Repair of formamidopyrimidines in DNA involves different glycosylases - Role of the OGG1, NTH1, and NEIL1 enzymes
Repair of formamidopyrimidines in DNA involves different glycosylases - Role of the OGG1, NTH1, and NEIL1 enzymes
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DOI：
10.1074/jbc.m508772200
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发表时间：
2005-12-09
影响因子：
4.8
通讯作者：
Bohr, VA
中科院分区：
文献类型：
--
作者：
Hu, JP;de Souza-Pinto, NC;Bohr, VA
氧化诱导的DNA损伤2，6-二氨基-4-羟基-5-甲酰胺基嘧啶（FapyG）和4，6-二氨基-5-甲酰胺基嘧啶（FapyA）在暴露于电离辐射或其他自由基产生系统的培养细胞或组织的DNA中大量形成。体外研究表明，这些病变是错误编码的，可以阻断DNA聚合酶的进展，并且是碱基切除修复的底物。然而，还没有研究解决这些病变如何在细胞提取物中代谢。最近报道了在限定位置含有FapyG和FapyA的寡核苷酸的合成。这些构建体使我们能够研究缺乏两种主要DNA糖基化酶（OGG 1和NTH 1）的野生型和基因敲除小鼠的细胞核和线粒体提取物中FRENTA损伤的修复。还测定了来自这些小鼠的DNA中FapyG/FapyA的背景水平。来自野生型小鼠的肝DNA中的内源性FapyG水平显著高于8-羟基鸟嘌呤水平。FapyG和FapyA在野生型动物的核和线粒体提取物中有效修复，但在糖基化酶缺陷小鼠中没有。我们的研究结果表明，OGG 1和NTH 1是主要的DNA糖基化酶的去除FapyG和FapyA，分别。组织特异性分析表明，当NTH 1表达不足时，其他DNA糖基化酶可能有助于FapyA修复。我们在肝线粒体中鉴定了NEIL 1，这可以解释在OGG 1和NTH 1不存在的情况下残留的切割活性。FapyG和FapyA水平在来自敲除小鼠的DNA中显著升高，强调了OGG 1和NTH 1在这些病变修复中的生物学作用。
The oxidatively induced DNA lesions 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) and 4,6-diamino-5-formamidopyrimidine (FapyA) are formed abundantly in DNA of cultured cells or tissues exposed to ionizing radiation or to other free radical-generating systems. In vitro studies indicate that these lesions are miscoding, can block the progression of DNA polymerases, and are substrates for base excision repair. However, no study has yet addressed how these lesions are metabolized in cellular extracts. The synthesis of oligonucleotides containing FapyG and FapyA at defined positions was recently reported. These constructs allowed us to investigate the repair of Fapy lesions in nuclear and mitochondrial extracts from wild type and knock-out mice lacking the two major DNA glycosylases for repair of oxidative DNA damage, OGG1and NTH1. The background level of FapyG/FapyA in DNA from these mice was also determined. Endogenous FapyG levels in liver DNA from wild type mice were significantly higher than 8-hydroxyguanine levels. FapyG and FapyA were efficiently repaired in nuclear and mitochondrial extracts from wild type animals but not in the glycosylase-deficient mice. Our results indicated that OGG1 and NTH1 are the major DNA glycosylases for the removal of FapyG and FapyA, respectively. Tissue-specific analysis suggested that other DNA glycosylases may contribute to FapyA repair when NTH1 is poorly expressed. We identified NEIL1 in liver mitochondria, which could account for the residual incision activity in the absence of OGG1 and NTH1. FapyG and FapyA levels were significantly elevated in DNA from the knock-out mice, underscoring the biological role of OGG1 and NTH1 in the repair of these lesions.