肉桂酸衍生物对铜绿假单胞菌PAO1 Ⅲ型分泌系统的影响  

The influence of cinnamic acid derivatives on the type Ⅲ secretion system of Pseudomonas aeruginosa PAO1

作　　者：单金明[1] 吴小刚 袁小琛 方力为 梁翠荣[1] 陈新[1] YANG Ching-Hong 

机构地区：[1]常州大学制药与生命科学学院,江苏常州213164 [2]Department of Biological Sciences, University of Wisconsin-Milwaukee

出　　处：《微生物学通报》2014年第10期2061-2069,共9页Microbiology China

基　　金：国家自然科学基金面上项目(No.21272029)

摘　　要：【目的】铜绿假单胞菌是引起医院获得性感染最常见的条件致病菌,而Ⅲ型分泌系统(Type Ⅲ secretion system,TTSS)是其致病的主要因子之一。本文从合成的21个肉桂酸衍生物中筛选影响TTSS效应子(Effector)产生的化合物,并初步研究其作用机制。【方法】将TTSS效应子合成基因exoS的转录报告质粒pAT-exoS转入菌株PAO1中,获得PAO1(pAT-exoS)。待筛选的化合物与PAO1(pAT-exoS)菌株共培养6 h后,检测exoS基因的表达,从中筛选影响exoS基因表达的化合物。【结果】筛选结果表明:21个化合物中,3个化合物抑制exoS基因表达,2个化合物则促进exoS基因表达。此外,化合物TS128、TS143和TS160对菌株生长有明显的抑制作用。Western blot实验进一步证实筛选得到的化合物TS108、TS128和TS165可抑制ExoS的产生;化合物TS139和TS143则促进ExoS的产生。为进一步研究抑制剂的作用机理,过量表达TTSS主要的调控因子exsA基因可部分消除抑制剂TS108和TS165的抑制效果;而rsmZ rsmY双基因突变体PAO6421中添加抑制剂TS108和TS165并不能显著抑制exoS基因的表达,同样,抑制剂TS108和TS165也不影响受Gac/Rsm信号传导系统调控的群体感应信号分子的产生。【结论】抑制剂TS108和TS165的作用机制可能主要是影响esxA基因,从而影响exoS基因表达及蛋白产量。[Objective] Pseudomonas aeruginosa, an opportunistic human pathogen, possesses a type Ⅲ secretion system（TTSS） as one of the major virulence factors by which it secrets and translocateseffector proteins into human host cells. In this study, five compounds which influence the expression of TTSS were identified from cinnamic acid derivatives. [Methods] The transcriptional reporter of exoS gene on plasmid pAT-exoS was used to monitor the exoS expression in the wild type strain PAO1 with the candidate compounds. [Results] Among 21 compounds, three compounds decreased the expression of exoS gene, whereas two compounds increased the expression of exoS gene. In addition, the compounds TS128, TS143 and TS160 showed inhibitory effect in bacterial growth.Western blot assay further showed that the level of ExoS was altered when the medium was added TS108 and TS165. The expression of exoS was partly restored to the wild-type when the plasmid p35-exsA which containing the exsA gene was introduced into PAO1. However, there is no difference of the exoS expression in the rsmY rsmZ double mutant with inhibitors TS108 and TS165. In addition, the production of N-Acyl homoserine lactones, which is positively regulated by the Gac/Rsm signaling pathway, is not affected with the inhibitors TS108 and TS165. [Conclusion] Our results suggest that the inhibitors TS108 and TS165 affect the expression of exoS through the transcriptional regulator ExsA instead of the Gac/Rsm regulatory pathway.

关 键 词：PSEUDOMONAS AERUGINOSA ExoS 抑制剂 诱导剂 Ⅲ型分泌系统 

分 类 号：R378[医药卫生—病原生物学]