ADAPTIVE RESISTANCE TO NITRIC OXIDE

对一氧化氮的适应性抵抗

• 批准号: 6377414
• 负责人: Bruce F. Demple
• 金额: $ 28.25万
• 依托单位: HARVARD SCHOOL OF PUBLIC HEALTH
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-17 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6377414
• 关键词: biological signal transduction; cell type; cytotoxicity; enzyme mechanism; fibroblasts; gene targeting; genetic regulation; genetic transcription; genetically modified animals; heme oxygenase; laboratory mouse; messenger RNA; mitogen activated protein kinase; neoplasm /cancer genetics; nitric oxide

Nitric oxide (NO) is produced in mammalian cells by several NO synthases, including an inducible form with high output during inflammation. Since NO may contribute to carcinogenesis and age- related neurodegeneration, we wish to understand how mammalian cells counteract NO toxicity. We have identified a group of 12 proteins induced in normal human fibroblasts by exposure to subtoxic levels of NO, including the antioxidant enzyme heme oxygenase 1 (HO-1). We have shown that the induction of HO-1 mRNA occurs via NO-dependent changes in its stability. We will explore the mechanism of this regulation and try to define the signal transduction pathway for post-transcriptional control of HO-1. Other genes (e.g., MAP kinase phosphatase MKP-1/CL100) are controlled transcriptionally, and we will test specific transcription regulators for their roles in this expression. We have identified an adaptive resistance to NO in the rodent motor neurons: cells exposed to low-level NO that induces HO-1 become resistant to much higher (usually cytotoxic) levels of NO. We will characterize this response in various cell types by identifying NO-inducible genes and determining their roles in protection against NO-mediated toxicity. HO-1 knockout mice will allow us to define the defensive role of HO-1 in fibroblasts, neurons, and other tissues. This project involves parallel efforts to follow biochemical markers of nitric oxide damage (e.g., nitrotyrosine), to use genetics to establish the role of HO-1 in inducible resistance to NO, to elucidate regulatory and signal transduction pathways, and to identify and clone novel defense genes for NO resistance.

一氧化氮(NO)在哺乳动物细胞中由几种NO合成酶产生，其中包括一种在炎症过程中高产量的诱导型NO合成酶。由于NO可能参与癌症的发生和与年龄相关的神经退变，我们希望了解哺乳动物细胞如何抵消NO的毒性。我们已经确定了一组12种蛋白质，通过暴露于亚毒性水平的NO，包括抗氧化酶血红素加氧酶1(HO-1)，在正常的人成纤维细胞中诱导。我们已经证明，HO-1mRNA的诱导是通过其稳定性的NO依赖的变化而发生的。我们将探索这种调控的机制，并试图确定HO-1转录后调控的信号转导途径。其他基因(如MAP激酶磷酸酶MKP-1/CL100)受转录控制，我们将测试特定的转录调节因子在这种表达中的作用。我们已经在啮齿动物运动神经元中发现了对NO的适应性抵抗：暴露于低水平NO的细胞会对更高水平的NO(通常是细胞毒性的)产生抵抗。我们将通过识别NO诱导的基因并确定它们在保护NO介导的毒性中的作用来表征不同类型的细胞的这种反应。HO-1基因敲除小鼠将使我们能够确定HO-1在成纤维细胞、神经元和其他组织中的防御作用。该项目包括追踪一氧化氮损伤的生化标记物(如硝基酪氨酸)，利用遗传学来确定HO-1在NO诱导抗性中的作用，阐明调控和信号转导途径，以及鉴定和克隆与NO抗性相关的新的防御基因。