Dataset Information

Microevolution of the mexT and lasR Reinforces the Bias of Quorum Sensing System in Laboratory Strains of Pseudomonas aeruginosa PAO1.

ABSTRACT: The Pseudomonas aeruginosa strain PAO1 has routinely been used as a laboratory model for quorum sensing (QS). However, the microevolution of P. aeruginosa laboratory strains resulting in genetic and phenotypic variations have caused inconsistencies in QS research. To investigate the underlying causes of these variations, we analyzed 5 Pseudomonas aeruginosa PAO1 sublines from our laboratory using a combination of phenotypic characterization, high throughput genome sequencing, and bioinformatic analysis. The major phenotypic variations among the sublines spanned across the levels of QS signals and virulence factors such as pyocyanin and elastase. Furthermore, the sublines exhibited distinct variations in motility and biofilm formation. Most of the phenotypic variations were mapped to mutations in the lasR and mexT, which are key components of the QS circuit. By introducing these mutations in the subline PAO1-E, which is devoid of such mutations, we confirmed their influence on QS, virulence, motility, and biofilm formation. The findings further highlight a possible divergent regulatory mechanism between the LasR and MexT in the P. aeruginosa. The results of our study reveal the effects of microevolution on the reproducibility of most research data from QS studies and further highlight mexT as a key component of the QS circuit of P. aeruginosa.

SUBMITTER: Liu Y

PROVIDER: S-EPMC9041413 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Microevolution of the mexT and lasR Reinforces the Bias of Quorum Sensing System in Laboratory Strains of Pseudomonas aeruginosa PAO1.

Liu Yang Y Ahator Stephen Dela SD Wang Huishan H Feng Qishun Q Xu Yinuo Y Li Chuhao C Zhou Xiaofan X Zhang Lian-Hui LH

Frontiers in microbiology 20220412

The Pseudomonas aeruginosa strain PAO1 has routinely been used as a laboratory model for quorum sensing (QS). However, the microevolution of P. aeruginosa laboratory strains resulting in genetic and phenotypic variations have caused inconsistencies in QS research. To investigate the underlying causes of these variations, we analyzed 5 Pseudomonas aeruginosa PAO1 sublines from our laboratory using a combination of phenotypic characterization, high throughput genome sequencing ...[more]

PMID: 35495693

Similar Datasets

Project description:Pseudomonas aeruginosa (Pa) is a ubiquitous bacterium that uses quorum sensing (QS), a cell-cell communication system that enables it to sense cell density and to alter gene expression. Pa has three complete QS circuits controlled by the transcriptional regulators LasR, RhlR, and PqsR (MvfR), that together control hundreds of genes, including virulence factors. In the well-described strain PAO1, QS is organized hierarchically, with PqsR and RhlR activity dependent on LasR. In PAO1, this hierarchy depends on the non-QS transcription factor MexT; by an unknown mechanism, deletion of mexT allows for RhlR activity in the absence of LasR. We aimed to identify how regulators such as MexT modulate the QS architecture in Pa. We compared the transcriptome of PAO1 to that of PAO1ΔmexT and identified 152 differentially expressed genes. MexT does not appear to regulate rhlR or pqsR directly; however, we identified two MexT-regulated operons that may affect the hierarchy in PAO1. These operons encode the drug efflux pump genes mexEF-oprN and the Pseudomonas quinolone signal (PQS) synthesis genes pqsABCDE. We performed genetic experiments to test whether the products of these genes affected the QS hierarchy. As with the mexT knockout mutant, we found that a PAO1 mexEF knockout mutant exhibited RhlR activity earlier, and to a higher magnitude, than wild-type PAO1. MexEF-OprN is known to export the PQS precursor HHQ, and we found that exogenous addition of PQS to PAO1 partially affects RhlR activity, resulting in earlier timing and higher magnitude compared to wild-type PAO1. We further elucidated that this is likely due to positive regulation by PqsE. These data link both the drug efflux pump MexEF-OprN and PQS QS to the regulation of the QS hierarchy in PAO1. We wondered if the same applied to QS architectures in Pa clinical isolates. We discovered that there are alternate QS architectures in clinical isolates, where RhlR activity is not fully dependent on LasR. In these isolates, surprisingly, MexT does not influence the relationship between LasR and RhlR, and this is indicative of a different QS architecture in the clinical isolates. Overall, we further elucidated the regulation of QS architecture in PAO1 and identified unique QS architectures in clinical isolates. Importantly, our work reveals a new suite of factors that regulate QS in Pa, with implications for a variety of Pa behaviors both in the laboratory and clinical settings.

Project description:Chronic Pseudomonas aeruginosa infections cause significant morbidity in patients with cystic fibrosis (CF). Over years to decades, P. aeruginosa adapts genetically as it establishes chronic lung infections. Nonsynonymous mutations in lasR, the quorum-sensing (QS) master regulator, are common in CF. In laboratory strains of P. aeruginosa, LasR activates transcription of dozens of genes, including that for another QS regulator, RhlR. Despite the frequency with which lasR coding variants have been reported to occur in P. aeruginosa CF isolates, little is known about their consequences for QS. We sequenced lasR from 2,583 P. aeruginosa CF isolates. The lasR sequences of 580 isolates (22%) coded for polypeptides that differed from the conserved LasR polypeptides of well-studied laboratory strains. This collection included 173 unique lasR coding variants, 116 of which were either missense or nonsense mutations. We studied 31 of these variants. About one-sixth of the variant LasR proteins were functional, including 3 with nonsense mutations, and in some LasR-null isolates, genes that are LasR dependent in laboratory strains were nonetheless expressed. Furthermore, about half of the LasR-null isolates retained RhlR activity. Therefore, in some CF isolates the QS hierarchy is altered such that RhlR quorum sensing is independent of LasR regulation. Our analysis challenges the view that QS-silent P. aeruginosa is selected during the course of a chronic CF lung infection. Rather, some lasR sequence variants retain functionality, and many employ an alternate QS strategy involving RhlR. Chronic Pseudomonas aeruginosa infections, such as those in patients with the genetic disease cystic fibrosis, are notable in that mutants with defects in the quorum-sensing transcription factor LasR frequently arise. In laboratory strains of P. aeruginosa, quorum sensing activates transcription of dozens of genes, many of which encode virulence factors, such as secreted proteases and hydrogen cyanide synthases. In well-studied laboratory strains, LasR-null mutants have a quorum-sensing-deficient phenotype. Therefore, the presence of LasR variants in chronic infections has been interpreted to indicate that quorum-sensing-regulated products are not important for those infections. We report that some P. aeruginosa LasR variant clinical isolates are not LasR-null mutants, and others have uncoupled a second quorum-sensing system, the RhlR system, from LasR regulation. In these uncoupled isolates, RhlR independently activates at least some quorum-sensing-dependent genes. Our findings suggest that quorum sensing plays a role in chronic P. aeruginosa infections, despite the emergence of LasR coding variants.

Dataset Information

Microevolution of the mexT and lasR Reinforces the Bias of Quorum Sensing System in Laboratory Strains of Pseudomonas aeruginosa PAO1.

Publications

Microevolution of the <i>mexT</i> and <i>lasR</i> Reinforces the Bias of Quorum Sensing System in Laboratory Strains of <i>Pseudomonas aeruginosa</i> PAO1.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets