Mitochondrial ROS microdomains and neuronal ischemia

线粒体 ROS 微区和神经元缺血

• 批准号: 9059782
• 负责人: Andrew Phillip Wojtovich
• 金额: $ 33.58万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9059782
• 关键词: Acute; Address; Animal Model; Antioxidants; Back; Behavioral Assay; Biochemistry; Bioenergetics; Biological; Biological Models; Biology; Biosensor; CRISPR/Cas technology; Caenorhabditis elegans; Calcium; Cell Death; Cell Line; Cells; Chimeric Proteins; Complex; Disease; Exhibits; Generations; Genes; Genetic; Genetic Epistasis; Genetic Models; Genome; Goals; Health; Homeostasis; Human; Huntington Disease; Hydrogen Peroxide; Individual; Injury; Ischemia; Lead; Light; Lighting; Location; Mammals; Membrane; Metabolic; Metabolism; Methodology; Mitochondria; Modeling; Modification; Mus; Nature; Neurodegenerative Disorders; Neurons; Nuclear; Optics; Outcome; Output; Oxidation-Reduction; Oxidative Stress; Parkinson Disease; Pathology; Pathway interactions; Phenotype; Physiological; Physiology; Positioning Attribute; Predisposition; Process; Production; Proteins; Reactive Oxygen Species; Resistance; Respiratory Chain; Role; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Stress; Stroke; Succinate Dehydrogenase; Superoxides; Techniques; Technology; Testing; Time; Transgenic Organisms; Translating; Translations; Variant; biological adaptation to stress; genome editing; innovative technologies; light effects; macromolecule; new technology; novel; novel strategies; novel therapeutic intervention; optogenetics; oxidative damage; research study; respiratory; response; second messenger; spatiotemporal; tool

 DESCRIPTION (provided by applicant): Reactive oxygen species (ROS) contribute to a diverse range of neurodegenerative diseases and the pathology of acute injuries such as stroke in mammals. However, not all ROS are equal, and the disease output depends strongly on the timing and location of ROS production. Furthermore, ROS in limited quantities have been shown to trigger adaptive signaling pathways and may contribute to mitohormesis. These facts lead to our central hypothesis that the biological impact of ROS, like other second messengers, depends on their timing, quantity and site of generation. However, due to their reactive nature, there is currently no means to control the site or timing of ROS production. This proposal addresses this gap by combining novel photo- inducible genetically-encoded ROS generating proteins with the power of C. elegans genetics to determine the evolutionarily-conserved role of ROS signaling in the context of neuronal ischemic sensitivity and stress resistance. In brief, new optogenetic tools for making ROS will be expressed as protein fusions using CRISPR/Cas9 technology with nuclear genes that encode mitochondrial respiratory chain subunits. The proposal focuses on the complex II subunit SDHC and utilizes expertise in biosensors, the model organism C. elegans, and optics to study ROS signaling with an unprecedented degree of precision. Parallel approaches utilizing cortical neurons in culture will address the evolutionarily conserved role of mitochondrial ROS in regulating neuronal sensitivity to ischemia. This approach could potentially yield new therapeutic strategies for diseases in which redox homeostasis has been disrupted. Overall, this novel approach will advance our understanding of mitochondrial redox signaling, allowing us to ask questions that have previously been unanswerable using conventional methodologies.

 描述(由申请人提供)：活性氧物种(ROS)有助于多种神经退行性疾病和哺乳动物急性损伤(如中风)的病理。然而，并不是所有的ROS都是相等的，疾病的产出强烈地依赖于ROS产生的时间和地点。此外，有限数量的ROS已被证明可以触发适应性信号通路，并可能有助于有丝分裂。这些事实导致了我们的中心假设，即ROS像其他第二信使一样，其生物学影响取决于它们的时间、数量和发生地点。然而，由于其反应性，目前还没有办法控制ROS生产的地点或时间。这一建议通过将新颖的光诱导遗传编码的ROS产生蛋白与线虫遗传学的力量相结合来解决这一差距，以确定ROS信号在神经元缺血敏感性和应激抵抗中的进化保守作用。简而言之，制造ROS的新的光遗传工具将表现为使用CRISPR/Cas9技术与编码线粒体呼吸链亚基的核基因的蛋白质融合。该提案侧重于复杂的II亚基SDHC，并利用生物传感器、模式生物线虫和光学方面的专业知识，以前所未有的精度研究ROS信号。利用培养的皮质神经元的平行方法将解决线粒体ROS在调节神经元对缺血的敏感性方面在进化上的保守作用。这种方法可能会为氧化还原动态平衡被破坏的疾病产生新的治疗策略。总体而言，这种新的方法将促进我们对线粒体氧化还原信号的理解，使我们能够提出以前使用传统方法无法回答的问题。