Mitchondria-Nuclear Crosstalk in Hypoxic Gene Induction

描述(由申请人提供)：细胞、组织和生物体感知和响应氧气水平变化的能力通常对它们的生存至关重要。细胞对低氧的适应就是一个很好的例子。间歇性低氧与人类多种病理生理状态(如睡眠呼吸暂停、中枢性低通气综合征和血管闭塞)有关；这些状态可以改变新陈代谢，诱导血管生成，并影响炎症反应。一些研究已经报道，对低氧的适应需要大量核基因的诱导，线粒体呼吸链在哺乳动物和酵母细胞中一些低氧基因的诱导中起着重要作用。我们研究的总体目标是了解线粒体在这一过程中的作用。这些研究结合了蛋白质组、基因组和遗传策略来分析酵母中低氧基因的诱导。此外，还将对培养中的哺乳动物细胞进行平行研究。目的1将使用基因图谱方法来鉴定在酵母线粒体呼吸链控制下的低氧核基因。目的2将重点放在线粒体依赖的低氧基因诱导途径本身。基因操作将被用于识别和排序酵母基因，这些基因对于诱导线粒体依赖的低氧核基因是必不可少的。目的3将评估线粒体产生的活性氧和一氧化氮在酵母低氧基因诱导中的重要性，重点了解蛋白质酪氨酸硝化和蛋白质羰化在低氧信号中的作用。目的4研究低氧基因在哺乳动物细胞中的诱导。它解决了两个有争议或未完全探讨的问题。首先，它将使用在酵母中取得成功的方法来询问哺乳动物细胞在缺氧或缺氧条件下是否经历短暂的氧化或亚硝化压力，如果是，则识别在缺氧或缺氧条件下发生羰化或酪氨酸硝化的蛋白质。其次，它将在RNAi击倒实验中使用，以评估这些碳化或硝化蛋白质在哺乳动物细胞低氧信号中的重要性。细胞不能对低氧做出正确的反应可导致多种病理状态(如贫血、心肌梗死、视网膜病变和肿瘤生长)。对缺氧信号通路的了解可能会为这些疾病带来新的治疗方法，并可能有助于我们对人类衰老的理解。

DESCRIPTION (provided by applicant): The ability of cells, tissues, and organisms to sense and respond to changes in oxygen levels is often crucial to their survival. Cellular adaptation to hypoxia is a good example of this. Intermittent episodes of hypoxia are associated with a variety of human patho-physiological states (e.g., sleep apnea, central hypoventilation syndrome, and vascular occlusion); these can alter metabolism, induce angiogenesis, and affect inflammatory responses. Several studies have reported that adaptation to hypoxia requires the induction of a large number of nuclear genes and that the mitochondrial respiratory chain plays an important role in the induction of some of these hypoxic genes in mammalian and yeast cells. The overall goal of our studies is to understand the role of the mitochondrion in this process. These studies combine proteomic, genomic, and genetic strategies to analyze hypoxic gene induction in yeast. In addition, parallel studies will be done with mammalian cells in culture. Aim 1 will use gene profiling methods to identify hypoxic nuclear genes that are under the control of the mitochondrial respiratory chain in yeast. Aim 2 will focus on the mitochondrial - dependent hypoxic gene induction pathway itself. Genetic manipulation will be used to identify and order yeast genes that are essential for the induction of mitochondrial-dependent hypoxic nuclear genes. Aim 3 will evaluate the importance of mitochondrially-generated reactive oxygen species and nitric oxide in yeast hypoxic gene induction, with emphasis on understanding the role of protein tyrosine nitration and protein carbonylation in hypoxic signaling. Aim 4 focuses on hypoxic gene induction in mammalian cells. It addresses two questions that are controversial or incompletely explored. First, it will use methodologies that have been successful in yeast to ask if mammalian cells experience transient oxidative or nitrosative stress when exposed to hypoxia or anoxia and, if so, identify proteins carbonylated or tyrosine nitrated under anoxic or hypoxic conditions. Second, it will use in RNAi knock down experiments to assess the importance of these carbonylated or nitrated proteins in hypoxic signaling in mammalian cells. The failure of cells to respond properly to hypoxia can lead to a variety of pathological states (e.g., anemia, myocardial infarction, retinopathy, and the growth of tumors). An understanding of hypoxic signaling pathways may lead to new therapies for these diseases and may help with our understanding of human aging.