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Role of Prdx6 in the activation of NADPH oxidase

Prdx6 在 NADPH 氧化酶激活中的作用

基本信息

批准号：
8432046
负责人：
Aron B. FISHER
金额：
$ 49.63万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-02-01 至 2016-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8432046
关键词：
Acute Lung Injury Agonist Alveolar Macrophages Angiotensin II Applications Grants Binding Biochemical Pathway Cell membrane Cell-Free System Cells Concanavalin A Cysteine Data Development Endothelial Cells Endothelium Family Generations Goals Human In Vitro Inflammation Inflammatory Injury Ischemia Kinetics Knock-out Knockout Mice Lung Lung diseases Lysophosphatidylcholines Measurement Mediating Membrane Methods Modeling Molecular Multienzyme Complexes Mus NADPH Oxidase Null Lymphocytes Oxidants Oxidative Stress Pathway interactions Peroxidases Phospholipase A2 Phospholipids Phosphorylation Phosphotransferases Physiological Production Proteinase 3 Proteins Publications Reactive Oxygen Species Recombinant Proteins Reperfusion Therapy Role Signal Transduction Stimulus Testing Tissues cell injury cell type in vivo inhibitor/antagonist insight lung injury lung ischemia member neutrophil cytosol factor 67K novel peroxiredoxin phospholipase A2 inhibitor prevent protein protein interaction public health relevance reconstitution response

项目摘要

DESCRIPTION (provided by applicant): The goal of this proposal is to evaluate a novel role for Prdx6 in the activation of NADPH oxidase (NOX2). Our hypothesis is that in response to a stimulus, Prdx6 is phosphorylated and translocates to the plasma membrane where it generates lysophosphatidylcholine (lysoPC) resulting in activation of NOX2. This hypothesis will be tested by 3 (of 5) specific aims. Specific Aim 1 will evaluate the effect of Prdx6 "knockout" on agonist-induced activation of NOX2 in alveolar macrophages (AM) and pulmonary microvascular endothelial cells in culture and in the isolated perfused lung. Activation is determined by translocation of cytosolic components to the membrane and generation of reactive oxygen species (ROS). Specific Aim 2 will evaluate the mechanism for Prdx6-mediated activation of NOX2; we propose that generation of lysoPC by the phospholipase A2 (PLA2) activity of Prdx6 is responsible. Specific Aim 3 will evaluate the requirement for Prdx6 phosphorylation in its translocation to the plasma membrane and NOX2 activation. We further propose that binding of one of the cytosolic components (p67phox) to Prdx6 inhibits its PLA2 activity and abrogates the NOX2-activation signal. Specific Aim 4 will study the interaction of Prdx6 with p67phox in intact cells and with recombinant protein in vitro with specific focus on the PLA2 activity of Prdx6 and the kinetics of the protein-protein interaction. Finally, Specific Aim 5 will investigate the effect of a Prdx6 PLA2 inhibitor, MJ33, in preventing oxidative stress with ischemia in the isolated lung and acute lung injury with ischemia-reperfusion in vivo. We postulate that this agent will maintain the protective peroxidase activity of Prdx6 while inhibiting the activation of NOX2. The proposed studies will provide a coordinated effort to investigate this novel role of Prdx6 and will provide the basic insights for development of new methods to inhibit the activation of the NOX2 enzyme complex and ameliorate ROS-mediated lung injury.

描述（由申请人提供）：该提案的目的是评估 Prdx6 在 NADPH 氧化酶（NOX2）激活中的新作用。我们的假设是，响应刺激，Prdx6 被磷酸化并转移到质膜，在质膜中生成溶血磷脂酰胆碱 (lysoPC)，从而激活 NOX2。该假设将通过 3 个（共 5 个）具体目标进行检验。具体目标 1 将评估 Prdx6“敲除”对培养物和离体灌注肺中的肺泡巨噬细胞 (AM) 和肺微血管内皮细胞中激动剂诱导的 NOX2 激活的影响。激活是通过胞质成分向膜的易位和活性氧 (ROS) 的产生来确定的。具体目标 2 将评估 Prdx6 介导的 NOX2 激活机制；我们认为，Prdx6 的磷脂酶 A2 (PLA2) 活性产生了 lysoPC。具体目标 3 将评估 Prdx6 磷酸化在质膜易位和 NOX2 激活中的需求。我们进一步提出，胞质成分之一 (p67phox) 与 Prdx6 的结合会抑制其 PLA2 活性并消除 NOX2 激活信号。具体目标 4 将研究 Prdx6 与 p67phox 在完整细胞中的相互作用以及与体外重组蛋白的相互作用，特别关注 Prdx6 的 PLA2 活性和蛋白质-蛋白质相互作用的动力学。最后，具体目标 5 将研究 Prdx6 PLA2 抑制剂 MJ33 在预防离体肺缺血引起的氧化应激和体内缺血再灌注引起的急性肺损伤中的作用。我们假设该试剂将维持 Prdx6 的保护性过氧化物酶活性，同时抑制 NOX2 的激活。拟议的研究将协调一致地研究 Prdx6 的这一新作用，并将为开发抑制 NOX2 酶复合物激活和改善 ROS 介导的肺损伤的新方法提供基本见解。