Hypoxic Regulation of Mitochondrial Function

低氧主要通过HIF1转录因子的诱导引起基因表达的变化，并且 这些变化中的许多被认为有助于适应氧气有限的不利环境。一 一类被广泛研究的低氧诱导基因是糖酵解酶。这些 当还原的氧气不支持时，分子被认为是维持能量产生所必需的 线粒体内的氧化磷酸化。虽然糖酵解对细胞在低氧条件下的生长很重要， 我们发现线粒体并不只是被动地停止功能。HIF-熟练细胞积极减少 缺氧时的耗氧量，而缺氧诱导因子缺乏的细胞则不是。因此，我们使用了表情分析和 在过去的资金周期中进行数据挖掘，以确定针对低氧诱导的蛋白 线粒体。这些假定的HIF-1调节的线粒体蛋白不会导致细胞凋亡，但我们的 功能数据支持这一新概念，即它们主动调节线粒体活动以响应 缺氧。因此，我们建议在此应用程序中解决以下四个问题。在具体目标1中， 我们将确定依赖于HIF的基因表达变化是否会导致脑组织氧耗的改变 线粒体通过靶基因BNip3/L和/或丙酮酸脱氢酶1(PDK1)的诱导 和/或缺氧诱导基因1(HIG1)。在特定目标2中，我们将检验药理学的假设 逆转这些依赖HIF-1的变化将增加氧气消耗，减少细胞内氧气 浓度，并导致对氧气依赖治疗的敏感性，如缺氧细胞毒素 替拉帕扎明(TPZ)。或二硝基苯甲酰胺芥末PR-104。在具体的目标3中，我们会问这个药理学 使肿瘤更缺氧的治疗也会使它们更具侵袭性，更有可能转移。 最后，在特定的目标4中，我们假设我们可以确定其他新的低氧调节因子 线粒体功能通过筛选酵母缺失文库在低氧中生长的非 可发酵碳源培养基。拟议的实验将使我们能够确定这些 分子有助于线粒体功能的低氧调节，它们有助于细胞的生长 模型肿瘤，以及它们对肿瘤对氧气依赖治疗的反应有什么影响。

Hypoxia causes gene expression changes largely through the induction of the HIF1 transcription factor, and many of these changes are thought to help adapt to the adverse environment where oxygen is limiting. One class of hypoxia-induced genes that have been extensively studied is the glycolytic enzymes. These molecules are thought to be necessary to maintain energy production when reduced oxygen will not support oxidative phosphorylation within the mitochondria. While glycolysis is important to cellular growth in hypoxia, we find that the mitochondrion does not just passively stop functioning. HIF-proficient cells actively reduce oxygen consumption in hypoxia while HIF-deficient cells do not. We therefore used expression profiling and data mining during the past funding cycle to identify hypoxia-induced proteins that are targeted to the mitochondria. These putative HIF-1 regulated mitochondrial proteins do not cause apoptosis, but our functional data supports the novel concept that they actively regulate mitochondrial activity in response to hypoxia. We therefore propose to address the following four questions in this application. In specific aim 1, we will determine if HIF-dependent gene expression changes result in altered oxygen consumption in the mitochondria through the induction of target genes BNip3/L, and/or pyruvate dehydrogenase kinase 1 (PDK1) and/or hypoxia-induced gene 1 (HIG1). In specific aim 2 we will test the hypothesis that pharmacologic reversal of these HIF-1 dependent changes will increase oxygen consumption, diminish intracellular oxygen concentrations, and result in sensitivity to oxygen-dependent therapies such as the hypoxic cytotoxins tirapazamine (TPZ).or dinitobenzamide mustard Pr-104. In specific aim 3 we will ask if this pharmacologic treatment that makes tumors more hypoxic also makes them more aggressive and likely to metastasize. Lastly in specific aim 4 we hypothesize that we can identify additional novel regulators of hypoxic mitochondrial function through a screen of the yeast deletion library using growth in hypoxia on non- fermentable carbon source media. The proposed experiments will allow us to determine how these molecules contribute to hypoxic regulation of mitochondrial function, what they contribute to the growth of model tumors, and what impact they have on the tumor's response to oxygen-dependent therapy.