CONTROL OF PHAGOCYTE NADPH OXIDASE BY CYTOSOLIC PROTEINS

胞浆蛋白对吞噬细胞 NADPH 氧化酶的控制

• 批准号: 6686322
• 负责人: Bernard M Babior
• 金额: $ 29.17万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-07-01 至 2005-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6686322
• 关键词: NAD(P)H dehydrogenase; actin binding protein; biological signal transduction; chronic granulomatous disease; clinical research; cytoplasm; enzyme activity; enzyme biosynthesis; enzyme inhibitors; fluorescence microscopy; guanine nucleotide exchange factors; guanosinetriphosphatase activating protein; guanosinetriphosphatases; human subject; immunofluorescence technique; inflammation; laboratory rabbit; neutrophil; phagocytosis; vesicle /vacuole

DESCRIPTION (Applicant's Description Verbatim): The superoxide(02-)-generating NADPH oxidase of phagocytes is a potent source of reactive oxygen species (ROS). Its importance to innate immunity is manifested by the inherited syndrome chronic granulomatous disease, in which oxidase activity is absent and patients are susceptible to life-threatening microbial infections. O2-and its derivatives also cause severe tissue damage, contributing to inflammatory diseases such as ischemia-reperfusion injury and acute respiratory distress syndrome, and their prolonged generation in chronic inflammation can lead to dysplasia and malignancy. ROS produced by homologous systems in non-phagocytes are implicated in abnormal cell growth. The long-term objectives of the proposed research are to understand the mechanisms that control NADPH oxidase assembly and activity, and to identify steps that can be inhibited to minimize tissue damage. These four specific aims address gaps in knowledge that impede achievement of these goals. (1) Identify the processes of molecular recognition that encode the complex of cytosolic oxidase proteins to associate with the flavocytochrome to form a isolated phagosomes and cellular fractions, will be used to study the formation of the active enzyme and determine the roles of coronin and cofilin in controlling its assembly. 2) Analyze interactions between NADPH oxidase components, and identify factors that promote their association and dissociation. A biosensor instrument will be used to quantitate interactions between oxidase-related proteins and study how they are modified by changes that mimic intracellular signals. Development of this technique will provide an assay for molecules designed to inhibit oxidase activation. 3) Identify and isolate GTPase activating proteins and guanine nucleotide exchange factors for Rac2 and study their involvement in controlling Rac and NADPH oxidase activity. The identities of the regulatory proteins that control Rae activity in the neutrophil by altering its GDP/GTP-bound state are not known. These proteins are likely to be important in the negative and positive regulation of 02- generation. 4) Identify signaling events and molecules that terminate superoxide generation. The processes that switch off the enzyme have important clinical implications for inflammatory disease, but they are not well understood. Cell-free oxidase systems and intact cells will be used to determine if a) phosphoinositides; b) changes in the phosphorylation state of phox proteins; and c) changes in the GDP/GTP-bound state of Rae are negative regulators of the enzyme.

描述(申请人逐字描述)：超氧化物(02-)-生成 吞噬细胞的NADPH氧化酶是一种有效的活性氧来源 (ROS)。它对先天免疫的重要性体现在遗传性 慢性肉芽肿性疾病，其中缺乏氧化酶活性和 患者容易受到危及生命的微生物感染。O2-及ITS 衍生物还会造成严重的组织损伤，导致炎症 缺血再灌注损伤和急性呼吸窘迫等疾病 综合症，以及它们在慢性炎症中的长期存在可导致 不典型增生和恶性。非吞噬细胞中同源系统产生的ROS 与细胞的异常生长有关。该计划的长远目标 建议的研究是为了了解控制NADPH氧化酶的机制 装配和活动，并确定可以被禁止以将其降至最低的步骤 组织损伤。这四个具体目标解决了阻碍知识差距的问题 实现这些目标。(1)识别分子识别过程 编码胞浆氧化酶蛋白的复合体，与 黄色素形成一个孤立的吞噬小体和细胞组分，将 用于研究活性酶的形成并确定其作用 冠状病毒和粘附素在控制其组装中的作用。2)分析交互 NADPH氧化酶组分之间的关系，并确定促进其 联想和解离。将使用生物传感器仪器来定量 氧化酶相关蛋白之间的相互作用及其修饰研究 通过模拟细胞内信号的变化。这项技术的发展将 提供一种检测旨在抑制氧化酶激活的分子的方法。3) GTP酶激活蛋白的鉴定及鸟嘌呤核苷酸交换 RAC2的影响因素及其在RAC和NADPH控制中的作用 氧化物酶活性。控制RAE的调控蛋白的特性 中性粒细胞通过改变其GDP/GTP结合状态的活性尚不清楚。 这些蛋白质很可能在阴阳两方面都很重要。 调控02代。4)识别信号事件和分子 终止超氧化物的产生。关闭这种酶的过程有 炎症性疾病的重要临床意义，但它们并不是很好 明白了。无细胞的氧化酶系统和完整的细胞将被用于 确定a)磷脂酰肌醇；b)磷酸化状态的变化 PHOX蛋白；以及c)RAE的GDP/GTP结合态的变化是负的 酶的调节器。