Mitochondrial Protein Misfolding and Aggregation after Hypoxia: Mechanisms and Mitigation

项目摘要 低氧和复氧对细胞造成严重破坏。如果不加以修复，这种破坏最终将导致细胞 心肌梗死和中风等疾病的功能障碍和死亡，这是导致死亡的头号原因 以及美国的残疾问题。不幸的是，对于缺氧性损伤没有有效的治疗方法，除非恢复氧合， 已经被批准，这表明缺氧损伤的一个未被认识的方面没有得到有效的治疗 通过以前的策略。线粒体长期以来一直被认为是缺氧性损伤的中心。线粒体是 氧在细胞中的主要利用者，将氧转化为细胞生长所需的化学势能 细胞和有机体的生存。线粒体也是细胞死亡过程的中心--特别是 细胞凋亡和钙介导的死亡形式，包括坏死。坏死和细胞凋亡都被认为是 是缺氧损伤后最常见的死亡机制。然而，充分了解缺氧是如何 损伤线粒体导致细胞死亡是缺乏的。我们最近报道了一种新型的缺氧-- 诱发线粒体病理--线粒体蛋白错误折叠。我们公布的数据显示， 线粒体蛋白的错误折叠发生在缺氧损伤级联反应的早期，在细胞死亡的任何证据之前。 这表明，线粒体蛋白质的错误折叠可能是缺氧和 缺氧性细胞损伤和死亡的原因。符合这一假说的，遗传的或药理学的 线虫中激活线粒体未折叠蛋白反应的操作 线粒体错误折叠蛋白(MitoUPR)是细胞内对错误折叠蛋白的一种稳态反应，可保护缺氧 伤害并提高动物的存活率。自从这篇文章发表以来，我们已经开发出新的荧光线粒体 线虫蛋白质报告工具，以便直接研究蛋白质错误折叠并有初步证据 线粒体蛋白质不仅错误折叠，而且在缺氧后聚集。该项目的目标是 对低氧诱导的线粒体蛋白错误折叠和 聚集，以确定减轻线粒体蛋白平衡中断的方法，并确定是否 类似的干扰可以在人类疾病的小鼠模型中检测到并得到缓解。我们的将军 策略是利用线虫速度快、成本低和专门的细胞生物学工具来 基础发现，并在可能的情况下将我们的发现应用于缺氧性疾病的哺乳动物模型。 我们的具体目标如下：目标1.确定错误折叠和聚集的线粒体的身份 蛋白质和聚集的动力学、决定因素和后果。目标2.确定遗传和 改善线虫线粒体蛋白质错误折叠的药物操作。目标3. 确定线粒体蛋白质错误折叠/聚集是否发生在疾病的小鼠模型中。 完成这些目标将增加我们对线粒体一种新的缺氧性病理的理解 并可能会找到缓解这种影响的方法。

Project Summary Hypoxia and reoxygenation create havoc in cells. This havoc if unrepaired will ultimately lead to cell dysfunction and death in diseases such as myocardial infarction and stroke, the number one causes of death and disability in the US. Unfortunately, no effective therapy for hypoxic injury, short of restoring oxygenation, has been approved, suggesting that an unrecognized aspect of hypoxic injury is not being effectively treated by previous strategies. Mitochondria have long been recognized as central to hypoxic injury. Mitochondria are the primary utilizer of oxygen in cells, converting oxygen to the chemical potential energy required for the survival of cells and the organism. Mitochondria also are central to cell death processes – in particular apoptosis and forms of calcium-mediated death including necrosis. Both necrosis and apoptosis are thought to be the most prevalent mechanisms of death after hypoxic injury. However, a full understanding of how hypoxia injures mitochondria leading to cell death is lacking. We have recently reported a novel type of hypoxia- induced mitochondrial pathology – mitochondrial protein misfolding. Our published data show that mitochondrial protein misfolding occurs early in the hypoxic injury cascade, prior to any evidence of cell death. This suggests the hypothesis that mitochondrial protein misfolding may be both a consequence of hypoxia and a cause of hypoxic cell injury and death. Consistent with this hypothesis, genetic or pharmacologic manipulations in the nematode C. elegans that activate the mitochondrial unfolded protein response (mitoUPR), an intracellular homeostatic response to mitochondrial misfolded proteins, protects from hypoxic injury and improves animal survival. Since this publication, we have developed new fluorescent mitochondrial protein reporter tools in C. elegans in order to study protein misfolding directly and have preliminary evidence that mitochondrial proteins not only misfold but aggregate after hypoxia. The goals of this project are to develop a fundamental understanding of hypoxia-induced mitochondrial protein misfolding and aggregation, to identify ways to mitigate disruption of mitochondrial proteostasis, and to determine if similar disruption can be detected and mitigated in mouse models of human disease. Our general strategy is to take advantage of the speed, low cost, and specialized cell biological tools of C. elegans for fundamental discovery and to apply where possible our discoveries to mammalian models of hypoxic disease. Our specific aims are as follows: Aim 1. Determine the identity of the misfolded and aggregated mitochondrial proteins and the kinetics, determinants, and consequences of aggregation. Aim 2. Identify genetic and pharmacological manipulations that ameliorate mitochondrial protein misfolding in C. elegans. Aim 3. Determine whether mitochondrial protein misfolding/aggregation occur in mouse models of disease. Completion of these aims will increase our understanding of a novel hypoxic pathology of the mitochondria and will potentially identify ways to mitigate it.