Fascin1 in Growth Cone Motility and Guidance

Fascin1 在生长锥运动和指导中的作用

基本信息

批准号：
10606165
负责人：
Katherine Rebecca Hardin
金额：
$ 4.77万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-05-15 至 2025-05-14
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10606165
关键词：
Actins Adult Axon Biological Assay Biological Models Brain Brain Diseases Bundling CRISPR/Cas technology Cell Culture Techniques Cells Central Nervous System Cues Cultured Cells Cytoskeleton Data Defect Development Developmental Process Drosophila genus Embryonic Development Environment Epilepsy F-Actin Fiber Filament Filopodia Fingers Genes Genetic Growth Cones Hippocampus Image Imaging Techniques Knock-out Knowledge Label Link Lobe Membrane Memory Microfilaments Modeling Molecular Molecular Biology Molecular and Cellular Biology Morphology Movement Mushroom Bodies Neurons Olfactory Learning Orthologous Gene Phosphorylation Process Protein Kinase C Proteins RNA Interference Rattus Regulation Role Sampling Sensory Side Signal Transduction Structure System Testing Traction Training Work autism spectrum disorder axon growth axon guidance axonal pathfinding cell motility extracellular fascin fly in vivo insight migration mutant nervous system disorder neural network neuronal circuitry pharmacologic protein crosslink receptor response spatiotemporal training opportunity

Actins Adult Axon Biological Assay Biological Models Brain Brain Diseases Bundling CRISPR/Cas technology Cell Culture Techniques Cells Central Nervous System Cues Cultured Cells Cytoskeleton Data Defect Development Developmental Process Drosophila genus Embryonic Development Environment Epilepsy F-Actin Fiber Filament Filopodia Fingers Genes Genetic Growth Cones Hippocampus Image Imaging Techniques Knock-out Knowledge Label Link Lobe Membrane Memory Microfilaments Modeling Molecular Molecular Biology Molecular and Cellular Biology Morphology Movement Mushroom Bodies Neurons Olfactory Learning Orthologous Gene Phosphorylation Process Protein Kinase C Proteins RNA Interference Rattus Regulation Role Sampling Sensory Side Signal Transduction Structure System Testing Traction Training Work autism spectrum disorder axon growth axon guidance axonal pathfinding cell motility extracellular fascin fly in vivo insight migration mutant nervous system disorder neural network neuronal circuitry pharmacologic protein crosslink receptor response spatiotemporal training opportunity

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项目摘要

PROJECT SUMMARY: The formation of the brain’s intricate neural network begins in embryogenesis. Axon guidance is a critical developmental stage in which precise wiring of the central nervous system is achieved when axonal fibers connect with specific target cells. Errors in axon guidance can result in wiring defects that are associated with neurological disorders including epilepsy. The tips of axons have highly motile structures called growth cones that can sense and respond to extracellular guidance cues to direct axon migration. Growth cones depend on actin-based structures called filopodia to sense their surrounding environment and detect external guidance cues. The formation of filopodia is dependent on the bundling of parallel actin filaments by actin cross-linking proteins including Fascin1. During the guidance response, growth cone filopodia undergo remodeling, stabilizing in the direction of attractive cues and collapsing in response to repulsive cues. I hypothesize that the loss of Fascin1 in developing hippocampal neurons will result in erroneous axon guidance due to an inability to regulate filopodia remodeling. The studies proposed here will utilize molecular, cellular biology, and imaging techniques to study the role and regulation of Fascin1 in axon growth cones using cultured rat hippocampal neurons and an in vivo Drosophila model. In Aim 1.1, the effects of Fascin1 depletion via CRISPR-Cas9 editing on filopodia extension, persistence, and retraction will be studied. Three different axon guidance assays will be utilized to determine if Fascin1 depletion alters the ability of growth cones to undergo the guidance response. Aim 1.2 will study the spatiotemporal dynamics of Fascin1 in growth cones of cultured rat hippocampal neurons undergoing guided migration and the regulation of neuronal Fascin1 by protein kinase C (PKC). Previous work established that phosphorylation of Ser-39 of Fascin1 by PKC abrogates Fascin1’s filament bundling ability and I hypothesize that PKC regulates growth cone filopodia stability during axon guidance via this role. Aim 2 uses a Drosophila- based approach to study the role of Fascin1 in axon guidance in vivo. Drosophila express an single ortholog of mammalian Fascin1 called Singed and my preliminary studies indicate that the presence of Singed is required for proper formation of the mushroom body, a neuronal structure that is dependent on axon guidance to form. The studies proposed here will further the knowledge of cytoskeletal regulation during axon guidance which is of importance due to the association of errors in axon guidance with neurological disorders, including epilepsy.

项目摘要：大脑错综复杂的神经网络的形成始于胚胎发生。轴突指导是关键发育阶段，当轴突纤维时，可以实现中枢神经系统的精确接线与特定的目标细胞连接。轴突指导中的错误可能导致与之相关的接线缺陷神经系统疾病，包括癫痫。轴突的尖端具有称为生长锥的高通力结构这可以感知并响应细胞外引导提示，以指导轴突迁移。生长锥取决于基于肌动蛋白的结构称为丝状虫，以感知其周围环境并检测外部指导提示。丝状形成取决于肌动蛋白交联的平行肌动蛋白丝的束包括fascin1的蛋白质。在指导响应期间，生长锥丝状疾病进行重塑，稳定在有吸引力的提示和响应排斥线索的方向上。我假设失去 fascin1在发展海马神经元时会导致错误的轴突指导，因为无法调节丝状重塑。这里提出的研究将利用分子，细胞生物学和成像技术使用培养的大鼠海马神经元和一个体内果蝇模型。在AIM 1.1中，Fascin1通过CRISPR-CAS9编辑耗尽对丝状扩展，持久性，缩回将进行研究。将利用三种不同的轴突指导测定法来确定fascin1是否耗尽会改变生长锥的能力进行指导反应。 AIM 1.2将研究 Fascin1在培养的大鼠海马神经元的生长锥中的时空动力学。蛋白激酶C（PKC）迁移和神经元fascin1的调节。以前的工作确定了 PKC对fascin1的Ser-39磷酸化消除了Fascin1的细丝捆绑能，我假设 PKC通过此角色在轴突指导期间调节生长锥丝状稳定性。 AIM 2使用果蝇 - 基于研究fascin1在体内轴突指导中的作用的方法。果蝇表达一个单一的直系同源哺乳动物Fascin1称为Singed，我的初步研究表明需要唱歌的存在为了正确形成肌肉体，一种依赖轴突指导形成的神经元结构。这里提出的研究将进一步了解轴突指导期间细胞骨架调节的了解由于轴突引导与神经系统疾病的误差的关联，包括癫痫病的重要性。