串联病变,由两个或多个连续受损的核苷酸定义,是电离辐射的标志.最近,发现了含有5-甲酰基-2'-脱氧尿苷(5-fdU)的串联病变,侧翼为5'-8-OxodGuo或Fapy•dG,它们在人类细胞中比孤立的病变更具诱变性。在目前的研究中,我们检查了这些串联病变在大肠杆菌中的复制。与孤立的病变相比,两个串联病变的旁路效率降低了30-40%。分离的8-OxodGuo和Fapy•dG的突变频率(MFs)较低,并且没有从5-fdU构建体的复制中分离出突变体。8-OxodGuo的突变类型是靶向G→T变性,而Fapy•dG主要产生G→T和G缺失。5\'-8-OxodGuo-5-fdU也仅给出G→T突变,增加了3倍和11倍,没有和有SOS感应,分别,与孤立的8-OxodGuo相比。在mutY/mutM细胞中,8-OxodGuo和5'-8-OxodGuo-5-fdU的MF增加了13倍和7倍,分别。在PolII和PolIV缺陷细胞中,5'-8-OxodGuo-5-fdU的MF增加了2倍和3倍,分别,这表明这些聚合酶在很大程度上没有错误的旁路。5'-Fapy•dG-5-fdU的MF相似,没有(13±1%)和(16±2%)SOS诱导。与先前在人类细胞中报道的5'-Fapy•dG-5-fdU的复杂突变谱不同,以G→T为主要类型的错误,在大肠杆菌中,突变主要来自5-fdU的缺失。我们推测,在串联5\'-8-OxodGuo-5-fdU中,Fpg和MutY修复蛋白对与8-OxodGuo相反的腺嘌呤的去除部分受损,导致G→T突变的增加,而滑移机制可能在5'-Fapy•dG-5-fdU诱变中运行。这项研究表明,这些串联病变不仅比分离的病变更具诱变性,而且在不同的生物体中也可能表现出不同类型的突变。
Tandem lesions, which are defined by two or more contiguously damaged nucleotides, are a hallmark of ionizing radiation. Recently, tandem lesions containing 5-formyl-2\'-
deoxyuridine (5-fdU) flanked by a 5\'-8-OxodGuo or Fapy•dG were discovered, and they are more mutagenic in human cells than the isolated lesions. In the current study, we examined replication of these tandem lesions in Escherichia coli. Bypass efficiency of both tandem lesions was reduced by 30-40% compared to the isolated lesions. Mutation frequencies (MFs) of isolated 8-OxodGuo and Fapy•dG were low, and no mutants were isolated from replication of a 5-fdU construct. The types of mutations from 8-OxodGuo were targeted G → T transversion, whereas Fapy•dG predominantly gave G → T and G deletion. 5\'-8-OxodGuo-5-fdU also gave exclusively G → T mutation, which was 3-fold and 11-fold greater, without and with SOS induction, respectively, compared to that of an isolated 8-OxodGuo. In mutY/mutM cells, the MF of 8-OxodGuo and 5\'-8-OxodGuo-5-fdU increased 13-fold and 7-fold, respectively. The MF of 5\'-8-OxodGuo-5-fdU increased 2-fold and 3-fold in Pol II- and Pol IV-deficient cells, respectively, suggesting that these polymerases carry out largely error-free bypass. The MF of 5\'- Fapy•dG-5-fdU was similar without (13 ± 1%) and with (16 ± 2%) SOS induction. Unlike the complex mutation spectrum reported earlier in human cells for 5\'- Fapy•dG-5-fdU, with G → T as the major type of errors, in E. coli, the mutations were predominantly from deletion of 5-fdU. We postulate that removal of adenine-incorporated opposite 8-OxodGuo by Fpg and MutY repair proteins is partially impaired in the tandem 5\'-8-OxodGuo-5-fdU, resulting in an increase in the G → T mutations, whereas a slippage mechanism may be operating in the 5\'- Fapy•dG-5-fdU mutagenesis. This study showed that not only are these tandem lesions more mutagenic than the isolated lesions but they may also exhibit different types of mutations in different organisms.