The
            <i>Pseudomonas aeruginosa</i>
            Vfr Regulator Controls Global Virulence Factor Expression through Cyclic AMP-Dependent and -Independent Mechanisms

Fuchs, Erin L.; Brutinel, Evan D.; Jones, Adriana K.; Fulcher, Nanette B.; Urbanowski, Mark L.; Yahr, Timothy L.; Wolfgang, Matthew C.

doi:10.1128/jb.00363-10

Cited by 102 publications

(130 citation statements)

References 56 publications

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“…1), showed reduced expression when transferred to a vfr mutant derivative of the PAO1 strain (Medina et al, 2003a). However, it was not determined whether this effect was a direct consequence of Vfr binding to the rhlR promoter region or an indirect effect, since Vfr activates lasR transcription (Fuchs et al, 2010) and in turn LasR/ 3O-C12-HSL directly activates rhlR transcription (Latifi et al, 1996;Medina et al, 2003a).…”

Section: Resultsmentioning

confidence: 99%

“…It remains to be determined whether the activation of rhlR transcription by Vfr is dependent on the interaction of this protein with cAMP or is independent of the binding of this cyclic nucleotide as has been shown for Vfr-dependent lasR activation (Fuchs et al, 2010). Fig.…”

Section: Integrative Model Of Rhlr Transcriptional Regulationmentioning

confidence: 99%

“…Vfr acts at the top of the regulatory QS cascade, since this is the only protein known to be directly involved in the transcriptional activation of lasR (Albus et al, 1997); the transcriptional activation of lasR by Vfr is independent of cAMP binding by this protein (Fuchs et al, 2010). It has been reported that RpoN is involved in Vfr transcription (Heurlier et al, 2003), so the effect of this sigma factor in rhlR transcription might be indirect.…”

Section: Introductionmentioning

confidence: 99%

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Transcriptional regulation of Pseudomonas aeruginosa rhlR: role of the CRP orthologue Vfr (virulence factor regulator) and quorum-sensing regulators LasR and RhlR

et al. 2011

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The production of many virulence factors by Pseudomonas aeruginosa is regulated by the quorum-sensing (QS) response. In this regulatory network LasR and RhlR, bound to their corresponding autoinducers, play a central role. The QS response has a hierarchical structure: LasR/3O-C12-HSL activates the transcription of rhlR, and RhlR/C4-HSL activates the transcription of several genes, including the rhlAB operon, which encodes the enzymes responsible for rhamnolipid synthesis. The rhlAB operon is located immediately upstream of the rhlR gene. rhlR has four transcription start sites, two of which are located in the rhlB coding region. Vfr directly activates transcription of lasR, and has been reported to be also involved in rhlR expression. The aim of this work was to characterize the details of the mechanism of rhlR transcriptional regulation. We show that Vfr directly regulates rhlR transcription through its binding to several Vfr-binding sites (VBSs) present in the rhlR promoter region, one of which has a negative effect on transcription. Two of the VBSs overlap with las boxes where LasR/3O-C12-HSL binds to activate rhlR transcription. We also show that rhlR transcription is subject to positive-feedback autoregulation through RhlR/C4-HSL activation of the rhlA promoter. This positive autoregulation plays a major role in rhlR expression.

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Section: Resultsmentioning

confidence: 99%

Section: Integrative Model Of Rhlr Transcriptional Regulationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Transcriptional regulation of Pseudomonas aeruginosa rhlR: role of the CRP orthologue Vfr (virulence factor regulator) and quorum-sensing regulators LasR and RhlR

et al. 2011

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“…Nevertheless, it has been reported recently that a fimV mutant was defective in cAMP synthesis and that cAMP was required for Vfr activity and vfr expression (Fuchs et al, 2010). Vfr has also been shown to control expression of the lasR promoter and thus QS in a cAMP-independent manner (Fuchs et al, 2010).…”

Section: Influence Of Fimv On Lasb Expressionmentioning

confidence: 99%

Role of fimV in type II secretion system-dependent protein secretion of Pseudomonas aeruginosa on solid medium

et al. 2011

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Although classical type II secretion systems (T2SSs) are widely present in Gram-negative bacteria, atypical T2SSs can be found in some species. In Pseudomonas aeruginosa, in addition to the classical T2SS Xcp, it was reported that two genes, xphA and xqhA, located outside the xcp locus were organized in an operon (PaQa) which encodes the orphan PaQa subunit. This subunit is able to associate with other components of the classical Xcp machinery to form a functional hybrid T2SS. In the present study, using a transcriptional lacZ fusion, we found that the PaQa operon was more efficiently expressed (i) on solid LB agar than in liquid LB medium, (ii) at 25 °C than at 37 °C and (iii) at an early stage of growth. These results suggested an adaptation of the hybrid system to particular environmental conditions. Transposon mutagenesis led to the finding that vfr and fimV genes are required for optimal expression of the orphan PaQa operon in the defined growth conditions used. Using an original culturing device designed to monitor secretion on solid medium, the ring-plate system, we found that T2SS-dependent secretion of exoproteins, namely the elastase LasB, was affected in a fimV deletion mutant. Our findings led to the discovery of an interplay between FimV and the global regulator Vfr triggering the modulation of the level of Vfr and consequently the modulation of T2SS-dependent secretion on solid medium.

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“…However, CRP is unable to rescue the protease-or exotoxin A-deficient phenotypes of the vfr mutant (40). Recent work suggests that this discrepancy might be due at least in part to cAMPindependent regulation at the lasR promoter (15).…”

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confidence: 94%

Crystal Structure of the Pseudomonas aeruginosa Virulence Factor Regulator

et al. 2011

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Virulence factor regulator (Vfr) enhances Pseudomonas aeruginosa pathogenicity through its role as a global transcriptional regulator. The crystal structure of Vfr shows that it is a winged-helix DNA-binding protein like its homologue cyclic AMP receptor protein (CRP). In addition to an expected primary cyclic AMP-binding site, a second ligand-binding site is nestled between the N-terminal domain and the C-terminal helix-turn-helix domain. Unlike CRP, Vfr is a symmetric dimer in the absence of DNA. Removal of seven disordered N-terminal residues of Vfr prevents the growth of P. aeruginosa.Pseudomonas aeruginosa is a ubiquitous, Gram-negative opportunistic pathogen that is a leading source of morbidity and mortality for individuals with compromised immune systems or cystic fibrosis. P. aeruginosa switches from an environmental organism to a pathogen through the regulation and elaboration of multiple virulence factors. Virulence factor regulator (Vfr) is a central player in a transcriptional cascade that controls this transition.Vfr is a 24-kDa protein belonging to the winged-helix family of transcription regulators (40). Vfr positively regulates quorum sensing by promoting the transcription of lasR, upregulates transcription of the type III secretion system, represses flagellar gene transcription, and positively regulates twitching motility (1, 2, 9, 41). Vfr binds cyclic adenosine 3Ј5Ј monophosphate (cyclic AMP [cAMP]) in vitro with a dissociation constant of 1.6 M (39). Deletion of both cytoplasmic P. aeruginosa adenyl cyclases results in a transcription profile similar to a vfr deletion (41). Furthermore, like the strain carrying the vfr deletion, the double adenyl cyclase mutant has reduced virulence compared to the wild-type strain in a mouse pneumonia model (38). Thus, cAMP is a biologically relevant ligand for Vfr in vivo.The amino acid sequence of Vfr is 67% identical to that of cAMP receptor protein (CRP) from Escherichia coli (40), a long-standing model for the study of transcription regulation and a structurally well-characterized protein (22). When overexpressed, Vfr complements the ␤-galactosidase and tryptophanase-deficient phenotypes of a crp deletion in E. coli (40). Conversely, CRP expressed in a vfr deletion strain of P. aeruginosa promotes sub-wild-type levels of lasR transcription. However, CRP is unable to rescue the protease-or exotoxin A-deficient phenotypes of the vfr mutant (40). Recent work suggests that this discrepancy might be due at least in part to cAMPindependent regulation at the lasR promoter (15).Mounting evidence supports a model for CRP function in which a dimer of CRP binds a cAMP ligand in the N-terminal domain of each monomer to achieve the protein conformation needed for high-affinity specific DNA binding and DNA bending that precedes transcriptional activation. CRP demonstrates low affinity for specific DNA sequences, with a K d of ϳ10 Ϫ4 M, in the absence of cAMP (16). Its specificity and affinity for its promoter sites increase to 10 Ϫ7 to 10 Ϫ8 M in cAMP concentrations...

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The Pseudomonas aeruginosa Vfr Regulator Controls Global Virulence Factor Expression through Cyclic AMP-Dependent and -Independent Mechanisms

Cited by 102 publications

References 56 publications

Transcriptional regulation of Pseudomonas aeruginosa rhlR: role of the CRP orthologue Vfr (virulence factor regulator) and quorum-sensing regulators LasR and RhlR

Transcriptional regulation of Pseudomonas aeruginosa rhlR: role of the CRP orthologue Vfr (virulence factor regulator) and quorum-sensing regulators LasR and RhlR

Role of fimV in type II secretion system-dependent protein secretion of Pseudomonas aeruginosa on solid medium

Crystal Structure of the Pseudomonas aeruginosa Virulence Factor Regulator

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