荧光假单胞菌2P24中retS对抗生素2，4-二乙酰基间苯三酚合成的影响

Effect of retS gene on biosynthesis of 2，4-diacetylphloroglucinol in Pseudomonas fluorescens 2P24

摘要:

摘要:假单胞菌中RetS是一个位于膜上的感应激酶，对多种基因的表达都有调控作用。在铜绿假单胞菌中，RetS可以与另一个感应激酶GacS直接互作，并抑制GacS的磷酸化。【目的】本文利用遗传学方法研究了荧光假单胞菌2P24中RetS对抗生素2，4-二乙酰基间苯三酚(2，4-DAPG)合成的影响，并对其可能的调控机制进行了初步探索。【方法】利用高压液相色谱法(HPLC)检测2P24及其衍生菌株中2，4-DAPG的产量。将Gac/Rsm信号途径中小RNA及调控蛋白的转录报告质粒转入到菌株2P24及其retS突变菌株中，检查RetS 对以上基因转录表达的影响。【结果】菌株2P24 中缺失retS后未知红色素和抗生素2，4-DAPG的产量较野生型均明显升高。进一步试验表明，RetS转录水平负调控小RNA RsmX和RsmZ的表达，这说明RetS可在转录后水平影响2，4-DAPG的合成。然而，同时缺失retS和gacS或同时缺失retS和gacA之后，由retS单基因缺失所造成的未知红色素和2，4-DAPG合成量升高、小RNA转录表达增强等性状消失，而与gacS或gacA单基因缺失突变体的表型一致。【结论】以上结果说明菌株2P24中RetS是2，4-DAPG及未知红色素合成的负调控因子，并且RetS对2，4-DAPG及未知红色素合成的调控依赖于Gac/Rsm信号传递路径。

Abstract:

Abstract:Regulator of exopolysaccharide and type Ⅲ secretion (RetS) is a hybrid sensor kinase/response regulator protein located on bacterial membrane，and essential for expression of numerous genes．In Pseudomonas aeruginosa，RetS modulates the phosphorylation state of another kinase GacS via a direct interaction．［Objective］The goal of this study is to study the effect of retS on the antibiotic 2，4-diacetyl-phloroglucinol (2，4-DAPG) production in P．fluorescens 2P24．［Methods］Production of 2，4-DAPG in strain 2P24 and its mutants was quantified by HPLC． To determine the effect of RetS on the Gac/Rsm pathway，the promoters of the small RNA genes rsmX/rsmY/rsmZ and regulatory genes rsmA/rsmE in the Rsm pathway were fused with a promoterless lacZ，and the promoter activities were measured in 2P24 and the retSdeficient mutants．［Results］Genetic inactivation of the retS in strain 2P24 increased the production of an uncharacterized red pigment and the antibiotic 2，4-DAPG．RetS negatively regulated the transcription of the small RNAs RsmX and RsmZ．In the retS and gacS double mutant or the retS and gacA double mutant，all the phenotypic changes caused by the retS deletion were reversed to the level of gacS or gacA single gene mutant．［Conclusion］In P．fluorescens 2P24，RetS negatively regulates the production of antibiotic 2，4-DAPG through the Gac/Rsm pathway.