Genetic Analysis of Autophagy in the Drosophila Nervous System

果蝇神经系统自噬的遗传分析

• 批准号: 7387693
• 负责人: KIM D. FINLEY
• 金额: $ 19.06万
• 依托单位: SAN DIEGO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7387693
• 关键词: Adult; Affect; Age; Aging; Alzheimer' s Disease; Animal Model; Animals; Autophagocytosis; Biological Assay; Brain; Clinical Trials; Complementary DNA; Complex; Cultured Cells; Defect; Degenerative Disorder; Development; Development, Other; Disease; Drosophila genus; Electron Microscopy; Employee Strikes; Eukaryotic Cell; Excision; Eye; Future; Gene Expression; Genes; Genetic; Genetic Models; Genetic Research; Goals; Head; Health; Human; Huntington Disease; Image; Immunofluorescence Immunologic; Individual; Inherited; Laboratories; Link; Location; Longevity; Lysosomes; Maintenance; Messenger RNA; Modeling; Mutation; Nerve Degeneration; Nervous system structure; Neurodegenerative Disorders; Neurons; Neurophysiology - biologic function; Oxidants; Oxidative Stress; Parkinson Disease; Pathology; Pathway interactions; Pattern; Peptides; Phenotype; Phosphorus; Play; Premature aging syndrome; Process; Proteins; Rate; Reagent; Relative (related person); Research; Research Proposals; Role; Severities; Sirolimus; Stress; System; Techniques; Time; Tissues; Toxic effect; Transgenic Organisms; Transport Vesicles; Ubiquitin; Western Blotting; Work; age related; biological adaptation to stress; cDNA Expression; cytotoxic; design; design and construction; expression vector; fly; gain of function; genetic analysis; human disease; human tissue; insight; loss of function; loss of function mutation; member; mutant; neuron loss; normal aging; polyglutamine; prevent; protein aggregate; relating to nervous system; response; trafficking

DESCRIPTION (provided by applicant): During normal aging and in the progression of common neurodegenerative disorders such as Alzheimer's and Parkinson's disease, both intra-cellular and extra-cellular protein aggregates containing ubiquitin can accumulate in neurons and neural tissues of humans. The underlying cellular pathways that prevent or suppress this accumulation from occurring are not well understood but a growing body of evidence indicates the macroautophagy pathway (autophagy) is involved with the removal of age-related cellular damage and protein aggregates. Autophagy is a highly conserved lysosomal trafficking pathway that functions by sequestering damaged cellular components or aggregated proteins into new vesicles that are transported to the lysosome for degradation. Using Drosophila genetics, we have shown that mutations in autophagy genes result in progressive neural degeneration that is accompanied by the accumulation of ubiquitinated protein aggregates. We also demonstrate that increasing the expression levels of individual autophagy genes in the mature CNS can dramatically extend adult lifespan. In this research proposal we will we continue our phenotypic characterization of new autophagy mutant strains for reduced longevity and the development of progressive neural defects. To determine which members of the autophagic pathway are essential for neuronal maintenance we will use genetic and transgenic techniques (Gal4/UAS) to suppress (UAS- dsRNAi) or enhance (UAS-cDNA) the expression levels of autophagy genes in the adult fly brain. Adult longevity profiles, formation of neural aggregates and accumulation insoluble ubiquitinated proteins (IUP) will be used as assays to detect changes in aging patterns, response to environmental stress (oxidant exposure) and the suppression of cytotoxic phenotypes by aggregate prone proteins (PolyQ). Immunofluorescence imaging and electron microscopy studies will be used to determine the timing and location of ubiquitinated inclusion formation in the CNS and the type and severity of intracellular trafficking defects occurring in neurons. The goal of this proposal is to determine the role that individual autophagy genes and the pathway in general has on neuronal aging and the elimination of cellular damage from neurons. The implications for human health are significant since defects in autophagic trafficking are found in many neural degenerative disorders and that upregulating the pathway by rapamycin treatment is being used in clinical trials on Huntington's disease patients.Relevance: The accumulation of age-dependent damage and the formation of neural aggregates are associated with degenerative disorders affecting millions of people. Recent genetic research in fruit flies has shown a cellular pathway that removes cellular damage from nerve cells can significantly affect longevity by protecting the aging nervous system. Insight from this genetic model of neural degeneration and protection it affords the nervous system will enhance our understanding of complex processes that occur in people and will direct future research that is designed to promote human health and longevity.

描述（由申请人提供）：在正常衰老和常见神经退行性疾病的进展过程中，例如阿尔茨海默氏病和帕金森氏病，含有泛素的细胞内和细胞外蛋白质聚集物都可以堆积在人类的神经元和神经组织中。尚不清楚阻止或抑制这种积累的潜在细胞途径，但越来越多的证据表明大噬菌途径（自噬）与去除年龄相关的细胞损伤和蛋白质聚集体有关。自噬是一种高度保守的溶酶体运输途径，通过将受损的细胞成分或聚集的蛋白质隔离到新囊泡中，从而发挥作用，这些蛋白质被转运到溶酶体以降解。使用果蝇遗传学，我们表明自噬基因的突变导致进行性神经变性，并伴随着泛素化蛋白聚集体的积累。我们还证明，在成熟的中枢神经系统中增加单个自噬基因的表达水平可以极大地延长成人的寿命。在这项研究建议中，我们将继续对新的自噬突变菌株的表型表征，以减少寿命和进行性神经缺陷的发展。为了确定自噬途径的哪些成员对于神经元维持至关重要，我们将使用遗传和转基因技术（GAL4/UAS）抑制（UAS-DSRNAI）或增强（UAS-CDNA）在成年蝇脑中自噬基因的表达水平。成人的寿命谱，神经聚集体的形成和积累不溶性泛素化蛋白（IUP）将用作检测衰老模式变化的测定，对环境应激的反应（氧化剂暴露）以及聚集型prone蛋白（Polyq）对细胞毒性表型的抑制。免疫荧光成像和电子显微镜研究将用于确定中枢神经系统中泛素化包容性形成的时间和位置以及神经元中发生的细胞内运输缺陷的类型和严重程度。该建议的目的是确定单个自噬基因和一般途径在神经元衰老以及消除神经元细胞损伤的作用。由于在许多神经退化性疾病中发现了自噬贩运的缺陷，因此对人类健康的影响很大，并且在Huntington疾病患者的临床试验中使用了通过雷帕霉素治疗来上调途径。RELELVANCE：年龄依赖性损害的积累，与神经聚集疾病的形成与米尔氏症的形成相关性米尔氏症症状构成米诺的疾病。果蝇中最近的遗传研究表明，通过保护衰老神经系统来消除神经细胞的细胞损伤可以显着影响寿命。从这种神经变性和保护的遗传模型中，它提供了神经系统的洞察力将增强我们对人们在人们中发生的复杂过程的理解，并将指导未来旨在促进人类健康和寿命的研究。