在神经系统功能的许多方面对EV流量的新见解奠定基础， 包括人类神经疾病。