Nitric oxide-induced damage to mtDNA and its subsequent repair
Nitric oxide-induced damage to mtDNA and its subsequent repair
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DOI：
10.1093/nar/27.22.4510
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发表时间：
1999-11-15
影响因子：
14.9
通讯作者：
Wilson, GL
中科院分区：
文献类型：
--
作者：
Grishko, VI;Druzhyna, N;Wilson, GL
线粒体DNA(MtDNA)突变最近被认为与多种人类疾病有关。细胞持续暴露于其中一些突变可能导致的一种潜在的DNA损伤剂是一氧化氮(NO)。到目前为止，关于这种气体对线粒体DNA造成的损害的信息还很少。因此，本研究旨在探讨NO对线粒体DNA的损伤及其修复作用。正常人成纤维细胞暴露于由1-propanamine，3-(2-hydroxy-2-nitroso-1-propylhydrazinc)快速分解产生的NO(Papa NONOate)，并用定量Southern印迹法检测其对线粒体DNA的损伤。这种气体被发现会对碱敏感的线粒体DNA造成损伤。用尿嘧啶-DNA糖基酶或3-甲基腺嘌呤DNA糖基酶处理DNA，没有发现额外的损伤，表明大多数损伤是由鸟嘌呤脱氨为黄嘌呤引起的。使用连接介导的聚合酶链式反应的研究支持这一发现。当分析暴露于NO的细胞的200个碱基的线粒体DNA序列时，发现鸟嘌呤是主要的受损碱基。然而，特定的腺嘌呤也受到了损害。在嘧啶部位未观察到损伤。NO引起的损伤的核苷酸模式不同于由活性氧物种生成器或甲基化化学物质甲基亚硝脲产生的损伤。NO所致的损伤大部分能迅速修复。然而，似乎有一部分损伤要么缓慢修复，要么根本不被线粒体修复。
Mutations in mitochondrial DNA (mtDNA) have recently been associated with a variety of human diseases. One potential DNA-damaging agent to which cells are continually exposed that could be responsible for some of these mutations is nitric oxide (NO). To date, little information has been forthcoming concerning the damage caused by this gas to mtDNA. Therefore, this study was designed to investigate damage to mtDNA induced by NO and to evaluate its subsequent repair. Normal human fibroblasts were exposed to NO produced by the rapid decomposition of 1-propanamine,3-(2-hydroxy-2-nitroso-1-propylhydrazinc) (PAPA NONOate) and the resultant damage to mtDNA was determined by quantitative Southern blot analysis. This gas was found to cause damage to mtDNA that was alkali-sensitive. Treatment of the DNA with uracil-DNA glycosylase or 3-methyladenine DNA glycosylase failed to reveal additional damage, indicating that most of the lesions produced were caused by the deamination of guanine to xanthine. Studies using ligation-mediated PCR supported this finding. When a 200 bp sequence of mtDNA from cells exposed to NO was analyzed, guanine was found to be the predominantly damaged base. However, there also was damage to specific adenines. No lesions were observed at pyrimidine sites. The nucleotide pattern of damage induced by NO was different from that produced by either a reactive oxygen species generator or the methylating chemical, methylnitrosourea. Most of the lesions produced by NO were repaired rapidly. However, there appeared to be a subset of lesions which were repaired either slowly or not at all by the mitochondria.