Mechanisms of Presynaptic Maintenance in C. elegans

线虫突触前维持机制

• 批准号: 10562841
• 负责人: Wendy Herbst
• 金额: $ 6.95万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-15 至 2026-02-14
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10562841
• 关键词: Adult; Aging; Auxins; Behavioral Assay; Biochemical; Biological; Biological Models; Biotinylation; Brain; Caenorhabditis elegans; Calcium; Cells; Cellular biology; Cognition; Communication; Development; Disparity; ELK1 gene; Fluorescence; Genetic; Genetic Screening; Image; Knowledge; Label; Laboratories; Life; Longevity; Maintenance; Mammals; Mass Spectrum Analysis; Measures; Memory; Microscopy; Movement; Nerve Degeneration; Nervous System; Neurodegenerative Disorders; Neurons; Neurophysiology - biologic function; Neurosciences; Organism; Phenotype; Phosphoric Monoester Hydrolases; Phosphotransferases; Physiologic pulse; Protein Biosynthesis; Proteins; Proteomics; Role; Scaffolding Protein; Structure; Synapses; Synaptic Vesicles; System; Testing; Therapeutic; Time; Universities; Visual; Visualization; Work; candidate identification; cell type; experience; gene conservation; genetic manipulation; insight; neural circuit; preservation; presynaptic; protein degradation; synaptogenesis; therapeutic target; tool; training opportunity; ubiquitin ligase

Project Summary While a growing body of work has contributed to our understanding of synapse assembly, less is known about how synapses are maintained throughout life. This long life of synapses is crucial for the sustained function of neural circuits, including those supporting cognition, movement, and other vital functions. However, maintaining long-lived synaptic connections presents a cell biological challenge, as synaptic proteins have finite lifetimes, synaptic vesicles turnover rapidly, and protein synthesis is scarce in the presynaptic compartment. This project will study the mechanisms of presynaptic maintenance, using C. elegans as a model system. In Aim 1, I will investigate the proteins involved in maintaining synaptic structures, using the auxin-inducible degron system to remove the candidate proteins SYD-2, SYD-1, SAD-1, CDK-5, and PCT-1 from the mature nervous system. Changes in synapse organization will be assessed using endogenous, cell-type specific markers of synaptic vesicles and active zone proteins. In Aim 2, I will identify regulators of the presynaptic scaffolding protein SYD-2. I will implement a visual forward genetic screen to identify candidates that regulate SYD-2 stability, using a pulse-chase SYD-2 HaloTag approach to visualize SYD-2 turnover. In Aim 3, I will identify regulators of SYD-2 through the use of Split-TurboID proximity biotinylation to detect interacting partners of SYD-2 in the presynaptic compartment. Candidate regulators of SYD-2 turnover will be assessed using the SYD-2 HaloTag system. This work will identify key synaptic maintenance proteins and their regulators. Understanding the mechanisms that maintain stable synapses provides therapeutic avenues for preserving synapses in aging and neurodegenerative diseases. This project, performed in the laboratory of Dr. Kang Shen at Stanford University, provides a strong training opportunity for me in the fields of cell biology and neuroscience, and I will gain new experience with the C. elegans model system, genetic manipulations, and microscopy.

项目摘要 虽然越来越多的工作有助于我们理解突触 组装，很少有人知道突触是如何维持整个生命。这漫长的生命 突触对于神经回路的持续功能至关重要，包括那些支持 认知、运动和其他重要功能。然而，维持长寿命的突触 连接提出了细胞生物学挑战，因为突触蛋白具有有限的寿命， 突触囊泡更新迅速，突触前蛋白质合成很少 车厢本课题将利用C. elegans作为一个模型系统。在目标1中，我将研究参与维持 突触结构，使用生长素诱导的降解决定子系统去除候选蛋白 来自成熟神经系统的SYD-2、SYD-1、SAD-1、CDK-5和PCT-1。变化 将使用突触的内源性、细胞类型特异性标记物来评估突触组织。 囊泡和活性区蛋白。在目标2中，我将确定突触前神经元的调节因子， 支架蛋白SYD-2。我会进行一个视觉遗传筛查来识别候选人 调节SYD-2稳定性，使用脉冲追踪SYD-2 HaloTag方法来可视化SYD-2 周转在目标3中，我将通过使用Split-TurboID接近度来识别SYD-2的调节器 生物素化以检测突触前区室中SYD-2的相互作用伴侣。候选 将使用SYD-2 HaloTag系统评估SYD-2周转的调节剂。这项工作将 识别关键的突触维持蛋白及其调节因子。了解机制 保持稳定的突触提供了在老化中保持突触的治疗途径 和神经退行性疾病。该项目在Kang Shen博士的实验室进行， 斯坦福大学，为我提供了一个很好的培训机会，在细胞生物学和 神经科学，我将获得新的经验与C。elegans模型系统，遗传的 操作和显微镜。