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

基本信息

批准号：
9022533
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
项目类别：
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/9022533
关键词：
Affect Animal Model Animals Axon Axonal Transport Binding Binding Proteins Biological Assay Body Weight decreased Calcium Cells 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 Health Human Hypercapnic respiratory failure Image Impairment Inherited Intracellular Transport Knock-in 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 Vesicle Vesicle Transport Pathway age related cell type dynactin fly genetic manipulation in vivo insight mutant nervous system disorder neurotransmission neurotransmitter release novel prevent protein complex retrograde transport synaptic function therapeutic development trafficking

项目摘要

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（由申请人提供）：轴突运输的破坏被认为发生在许多神经系统疾病（包括周围神经病和神经退行性疾病）的早期。运输缺陷导致疾病的机制包括细胞器向远端轴突的传递受损、逆行神经营养信号传导缺陷以及突触末端内突触小泡循环的破坏。然而，轴突运输的改变导致疾病的方式尚不清楚。果蝇、果蝇等简单的模型生物可以将基因操作与轴突运输和突触生理学分析相结合，为周围神经和神经退行性疾病的病理生理学提供见解。该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如何改变以确定治疗开发的新靶点。逆行轴突运输由动力蛋白/动力蛋白复合物介导。 p150Glued dynactin 亚基在两种不同的、非重叠的常染色体显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 7B 型 (HMN7B) 的运动神经元疾病，另一种称为佩里综合征，其特征为帕金森病、抑郁症、通气不足和体重减轻。 HMN7B 和 Perry 突变在 p150Glued CAP-Gly 微管 (MT) 结合域内相距 12 个残基。我们的初步结果表明，这些突变对 p150 与 MT 的相互作用以及 p150 在突触的功能产生不同的影响。此外，我们发现 HMN7B 突变会破坏突触最远端（称为末端 boutons）逆行 MT 介导的运输的启动，并阻止神经肌肉接头（NMJ）处的神经递质释放。在目标 1 中，我们将确定 HMN7B 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缺陷。在目标 2 中，我们将研究 NMJ TB 逆行转运的启动，以确定当动态 MT plus 末端通过 p150 和末端结合蛋白 1 (EB1) 之间的相互作用捕获囊泡时是否会发生启动，EB1 是“MT plus 的主调节因子”。结束”。我们还将检验以下假设：p150 中的 HMN7B 突变（而非 Perry 突变）会破坏 TB 处的逆行起始，并确定这种缺陷是否是由于 TB 处微管动力学的改变所致。在 Aim3 中，我们将调查 p150 突变如何通过结合钙和 FM1-43 成像、超微结构分析和电生理学来改变突触传递。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。