Kinetochore Protein Functions in Synaptogenesis

动粒蛋白在突触发生中的功能

• 批准号: 10248433
• 负责人: Thomas L. Schwarz
• 金额: $ 55.38万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10248433
• 关键词: Alzheimer' s Disease; Axon; Axonal Transport; Binding; Binding Sites; Brain; Cell Nucleus; Cell division; Cells; Centromere; Chromosome Segregation; Chromosome abnormality; Chromosomes; Competence; Complement; Complex; Cytoskeleton; Defect; Dendrites; Dendritic Spines; Development; Drosophila genus; Electron Microscopy; Electrophysiology (science); Embryo; Embryonic Development; Genetic; Growth; Growth Cones; Hippocampus (Brain); Kinetochores; Knowledge; Mechanics; Microtubule Stabilization; Microtubules; Mitosis; Mitotic; Morphology; Motor Neurons; Muscle; Mutation; Nature; Neurites; Neurodegenerative Disorders; Neuromuscular Junction; Neurons; Phenotype; Plus End of the Microtubule; Post-Translational Protein Processing; Presynaptic Terminals; Process; Property; Proteins; RNA Interference; Rattus; Research; Role; Sensory; Shapes; Signal Transduction; Structure; Surface; Swelling; Synapses; System; Testing; Thinness; Vertebral column; daughter cell; density; fly; knock-down; light microscopy; mutant; neurodevelopment; novel; presynaptic; protein complex; protein function; synaptogenesis

From a Drosophila mutant screen for defects in the formation of the embryonic neuromuscular junction (NMJ), we have learned that mutations in components of the kinetochore complex are needed for the transformation of a growth cone into a correctly shaped synaptic connection. Loss of these proteins also alters the structure of sensory dendrites. In cultured mammalian neurons, kinetochore proteins also appear to guide development: their knockdown by RNAi causes an excess of filipodia like protrusions to form on hippocampal dendrites. This is a completely novel function for the kinetochore, a protein complex previously known only to function at the centromere of chromosomes where it is required in dividing cells to “catch” and stabilize spindle microtubules and thereby enable the segregation of chromosomes to the daughter cells. The neuronal phenptypes cannot be explained by defects in chromosome mechanics and therefore we hypothesize a postmitotic function for a “neuro-kinetochore”, a function that is likely to involve the same core property of the centromeric kinetochore: the ability to bind to the plus-ends of microtubules and stabilize them. We propose to test the hypothesis that a complex very much akin to that found at centromeres will function locally in post-mitotic neurons to assist in the transformation of dynamic growth cone microtubules into stable bundles of synaptic microtubules and similarly to stabilize dendritic microtubules and thereby arrest dendrite growth. The proposal makes use of the advantages of both Drosophila and mammalian systems. Aim 1 of this proposal therefore seeks to characterize in greater depth the nature of the defects at the embryonic fly NMJ and in hippocampal dendrites. Aim 2 delves into the mechanism underlying the phenotype by asking whether structure function studies support the hypothesis of a kinetochore-like structure that must bind to microtubules. Aim 3 focuses on the microtubule cytoskeleton and asks whether there are defects in microtubule dynamics and stabilization in the mutants, and how this novel role for kinetochore proteins fits into our knowledge of the processes that transform growth cones into mature endings and determine the morphology of dendrites.

通过果蝇突变体筛选胚胎神经肌肉接头（NMJ）形成的缺陷， 我们了解到，转变需要动粒复合体成分的突变 将生长锥形成正确形状的突触连接。这些蛋白质的损失也会改变 感觉树突。在培养的哺乳动物神经元中，动粒蛋白似乎也指导发育： RNAi 抑制它们会导致海马树突上形成过多的丝状伪足样突起。这 是动粒的一个全新功能，动粒是一种蛋白质复合物，以前只知道在动粒上发挥作用 染色体的着丝粒，分裂细胞需要它“捕获”并稳定纺锤体微管 从而使染色体能够分离到子细胞。神经元表型不能 可以用染色体力学的缺陷来解释，因此我们假设有丝分裂后功能 “神经着丝粒”，一种可能涉及着丝粒着丝粒相同核心特性的功能： 与微管正端结合并稳定它们的能力。我们建议检验以下假设： 与着丝粒中发现的非常相似的复合物将在有丝分裂后神经元中局部发挥作用，以协助 将动态生长锥微管转化为稳定的突触微管束等 稳定树突微管，从而阻止树突生长。该提案利用了 果蝇和哺乳动物系统的优点。因此，本提案的目标 1 旨在描述 更深入地了解果蝇胚胎 NMJ 和海马树突缺陷的性质。瞄准2个洞 通过询问结构功能研究是否支持表型背后的机制 必须与微管结合的类似动粒结构的假设。目标 3 关注微管 细胞骨架并询问突变体的微管动力学和稳定性是否存在缺陷，以及 动粒蛋白的这种新作用如何适应我们对改变生长过程的了解 锥体进入成熟的末端并确定树突的形态。