Screening for drug targets in a Drosophila model of muscle degeneration

在果蝇肌肉变性模型中筛选药物靶点

• 批准号: 7675655
• 负责人: Edward Owusu-Ansah
• 金额: $ 4.88万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7675655
• 关键词: Acquired Immunodeficiency Syndrome; Address; Apoptosis; Atrophic; Attenuated; Autophagocytosis; Biochemical; Biological Assay; Biological Markers; Chromatin; Clinical; Code; Data; Degradation Pathway; Diabetes Mellitus; Drosophila genus; Drug Delivery Systems; Dystrophin; Ecdysone; Event; Functional disorder; Genes; Genetic; Genetic Screening; Goals; Heart failure; Heat shock proteins; Histones; Human; Human Genome; Inherited; Larva; Link; Luciferases; Malignant Neoplasms; Mediating; Messenger RNA; Modeling; Molecular; Molecular Profiling; Molecular Target; Muscle; Muscle Weakness; Muscular Atrophy; Muscular Dystrophies; Myopathy; Organism; Pan Genus; Parkinsonian Disorders; Pathway interactions; Phosphoric Monoester Hydrolases; Phosphotransferases; Portraits; Process; Protein Isoforms; Proteins; Proteomics; RNA Interference; Relative (related person); Reporter; Ribosomal Proteins; Role; Screening procedure; Sepsis; Signal Pathway; Signal Transduction; Sporadic Inclusion Body Myopathy; Symptoms; System; Techniques; Testing; Transgenic Organisms; Ubiquitin; Uremia; base; ecdysone receptor; fly; genome-wide; in vivo; in vivo Model; inhibitor/antagonist; insulin signaling; multicatalytic endopeptidase complex; muscle degeneration; muscle form; parkin gene/protein; protein expression; public health relevance; respiratory; small molecule; wasting

DESCRIPTION (provided by applicant): Muscular dystrophy refers to a diverse group of debilitating hereditary muscle disorders typified by a host of symptoms, including progressive muscular weakness and muscle wasting. To examine the molecular mechanisms that contribute to muscle loss, we have established a potent model of muscle degeneration in Drosophila. Constitutive expression of the ecdysone receptor (EcR) in somatic muscles of Drosophila larvae triggers their degeneration, thereby serving as a model for elucidating the molecular underpinnings of muscle atrophy/degeneration in a genetically tractable organism. Using this Drosophila system, we propose the following: Aim 1. Establish the relative contribution of catabolic and anabolic signals to EcR-mediated muscle wasting: Using a combination of genetic and biochemical techniques, we will examine the effect of inhibiting apoptosis, autophagy and degradation via the proteasomal pathway (i.e catabolic signals) as well as hyperstimulation of the insulin signaling network (anabolic signals) on EcR-mediated muscle degeneration; Aim 2. Identify universal biomarkers of muscle degeneration in flies and generate a "muscle degeneration reporter" for subsequent genetic screens: We will compare the mRNA and proteomic expression profiles of EcR-mediated muscle degeneration with that of three other well-established fly models of muscle degeneration (i.e. the dystrophin model, parkin/pinkl model and a sporadic inclusion-body myositis model) to identify several muscle pan-degeneration markers. Finally, we will develop a suitable assay for muscle degeneration, most likely a Luciferase transcriptional reporter based on one of the pan-degeneration markers; and Aim 3. Identify drug targets via a transgenic RNAi screen in muscles to identify suppressors of EcR- mediated muscle degeneration: Using publicly available RNAi transgenic lines, we will carry out a genome- wide in vivo RNAi screen (using the Luciferase-based reporter developed in Aim 2) in larval muscles to identify genes/pathways whose inhibition can ameliorate EcR-mediated muscle degeneration. PUBLIC HEALTH RELEVANCE: Given the high degree of conservation between the Drosophila and human genomes, we anticipate that molecular targets identified by this study will eventually provide leads for addressing muscle wasting/degeneration in humans. In summary, we seek to provide a genome-wide portrait of the molecular mechanisms that either trigger or suppress ecdysone-mediated muscle wasting in Drosophila, with the ultimate goal of identifying suitable drug targets for attenuating muscle degeneration in humans.

描述（由申请人提供）：肌营养不良症是指多种使人衰弱的遗传性肌肉疾病，其典型症状包括进行性肌肉无力和肌肉萎缩。为了研究导致肌肉损失的分子机制，我们建立了果蝇肌肉退化的有效模型。果蝇幼虫体肌中蜕皮激素受体（EcR）的组成型表达触发了它们的退化，从而可以作为阐明遗传易驯化生物体中肌肉萎缩/退化的分子基础的模型。使用该果蝇系统，我们提出以下建议： 目标 1. 确定分解代谢和合成代谢信号对 EcR 介导的肌肉萎缩的相对贡献：结合遗传和生化技术，我们将检查抑制细胞凋亡、自噬和降解的效果通过蛋白酶体途径（即分解代谢信号）以及胰岛素信号网络（合成代谢信号）对 EcR 介导的肌肉变性的过度刺激；目标 2. 鉴定果蝇肌肉变性的通用生物标志物，并生成“肌肉变性报告基因”用于后续基因筛选：我们将比较 EcR 介导的肌肉变性的 mRNA 和蛋白质组表达谱与其他三种成熟的果蝇模型的 mRNA 和蛋白质组表达谱。肌肉变性（即肌营养不良蛋白模型、parkin/pinkl 模型和散发性包涵体肌炎模型），以确定几种肌肉泛变性标记物。最后，我们将开发一种合适的肌肉变性检测方法，很可能是基于一种泛变性标记物的荧光素酶转录报告基因；目标 3. 通过肌肉中的转基因 RNAi 筛选来识别药物靶标，以识别 EcR 介导的肌肉变性的抑制因子：使用公开的 RNAi 转基因系，我们将进行全基因组体内 RNAi 筛选（使用基于荧光素酶的报告基因）在目标 2）中开发了幼虫肌肉，以确定其抑制可以改善 EcR 介导的肌肉退化的基因/途径。公共健康相关性：鉴于果蝇和人类基因组之间的高度保守性，我们预计本研究确定的分子靶点最终将为解决人类肌肉萎缩/退化问题提供线索。总之，我们寻求提供触发或抑制果蝇蜕皮激素介导的肌肉萎缩的分子机制的全基因组图谱，最终目标是确定减轻人类肌肉退化的合适药物靶点。