Functional characterization of quorum sensing LuxR-type transcriptional regulator, EasR in Enterobacter asburiae strain L1

Abstract

Over the past decades, Enterobacter spp. have been identified as challenging and important pathogens. The emergence of multidrug-resistant Enterobacteria especially those that produce Klebsiella pneumoniae carbapenemase has been a very worrying health crisis. Although efforts have been made to unravel the complex mechanisms that contribute to the pathogenicity of different Enterobacter spp., there is very little information associated with AHL-type QS mechanism in Enterobacter spp. Signaling via N-acyl homoserine lactone (AHL) is the most common quorum sensing (QS) mechanism utilized by Proteobacteria. A typical AHL-based QS system involves two key players: a luxI gene homolog to synthesize AHLs and a luxR gene homolog, an AHL-dependent transcriptional regulator. These signaling molecules enable inter-species and intra-species interaction in response to external stimuli according to population density. In our recent study, we reported the genome of AHL-producing bacterium, Enterobacter asburiae strain L1. Whole genome sequencing and in silico analysis revealed the presence of a pair of luxI/R genes responsible for AHL-type QS, designated as easI/R, in strain L1. In a QS system, a LuxR transcriptional protein detects and responds to the concentration of a specific AHL controlling gene expression. In E. asburiae strain L1, EasR protein binds to its cognate AHLs, N-butanoyl homoserine lactone (C4-HSL) and N–hexanoyl homoserine lactone (C6-HSL), modulating the expression of targeted genes. In this current work, we have cloned the 693 bp luxR homolog of strain L1 for further characterization. The functionality and specificity of EasR protein in response to different AHL signaling molecules to activate gene transcription were tested and validated with β-galactosidase assays. Higher β-galactosidase activities were detected for cells harboring EasR, indicating EasR is a functional transcriptional regulator. This is the first report documenting the cloning and characterization of transcriptional regulator, luxR homolog of E. asburiae.

Essential properties and pitfalls of colorimetric Reverse Transcription Loop-mediated Isothermal Amplification as a point-of-care test for SARS-CoV-2 diagnosis

“…Samples that are undetected for the target and for the internal control are invalidated, preventing false-negative results. Some SARS-CoV-2 RT-LAMP studies use an internal control from RT-qPCR (Lau et al 2020 ; Osterdahl et al 2020 ; Zhang and Tanner 2020 ) however, the objective is not performing any sophisticated additional technique and validation for the point-of-care application. With that in mind, the present study showed for the first time the essential role of the internal control in RT-LAMP context, evaluating the performance of the primers and the clinical relevance of the method.…”

Quorum Sensing in ESKAPE Bugs: A Target for Combating Antimicrobial Resistance and Bacterial Virulence

“…These are generated by a LuxR homolog, which has been found to decrease bacterial adhesion and downregulate biofilm development [ 57 ]. In E. asburiae , the QS pathway is triggered by C4-HSL and C6-HSL, which bind to their cognate EasR receptor protein (the transcriptional regulator lux R homolog), thus triggering QS-associated gene transcription, as well as its related phenotypes and biofilm formation [ 58 ].…”

VfqI-VfqR quorum sensing circuit modulates type VI secretion system VflT6SS2 in Vibrio fluvialis

“…Domain analysis (Pfam) revealed two principal conserved domains in VfqR: N-terminal AHL-binding domain (residues 16–163) and C-terminal DNA binding HTH domain (residues 174–230). Sequence alignments of VfqR with canonical LuxR homologs revealed the presence of strongly conserved residues both in the AHL-binding domain (W61, Y65, D74, P75, W89, G114) and the DNA-binding domain (E180, W186, G190) which are present in at least 95% of LuxR-type proteins [ 35 , 36 ] ( Fig. 2 A).…”