Mechanism of age-dependent neuromuscular degeneration caused by protein misfolding on the inner mitochondrial membrane

线粒体内膜蛋白质错误折叠引起的年龄依赖性神经肌肉变性的机制

基本信息

批准号：
9980255
负责人：
Liam Patrick Coyne
金额：
$ 5.05万
依托单位：
UPSTATE MEDICAL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-30 至 2022-08-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9980255
关键词：
ATP Synthesis Pathway Acute Address Adenine Nucleotide Translocase Affect Aging Alleles Ataxia Calcium Signaling Cell Death Cell physiology Cells Chronic Chronic progressive external ophthalmoplegia Clinical Complex Cytosol Defect Degenerative Disorder Degradation Pathway Disease Exhibits Failure Histologic Histology Homeostasis Immunohistochemistry Inherited Inner mitochondrial membrane Intervention Knock-in Knock-in Mouse Mammals Mass Spectrum Analysis Measures Membrane Membrane Potentials Mitochondria Mitochondrial Membrane Protein Mitochondrial Myopathies Mitochondrial Proteins Modeling Mus Muscle Muscle Mitochondria Muscle Weakness Muscle function Mutation Nerve Degeneration Neurodegenerative Disorders Neuromuscular Diseases Organelles Organism Orthologous Gene Oxidation-Reduction Oxidative Phosphorylation Paralysed Paraplegia Parkinson Disease Pathogenicity Pathology Pathway interactions Peptide Hydrolases Phenotype Physiological Process Production Protein Import Protein Precursors Proteins Proteomics Quality Control Respiration Role Skeletal Muscle Spastic Paraplegia Spinocerebellar Ataxias Stress Study models Syndrome System Testing Time Transmission Electron Microscopy Validation Variant Western Blotting Yeasts age related base density disease-causing mutation effective therapy exercise intolerance in vivo insight misfolded protein mitochondrial membrane mouse model muscle degeneration mutant neuromuscular novel prevent protein aggregation protein degradation protein misfolding proteostasis respiratory response skeletal muscle weakness spasticity tool

项目摘要

PROJECT SUMMARY Mitochondria are essential organelles responsible for many cellular processes including energy production through oxidative phosphorylation (OXPHOS), calcium signaling, redox homeostasis, and cell death. Many of these functions are executed by a densely packed network of proteins in the inner mitochondrial membrane (IMM). As such, mitochondrial functions depend on protein homeostasis (proteostasis) in the IMM. IMM proteostasis is maintained by an extensive protein quality control system composed, in part, of IMM proteases that degrade misfolded proteins. Persistent IMM protein misfolding is speculated to cause a growing number of age-dependent neurodegenerative and neuromuscular degenerative diseases. For example, mutations in IMM proteases cause Parkinson's disease, spinocerebellar ataxia, spastic ataxia syndrome, spastic paraplegia, and other neurodegenerative diseases. Does a failure to degrade misfolded IMM proteins cause these diseases? This is unknown. In addition, misfolded variants of the ADP/ATP exchanger in the IMM, Ant1, cause autosomal dominant Progressive External Ophthalmoplegia and a mitochondrial myopathy. We used one such misfolded Ant1 variant to dissect the cellular consequences of IMM protein misfolding in yeast. Unexpectedly, IMM protein misfolding does not kill yeast cells by affecting essential mitochondrial functions in the IMM, but instead by disrupting mitochondrial protein import causing the toxic accumulation of mitochondrial precursor proteins in the cytosol. This novel mechanism was termed mitochondrial precursor overaccumulation stress (mPOS). It is not known if mPOS occurs in higher organisms, or if it can contribute to disease. Moreover, despite the significant clinical implications, the physiological consequences of IMM protein misfolding are poorly understood. To address these issues, we generated a novel knock-in (KI) mouse model expressing a pathogenic misfolded variant of Ant1. A fraction of KI mice undergo drastic neurodegeneration culminating in paralysis, thus confirming the pathogenic potential of IMM protein misfolding and providing a tool for in vivo mechanistic studies. Non-paralyzed KI mice exhibit exercise intolerance and reduced skeletal muscle mitochondrial respiration in skeletal muscle. This raises the possibility that IMM protein misfolding causes damage to specific components in the OXPHOS pathway leading to muscle weakness. We test this hypothesis in Aim 1. In addition to OXPHOS deficiency, cytosolic protein degradation pathways are activated in skeletal muscle of “asymptomatic” KI mice. The activation of cytosolic protein degradation pathways suggests that cytosolic proteostasis is challenged, possibly via mPOS. Therefore, in Aim 2, we test the hypothesis that IMM protein misfolding induces mPOS in KI mouse muscle to cause muscle weakness. The mechanism(s) revealed in this proposal will be important for understanding the role of IMM protein misfolding in Ant1-induced pathologies and neuromuscular degenerative diseases alike. In long term, interventions alleviating the identified mechanism(s) may be effective treatments for these diseases, of which there are currently none. !

项目摘要线粒体是负责许多细胞过程（包括能量产生）的必不可少的细胞器通过氧化物磷酸化（OXPHOS），钙信号传导，氧化还原稳态和细胞死亡。许多这些功能由内部线粒体膜中的密度填充的蛋白质网络执行（imm）。因此，线粒体功能取决于IMM中的蛋白质稳态（蛋白抑制）。 imm 蛋白质的蛋白质由广泛的蛋白质质量控制系统组成，部分是由Imm蛋白酶组成的那降解了错误折叠的蛋白质。据推测，持续的IMM蛋白蛋白质折叠折叠率误折录会导致越来越多的年龄依赖性神经退行性和神经肌肉退化性疾病。例如，IMM中的突变蛋白酶会引起帕金森氏病，脊髓脑性共济失调，痉挛性共济失调综合征，痉挛性截瘫和其他神经退行性疾病。未能降解错误折叠的IMM蛋白会引起这些疾病吗？这是未知的。此外，IMM，ANT1中ADP/ATP交换器的错误折叠变体原因常染色体主导的进行性外科治疗和线粒体肌病。我们使用了这样的错误折叠的ANT1变体以剖析酵母中Imm蛋白折叠的细胞后果。不料， IMM蛋白错误折叠不会通过影响IMM中必需的线粒体功能而杀死酵母细胞，但是相反，通过破坏线粒体蛋白导入导致线粒体前体的毒性积累细胞质中的蛋白质。这种新型机制称为线粒体前体过度累积应力（MPO）。尚不清楚MPO是否发生在较高的生物体中，或者它是否可以导致疾病。而且，尽管具有重大的临床意义，但IMM蛋白质错误折叠的物理后果是理解不佳。为了解决这些问题，我们生成了一种新颖的敲入（Ki）小鼠模型，表达了 ANT1的致病性错误折叠变体。一小部分Ki小鼠经历了急剧的神经变性最终瘫痪，从而确认了Imm蛋白错误折叠的致病潜力，并为体内提供了工具机械研究。非分析的Ki小鼠暴露了运动的运动和骨骼肌肉减少骨骼肌的线粒体呼吸。这增加了IMM蛋白异常折叠的可能性 OXPHOS途径中特定组件的损害导致肌肉无力。我们对此进行测试 AIM 1中的假设。除了Oxphos缺乏外，胞质蛋白降解途径在 “无症状” Ki小鼠的骨骼肌。胞质蛋白降解途径的激活表明通过MPO挑战了胞质蛋白抑制作用。因此，在AIM 2中，我们检验了以下假设 IMM蛋白错误折叠会诱导MPO在Ki小鼠肌肉中引起肌肉无力。机制在该提案中揭示的对于理解IMM蛋白错误折叠的作用在ANT1诱导的病理和神经肌肉退化性疾病。从长远来看，干预措施减轻了确定的机制可能是对这些疾病的有效治疗方法，目前没有。呢