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Oxidative DNA damage-induced tumorigenesis and its avoidance mechanism

DNA氧化损伤诱导肿瘤发生及其避免机制

基本信息

批准号：
12213098
负责人：
TSUZUKI Teruhisa
金额：
$ 49.66万
依托单位：
Kyushu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2004
项目状态：
已结题

项目摘要

Oxygen radicals, which can be produced through normal cellular metabolism, are thought to play an important role in mutagenesis and tumorigenesis. Among various classes of oxidative DNA damage, 8-oxo-7,8-dihydroguanine (8-oxoG) is the most abundant, and appears to play important roles in mutagenesis and carcinogenesis. Studies with Escherichia coli mutator mutants revealed that organisms possess elaborate mechanisms that prevent mutations caused by oxidation of the guanine base, in both DNA and free nucleotide forms. Enzymatic activities which may be responsible for preventing 8-oxoG-evoked mutations were identified in mammalian cells. We have focused on following the two enzymes. MTH1 (Mthl) protein is the mammalian counterpart of E. coli MutT protein, which hydrolyzes 8-oxo-dGTP to monophosphate in the nucleotide pool, thereby preventing occurrence of transversion mutations. On the other hand, MUTYH (Mutyh) protein, a counterpart of E. coli MutY protein, having adenine/2-hydroxyadeni … More ne DNA glycosylase activity, is expected to prevent G : C to T : A transversions, by excising adenine from G : A mismatches induced by 8-oxoG and 2-OH-A. To analyze the function of the mammalian Mthl and Mutyh proteins in vivo, we established gene-knockout mice for these two enzymes by gene targeting, and investigated spontaneous tumorigenesis as well as mutagenesis.When examined 18 months after birth, a greater number of tumors were formed in the lungs, livers of Mthl-deficient mice, as compared with wild-type mice (Tsuzuki, T. et al., 2001). Mutation frequencies on the rpsL transgene in spleen samples recovered at the age of 4 and 24 weeks, were determined. The spontaneous mutation frequency observed in spleen samples from Mthl-deficient mice showed no dramatic increase compared to the value of the one in wild-type mice. The site distribution of the mutations occurred on rpsL gene was slightly different between these to Mthl genotypes in spleen samples. In Mthl-deficient mice, there are 1-basepair frameshift mutations at the mononucleotide repeats those were not found in wild-type mice (Egashira, A. et al., 2002). When examined 18 months after birth, a greater number of tumors had formed in various tissues of Mutyh-deficient mice, as compared with wild-type mice. Especially, more small intestinal tumors were formed in Mutyh-deficient mice than in wild-type mice (in preparation). Mutation frequency observed in spleen samples from the Mutyh-deficient mice, at the age of 24 weeks, showed no apparent increase compared to the value of samples from wild-type mice. However, the site distribution of the mutations that occurred in the rpsL gene was significantly different between these two Mutyh genotypes ; an increase in frequency of G : C to T : A transversions was evident in Mutyh nullizygous mice (in preparation).Thus, both the Mthl-and Mutyh-deficient mice will provide useful models for investigating the roles of oxidative stress in human health. Less

氧自由基可以通过正常的细胞代谢产生，被认为在诱变和肿瘤发生中起重要作用。在各种类型的氧化性DNA损伤中，8-氧代-7，8-二氢鸟嘌呤（8-oxoG）是最丰富的，并且似乎在致突变和致癌中起重要作用。对大肠杆菌突变体的研究表明，生物体具有复杂的机制，可以防止DNA和游离核苷酸形式的鸟嘌呤碱基氧化引起的突变。在哺乳动物细胞中鉴定了可能负责防止8-oxoG诱发突变的酶活性。我们重点关注这两种酶。MTH 1（Mth 1）蛋白是大肠杆菌的哺乳动物对应物。coli MutT蛋白，其将核苷酸库中的8-氧代-dGTP水解为单磷酸，从而防止颠换突变的发生。另一方面，MUTYH（Mutyh）蛋白，一种与E.大肠杆菌MutY蛋白，具有腺嘌呤/2-羟基腺嘌呤 ...更多信息新的DNA糖基化酶活性，预期通过从由8-oxoG和2-OH-A诱导的G：A错配中切除腺嘌呤来防止G：C到T：A颠换。为了分析哺乳动物Mthl和Mutyh蛋白在体内的功能，我们通过基因靶向建立了这两种酶的基因敲除小鼠，并研究了自发肿瘤发生以及诱变。例如，2001年）的报告。测定在4周龄和24周龄时回收的脾样品中rpsL转基因的突变频率。在Mthl缺陷小鼠的脾样品中观察到的自发突变频率与野生型小鼠中的值相比没有显示出显著增加。rpsL基因突变的位点分布与Mthl基因型之间略有不同。在Mthl缺陷型小鼠中，在单核苷酸重复序列处存在1个碱基对移码突变，这些突变在野生型小鼠中未发现（Egashira，A.例如，2002年）。出生后18个月检查时，与野生型小鼠相比，Mutyh缺陷小鼠的各种组织中形成了更多的肿瘤。特别是，在Mutyh缺陷型小鼠中形成的小肠肿瘤比野生型小鼠中形成的小肠肿瘤更多（制备中）。在24周龄时，在Mutyh缺陷小鼠的脾样品中观察到的突变频率与野生型小鼠的样品值相比没有明显增加。然而，这两种Mutyh基因型之间的rpsL基因中发生的突变的位点分布显著不同;在Mutyh缺失小鼠中G：C到T：A颠换的频率增加是明显的（在制备中）。因此，Mthl和Mutyh缺陷小鼠将为研究氧化应激在人类健康中的作用提供有用的模型。少