Kinetochore Protein Functions in Synaptogenesis

动粒蛋白在突触发生中的功能

• 批准号: 10017352
• 负责人: Thomas L. Schwarz
• 金额: $ 54.47万
• 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/10017352
• 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.

从果蝇突变体筛选中，在形成胚胎神经肌肉连接（NMJ）的缺陷中， 我们了解到，转化需要动力学复合体组件中的突变 生长锥到正确形状的突触连接中。这些蛋白质的丧失也改变了 感官树突。在培养的哺乳动物神经元中，动力学蛋白似乎也指导开发： 它们被RNAi击倒会导致过量的菲律宾像突起在海马树突上形成。这 是动力学的完全新颖的功能，这是一种以前已知的蛋白质复合物 染色体的着丝粒，在分裂细胞以“捕获”和稳定纺锤微管时需要它 从而使染色体隔离到子细胞。神经元现象不能 可以通过染色体力学的缺陷来解释，因此我们假设A “ Neuro-Kinetochore”，该功能可能涉及centromeric kinetochore的相同核心特性： 与微管的加末端结合并稳定它们的能力。我们建议检验以下假设 相似的复合物类似于在丝粒处发现的复合物将在有丝分裂后神经元中局部发挥作用，以帮助 将动态生长锥微管转化为稳定的合成微管束，并类似地转化 稳定树突微管，从而阻止树突生长。该提案利用 果蝇和哺乳动物系统的优势。因此，本提案的目标1试图表征 在更深入的胚胎蝇NMJ和海马树突中缺陷的性质。目标2钻探 通过询问结构功能研究是否支持表型的基础机制 必须与微管结合的动力学样结构的假设。 AIM 3专注于微管 细胞骨架并询问微管动力学和突变体的稳定性缺陷，并且 动力学蛋白的这种新颖作用如何符合我们对改变增长的过程的知识 锥体成熟的结尾并确定树突的形态。