Organization and function of the periactive zone

周围活动区的组织和功能

• 批准号: 10381522
• 负责人: Avital Adah Rodal
• 金额: $ 38万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-15 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10381522
• 关键词: Actins; Affect; Architecture; Biochemical Genetics; Biological; Biological Assay; Cell Adhesion; Cell Adhesion Molecules; Cell physiology; Cells; Clathrin; Complex; Cytoskeleton; Data; Disease; Drosophila genus; Endocytosis; Etiology; Event; Geometry; Goals; Homologous Gene; Image; Individual; Learning; Link; Maps; Mediating; Membrane; Methods; Microfilaments; Modeling; Monitor; Morphogenesis; Neuroglia; Neuromuscular Junction; Neurons; Neurotransmitters; Probability; Process; Property; Proteins; Recycling; Regulation; Resolution; Rest; Reticulum; Role; Site; Structure; Synapses; Synaptic Membranes; Synaptic Transmission; Synaptic Vesicles; System; Testing; Time; Work; experimental study; genetic testing; high resolution imaging; human disease; imaging modality; in vivo; mutant; nervous system disorder; particle; polymerization; presynaptic; receptor; response; segregation; spatiotemporal; synaptic function; target SNARE proteins; tool; transmission process; vesicular release

PROJECT SUMMARY The goal of this proposal is to understand the spatial and functional organization of the presynaptic periactive zone (PAZ), which is found adjacent to sites of synaptic vesicle release. The PAZ is a micron-scale structure, occupied by dozens of proteins that work together in multivalent assemblies to couple membrane remodeling to force-generating actin polymerization. Studies of PAZ proteins in many systems have suggested that these proteins act at multiple steps of the synaptic vesicle cycle as well as in other synaptic membrane functions (e.g. synaptic morphogenesis and receptor traffic). It remains unknown how the micron-scale organization and regulation of PAZ proteins direct their membrane and cytoskeleton remodeling activities to these different neuron-specific functions. We will use the Drosophila larval neuromuscular junction (NMJ), a powerful model synapse, to decipher how PAZ protein assemblies, activities, and cellular functions are linked. Using high- resolution imaging, we recently found that PAZ proteins occupy both overlapping and distinct domains within the PAZ, and that proper segregation of PAZ proteins between these domains depends on their multivalent interactions with each other. We have also recently described PAZ-dependent dynamic actin filament structures, which represent a direct readout of PAZ protein activities in these different domains. Using these tools, we will ask how synapses regulate PAZ protein activities and interactions in space and time, and how PAZ organization underlies its diverse neuron-specific functions, in response to synaptic activity and transmission. In Aim 1, we will determine how PAZ proteins are organized at resting and active synapses using complementary fixed, live, and super-resolution imaging methods, and develop new quantitative methods to describe their geometric relationships in PAZ domains. In Aim 2, we will test the hypothesis that synaptic actin patches represent clathrin-dependent synaptic vesicle recycling events, and identify the determinants of synaptic actin patch assembly and dynamics. In Aim 3, we will ask how PAZ organization and synaptic activity control diverse cell biological PAZ functions, including release site clearance and organization of cell adhesion complexes. Overall, our experiments will explain how organization of the PAZ into distinct subdomains underlies multiple functional and structural properties of synapses. PAZ proteins are implicated in multiple neurological disorders, so deciphering their in vivo functions will be critical for understanding the etiology of these human diseases.

项目摘要 该建议的目的是了解突触前周期的空间和功能组织 区域（PAZ），发现与突触囊泡释放部位相邻。 PAZ是微米尺度的结构， 被数十种蛋白质占据 产生肌动蛋白聚合。在许多系统中对PAZ蛋白的研究表明这些 蛋白质在突触囊泡周期的多个步骤以及其他突触膜功能中起作用 （例如，突触形态发生和受体流量）。微米级组织和 PAZ蛋白的调节将其膜和细胞骨架重塑活性引导到这些不同 神经元特异性功能。我们将使用果蝇幼虫神经肌肉连接（NMJ），这是一个强大的模型 突触，破译PAZ蛋白质组件，活性和细胞功能如何联系起来。使用高 分辨率成像，我们最近发现，PAZ蛋白既占据重叠和不同的领域 PAZ，以及这些域之间PAZ蛋白的适当分离取决于其多价 彼此相互作用。我们最近还描述了依赖PAZ的动态肌动蛋白丝 结构，代表了这些不同领域中PAZ蛋白活性的直接读数。使用这些 工具，我们将询问突触如何调节PAZ蛋白质活动和时空的互动，以及如何 PAZ组织是针对突触活动和 传播。在AIM 1中，我们将确定如何在使用静止和主动突触中组织PAZ蛋白 互补固定，现场和超分辨率成像方法，并开发新的定量方法 描述他们在PAZ域中的几何关系。在AIM 2中，我们将测试突触肌动蛋白的假设 斑块代表网格蛋白依赖性突触囊泡回收事件，并确定的决定因素 突触肌动蛋白斑块组件和动力学。在AIM 3中，我们将询问PAZ如何组织和突触活动 控制各种细胞生物PAZ功能，包括释放位点清除和细胞粘附的组织 复合物。总体而言，我们的实验将解释如何将PAZ组织成不同的子域 突触的多个功能和结构特性是突触的基础。 PAZ蛋白与多个有关 神经系统疾病，因此解密其体内功能对于理解的病因至关重要 这些人类疾病。