Metabolic Mechanisms of Copper-Dependent Neurodegeneration and Excitability in Menkes Disease

门克斯病铜依赖性神经变性和兴奋性的代谢机制

• 批准号: 10462355
• 负责人: Alicia R Lane
• 金额: $ 4.68万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-06 至 2025-04-05
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10462355
• 关键词: Acute; Affect; Animal Model; Binding; Bioenergetics; Biological; Birth; Brain; Cell Death; Cells; Copper; Data; Disease; Engineering; Epilepsy; Equilibrium; Exhibits; Gene Expression; Genes; Genetic Diseases; Genetic Models; Genetic Transcription; Glycolysis; Goals; Homeostasis; Human; Hypoxia; Infant; Intake; Intellectual functioning disability; Knock-out; Knowledge; Lead; Link; Menkes Kinky Hair Syndrome; Metabolic; Metabolic Pathway; Metabolism; Metals; Mitochondria; Modeling; Molecular; Molecular Target; Mus; Mutant Strains Mice; Mutation; National Research Service Awards; Nerve Degeneration; Nervous system structure; Neurologic; Neurologic Symptoms; Neurons; Neurotransmitters; Nutrient; Oxidative Phosphorylation; Oxygen; Oxygen saturation measurement; Parkinson Disease; Pathology; Pathway interactions; Phenotype; Play; Production; Reactive Oxygen Species; Regulation; Research; Respiration; Role; Testing; Therapeutic; base; calcium indicator; cell injury; cellular engineering; chromatin immunoprecipitation; cytochrome c oxidase; enzyme activity; epileptic encephalopathies; experimental study; extracellular; inorganic phosphate; metabolic phenotype; mitochondrial metabolism; molecular phenotype; mouse model; nano-string; neuroblastoma cell; neuron loss; neuronal excitability; neuropathology; neurotransmission; protein expression; rare genetic disorder; response; transcription factor; transcriptome sequencing; transcriptomics

PROJECT SUMMARY Menkes disease is a rare genetic condition in which the disruption of copper homeostasis induces neurodegeneration and other neurological symptoms soon after birth. The underlying mechanisms of Menkes neuropathology remain unclear, but the metabolic changes observed in Menkes disease and the crucial role of mitochondria in neurons point to dysregulation of cellular bioenergetics as a possible factor. Preliminary data in human cells indicates that copper depletion decreases expression of genes regulated by hypoxia induced factor 1 alpha (HIF-1α). HIF-1α is a transcription factor sensitive to metals and oxygen that regulates cellular bioenergetics by switching metabolism from mitochondrial oxidative phosphorylation to glycolysis. Further, these copper depleted cells exhibit increased mitochondrial respiration. Resolving the newly identified role of HIF-1α in regulating mitochondrial function is central to understanding how copper dyshomeostasis elicits neurodegeneration in Menkes disease. Thus, the overall objective of this F31 NRSA application is to test how copper depletion influences the HIF-1α pathway in neurons to regulate cellular metabolism and influence cell excitability and survival. The central hypothesis that will be tested in this proposal is that neuronal copper depletion selectively downregulates transcriptional activity of the HIF-1α pathway to redirect nutrients through mitochondrial respiration rather than glycolysis, rendering cells hyperexcitable due to production of reactive oxygen species by mitochondria and thus susceptible to cell death. In Aim 1, the HIF-1α pathway will be stimulated in copper depleted and control neuroblastoma cells or primary cultured neurons in order to comprehensively assess gene expression, determine binding of HIF-1α to target genes, and quantify mitochondrial respiration and glycolysis in the context of HIF-1α activity. In Aim 2, genetically encoded calcium indicators will be used in primary neuronal cultures from wildtype or neuronal-specific copper depleted mice while stimulating the HIF-1α pathway to assess how copper depletion affects cell excitability and determine the effect of HIF-1α on these phenotypes. Completion of these aims will clarify the metabolic pathways responsive to copper and their effects on neuronal function. The application of this knowledge will inform our understanding, research, and treatment of neuropathology of diseases known to be associated with dysregulated metals and/or metabolism for which there are currently limited therapeutics.

项目摘要 Menkes病是一种罕见的遗传状况，其中铜稳态的破坏会诱导 出生后不久，神经变性和其他神经系统符号。 Menkes的基本机制 神经病理学尚不清楚，但是在Menkes疾病中观察到的代谢变化和至关重要的作用 神经元中的线粒体指出细胞生物能剂的失调是可能的因素。初步数据 人类细胞表明，铜的耗竭降低了由缺氧诱导因子调节的基因的表达 1个α（HIF-1α）。 HIF-1α是对金属和氧气敏感的转录因子，可调节细胞 通过将代谢从线粒体氧化磷酸化转换为糖酵解的生物能学。此外，这些 铜部署细胞暴露的细胞增加了线粒体呼吸。解决HIF-1α的新鉴定的作用 在调节线粒体功能方面对于了解铜dyshomeostosis是如何引起的核心 Menkes病的神经变性。这是此F31 NRSA应用的总体目标是测试 铜部署影响神经元中的HIF-1α途径，以调节细胞代谢并影响细胞 兴奋性和生存。该提议中将测试的中心假设是神经元铜 耗尽选择性下调HIF-1α途径的转录活性通过 线粒体呼吸而不是糖酵解，由于反应性的产生而使细胞过度兴奋 线粒体通过线粒体氧气，因此易于细胞死亡。在AIM 1中，HIF-1α途径将是 在铜耗尽并控制神经母细胞瘤细胞或原代培养神经元中刺激，以便 全面评估基因表达，确定HIF-1α与靶基因的结合，然后定量 在HIF-1α活性的背景下，线粒体呼吸和糖酵解。在AIM 2中，遗传编码的钙 指标将用于来自野生型或神经元特异性铜的原发性神经元培养物，而小鼠则使用 刺激HIF-1α途径，以评估铜部署如何影响细胞兴奋并确定效果 在这些表型上的HIF-1α。这些目标的完成将阐明反应的代谢途径 铜及其对神经元功能的影响。这些知识的应用将为我们的理解提供信息， 研究和治疗已知与金属失调相关的疾病的神经病理学和/或 目前治疗有限的代谢。