scholarly journals Listeria monocytogenes σB Regulates Stress Response and Virulence Functions

2003 ◽  
Vol 185 (19) ◽  
pp. 5722-5734 ◽  
Author(s):  
Mark J. Kazmierczak ◽  
Sharon C. Mithoe ◽  
Kathryn J. Boor ◽  
Martin Wiedmann
Keyword(s):  
Gene Expression ◽  
Listeria Monocytogenes ◽  
Stress Response ◽  
Consensus Sequence ◽  
Virulence Gene ◽  
Wild Type ◽  
Promoter Regions ◽  
Virulence Gene Expression ◽  
Stress Response Genes ◽  
In The Wild

ABSTRACT While the stress-responsive alternative sigma factor σB has been identified in different species of Bacillus, Listeria, and Staphylococcus, theσ B regulon has been extensively characterized only in B. subtilis. We combined biocomputing and microarray-based strategies to identify σB-dependent genes in the facultative intracellular pathogen Listeria monocytogenes. Hidden Markov model (HMM)-based searches identified 170 candidateσ B-dependent promoter sequences in the strain EGD-e genome sequence. These data were used to develop a specialized, 208-gene microarray, which included 166 genes downstream of HMM-predicted σB-dependent promoters as well as selected virulence and stress response genes. RNA for the microarray experiments was isolated from both wild-type and ΔsigB null mutant L. monocytogenes cells grown to stationary phase or exposed to osmotic stress (0.5 M KCl). Microarray analyses identified a total of 55 genes with statistically significantσ B-dependent expression under the conditions used in these experiments, with at least 1.5-fold-higher expression in the wild type over the sigB mutant under either stress condition (51 genes showed at least 2.0-fold-higher expression in the wild type). Of the 55 genes exhibiting σB-dependent expression, 54 were preceded by a sequence resembling the σB promoter consensus sequence. Rapid amplification of cDNA ends-PCR was used to confirm the σB-dependent nature of a subset of eight selected promoter regions. Notably, theσ B-dependent L. monocytogenes genes identified through this HMM/microarray strategy included both stress response genes (e.g., gadB, ctc, and the glutathione reductase gene lmo1433) and virulence genes (e.g., inlA, inlB, and bsh). Our data demonstrate that, in addition to regulating expression of genes important for survival under environmental stress conditions, σB also contributes to regulation of virulence gene expression in L. monocytogenes. These findings strongly suggest thatσ B contributes to L. monocytogenes gene expression during infection.

scholarly journals Contributions of Listeria monocytogenes σ B and PrfA to expression of virulence and stress response genes during extra- and intracellular growth

Microbiology ◽  
2006 ◽  
Vol 152 (6) ◽  
pp. 1827-1838 ◽  
Author(s):  
Mark J. Kazmierczak ◽  
Martin Wiedmann ◽  
Kathryn J. Boor
Keyword(s):  
Listeria Monocytogenes ◽  
Stress Response ◽  
Stationary Phase ◽  
Virulence Gene ◽  
Exponential Phase ◽  
Promoter Regions ◽  
Transcript Levels ◽  
Virulence Gene Expression ◽  
Qrt Pcr ◽  
Relative Contribution

Listeria monocytogenes σ B and PrfA are pleiotropic regulators of stress response and virulence gene expression. Quantitative RT-PCR (qRT-PCR) was used to measure transcript levels of σ B- and PrfA-dependent genes in exponential-phase L. monocytogenes wild-type and ΔsigB strains as well as in bacteria exposed to environmental stresses (0.3 M NaCl or growth to stationary phase) or present in the vacuole or cytosol of human intestinal epithelial cells. Stationary-phase or NaCl-exposed L. monocytogenes showed σ B-dependent increases in opuCA (10- and 17-fold higher, respectively) and gadA transcript levels (77- and 14-fold higher, respectively) as compared to non-stressed, exponential-phase bacteria. While PrfA activity, as reflected by plcA transcript levels, was up to 95-fold higher in intracellular L. monocytogenes as compared to non-stressed bacteria, σ B activity was only slightly higher in intracellular than in non-stressed bacteria. Increased plcA transcript levels, which were similar in both host cell vacuole and cytosol, were associated with increases in both prfA expression and PrfA activity. qRT-PCR assays were designed to measure expression of prfA from each of its three promoter regions. Under all conditions, readthrough transcription from the upstream plcA promoter was very low. The relative contribution to total prfA transcription from the σ A-dependent P1prfA promoter ranged from ∼17 % to 30 %, while the contribution of the P2prfA region, which appears to be transcribed by both σ A and σ B, ranged from ∼70 % to 82 % of total prfA transcript levels. In summary (i) σ B is primarily activated during environmental stress and does not contribute to PrfA activation in intracellular L. monocytogenes and (ii) the partially σ B-dependent P2prfA promoter region contributes the majority of prfA transcripts in both intra- and extracellular bacteria.

scholarly journals Involvement of Agrobacterium tumefaciens Galacturonate Tripartite ATP-Independent Periplasmic (TRAP) Transporter GaaPQM in Virulence Gene Expression

2015 ◽  
Vol 82 (4) ◽  
pp. 1136-1146 ◽  
Author(s):  
Jinlei Zhao ◽  
Andrew N. Binns
Keyword(s):  
Gene Expression ◽  
Agrobacterium Tumefaciens ◽  
Galacturonic Acid ◽  
Virulence Gene ◽  
Promoter Regions ◽  
Content Type ◽  
Virulence Gene Expression ◽  
Sugar Binding ◽  
Neutral Sugars ◽  
High Ligand

ABSTRACTMonosaccharides capable of serving as nutrients for the soil bacteriumAgrobacterium tumefaciensare also inducers of thevirregulon present in the tumor-inducing (Ti) plasmid of this plant pathogen. One such monosaccharide is galacturonate, the predominant monomer of pectin found in plant cell walls. This ligand is recognized by the periplasmic sugar binding protein ChvE, which interacts with the VirA histidine kinase that controlsvirgene expression. Although ChvE is also a member of the ChvE-MmsAB ABC transporter involved in the utilization of many neutral sugars, it is not involved in galacturonate utilization. In this study, a putative tripartite ATP-independent periplasmic (TRAP) transporter, GaaPQM, is shown to be essential for the utilization of galacturonic acid; we show that residue R169 in the predicted sugar binding site of the GaaP is required for activity. The gene upstream ofgaaPQM(gaaR) encodes a member of the GntR family of regulators. GaaR is shown to repress the expression ofgaaPQM, and the repression is relieved in the presence of the substrate for GaaPQM. Moreover, GaaR is shown to bind putative promoter regions in the sequences required for galacturonic acid utilization. Finally,A. tumefaciensstrains carrying a deletion ofgaaPQMare more sensitive to galacturonate as an inducer ofvirgene expression, while the overexpression ofgaaPQMresults in strains being less sensitive to thisvirinducer. This supports a model in which transporter activity is crucial in ensuring thatvirgene expression occurs only at sites of high ligand concentration, such as those at a plant wound site.

scholarly journals Listeria monocytogenes σB Modulates PrfA-Mediated Virulence Factor Expression

2009 ◽  
Vol 77 (5) ◽  
pp. 2113-2124 ◽  
Author(s):  
Juliane Ollinger ◽  
Barbara Bowen ◽  
Martin Wiedmann ◽  
Kathryn J. Boor ◽  
Teresa M. Bergholz
Keyword(s):  
Gene Expression ◽  
Listeria Monocytogenes ◽  
Transcriptional Activation ◽  
Virulence Gene ◽  
Transcript Profiling ◽  
Virulence Gene Expression ◽  
Qrt Pcr ◽  
Genome Wide

ABSTRACT Listeria monocytogenes σB and positive regulatory factor A (PrfA) are pleiotropic transcriptional regulators that coregulate a subset of virulence genes. A positive regulatory role for σB in prfA transcription has been well established; therefore, observations of increased virulence gene expression and hemolytic activity in a ΔsigB strain initially appeared paradoxical. To test the hypothesis that L. monocytogenes σB contributes to a regulatory network critical for appropriate repression as well as induction of virulence gene expression, genome-wide transcript profiling and follow-up quantitative reverse transcriptase PCR (qRT-PCR), reporter fusion, and phenotypic experiments were conducted using L. monocytogenes prfA*, prfA* ΔsigB, ΔprfA, and ΔprfA ΔsigB strains. Genome-wide transcript profiling and qRT-PCR showed that in the presence of active PrfA (PrfA*), σB is responsible for reduced expression of the PrfA regulon. σB-dependent modulation of PrfA regulon expression reduced the cytotoxic effects of a PrfA* strain in HepG2 cells, highlighting the functional importance of regulatory interactions between PrfA and σB. The emerging model of the role of σB in regulating overall PrfA activity includes a switch from transcriptional activation at the P2 prfA promoter (e.g., in extracellular bacteria when PrfA activity is low) to posttranscriptional downregulation of PrfA regulon expression (e.g., in intracellular bacteria when PrfA activity is high).

scholarly journals The SloR/Dlg Metalloregulator Modulates Streptococcus mutans Virulence Gene Expression

2006 ◽  
Vol 188 (14) ◽  
pp. 5033-5044 ◽  
Author(s):  
Elizabeth Rolerson ◽  
Adam Swick ◽  
Lindsay Newlon ◽  
Cameron Palmer ◽  
Yong Pan ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Streptococcus Mutans ◽  
Metal Ion ◽  
Ion Homeostasis ◽  
Virulence Gene ◽  
Wild Type ◽  
Genetic Competence ◽  
Virulence Gene Expression ◽  
Metal Ion Homeostasis

ABSTRACT Metal ion availability in the human oral cavity plays a putative role in Streptococcus mutans virulence gene expression and in appropriate formation of the plaque biofilm. In this report, we present evidence that supports such a role for the DtxR-like SloR metalloregulator (called Dlg in our previous publications) in this oral pathogen. Specifically, the results of gel mobility shift assays revealed the sloABC, sloR, comDE, ropA, sod, and spaP promoters as targets of SloR binding. We confirmed differential expression of these genes in a GMS584 SloR-deficient mutant versus the UA159 wild-type progenitor by real-time semiquantitative reverse transcriptase PCR experiments. The results of additional expression studies support a role for SloR in S. mutans control of glucosyltransferases, glucan binding proteins, and genes relevant to antibiotic resistance. Phenotypic analysis of GMS584 revealed that it forms aberrant biofilms on an abiotic surface, is compromised for genetic competence, and demonstrates heightened incorporation of iron and manganese as well as resistance to oxidative stress compared to the wild type. Taken together, these findings support a role for SloR in S. mutans adherence, biofilm formation, genetic competence, metal ion homeostasis, oxidative stress tolerance, and antibiotic gene regulation, all of which contribute to S. mutans-induced disease.

scholarly journals The Transcriptional Regulator RovS Controls the Attachment of Streptococcus agalactiae to Human Epithelial Cells and the Expression of Virulence Genes

2006 ◽  
Vol 74 (10) ◽  
pp. 5625-5635 ◽  
Author(s):  
Ulrike M. Samen ◽  
Bernhard J. Eikmanns ◽  
Dieter J. Reinscheid
Keyword(s):  
Gene Expression ◽  
Streptococcus Agalactiae ◽  
Bacterial Infections ◽  
Virulence Genes ◽  
Consensus Sequence ◽  
Transcriptional Regulator ◽  
Epithelial Cell Line ◽  
Promoter Regions ◽  
Mobility Shift ◽  
Virulence Gene Expression

ABSTRACT Streptococcus agalactiae is part of the normal flora of the human gastrointestinal tract and also the leading cause of bacterial infections in human newborns and immunocompromised adults. The colonization and infection of different regions within the human host require a regulatory network in S. agalactiae that senses environmental stimuli and controls the formation of specific virulence factors. In the present study, we characterized an Rgg-like transcriptional regulator, designated RovS (regulator of virulence in Streptococcus agalactiae). Deletion of the rovS gene in the genome of S. agalactiae resulted in strain 6313 ΔrovS, which exhibited an increased attachment to immobilized fibrinogen and a significant increase in adherence to the eukaryotic lung epithelial cell line A549. Quantification of expression levels of known and putative S. agalactiae virulence genes by real-time PCR revealed that RovS influences the expression of fbsA, gbs0230, sodA, rogB, and the cyl operon. The altered gene expression in mutant 6313 ΔrovS was restored by plasmid-mediated expression of rovS, confirming the RovS deficiency as the cause for the observed changes in virulence gene expression in S. agalactiae. DNA electrophoretic mobility shift assays showed that RovS specifically binds to the promoter regions of fbsA, gbs0230, sodA, and the cyl operon, indicating that RovS directly regulates their expression. Deletion and mutation studies in the promoter region of fbsA, encoding the main fibrinogen receptor in S. agalactiae, identified a RovS DNA motif. Similar motifs were also found in the promoter regions of gbs0230, sodA, and the cyl operon, and alignments allowed us to propose a consensus sequence for the DNA-binding site of RovS.

scholarly journals The response regulator ResD modulates virulence gene expression in response to carbohydrates in Listeria monocytogenes

2006 ◽  
Vol 61 (6) ◽  
pp. 1622-1635 ◽  
Author(s):  
Marianne H. Larsen ◽  
Birgitte H. Kallipolitis ◽  
Janne K. Christiansen ◽  
John E. Olsen ◽  
Hanne Ingmer
Keyword(s):  
Gene Expression ◽  
Listeria Monocytogenes ◽  
Response Regulator ◽  
Virulence Gene ◽  
Virulence Gene Expression

scholarly journals The bvr Locus of Listeria monocytogenes Mediates Virulence Gene Repression by β-Glucosides

1999 ◽  
Vol 181 (16) ◽  
pp. 5024-5032 ◽  
Author(s):  
Klaus Brehm ◽  
María-Teresa Ripio ◽  
Jürgen Kreft ◽  
José-Antonio Vázquez-Boland
Keyword(s):  
Listeria Monocytogenes ◽  
Virulence Genes ◽  
Natural Habitat ◽  
Sensory System ◽  
Virulence Gene ◽  
Gene Repression ◽  
Signal Molecules ◽  
Wild Type ◽  
Phosphotransferase System ◽  
Virulence Gene Expression

ABSTRACT The β-glucoside cellobiose has been reported to specifically repress the PrfA-dependent virulence genes hly andplcA in Listeria monocytogenes NCTC 7973. This led to the hypothesis that β-glucosides, sugars of plant origin, may act as signal molecules, preventing the expression of virulence genes if L. monocytogenes is living in its natural habitat (soil). In three other laboratory strains (EGD, L028, and 10403S), however, the effect of cellobiose was not unique, and all fermentable carbohydrates repressed hly. This suggested that the downregulation of virulence genes by β-glucosides is not a specific phenomenon but, rather, an aspect of a global regulatory mechanism of catabolite repression (CR). We assessed the effect of carbohydrates on virulence gene expression in a panel of wild-type isolates of L. monocytogenes by using the PrfA-dependent phospholipase C geneplcB as a reporter. Utilization of any fermentable sugar caused plcB repression in wild-type L. monocytogenes. However, an EGD variant was identified in which, as in NCTC 7973, plcB was only repressed by β-glucosides. Thus, the regulation of L. monocytogenes virulence genes by sugars appears to be mediated by two separate mechanisms, one presumably involving a CR pathway and another specifically responding to β-glucosides. We have identified in L. monocytogenes a 4-kb operon, bvrABC, encoding an antiterminator of the BglG family (bvrA), a β-glucoside-specific enzyme II permease component of the phosphoenolpyruvate-sugar phosphotransferase system (bvrB), and a putative ADP-ribosylglycohydrolase (bvrC). Low-stringency Southern blots showed that this locus is absent from other Listeria spp. Transcription ofbvrB was induced by cellobiose and salicin but not by arbutin. Disruption of the bvr operon by replacing part ofbvrAB with an interposon abolished the repression by cellobiose and salicin but not that by arbutin. Our data indicate that the bvr locus encodes a β-glucoside-specific sensor that mediates virulence gene repression upon detection of cellobiose and salicin. Bvr is the first sensory system found in L. monocytogenes that is involved in environmental regulation of virulence genes.

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