Cellular and Molecular Mechanisms of FUS-related Amyotrophic Lateral Sclerosis

FUS相关肌萎缩侧索硬化症的细胞和分子机制

基本信息

批准号：
8811504
负责人：
Udai B Pandey
金额：
$ 34.05万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-03-01 至 2018-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8811504
关键词：
ATP Synthesis Pathway Alternative Splicing Amyotrophic Lateral Sclerosis Behavioral Assay Binding Biochemical Biological Models Biology Cell Culture Techniques Cell Line Cell Nucleus Cells Clinical Cytoplasm DNA Data Disease Drosophila eye Drosophila genus Ectopic Expression Enzymes Exons Eye Functional disorder Genes Genetic Genetic Screening Goals Health Homologous Gene Human Hypersensitivity Lead Link Mammalian Cell Mammals Mediating Messenger RNA Metabolism Mission Mitochondria Modeling Molecular Morphology Motor Neurons Muscle Mutate Mutation Myotonic Dystrophy Nematoda Nerve Degeneration Neurodegenerative Disorders Neurons Nuclear Export Paralysed Paraquat Pathogenesis Pathway interactions Patients Phenotype Protein Binding Proteins Publishing RNA RNA Binding RNA Splicing RNA-Binding Proteins Relative (related person)Research Research Personnel Role Rotenone Series Signal Transduction Testing Therapeutic Intervention Toxic effect Toxin Transgenic Organisms Translating Ubiquitin United States National Institutes of Health Validation Wing abstracting disease-causing mutation effective therapy fly human disease in vivo insight mRNA Precursor motor neuron degeneration muscle degeneration mutant neurodegenerative phenotype novel novel therapeutics nucleocytoplasmic transport polyglutamine protein TDP-43 research study trafficking

项目摘要

DESCRIPTION (provided by applicant): Abstract Amyotrophic lateral sclerosis (ALS) is a late onset neurodegenerative disorder characterized by the loss of upper and lower motor neurons. FUS and TDP-43 are DNA/RNA binding proteins found to be mutated in both sporadic and familial forms of ALS. To investigate the pathogenesis of ALS caused by FUS/TLS mutations, we established a series of transgenic Drosophila lines that ectopically express human wild type and mutant FUS/TLS. Targeted expression of mutant, but not wild type FUS, in Drosophila eyes causes the formation of ubiquitinated aggregates and an external eye degenerative phenotype. Interestingly, ectopic expression of mutant FUS in motor neurons resulted in larval paralysis, pupal lethality, whereas wild type FUS expression had minimal effect. Mutant FUS localized to both the cytoplasm and nucleus, whereas wild type FUS localized only to the nucleus, suggesting that the cytoplasmic localization of FUS/TLS is required for toxicity. Furthermore, we found that deletion of the nuclear export signal strongly suppressed toxicity, suggesting that cytoplasmic localization is necessary for neurodegeneration. We found that expression of mutant FUS in the fly muscles leads to held-up wing phenotype, muscle degeneration and mitochondrial degeneration. We also found that mutating RNA binding residues of FUS strongly suppress mutant FUS toxicity in vivo. While doing an unbiased genetic screening, we discovered muscleblind as a novel modifier of mutant FUS toxicity. In our proposed studies, we will determine the role of mitochondrial functions in mediating FUS- related ALS and which functions of mitochondria are disrupted by the ALS causing mutations. We will systematically determine the role of RNA binding ability of FUS in causing ALS pathogenesis. We will also investigate if muscleblind regulates alternative splicing and sub- cellular distribution of FUS and contributes to the onset and progression of FUS-related ALS. Our proposed studies are expected to provide novel insights into the molecular mechanism of FUS-related ALS that would help in developing a therapeutic interventions for ALS for which currently no cure available.

描述（由申请人提供）：摘要肌萎缩侧索硬化症（ALS）是一种迟发性神经退行性疾病，其特征是上运动神经元和下运动神经元的丧失。 FUS 和 TDP-43 是 DNA/RNA 结合蛋白，在散发性和家族性 ALS 中均发现突变。为了研究 FUS/TLS 突变引起的 ALS 的发病机制，我们建立了一系列异位表达人类野生型和突变型 FUS/TLS 的转基因果蝇系。果蝇眼睛中突变型而非野生型 FUS 的靶向表达会导致泛素化聚集体的形成和外眼退化表型。有趣的是，运动神经元中突变型 FUS 的异位表达导致幼虫麻痹、蛹死亡，而野生型 FUS 表达的影响很小。突变型 FUS 定位于细胞质和细胞核，而野生型 FUS 仅定位于细胞核，这表明 FUS/TLS 的细胞质定位是毒性所必需的。此外，我们发现核输出信号的缺失强烈抑制了毒性，这表明细胞质定位对于神经变性是必要的。我们发现，突变型 FUS 在果蝇肌肉中的表达会导致翅膀滞留表型、肌肉变性和线粒体变性。我们还发现，FUS 的 RNA 结合残基突变可强烈抑制突变型 FUS 的体内毒性。在进行公正的基因筛查时，我们发现肌肉盲是突变型 FUS 毒性的一种新型修饰剂。在我们提出的研究中，我们将确定线粒体功能在介导 FUS 相关 ALS 中的作用，以及线粒体的哪些功能被引起突变的 ALS 破坏。我们将系统地确定FUS的RNA结合能力在引起ALS发病机制中的作用。我们还将研究muscleblind是否调节FUS和FUS的选择性剪接和亚细胞分布有助于 FUS 相关 ALS 的发生和进展。我们提出的研究预计将为 FUS 相关 ALS 的分子机制提供新的见解，这将有助于开发针对目前尚无治愈方法的 ALS 的治疗干预措施。