scholarly journals Mechanism underlying autoinducer recognition in the Vibrio cholerae DPO-VqmA quorum-sensing pathway

2019 ◽  
Author(s):  
Xiuliang Huang ◽  
Olivia P. Duddy ◽  
Justin E. Silpe ◽  
Jon E. Paczkowski ◽  
Jianping Cong ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Conformational Changes ◽  
Site Directed Mutagenesis ◽  
Communication Process ◽  
Linear Molecule ◽  
Gene Encoding ◽  
Collective Behaviors ◽  
Existing Structures ◽  
Extracellular Signaling

ABSTRACTQuorum sensing is a bacterial communication process whereby bacteria produce, release and detect the accumulation of extracellular signaling molecules called autoinducers to coordinate collective behaviors. In Vibrio cholerae, the quorum-sensing autoinducer, DPO (3,5-dimethyl-pyrazin-2-ol), binds the receptor-transcription factor, VqmA. In response, the DPO-VqmA complex activates transcription of the vqmR gene encoding the VqmR small RNA. VqmR represses genes required for biofilm formation and virulence factor production. Here, we show that VqmA has DPO-dependent and DPO-independent activity. We solved the DPO-VqmA crystal structure and compared it to existing structures to understand the conformational changes the protein undergoes upon DNA binding. Analysis of DPO analogs reveals that a hydroxyl or carbonyl group at the 2’ position is critical for binding. The proposed DPO precursor, a linear molecule, Ala-AA (N-alanyl-aminoacetone), also binds and activates VqmA. DPO and Ala-AA occupy the same binding site as judged by site-directed mutagenesis and competitive ligand binding analyses.

scholarly journals Mechanism underlying autoinducer recognition in the Vibrio cholerae DPO-VqmA quorum-sensing pathway

2020 ◽  
Vol 295 (10) ◽  
pp. 2916-2931 ◽  
Author(s):  
Xiuliang Huang ◽  
Olivia P. Duddy ◽  
Justin E. Silpe ◽  
Jon E. Paczkowski ◽  
Jianping Cong ◽  
...  
Keyword(s):  
Dna Binding ◽  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Conformational Changes ◽  
Transcriptional Activity ◽  
Function Analysis ◽  
Site Directed Mutagenesis ◽  
Communication Process ◽  
Linear Molecule ◽  
Collective Behaviors

Quorum sensing is a bacterial communication process whereby bacteria produce, release, and detect extracellular signaling molecules called autoinducers to coordinate collective behaviors. In the pathogen Vibrio cholerae, the quorum-sensing autoinducer 3,5-dimethyl-pyrazin-2-ol (DPO) binds the receptor and transcription factor VqmA. The DPO-VqmA complex activates transcription of vqmR, encoding the VqmR small RNA, which represses genes required for biofilm formation and virulence factor production. Here, we show that VqmA is soluble and properly folded and activates basal-level transcription of its target vqmR in the absence of DPO. VqmA transcriptional activity is increased in response to increasing concentrations of DPO, allowing VqmA to drive the V. cholerae quorum-sensing transition at high cell densities. We solved the DPO-VqmA crystal structure to 2.0 Å resolution and compared it with existing structures to understand the conformational changes VqmA undergoes upon DNA binding. Analysis of DPO analogs showed that a hydroxyl or carbonyl group at the 2′-position is critical for binding to VqmA. The proposed DPO precursor, a linear molecule, N-alanyl-aminoacetone (Ala-AA), also bound and activated VqmA. Results from site-directed mutagenesis and competitive ligand-binding analyses revealed that DPO and Ala-AA occupy the same binding site. In summary, our structure-function analysis identifies key features required for VqmA activation and DNA binding and establishes that, whereas VqmA binds two different ligands, VqmA does not require a bound ligand for folding or basal transcriptional activity. However, bound ligand is required for maximal activity.

scholarly journals The Vibrio cholerae quorum-sensing protein VqmA integrates cell density, environmental, and host-derived cues into the control of virulence

2020 ◽  
Author(s):  
Ameya A. Mashruwala ◽  
Bonnie L. Bassler
Keyword(s):  
Small Intestine ◽  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Cell Density ◽  
Chemical Communication ◽  
Disulfide Bonds ◽  
Redox Status ◽  
Communication Process ◽  
Signal Molecules ◽  
Collective Behaviors

Scientific AbstractQuorum sensing is a chemical communication process in which bacteria use the production, release, and detection of signal molecules called autoinducers to orchestrate collective behaviors. The human pathogen Vibrio cholerae requires quorum sensing to infect the small intestine. There, V. cholerae encounters the absence of oxygen and the presence of bile. We show that these two stimuli differentially affect quorum sensing function and, in turn, V. cholerae pathogenicity. The quorum-sensing receptor-transcription factor called VqmA, that detects the autoinducer called DPO, also detects the lack of oxygen and the presence of bile. Detection occurs via DPO-, oxygen-, bile-, and redox-responsive disulfide bonds that alter VqmA DNA binding activity. We propose that VqmA serves as an information processing hub that integrates quorum- sensing information, redox status, the presence or absence of oxygen, and host cues. In response to the information acquired through this mechanism, V. cholerae appropriately modulates its virulence output.Lay AbstractQuorum sensing (QS) is a process of chemical communication bacteria use to orchestrate collective behaviors. QS communication relies on chemical signal molecules called autoinducers. QS regulates virulence in Vibrio cholerae, the causative agent of the disease cholera. Transit into the human small intestine, the site of cholera infection, exposes V. cholerae to the host environment. In this study, we show that the combination of two stimuli encountered in the small intestine, the absence of oxygen and the presence of host-produced bile, impinge on V. cholerae QS function and, in turn, pathogenicity. We suggest that possessing a QS system that is responsive to multiple environmental, host, and cell density cues enables V. cholerae to fine-tune its virulence capacity in the human intestine.

scholarly journals The Virulence Activator AphA Links Quorum Sensing to Pathogenesis and Physiology in Vibrio cholerae by Repressing the Expression of a Penicillin Amidase Gene on the Small Chromosome

2003 ◽  
Vol 185 (16) ◽  
pp. 4825-4836 ◽  
Author(s):  
Gabriela Kovacikova ◽  
Wei Lin ◽  
Karen Skorupski
Keyword(s):  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Cell Density ◽  
Transcriptional Activation ◽  
Small Chromosome ◽  
Site Directed Mutagenesis ◽  
Base Pairs ◽  
Penicillin Amidase ◽  
Expression Site ◽  
Dyad Symmetry

ABSTRACT Activation of the tcpPH promoter on the Vibrio pathogenicity island by AphA and AphB initiates the Vibrio cholerae virulence cascade and is regulated by quorum sensing through the repressive action of HapR on aphA expression. To further understand how the chromosomally encoded AphA protein activates tcpPH expression, site-directed mutagenesis was used to identify the base pairs critical for AphA binding and transcriptional activation. This analysis revealed a region of partial dyad symmetry, TATGCA-N6-TNCNNA, that is important for both of these activities. Searching the V. cholerae genome for this binding site permitted the identification of a second one upstream of a penicillin V amidase (PVA) gene on the small chromosome. AphA binds to and footprints this site, which overlaps the pva transcriptional start, consistent with its role as a repressor at this promoter. Since aphA expression is under quorum-sensing control, the response regulators LuxO and HapR also influence pva expression. Thus, pva is repressed at low cell density when AphA levels are high, and it is derepressed at high cell density when AphA levels are reduced. Penicillin amidases are thought to function as scavengers for phenylacetylated compounds in the nonparasitic environment. That AphA oppositely regulates the expression of pva from that of virulence, together with the observation that PVA does not play a role in virulence, suggests that these activities are coordinated to serve V. cholerae in different biological niches.

scholarly journals Analysis of Activator and Repressor Functions Reveals the Requirements for Transcriptional Control by LuxR, the Master Regulator of Quorum Sensing in Vibrio harveyi

mBio ◽  
2013 ◽  
Vol 4 (4) ◽  
Author(s):  
Julia C. van Kessel ◽  
Luke E. Ulrich ◽  
Igor B. Zhulin ◽  
Bonnie L. Bassler
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Dna Binding ◽  
Quorum Sensing ◽  
Binding Site ◽  
Binding Sites ◽  
Vibrio Harveyi ◽  
Communication Process ◽  
Nucleotide Sequencing ◽  
Collective Behaviors

ABSTRACT LuxR-type transcription factors are the master regulators of quorum sensing in vibrios. LuxR proteins are unique members of the TetR superfamily of transcription factors because they activate and repress large regulons of genes. Here, we used chromatin immunoprecipitation and nucleotide sequencing (ChIP-seq) to identify LuxR binding sites in the Vibrio harveyi genome. Bioinformatics analyses showed that the LuxR consensus binding site at repressed promoters is a symmetric palindrome, whereas at activated promoters it is asymmetric and contains only half of the palindrome. Using a genetic screen, we isolated LuxR mutants that separated activation and repression functions at representative promoters. These LuxR mutants exhibit sequence-specific DNA binding defects that restrict activation or repression activity to subsets of target promoters. Altering the LuxR DNA binding site sequence to one more closely resembling the ideal LuxR consensus motif can restore in vivo function to a LuxR mutant. This study provides a mechanistic understanding of how a single protein can recognize a variety of binding sites to differentially regulate gene expression. IMPORTANCE Bacteria use the cell-cell communication process called quorum sensing to regulate collective behaviors. In vibrios, LuxR-type transcription factors control the quorum-sensing gene expression cascade. LuxR-type proteins are structural homologs of TetR-type transcription factors. LuxR proteins were assumed to function analogously to TetR proteins, which typically bind to a single conserved binding site to repress transcription of one or two genes. We find here that unlike TetR proteins, LuxR acts a global regulator, directly binding upstream of and controlling more than 100 genes. Again unlike TetR, LuxR functions as both an activator and a repressor, and these two activities can be separated by mutagenesis. Finally, the consensus binding motifs driving LuxR-activated and -repressed genes are distinct. This work shows that LuxR, although structurally similar to TetR, has evolved unique features enabling it to differentially control a large regulon of genes in response to quorum-sensing cues.

scholarly journals The PqsE-RhlR Interaction Regulates RhlR DNA Binding to Control Virulence Factor Production in Pseudomonas aeruginosa

2022 ◽  
Author(s):  
Kayla A. Simanek ◽  
Isabelle R. Taylor ◽  
Erica K. Richael ◽  
Erica Lasek-Nesselquist ◽  
Bonnie L. Bassler ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Dna Binding ◽  
Quorum Sensing ◽  
Virulence Factor ◽  
Cell Communication ◽  
Communication Process ◽  
Collective Behaviors ◽  
Factor Production ◽  
A Cell ◽  
Virulence Factor Production

Bacteria use a cell-cell communication process called quorum sensing (QS) to orchestrate collective behaviors. QS relies on the group-wide detection of molecules called autoinducers (AI).

scholarly journals Mechanism underlying the DNA-binding preferences of the Vibrio cholerae and vibriophage VP882 VqmA quorum-sensing receptors

2021 ◽  
Author(s):  
Bonnie L. Bassler ◽  
Olivia Duddy ◽  
Xiuliang Huang ◽  
Justin Silpe
Keyword(s):  
Dna Binding ◽  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Dna Sequences ◽  
Genetic Screen ◽  
Binding Domain ◽  
Communication Process ◽  
Dna Binding Domain ◽  
Phage Gene ◽  
Small Regulatory Rna

Quorum sensing is a chemical communication process that bacteria use to coordinate group behaviors. In the global pathogen Vibrio cholerae, one quorum-sensing receptor and transcription factor, called VqmA (VqmAVc), activates expression of the vqmR gene encoding the small regulatory RNA VqmR, which represses genes involved in virulence and biofilm formation. Vibriophage VP882 encodes a VqmA homolog called VqmAPhage that activates transcription of the phage gene qtip, and Qtip launches the phage lytic program. Curiously, VqmAPhage can activate vqmR expression but VqmAVc cannot activate expression of qtip. Here, we investigate the mechanism underlying this asymmetry. We find that promoter selectivity is driven exclusively by each VqmA DNA-binding domain and key DNA sequences in the vqmR and qtip promoters are required to maintain specificity. A protein sequence-guided mutagenesis approach revealed that the residue E194 of VqmAPhage and A192, the equivalent residue in VqmAVc, in the helix-turn-helix motifs contribute to promoter-binding specificity. A genetic screen to identify VqmAPhage mutants that are incapable of binding the qtip promoter but maintain binding to the vqmR promoter delivered additional VqmAPhage residues located immediately C-terminal to the helix-turn-helix motif as required for binding the qtip promoter. Surprisingly, these residues are conserved between VqmAPhage and VqmAVc. A second, targeted genetic screen revealed a region located in the VqmAVc DNA-binding domain as necessary to prevent VqmAVc from binding the qtip promoter, thus restricting DNA-binding to the vqmR promoter. We propose that the VqmAVc helix-turn-helix motif and the C-terminal flanking residues function together to prohibit VqmAVc from binding the qtip promoter.

scholarly journals The Vibrio cholerae Quorum-Sensing Protein VqmA Integrates Cell Density, Environmental, and Host-Derived Cues into the Control of Virulence

mBio ◽  
2020 ◽  
Vol 11 (4) ◽  
Author(s):  
Ameya A. Mashruwala ◽  
Bonnie L. Bassler
Keyword(s):  
Small Intestine ◽  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Cell Density ◽  
Chemical Communication ◽  
Bile Salts ◽  
Anaerobic Growth ◽  
Signal Molecules ◽  
Content Type ◽  
Collective Behaviors

ABSTRACT Quorum sensing is a chemical communication process in which bacteria use the production, release, and detection of signal molecules called autoinducers to orchestrate collective behaviors. The human pathogen Vibrio cholerae requires quorum sensing to infect the small intestine. There, V. cholerae encounters the absence of oxygen and the presence of bile salts. We show that these two stimuli differentially affect quorum-sensing function and, in turn, V. cholerae pathogenicity. First, during anaerobic growth, V. cholerae does not produce the CAI-1 autoinducer, while it continues to produce the DPO autoinducer, suggesting that CAI-1 may encode information specific to the aerobic lifestyle of V. cholerae. Second, the quorum-sensing receptor-transcription factor called VqmA, which detects the DPO autoinducer, also detects the lack of oxygen and the presence of bile salts. Detection occurs via oxygen-, bile salt-, and redox-responsive disulfide bonds that alter VqmA DNA binding activity. We propose that VqmA serves as an information processing hub that integrates quorum-sensing information, redox status, the presence or absence of oxygen, and host cues. In response to the information acquired through this mechanism, V. cholerae appropriately modulates its virulence output. IMPORTANCE Quorum sensing (QS) is a process of chemical communication that bacteria use to orchestrate collective behaviors. QS communication relies on chemical signal molecules called autoinducers. QS regulates virulence in Vibrio cholerae, the causative agent of the disease cholera. Transit into the human small intestine, the site of cholera infection, exposes V. cholerae to the host environment. In this study, we show that the combination of two stimuli encountered in the small intestine, the absence of oxygen and the presence of host-produced bile salts, impinge on V. cholerae QS function and, in turn, pathogenicity. We suggest that possessing a QS system that is responsive to multiple environmental, host, and cell density cues enables V. cholerae to fine-tune its virulence capacity in the human intestine.

scholarly journals Ethanolamine regulates CqsR quorum-sensing signaling inVibrio cholerae

2019 ◽  
Author(s):  
Samit Watve ◽  
Kelsey Barrasso ◽  
Sarah A. Jung ◽  
Kristen J. Davis ◽  
Lisa A. Hawver ◽  
...  
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Cell Density ◽  
High Cell Density ◽  
Chemical Signals ◽  
Communication Process ◽  
Control Group ◽  
Dual Function ◽  
High Cell

ABSTRACTThe pathogen that causes cholera,Vibrio cholerae, uses the cell-cell communication process known as quorum sensing (QS) to regulate virulence factor production and biofilm formation in response to changes in population density and complexity. QS is mediated through the detection of extracellular chemical signals called autoinducers. Four histidine kinases, LuxPQ, CqsS, CqsR and VpsS, have been identified as receptors to activate the key QS regulator LuxO at low cell density. At high cell density, detection of autoinducers by these receptors leads to deactivation of LuxO, resulting in population-wide gene expression changes. While the cognate autoinducers that regulate the activity of CqsS and LuxQ are known, the signals that regulate CqsR have not been determined. Here we show that the common metabolite ethanolamine specifically interacts with the ligand-binding CACHE domain of CqsRin vitroand induces the high cell-density QS response through CqsR kinase inhibition inV. choleraecells. We also identified residues in the CqsR CACHE domain important for ethanolamine detection and signal transduction. Moreover, mutations disrupting endogenous ethanolamine production inV. choleraedelay the onset of, but do not abolish, the high cell-density QS gene expression. Finally, we demonstrate that modulation of CqsR QS response by ethanolamine occurs inside animal hosts. Our findings suggest thatV. choleraeuses CqsR as a dual-function receptor to integrate information from the self-made signals as well as exogenous ethanolamine as an environmental cue to modulate QS response.IMPORTANCEMany bacteria use quorum sensing to regulate cellular processes that are important for their survival and adaptation to different environments. Quorum sensing usually depends on the detection on chemical signals called autoinducers made endogenously by the bacteria. We show here ethanolamine, a common metabolite made by various bacteria and eukaryotes, can modulate the activity of one of the quorum-sensing receptors inVibrio cholerae, the etiological agent of the disease cholera. Our results raise the possibility thatV. choleraeor other quorum-sensing bacteria can combine environmental sensing and quorum sensing to control group behaviors.

scholarly journals Secreted Proteases Control the Timing of Aggregative Community Formation in Vibrio cholerae

mBio ◽  
2021 ◽  
Author(s):  
Matthew Jemielita ◽  
Ameya A. Mashruwala ◽  
Julie S. Valastyan ◽  
Ned S. Wingreen ◽  
Bonnie L. Bassler
Keyword(s):  
Quorum Sensing ◽  
Vibrio Cholerae ◽  
Chemical Communication ◽  
Communication Process ◽  
Community Formation ◽  
Aggregate Formation ◽  
Content Type ◽  
Secreted Proteases

Bacteria can work as collectives to form multicellular communities. Vibrio cholerae , the bacterium that causes the disease cholera in humans, forms aggregated communities in liquid. Aggregate formation relies on a chemical communication process called quorum sensing.

scholarly journals Sodium and proton coupling in the conformational cycle of a MATE antiporter from Vibrio cholerae

2018 ◽  
Vol 115 (27) ◽  
pp. E6182-E6190 ◽  
Author(s):  
Derek P. Claxton ◽  
Kevin L. Jagessar ◽  
P. Ryan Steed ◽  
Richard A. Stein ◽  
Hassane S. Mchaourab
Keyword(s):  
Vibrio Cholerae ◽  
Conformational Changes ◽  
Conformational Dynamics ◽  
Site Directed Mutagenesis ◽  
Spin Labels ◽  
Release Mechanism ◽  
Conserved Residues ◽  
Terminal Domain ◽  
Mate Transporter

Secondary active transporters belonging to the multidrug and toxic compound extrusion (MATE) family harness the potential energy of electrochemical ion gradients to export a broad spectrum of cytotoxic compounds, thus contributing to multidrug resistance. The current mechanistic understanding of ion-coupled substrate transport has been informed by a limited set of MATE transporter crystal structures from multiple organisms that capture a 12-transmembrane helix topology adopting similar outward-facing conformations. Although these structures mapped conserved residues important for function, the mechanistic role of these residues in shaping the conformational cycle has not been investigated. Here, we use double-electron electron resonance (DEER) spectroscopy to explore ligand-dependent conformational changes of NorM from Vibrio cholerae (NorM-Vc), a MATE transporter proposed to be coupled to both Na+ and H+ gradients. Distance measurements between spin labels on the periplasmic side of NorM-Vc identified unique structural intermediates induced by binding of Na+, H+, or the substrate doxorubicin. The Na+- and H+-dependent intermediates were associated with distinct conformations of TM1. Site-directed mutagenesis of conserved residues revealed that Na+- and H+-driven conformational changes are facilitated by a network of polar residues in the N-terminal domain cavity, whereas conserved carboxylates buried in the C-terminal domain are critical for stabilizing the drug-bound state. Interpreted in conjunction with doxorubicin binding of mutant NorM-Vc and cell toxicity assays, these results establish the role of ion-coupled conformational dynamics in the functional cycle and implicate H+ in the doxorubicin release mechanism.

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