UVB-and sunlight-induced mutagenesis in phage M13mp2 and a possible role of guanine modification in its DNA

UVB 和阳光诱导的噬菌体 M13mp2 突变及其 DNA 中鸟嘌呤修饰的可能作用

基本信息

批准号：
05807208
负责人：
NEGISHI Kazuo
金额：
$ 1.15万
依托单位：
Okayama University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

项目摘要

Sunlight causes predominantly G-to-C mutation in lacZd gene of phage M13mp2. This result indicated a specific guanine modification of the DNA produced by sunlight. UVB is believed to be a toxic principle in sunlight. We analyzed the spectra of mutations induced by UVB.When SOS-induced E.coli were used as host cells, UVB induced the mutations of M13mp2 effieciently. In a host lacking the induction of SOS function, the induction of mutations was not very efficient. The induction was, however, still significant. The sequencing analysis showed the induction of transitions at pyrimidine bases on assay using SOS-induced E.coli as host cells. In contrast, mutations at guanine were seen most frequently in the experiments on a SOS-deficient host. These results indicate that UVB and sunlight can produce a damage on guanine of M13mp2DNA.The modification of guanine might be caused by active oxygens. To know whether sunlight and UVB can generate active oxygens in M13mp2 and DNA itself, we analyzed the formation of 8-oxo-guanine. Upon irradiation by sunlight or UVB,8-oxo-guanine was formed depending on the presence of oxygens. 8-Oxo-guanine itself seems not to cause the mutagenesis, because it is known to induce G-to-T transversions. Some other oxidative modification seems to occur in the DNA and induce these mutations.To compare with mutations induced by a chemical oxidative reagent, we analyzed spectra of E.coli mutations caused by superoxide generated by menadion. In wild-type cells, all possible base-pair changes were observed. In a mutM strain, however, GC-to-CG transversions were most frequent. This result suggests that damages removable by the action of MutM protein, a ring-opened guanine, for example, might cause this type of mutations both in sunlight-or UVB-treated M13mp2 and superoxide-mutagenesis in E.coli.

阳光主要导致噬菌体M13MP2的LACZD基因中的G-TO-C突变。该结果表明阳光产生的DNA的特异性鸟嘌呤修饰。 UVB被认为是阳光下的有毒原则。我们分析了UVB诱导的突变光谱。当SOS诱导的大肠杆菌用作宿主细胞时，UVB效应地诱导了M13MP2的突变。在缺乏SOS功能诱导的宿主中，突变的诱导不是很好。但是，诱导仍然很重要。测序分析表明，使用SOS诱导的大肠杆菌作为宿主细胞，在测定时在嘧啶基碱基处的过渡诱导。相反，在SOS缺陷宿主的实验中，最常看到鸟嘌呤的突变。这些结果表明，UVB和阳光会对M13MP2DNA的鸟嘌呤造成损害。鸟嘌呤的修饰可能是由活性氧引起的。为了知道阳光和UVB是否可以在M13MP2和DNA本身中产生活性氧，我们分析了8-氧气 - 瓜氨酸的形成。在阳光或UVB辐照后，根据氧气的存在形成8-氧气的瓜氨酸。 8-氧气 - 瓜氨酸本身似乎并没有引起诱变，因为已知它会诱导G-T-T转移。 DNA中似乎发生了其他一些氧化修饰并诱导这些突变。要与化学氧化试剂诱导的突变进行比较，我们分析了由MANADION产生的超氧化物引起的E.coli突变的光谱。在野生型细胞中，观察到所有可能的碱基对变化。然而，在MUTM菌株中，GC-CG横向横向最常见。该结果表明，例如，在阳光或UVB处理的M13MP2和E.Coli中的超氧化物 - 氧化物中，通过MUTM蛋白的作用（一种响环的鸟嘌呤）可消除的损害可能导致这种突变。