Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología-Consejo Superior de Investigaciones Científicas (CSIC), Madrid, Spain.
PLoS Genet. 2010 May 6;6(5):e1000931. doi: 10.1371/journal.pgen.1000931.
Hypermutation may accelerate bacterial evolution in the short-term. In the long-term, however, hypermutators (cells with an increased rate of mutation) can be expected to be at a disadvantage due to the accumulation of deleterious mutations. Therefore, in theory, hypermutators are doomed to extinction unless they compensate the elevated mutational burden (deleterious load). Different mechanisms capable of restoring a low mutation rate to hypermutators have been proposed. By choosing an 8-oxoguanine-repair-deficient (GO-deficient) Escherichia coli strain as a hypermutator model, we investigated the existence of genes able to rescue the hypermutable phenotype by multicopy suppression. Using an in vivo-generated mini-MudII4042 genomic library and a mutator screen, we obtained chromosomal fragments that decrease the rate of mutation in a mutT-deficient strain. Analysis of a selected clone showed that the expression of NorM is responsible for the decreased mutation rate in 8-oxoguanine-repair-deficient (mutT, mutY, and mutM mutY) strains. NorM is a member of the multidrug and toxin extrusion (MATE) family of efflux pumps whose role in E. coli cell physiology remains unknown. Our results indicate that NorM may act as a GO-system backup decreasing AT to CG and GC to TA transversions. In addition, the ability of NorM to reduce the level of intracellular reactive oxygen species (ROS) in a GO-deficient strain and protect the cell from oxidative stress, including protein carbonylation, suggests that it can extrude specific molecules-byproducts of bacterial metabolism-that oxidize the guanine present in both DNA and nucleotide pools. Altogether, our results indicate that NorM protects the cell from specific ROS when the GO system cannot cope with the damage.
超突变可能会在短期内加速细菌进化。然而,从长期来看,由于有害突变的积累,高突变体(突变率增加的细胞)可能处于劣势。因此,从理论上讲,除非高突变体能够补偿增加的突变负担(有害负荷),否则它们注定会灭绝。已经提出了不同的机制来使高突变体恢复到低突变率。通过选择缺乏 8-氧鸟嘌呤修复(GO 缺陷)的大肠杆菌菌株作为高突变体模型,我们研究了通过多拷贝抑制恢复高突变表型的存在的基因。我们使用体内生成的 mini-MudII4042 基因组文库和突变体筛选,获得了降低 mutT 缺陷菌株突变率的染色体片段。对选定的克隆进行分析表明,NorM 的表达负责降低 8-氧鸟嘌呤修复缺陷(mutT、mutY 和 mutM mutY)菌株中的突变率。NorM 是多药和毒素外排(MATE)家族外排泵的成员,其在大肠杆菌细胞生理学中的作用尚不清楚。我们的结果表明,NorM 可能作为 GO 系统的备份,降低 AT 到 CG 和 GC 到 TA 的颠换。此外,NorM 能够降低 GO 缺陷菌株中的细胞内活性氧(ROS)水平并保护细胞免受氧化应激,包括蛋白质羰基化,表明它可以排出特定的分子——细菌代谢的副产物——氧化存在于 DNA 和核苷酸池中的鸟嘌呤。总的来说,我们的结果表明,当 GO 系统无法应对损伤时,NorM 可以保护细胞免受特定的 ROS 损伤。