Departamento de Biofísica e Biometria, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ, Brazil.
Redox Rep. 2011;16(5):187-92. doi: 10.1179/1351000211Y.0000000010.
We previously demonstrated that reactive oxygen species (ROS) could be involved in the DNA damage induced by ultraviolet-C (UVC). In this study, we evaluated singlet oxygen ((1)O(2)) involvement in UVC-induced mutagenesis in Escherichia coli cells. First, we found that treatment with sodium azide, an (1)O(2) chelator, protected cells against UVC-induced lethality. The survival assay showed that the fpg mutant was more resistant to UVC lethality than the wild-type strain. The rifampicin mutagenesis assay showed that UVC mutagenesis was inhibited five times more in cells treated with sodium azide, and stimulated 20% more fpg mutant. These results suggest that (1)O(2) plays a predominant role in UVC-induced mutagenesis. (1)O(2) generates a specific mutagenic lesion, 8-oxoG, which is repaired by Fpg protein. This lesion was measured by GC-TA reversion in the CC104 strain, its fpg mutant (BH540), and both CC104 and BH540 transformed with the plasmid pFPG (overexpression of Fpg protein). This assay showed that mutagenesis was induced 2.5-fold in the GC-TA strain and 7-fold in the fpg mutant, while the fpg mutant transformed with pFPG was similar to GC-TA strain. This suggests that UVC can also cause ROS-mediated mutagenesis and that the Fpg protein may be involved in this repair.
我们之前已经证明，活性氧（ROS）可能参与了紫外线 C（UVC）诱导的 DNA 损伤。在这项研究中，我们评估了单线态氧（(1)O(2)）在大肠杆菌细胞中 UVC 诱导突变中的作用。首先，我们发现，使用叠氮化钠（一种 (1)O(2)螯合剂）处理可以保护细胞免受 UVC 诱导的致死性。存活实验表明，fpg 突变体比野生型菌株对 UVC 致死性的抗性更强。利福平诱变实验表明，用叠氮化钠处理的细胞中 UVC 诱变的抑制作用增加了五倍，并且 fpg 突变体的刺激作用增加了 20%。这些结果表明，(1)O(2)在 UVC 诱导的突变中起着主要作用。(1)O(2)产生一种特定的诱变损伤，8-氧鸟嘌呤（8-oxoG），由 Fpg 蛋白修复。这种损伤通过 CC104 菌株、其 fpg 突变体（BH540）以及 CC104 和 BH540 转化的质粒 pFPG（Fpg 蛋白的过表达）中的 GC-TA 回复来测量。该实验表明，GC-TA 菌株的突变诱导增加了 2.5 倍，fpg 突变体的突变诱导增加了 7 倍，而 pFPG 转化的 fpg 突变体与 GC-TA 菌株相似。这表明 UVC 也可以引起 ROS 介导的突变，并且 Fpg 蛋白可能参与这种修复。
