Dynactin function in axons, synapses, and neurodegenerative disease

Dynactin 在轴突、突触和神经退行性疾病中的功能

• 批准号: 8482528
• 负责人: Thomas E. Lloyd
• 金额: $ 35.44万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-15 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8482528
• 关键词: Affect; Animal Model; Animals; Axon; Axonal Transport; Binding; Binding Proteins; Biological Assay; Body Weight decreased; Calcium; Complex; Cytoskeleton; Data; Defect; Disease; Distal; Drosophila genus; Drosophila melanogaster; Dyes; Dynein ATPase; Electron Microscopy; Electrophysiology (science); Event; Exhibits; Exocytosis; Functional disorder; Genomics; Glues; Goals; Human; Hypercapnic respiratory failure; Image; Impairment; Inherited; Intracellular Transport; Larva; Lead; Life; Light; Mediating; Mental Depression; Microtubules; Mitochondria; Morphology; Motor; Motor Neuron Disease; Mutate; Mutation; Nerve; Neurodegenerative Disorders; Neuromuscular Junction; Neurons; Neuropathy; Organelles; Parkinsonian Disorders; Peripheral Nerves; Peripheral Nervous System Diseases; Phenotype; Physiology; Play; Plus End of the Microtubule; Presynaptic Terminals; Process; Proteins; Role; Signal Transduction; Sorting - Cell Movement; Specificity; Synapses; Synaptic Transmission; Synaptic Vesicles; Syndrome; Testing; Transgenic Organisms; Transport Vesicles; Vesicle; age related; cell type; dynactin; fly; genetic manipulation; in vivo; insight; mutant; nervous system disorder; neuronal cell body; neurotransmitter release; novel; prevent; protein complex; public health relevance; retrograde transport; synaptic function; therapeutic development; trafficking

DESCRIPTION (provided by applicant): Disruption of axonal transport is thought to occur early in the course of many neurological diseases including peripheral neuropathies and neurodegenerative diseases. Proposed mechanisms whereby transport defects lead to disease include impairment of organelle delivery to distal axons, defective retrograde neurotrophic signaling, and disruption of the synaptic vesicle cycle within the synaptic terminal. However, the way alterations in axonal transport cause disease is unclear. Simple model organisms such as the fruitfly, Drosophila melanogaster, allow genetic manipulations to be combined with analysis of axonal transport and synaptic physiology, and they are providing insights into the pathophysiology of peripheral nerve and neurodegenerative diseases. The long-term goal of this proposal is to understand how intracellular trafficking events are altered in neurological diseases in order to identify novel targets for therapeutic development. Retrograde axonal transport is mediated by the dynein/dynactin protein complex. The p150Glued dynactin subunit is mutated in two distinct, non-overlapping autosomal dominant neurodegenerative diseases: one that causes a motor neuron disease called Hereditary Motor Neuropathy type 7B (HMN7B), and the other that is called Perry Syndrome, characterized by parkinsonism, depression, hypoventilation, and weight loss. The HMN7B and Perry mutations are as close as 12 residues apart within the p150Glued CAP-Gly microtubule (MT)-binding domain. Our Preliminary Results suggest that these mutations differentially affect p150 interactions with MTs and p150 function at synapses. Furthermore, we find that HMN7B mutations disrupt the initiation of retrograde MT-mediated transport at the distal-most end of synapses (called terminal boutons) and block neurotransmitter release at the neuromuscular junction (NMJ). In Aim 1, we will determine if HMN7B and Perry Syndrome mutations disrupt distinct protein interactions with p150 binding partners and also whether they cause defects in axonal transport or retrograde signaling. In Aim 2, we will study the initiation of retrograde transport at NMJ TBs to determine if initiation occur when dynamic MT plus-ends capture vesicles through interactions between p150 and end binding protein-1 (EB1), a "master regulator of MT plus-ends". We will also test the hypothesis that HMN7B but not Perry mutations in p150 disrupt retrograde initiation at TBs, and determine if this defect is due to an alteration in microtubule dynamics at TBs. In Aim3, we will investigate how mutations in p150 alter synaptic transmission by combining calcium and FM1-43 imaging, ultrastructural analysis and electrophysiology. Together these studies will help elucidate the function of the p150 CAP-Gly domain in neurons, and hold promise for shedding light on the mechanisms of cell-type specificity of neurodegenerative disease.

描述（由申请方提供）：轴突运输中断被认为发生在许多神经系统疾病（包括周围神经病变和神经退行性疾病）病程的早期。运输缺陷导致疾病的机制包括细胞器传递到远端轴突的损伤、有缺陷的逆行神经营养信号传导和突触末端内突触囊泡周期的破坏。然而，轴突运输的改变导致疾病的方式尚不清楚。简单的模式生物，如果蝇，黑腹果蝇，允许基因操作与轴突运输和突触生理学的分析相结合，它们提供了对周围神经和神经退行性疾病的病理生理学的见解。这项提案的长期目标是了解细胞内运输事件在神经系统疾病中是如何改变的 以确定治疗开发的新靶点。 逆行轴突运输由动力蛋白/动力肌动蛋白蛋白复合物介导。p150 Glued dynactin亚基在两种不同的、非重叠的常染色体显性遗传性神经退行性疾病中发生突变：一种导致称为遗传性运动神经病7 B型（HMN 7 B）的运动神经元疾病，另一种称为佩里综合征，其特征为帕金森综合征、抑郁症、换气不足和体重减轻。HMN 7 B和佩里突变在p150 Glued CAP-Gly微管（MT）结合结构域内相隔12个残基。我们的初步结果表明，这些突变差异影响p150与MT的相互作用和p150在突触的功能。此外，我们发现HMN 7 B突变破坏了突触最远端（称为终末终扣）逆行MT介导的转运的启动，并阻断了神经肌肉接头（NMJ）的神经递质释放。在目标1中，我们将确定HMN 7 B和佩里综合征突变是否会破坏与p150结合伴侣的不同蛋白质相互作用，以及它们是否会导致轴突运输或逆行信号传导的缺陷。在目标2中，我们将研究在NMJ TB的逆行转运的起始，以确定是否发生启动时，动态MT加结束捕获囊泡通过p150和末端结合蛋白-1（EB 1），一个“MT加结束的主调节器”之间的相互作用。我们还将检验p150中HMN 7 B而不是佩里突变破坏TB处的逆行起始的假设，并确定这种缺陷是否是由于TB处微管动力学的改变。在AIM 3中，我们将研究 结合钙离子和FM 1 -43成像、超微结构分析和电生理学，研究p150突变如何改变突触传递。这些研究将有助于阐明p150 CAP-Gly结构域在神经元中的功能，并有望揭示神经退行性疾病细胞类型特异性的机制。