Cellular Redox Balancing and Oxidative Stress: Assembling a Global Model

描述（由申请人提供）：活性氧（ROS）是有氧代谢的必然结果。ROS的积累和由此产生的氧化应激似乎是从阿尔茨海默病到糖尿病的各种人类病理学的基础。相比之下，健康个体的细胞通过合成抗氧化剂化合物、快速修复氧化损伤和有效平衡细胞还原剂库来有效管理ROS。这些策略中的后者，氧化还原平衡，是知之甚少，主要是由于缺乏生理相关的动物模型来研究这个动态过程。我们最近的工作表明，简单的，生物现实的操作无机氮营养拟南芥可以迅速和可预测地改变细胞的氧化还原状态，导致适应性的变化，在线粒体呼吸链中的几个氧化还原平衡酶的表达和活性。这些结果表明，A. thaliana可以作为一个独特的模式，氧化还原平衡的研究和ROS管理中的作用。因此，本项目的主要目标是描述A.拟南芥细胞对氮源诱导的细胞氧化还原状态的变化。为此，我们将利用基因组微阵列直接研究根细胞的转录组如何受到细胞氧化还原状态改变的影响（具体目标2）。微阵列数据将使用设计用于识别和统计量化生化途径的协调调节的软件进行分析，我们特别感兴趣的是检查参与还原剂的产生、细胞内分配和氧化的途径（即潜在的氧化还原平衡途径）。除了研究基因表达水平的氧化还原平衡外，我们还将直接研究氮源如何影响细胞ROS水平（H2O2），脂质氧化以及抗氧化剂库（谷胱甘肽和抗坏血酸盐）的大小和氧化状态（具体目标1）。总的来说，这些研究将使我们能够将细胞氧化还原状态的生理现实变化与氧化应激的定量评估和定义的转录反应联系起来。最终，这项工作将为开发一个全球性的、实验支持的细胞氧化还原平衡模型提供信息，这一过程对维护人类健康至关重要。 从阿尔茨海默氏症到糖尿病，各种疾病都与细胞微妙的“电子经济”中断引起的氧化应激有关。拟议的项目将阐明健康细胞如何动态调整其生理机能，以防止在不断变化的环境条件下的氧化应激，使用植物拟南芥作为实验模型。

DESCRIPTION (provided by applicant): Reactive oxygen species (ROS) are an inevitable consequence of aerobic metabolism. The accumulation of ROS, and resulting oxidative stress, appear to underlie a diverse array of human pathologies, from Alzheimer's disease to diabetes. In contrast, the cells of healthy individuals effectively manage ROS through the synthesis of antioxidant compounds, the rapid repair of oxidative damage, and the efficient balancing of cellular reductant pools. The latter of these strategies, redox balancing, is poorly understood, primarily due to the lack of physiologically relevant animal models to study this dynamic process. Our recent work has shown that simple, biologically realistic manipulations of inorganic nitrogen nutrition in Arabidopsis thaliana can rapidly and predictably alter cellular redox status, leading to adaptive changes in the expression and activity of several redox balancing enzymes in the mitochondrial respiratory chain. These results demonstrate that A. thaliana can serve as a unique model for studies of redox balancing and its role in ROS management. Thus, the primary objective of this project is to characterize the global response of A. thaliana cells to nitrogen source-induced changes in cellular redox status. Toward this end, we will utilize genome microarrays to directly examine how the transcriptome of root cells is affected by alterations in cellular redox status (Specific Aim 2). Microarray data will be analyzed using software designed to identify and statistically quantify the coordinated regulation of biochemical pathways, and we are particularly interested in examining the pathways involved in the production, intracellular partitioning, and oxidation of reductant (i.e. potential redox balancing pathways). In addition to studying redox balancing at the level of gene expression, we will also directly examine how nitrogen source affects cellular ROS levels (H2O2), lipid per oxidation, and the size and oxidation state of antioxidant pools (glutathione and ascorbate) (Specific Aim 1). Overall, these studies will allow us to link a physiologically realistic change in cellular redox state to quantitative assessments of oxidative stress and a defined transcriptional response. Ultimately, this work will inform the development of a global, experimentally-supported model of cellular redox balancing, a process which is central to the maintenance of human health. A variety of diseases, from Alzheimer's to diabetes, are related to the oxidative stress that arises from disruptions in the cell's delicate "electron economy". The proposed project will elucidate how healthy cells dynamically adjust their physiology to prevent oxidative stress under changing environmental conditions, using the plant Arabidopsis thaliana as an experimental model.