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（由申请人提供）：轴突运输的破坏被认为发生在许多神经系统疾病的早期，包括周围神经病和神经退行性疾病。提出的机制导致疾病的运输缺陷包括细胞器递送到远端轴突的损害，有缺陷的逆行神经营养信号传导以及突触末端内突触囊泡周期的破坏。但是，轴突运输导致疾病的改变的方式尚不清楚。简单的模型生物，例如果蝇，果蝇黑色素蛋白酶，允许遗传操作与轴突运输和突触生理学的分析结合使用，它们正在为外围神经和神经退行性疾病的病理生理提供见解。该提案的长期目标是了解神经疾病的细胞内贩运事件如何改变 为了确定治疗性发育的新颖目标。 逆行轴突转运是由动力蛋白/dynactin蛋白复合物介导的。 P150粘结的Dynactin亚基在两种不同的非重叠常染色体显性神经沉着疾病中突变：一种引起一种称为遗传性运动神经病7B（HMN7B）的运动神经元疾病（HMN7B），另一种称为Perry综合征，parkinsonismissismissismissismissism，抑郁，弱化，体重和体重。 HMN7B和Perry突变在P150GLOUD CAP-GLY微管（MT）结合域内的12个残基与12个残基相距甚远。我们的初步结果表明，这些突变会差异地影响P150与MTS的相互作用，而P150功能在突触下。此外，我们发现HMN7B突变破坏了突触远端最远端（称为末端胸子）的逆行MT介导的转运的启动，并阻止了神经肌肉交界处的神经递质释放（NMJ）。在AIM 1中，我们将确定HMN7B和Perry综合征突变是否破坏了与P150结合伙伴的不同蛋白质相互作用，以及它们是否在轴突转运或逆行信号传导中引起缺陷。在AIM 2中，我们将研究NMJ TBS在NMJ TBS上的逆行转运的启动，以确定当动态MT加端端通过P150与终端结合蛋白-1（EB1）之间的相互作用捕获囊泡时是否发生启动，这是“ MT Plus-End的主要调节剂”。我们还将检验以下假设：p150中的HMN7B而不是佩里突变破坏了TBS处的逆行起始，并确定此缺陷是否是由于TBS时微管动力学的改变引起的。在AIM3中，我们将调查 P150中的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播。这些研究共同有助于阐明神经元中P150胶囊结构域的功能，并有望阐明神经退行性疾病的细胞类型特异性机理。