The Role of phosphoenolpyruvate carboxykinase and reactive oxygen species in the

描述(申请人提供)：地球上几乎所有的生物都共享支持无氧代谢的基本代谢途径，然而许多生物，包括大多数脊椎动物，如果没有分子氧，就不能长时间存活。一年生蓝鳍金枪鱼的胚胎是研究脊椎动物耐缺氧和缺氧敏感性的机制基础的极佳模型。斑潜蝇的胚胎经历了一段独特的发育休眠期，称为滞育期。最近的证据表明，斑腿猴胚胎具有一些非常独特的生理适应能力，这与其对长期缺氧的耐受有关。在250℃的温度下，休眠胚胎和发育活跃的胚胎都可以在没有氧气的情况下存活数月。在缺氧暴露的最初几个小时内，斑马鱼的胚胎表现出大量的ATP耗竭，并在同一时间段内失去了线粒体膜电位。这两个事件通常与其他脊椎动物细胞的细胞死亡有关，但斑马鱼的胚胎很快就会从线粒体生理学和能量学的这些剧烈变化中恢复过来。这些观察结果表明，与其他脊椎动物，甚至与其他表现出大量耐缺氧能力的脊椎动物相比，利姆奈斯胚胎的细胞具有一些非同寻常的特征。我将确定支持红曲霉分离细胞耐缺氧的代谢途径，评估缺氧和缺氧恢复过程中线粒体的功能和能量，并测试由磷酸烯醇式丙酮酸羧基激酶支持的替代代谢途径对缺氧生存至关重要的假设。通过使用提案中概述的综合方法，我们有望创建细胞耐缺氧的细胞生理学更完整的图景，如果这种特殊的脊椎动物极端嗜好的话。 与公共卫生相关：在发达国家，心脏病和中风是造成死亡的绝大多数原因。人类心脏和大脑组织对缺氧的极端敏感性在细胞水平上知之甚少。通过了解支持一年生金枪鱼胚胎极端耐缺氧的细胞机制，我们可能能够开发出治疗方法来调解或防止心脏病发作和中风对人类的破坏性影响。

DESCRIPTION (provided by applicant): Almost all organisms on Earth share the basic metabolic pathways that support anaerobic metabolism, and yet many organisms, including most vertebrates, cannot survive for long without molecular oxygen. Embryos of the annual killifish Austrofundulus limnaeus are an excellent model for investigating the mechanistic basis of anoxia-tolerance and anoxia-sensitivity in vertebrates. Embryos of A. limnaeus undergo a unique period of developmental dormancy called diapause. Recent evidence suggests that embryos of A. limnaeus have some very unique physiological adaptations that are associated with tolerance of long-term anoxia. Both dormant and actively developing embryos of A. limnaeus can survive for months without oxygen at 250C. Embryos of A. limnaeus display a massive depletion of ATP during the initial hours of anoxic exposure and lose their mitochondrial membrane potential during this same time frame. These two events are typically associated with cell death in other vertebrate cells, but embryos of A. limnaeus quickly recover from these drastic changes in mitochondrial physiology and energetics. These observations imply that cells of A. limnaeus embryos have some extraordinary characteristics compared to other vertebrates, and even to other vertebrates that exhibit substantial tolerance of anoxia. I will identify the metabolic pathways that support anoxia tolerance in isolated cells of A. limnaeus, assess mitochondrial function and energetics during anoxia and recovery from anoxia, and test the hypothesis that an alternate metabolic pathways supported by the enzyme phosphoenolpyruvate carboxykinase is critical for the survival of anoxia. By using the integrative approaches outlined in the proposal we can hopefully create a more complete picture of the cellular physiology of anoxia-tolerance in the cells if this exceptional vertebrate extremophile. PUBLIC HEALTH RELEVANCE: Heart disease and stroke are responsible for the vast majority of deaths in the developed world. The extreme sensitivity of human heart and brain tissue to lack of oxygen is poorly understood at the cellular level. By understanding the cellular mechanisms that support extreme anoxia tolerance in embryos of the annual killifish, Austrofundulus limnaeus, we may be able to develop treatments to mediate or prevent the damaging effects of heart attacks and strokes to humans.