Department of Genetics, University of Leicester, Leicester, UK.
Mol Microbiol. 2013 Jun;88(5):951-69. doi: 10.1111/mmi.12237. Epub 2013 May 7.
Previous analyses of luxS in Escherichia coli have used different strain backgrounds and design formats to produce the luxS mutation, resulting in luxS mutants with confusingly dissimilar phenotypes. This study therefore investigates the roles that strain background and mutational design strategy have upon the phenotype of the pathogenic E. coli luxS mutant. We inactivated luxS in three E. coli backgrounds: enteropathogenic E. coli E2348-69, and enterohaemorrhagic strains Sakai and NCTC12900. To investigate the influence of mutational design strategy, four mutation formats were used: antibiotic resistance insertion methodologies as previously employed, using tetracycline and chloramphenicol resistance cassettes, and non-polar strategies creating deletion and premature termination mutations. Our study showed that the E. coli luxS phenotype was markedly dependent on strain background: in some strains disruption of luxS caused significant metabolic stress or no stress at all. How the luxS mutation was constructed also shaped its phenotype: non-polar mutants were very similar to wild type, while mutations made using the antibiotic insertion methodologies produced phenotypes defective in growth and virulence. Proteomic profiling of our luxS mutants showed only a few proteins were differentially expressed and those that were altered suggested a metabolic rather than communication role for the E. coli luxS gene product.
先前对大肠杆菌 luxS 的分析使用了不同的菌株背景和设计格式来产生 luxS 突变，导致 luxS 突变体表现出令人困惑的不同表型。因此，本研究调查了菌株背景和突变设计策略对致病性大肠杆菌 luxS 突变体表型的影响。我们在三种大肠杆菌背景中失活了 luxS：肠致病性大肠杆菌 E2348-69 以及肠出血性菌株 Sakai 和 NCTC12900。为了研究突变设计策略的影响，我们使用了四种突变格式：以前使用的抗生素抗性插入方法，使用四环素和氯霉素抗性盒，以及创建缺失和过早终止突变的非极性策略。我们的研究表明，大肠杆菌 luxS 表型明显依赖于菌株背景：在某些菌株中，luxS 的破坏会导致显著的代谢应激或根本没有应激。突变的构建方式也影响了其表型：非极性突变体与野生型非常相似，而使用抗生素插入方法构建的突变则导致生长和毒力缺陷的表型。我们的 luxS 突变体的蛋白质组学分析表明，只有少数蛋白质表达水平存在差异，而改变的蛋白质表明大肠杆菌 luxS 基因产物具有代谢而非通讯作用。
