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。活性氧的积累和由此产生的氧化应激，似乎是一系列人类疾病的基础，从阿尔茨海默病到糖尿病。相比之下，健康个体的细胞通过抗氧化化合物的合成、氧化损伤的快速修复和细胞还原剂池的有效平衡来有效地管理ROS。后一种策略，氧化还原平衡，目前尚不清楚，主要是由于缺乏生理学上相关的动物模型来研究这一动态过程。我们最近的工作表明，拟南芥中无机氮营养的简单，生物学上现实的操作可以快速和可预测地改变细胞氧化还原状态，导致线粒体呼吸链中几种氧化还原平衡酶的表达和活性的适应性变化。这些结果表明，拟南芥可以作为研究氧化还原平衡及其在ROS管理中的作用的独特模型。因此，本项目的主要目的是表征拟南芥细胞对氮源诱导的细胞氧化还原状态变化的全球反应。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（Specific Aim 2）。微阵列数据将使用旨在识别和统计量化生化途径协调调节的软件进行分析，我们特别感兴趣的是检查涉及生产，细胞内分配和还原剂氧化的途径（即潜在的氧化还原平衡途径）。除了研究基因表达水平上的氧化还原平衡外，我们还将直接研究氮源如何影响细胞ROS水平（H2O2）、每次氧化的脂质以及抗氧化剂池（谷胱甘肽和抗坏血酸）的大小和氧化状态（Specific Aim 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和定义的转录反应的定量评估联系起来。最终，这项工作将为建立一个全球性的、实验支持的细胞氧化还原平衡模型提供信息，这一过程对维持人类健康至关重要。

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.