Differential Regulation of the Phenazine Biosynthetic Operons by Quorum Sensing in Pseudomonas aeruginosa PAO1-N.
Differential Regulation of the Phenazine Biosynthetic Operons by Quorum Sensing in Pseudomonas aeruginosa PAO1-N.
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DOI：
10.3389/fcimb.2018.00252
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发表时间：
2018
影响因子：
5.7
通讯作者：
Cámara M
中科院分区：
文献类型：
--
作者：
Higgins S;Heeb S;Rampioni G;Fletcher MP;Williams P;Cámara M
铜绿假单胞菌群体感应（QS）网络在铜绿假单胞菌适应环境变化和控制毒力因子产生中起着关键作用。三个相互关联的QS系统，即las， rhl和pqs，是产生pyocyanin的核心，pyocyanin是一种吩那嗪毒力因子，通常用作分析QS的表型标记。P. aeruginosa的花青素生产是一个复杂的过程，涉及两个几乎相同的操纵子phzA1B1C1D1E1F1G1 （phz1）和phzA2B2C2D2E2F2G2 (phz2)，它们驱动吩嗪-1-羧酸（PCA）的生产，PCA通过两个修饰酶PhzM和PhzS进一步转化为花青素。由于phz1和phz2操纵子之间的高度序列保守性（核苷酸同源性> 98%），通过RNA杂交，qRT-PCR或转录组学分析它们的个体表达具有挑战性。为了克服这一困难，我们利用基于发光的启动子融合每个非那嘧啶操纵子来测量浮游培养中P. aeruginosa PAO1-N基因背景在las， rhl和pqs QS系统的不同组分中受损的转录活性，在不同QS信号分子存在或不存在的情况下。利用这种方法，我们发现所有三个QS系统都在差异调节phz1和phz2非那嗪操纵子中发挥作用，从而揭示了P. aeruginosa中PCA生物合成的QS调节的复杂性比之前所认识的要高。铜绿假单胞菌QS调控网络相互交织的方式在分析特定QS调控性状的调控机制时带来了挑战。多个QS调节因子和信号与非那嗪毒力因子的产生有关。在这项工作中，我们设计了实验，利用个体突变和互补策略剖析了特定QS开关的贡献，以进一步了解这些QS元件在控制P. aeruginosa phenazine操纵子的表达中的具体作用。利用这种方法，我们梳理出哪些QS调节剂对两种非那嗪生物合成操纵子的调节有间接或直接的影响。获得的数据突出了铜绿假单胞菌QS级联的复杂性以及分析控制非那嗪次级代谢物的挑战。
The Pseudomonas aeruginosa quorum sensing (QS) network plays a key role in the adaptation to environmental changes and the control of virulence factor production in this opportunistic human pathogen. Three interlinked QS systems, namely las, rhl, and pqs, are central to the production of pyocyanin, a phenazine virulence factor which is typically used as phenotypic marker for analysing QS. Pyocyanin production in P. aeruginosa is a complex process involving two almost identical operons termed phzA1B1C1D1E1F1G1 (phz1) and phzA2B2C2D2E2F2G2 (phz2), which drive the production of phenazine-1-carboxylic acid (PCA) which is further converted to pyocyanin by two modifying enzymes PhzM and PhzS. Due to the high sequence conservation between the phz1 and phz2 operons (nucleotide identity > 98%), analysis of their individual expression by RNA hybridization, qRT-PCR or transcriptomics is challenging. To overcome this difficulty, we utilized luminescence based promoter fusions of each phenazine operon to measure in planktonic cultures their transcriptional activity in P. aeruginosa PAO1-N genetic backgrounds impaired in different components of the las, rhl, and pqs QS systems, in the presence or absence of different QS signal molecules. Using this approach, we found that all three QS systems play a role in differentially regulating the phz1 and phz2 phenazine operons, thus uncovering a higher level of complexity to the QS regulation of PCA biosynthesis in P. aeruginosa than previously appreciated. The way the P. aeruginosa QS regulatory networks are intertwined creates a challenge when analysing the mechanisms governing specific QS-regulated traits. Multiple QS regulators and signals have been associated with the production of phenazine virulence factors. In this work we designed experiments where we dissected the contribution of specific QS switches using individual mutations and complementation strategies to gain further understanding of the specific roles of these QS elements in controlling expression of the two P. aeruginosa phenazine operons. Using this approach we have teased out which QS regulators have either indirect or direct effects on the regulation of the two phenazine biosynthetic operons. The data obtained highlight the sophistication of the QS cascade in P. aeruginosa and the challenges in analysing the control of phenazine secondary metabolites.