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：
8529133
负责人：
Udai B Pandey
金额：
$ 32.49万
依托单位：
LSU HEALTH SCIENCES CENTER
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 2013-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8529133
关键词：
ATP Synthesis Pathway Amyotrophic Lateral Sclerosis Behavioral Assay Binding Biochemical Biological Models Biology Cell Culture System Cell Culture Techniques Cell Line Cell Nucleus Cells Clinical Cytoplasm DNA Data Disease Drosophila eye Drosophila genus Ectopic Expression Enzymes Exons Exposure to Eye Figs - dietary Functional disorder Genes Genetic Genetic Screening Goals Homologous Gene Human Hypersensitivity Lead Link Mammalian Cell Mammals Mediating Messenger RNA Metabolism Mission Mitochondria Modeling Molecular Morphology Motor Neurons Muscle Muscle Cells 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 Stress Testing Therapeutic Intervention Toxic effect Toxin Transgenic Organisms Translating Ubiquitin United States National Institutes of Health Validation Wing 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 public health relevance research study trafficking

项目摘要

DESCRIPTION (provided by applicant): 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. 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 as seen in human ALS patients with FUS mutations, suggesting that the cytoplasmic localization of FUS 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. Following up these observations, we performed an unbiased genetic screening and 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 molecular mechanisms of muscleblind mediated suppression of mutant FUS toxicity in our fly model followed by validation in mammalian cell culture system and human patient cells carrying FUS mutations. 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）：肌萎缩性外侧硬化症（ALS）是一种晚期神经退行性疾病，其特征是上下运动神经元的丧失。 FUS和TDP-43是DNA/RNA结合蛋白，发现在ALS的零星和家族形式中都突变。为了研究由FUS/TLS突变引起的ALS的发病机理，我们建立了一系列转基因果蝇线，这些果蝇线可以异位地表达人类的野生型和突变型FUS。果蝇眼中突变体的靶向表达，但不是野生型FU，导致泛素化骨料的形成和外眼变性表型。有趣的是，运动神经元中突变型FUS的异位表达导致幼虫瘫痪，pupal致死性，而野生型FUS表达的作用最少。突变的FUS位于细胞质和细胞核上，而野生型FU仅位于核中的野生型FUS中，如在人类ALS患有FUS突变的患者中所见，这表明FUS的细胞质定位是毒性的。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。我们发现突变型在蝇肌中的表达会导致固定的机翼表型，肌肉变性和线粒体变性。我们还发现，FUS的突变RNA结合残基强烈抑制体内突变的FUS毒性。遵循这些观察结果，我们进行了无偏的遗传筛查，并发现了肌肉闪烁是突变FUS毒性的新型修饰符。在我们提出的研究中，我们将确定线粒体功能在介导相关的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 RNA结合能力在引起ALS发病机理中的作用。我们还将研究在我们的蝇模型中肌肉盲介导的突变毒毒性抑制的分子机制，然后在哺乳动物细胞培养系统和携带FUS突变的人类患者细胞中进行验证。我们提出的研究有望提供有关FUS相关ALS的分子机制的新见解，这将有助于为目前无法治愈的ALS开发治疗干预措施。