Control of the E. coli SOS regulon expression by intracellular dNTP pool changes. Escherichia coli

We studied SOS mutator effect mediated by recA730 and changes in the dNTP pool. We found that established dNTP pool changes resulting from deficiencies in the ndk or dcd genes, had a strongly suppressive effect on the recA730 mutator effect. To investigate whether the observed reduction in SOS mutator effect is due to lowered expression of the entire SOS regulon, we performed microarray analysis of gene expression profiles in each of the single ndk, dcd, and recA730 strains, as well as the double recA730 ndk and recA730 dcd strains. Overall design: Comparison of transcriptomes of the bacterium Escherichia coli six strains: wt, dcd, ndk, recA730, recA730 dcd and recA730 ndk. All strains were derivatives of the MC4100 strain, carrying a sulA366 allele (Δ(argF-lac)169 sulA366). dcd and ndk alleles used were dcd::kan and ndk::cam, respectively. Strains were compared in pairs: wt vs dcd, wt vs ndk, wt vs recA730, recA730 vs recA730 dcd and recA730 vs recA730 ndk. Two or three biological replicates for each strain were used. Each biological replicate had two technical replicates with dye swapping.

本研究针对recA730介导的SOS诱变因子效应（SOS mutator effect）以及脱氧核苷三磷酸（dNTP）池的变化展开探究。研究发现，由ndk或dcd基因缺陷所导致的稳定dNTP池变化，可对recA730诱变因子效应产生显著的抑制作用。为探明本次观测到的SOS诱变因子效应减弱是否源于整个SOS调节子（SOS regulon）的表达水平下调，我们分别对单突变株ndk、dcd及recA730，以及双突变株recA730 ndk和recA730 dcd的基因表达谱开展了基因芯片分析（microarray analysis）。总体实验设计：本研究对6株大肠杆菌（Escherichia coli）的转录组进行比较，受试菌株分别为野生型（wt）、dcd突变株、ndk突变株、recA730突变株、recA730 dcd双突变株以及recA730 ndk双突变株。所有菌株均为MC4100菌株（MC4100 strain）的衍生株，均携带sulA366等位基因（sulA366 allele），基因型为Δ(argF-lac)169 sulA366。本研究所用的dcd和ndk等位基因分别为dcd::kan与ndk::cam。菌株间以配对方式开展比较：野生型与dcd、野生型与ndk、野生型与recA730、recA730与recA730 dcd以及recA730与recA730 ndk。每个菌株设置2~3个生物学重复（biological replicates），每个生物学重复搭配2个技术重复（technical replicates）并进行染料互换（dye swapping）。