Home Alone: Elimination of All but One Alternative Sigma Factor in Listeria monocytogenes Allows Prediction of New Roles for σB

ABSTRACT Transcription of the Listeria monocytogenes positive regulatory factor A protein (PrfA) is initiated from either of two promoters immediately upstream of prfA (prfAp 1 and prfAp 2) or from the upstream plcA promoter. We demonstrate that prfAp 2 is a functional σB-dependent promoter and that a sigB deletion mutation affects the virulence phenotype of L. monocytogenes. Thus, the alternative sigma factor σB contributes to virulence in L. monocytogenes.

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Listeria monocytogenes σHContributes to Expression of Competence Genes and Intracellular Growth

Journal of Bacteriology ◽

10.1128/jb.00718-15 ◽

2016 ◽

Vol 198 (8) ◽

pp. 1207-1217 ◽

Cited By ~ 4

Author(s):

Veronica Medrano Romero ◽

Kazuya Morikawa

Keyword(s):

Transcription Factor ◽

Listeria Monocytogenes ◽

Sigma Factor ◽

Physiological Role ◽

Population Level ◽

Alternative Sigma Factor ◽

Intracellular Growth ◽

Content Type ◽

Gram Positive Bacteria ◽

Regulation Scheme

ABSTRACTThe alternative sigma factor σHhas two functions in Gram-positive bacteria: it regulates sporulation and the development of genetic competence.Listeria monocytogenesis a nonsporulating species in which competence has not yet been detected. Nevertheless, the main competence regulators and a series of orthologous genes that form the competence machinery are present in its genome; some of the competence genes play a role in optimal phagosomal escape. In this study, strains overexpressing σHand strains with a σHdeletion were used to elucidate the contribution of σHto the expression of the competence machinery genes inL. monocytogenes. Gene expression analysis showed that σHis, indeed, involved incomGandcomEregulation. Unexpectedly, we observed a unique regulation scheme in which σHand the transcription factor ComK were involved. Population-level analysis showed that even with the overexpression of both factors, only a fraction of the cells expressed the competence machinery genes. Although we could not detect competence, σHwas crucial for phagosomal escape, which implies that this alternative sigma factor has specifically evolved to regulate theL. monocytogenesintracellular life cycle.IMPORTANCEListeria monocytogenescan be an intracellular pathogen capable of causing serious infections in humans and animal species. Recently, the competence machinery genes were described as being necessary for optimal phagosomal escape, in which the transcription factor ComK plays an important role. On the other hand, our previous phylogenetic analysis suggested that the alternative sigma factor σHmight play a role in the regulation of competence genes. The present study shows that some of the competence genes belong to the σHregulon and, importantly, that σHis essential for intracellular growth, implying a unique physiological role of σHamongFirmicutes.

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The Alternative Sigma Factor σB and the Virulence Gene Regulator PrfA Both Regulate Transcription of Listeria monocytogenes Internalins

Applied and Environmental Microbiology ◽

10.1128/aem.02664-06 ◽

2007 ◽

Vol 73 (9) ◽

pp. 2919-2930 ◽

Cited By ~ 47

Author(s):

Patrick McGann ◽

Martin Wiedmann ◽

Kathryn J. Boor

Keyword(s):

Listeria Monocytogenes ◽

Sigma Factor ◽

Virulence Genes ◽

Virulence Gene ◽

Alternative Sigma Factor ◽

Transcript Levels ◽

Virulence Gene Expression ◽

Reverse Transcription Pcr ◽

Culture Cells ◽

Tissue Culture Cells

ABSTRACT Some Listeria monocytogenes internalins are recognized as contributing to invasion of mammalian tissue culture cells. While PrfA is well established as a positive regulator of L. monocytogenes virulence gene expression, the stress-responsive σB has been recognized only recently as contributing to expression of virulence genes, including some that encode internalins. To measure the relative contributions of PrfA and σB to internalin gene transcription, we used reverse transcription-PCR to quantify transcript levels for eight internalin genes (inlA, inlB, inlC, inlC2, inlD, inlE, inlF, and inlG) in L. monocytogenes 10403S and in isogenic ΔprfA, ΔsigB, and ΔsigB ΔprfA strains. Strains were grown under defined conditions to produce (i) active PrfA, (ii) active σB and active PrfA, (iii) inactive PrfA, and (iv) active σB and inactive PrfA. Under the conditions tested, σB and PrfA contributed differentially to the expression of the various internalins such that (i) both σB and PrfA contributed to inlA and inlB transcription, (ii) only PrfA contributed to inlC transcription, (iii) only σB contributed to inlC2 and inlD transcription, and (iv) neither σB nor PrfA contributed to inlF and inlG transcription. inlE transcript levels were undetectable. The important role for σB in regulating expression of L. monocytogenes internalins suggests that exposure of this organism to environmental stress conditions, such as those encountered in the gastrointestinal tract, may activate internalin transcription. Interplay between σB and PrfA also appears to be critical for regulating transcription of some virulence genes, including inlA, inlB, and prfA.

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The Listeria monocytogenes σB Regulon and Its Virulence-Associated Functions Are Inhibited by a Small Molecule

mBio ◽

10.1128/mbio.00241-11 ◽

2011 ◽

Vol 2 (6) ◽

Cited By ~ 27

Author(s):

M. Elizabeth Palmer ◽

Soraya Chaturongakul ◽

Martin Wiedmann ◽

Kathryn J. Boor

Keyword(s):

Bacillus Subtilis ◽

Listeria Monocytogenes ◽

Small Molecule ◽

Sigma Factor ◽

Pathogen Transmission ◽

Bile Salt Hydrolase ◽

Alternative Sigma Factor ◽

Bacterial Survival ◽

Gram Positive ◽

Content Type

ABSTRACTThe stress-responsive alternative sigma factor σBis conserved across diverse Gram-positive bacterial genera. InListeria monocytogenes, σBregulates transcription of >150 genes, including genes contributing to virulence and to bacterial survival under host-associated stress conditions, such as those encountered in the human gastrointestinal lumen. An inhibitor ofL. monocytogenesσBactivity was identified by screening ~57,000 natural and synthesized small molecules using a high-throughput cell-based assay. The compound fluoro-phenyl-styrene-sulfonamide (FPSS) (IC50= 3.5 µM) downregulated the majority of genes previously identified as members of the σBregulon inL. monocytogenes10403S, thus generating a transcriptional profile comparable to that of a 10403S ΔsigBstrain. Specifically, of the 208 genes downregulated by FPSS, 75% had been identified previously as positively regulated by σB. Downregulated genes included key virulence and stress response genes, such asinlA,inlB,bsh,hfq,opuC, andbilE. From a functional perspective, FPSS also inhibitedL. monocytogenesinvasion of human intestinal epithelial cells and bile salt hydrolase activity. The ability of FPSS to inhibit σBactivity in bothL. monocytogenesandBacillus subtilisindicates its utility as a specific inhibitor of σBacross multiple Gram-positive genera.IMPORTANCEThe σBtranscription factor regulates expression of genes responsible for bacterial survival under changing environmental conditions and for virulence; therefore, this alternative sigma factor is important for transmission ofL. monocytogenesand other Gram-positive bacteria. Regulation of σBactivity is complex and tightly controlled, reflecting the key role of this factor in bacterial metabolism. We present multiple lines of evidence indicating that fluoro-phenyl-styrene-sulfonamide (FPSS) specifically inhibits activity of σBacross Gram-positive bacterial genera, i.e., in bothListeria monocytogenesandBacillus subtilis. Therefore, FPSS is an important new tool that will enable novel approaches for exploring complex regulatory networks inL. monocytogenesand other Gram-positive pathogens and for investigating small-molecule applications for controlling pathogen transmission.

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Identification of a σB-Dependent Small Noncoding RNA in Listeria monocytogenes

Journal of Bacteriology ◽

10.1128/jb.00740-08 ◽

2008 ◽

Vol 190 (18) ◽

pp. 6264-6270 ◽

Cited By ~ 34

Author(s):

Jesper Sejrup Nielsen ◽

Anders Steno Olsen ◽

Mette Bonde ◽

Poul Valentin-Hansen ◽

Birgitte H. Kallipolitis

Keyword(s):

Listeria Monocytogenes ◽

Stress Tolerance ◽

Sigma Factor ◽

Noncoding Rna ◽

Noncoding Rnas ◽

Alternative Sigma Factor ◽

Dependent Manner ◽

Small Noncoding Rna ◽

Small Noncoding Rnas ◽

Regulatory Circuits

ABSTRACT In Listeria monocytogenes, the alternative sigma factor σB plays important roles in stress tolerance and virulence. Here, we present the identification of SbrA, a novel small noncoding RNA that is produced in a σB-dependent manner. This finding adds the σB regulon to the growing list of stress-induced regulatory circuits that include small noncoding RNAs.

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Activation of the Listeria monocytogenes stressosome in the intracellular eukaryotic environment

Applied and Environmental Microbiology ◽

10.1128/aem.00397-21 ◽

2021 ◽

Author(s):

Charlotte Dessaux ◽

M. Graciela Pucciarelli ◽

Duarte N. Guerreiro ◽

Conor P. O’Byrne ◽

Francisco García-del Portillo

Keyword(s):

Listeria Monocytogenes ◽

Sigma Factor ◽

Environmental Changes ◽

Core Protein ◽

Structural Data ◽

Host Cells ◽

Intracellular Bacteria ◽

Macromolecular Complex ◽

Alternative Sigma Factor ◽

Signal Perception

Listeria monocytogenes is a ubiquitous environmental bacterium and intracellular pathogen that responds to stress using predominantly the alternative sigma factor SigB. Stress is sensed by a multiprotein complex, the stressosome, extensively studied in bacteria grown in nutrient media. Following signal perception, the stressosome triggers a phosphorylation cascade that releases SigB from its anti-sigma factor. Whether the stressosome is activated during the intracellular infection, is unknown. Here, we analysed the subcellular distribution of stressosome proteins in L. monocytogenes located inside epithelial cells following their immunodetection in membrane and cytosolic fractions prepared from intracellular bacteria. Unlike bacteria in laboratory media, intracellular bacteria have a large proportion of the core stressosome protein RsbR1 associated with the membrane. Another core protein, RsbS, is however undetectable. Despite the absence of RsbS, a SigB-dependent reporter revealed that SigB activity increases gradually from early (1 h) to late (6 h) post-infection times. We also found that RsbR1 paralogues attenuate the intensity of the SigB response and that the miniprotein Prli42, reported to tether the stressosome to the membrane in response to oxidative stress, plays no role in associating RsbR1 to the membrane of intracellular bacteria. Altogether, these data indicate that, once inside host cells, the L. monocytogenes stressosome may adopt a unique configuration to sense stress and to activate SigB in the intracellular eukaryotic niche. IMPORTANCE The response to stress mediated by the alternative sigma factor SigB has been extensively characterized in Bacillus subtilis and Listeria monocytogenes. These bacteria sense stress using a supra-macromolecular complex, the stressosome, which triggers a cascade that releases SigB from its anti-sigma factor. Despite much structural data of the complex available and analyses performed in mutants lacking components of the stressosome or the signalling cascade, the integration of the stress signal and the dynamics of stressosome proteins following environmental changes remain poorly understood. Our study provides data at the protein level on essential stressosome components and SigB activity when L. monocytogenes, normally a saprophytic bacterium, adapts to an intracellular lifestyle. Our results support activation of the stressosome complex in intracellular bacteria. The apparent loss of the stressosome core protein RsbS in intracellular L. monocytogenes also challenges current models, favouring the idea of a unique stressosome architecture responding to intracellular host cues.

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Faculty Opinions recommendation of A DNA damage response in Escherichia coli involving the alternative sigma factor, RpoS.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1146901.603997 ◽

2009 ◽

Author(s):

Simon Andrews

Keyword(s):

Escherichia Coli ◽

Dna Damage ◽

Dna Damage Response ◽

Sigma Factor ◽

Alternative Sigma Factor ◽

Damage Response ◽

Sigma Factor Rpos

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Global transcriptional response to vancomycin in Mycobacterium tuberculosis

Microbiology ◽

10.1099/mic.0.024802-0 ◽

2009 ◽

Vol 155 (4) ◽

pp. 1093-1102 ◽

Cited By ~ 78

Author(s):

Roberta Provvedi ◽

Francesca Boldrin ◽

Francesco Falciani ◽

Giorgio Palù ◽

Riccardo Manganelli

Keyword(s):

Mycobacterium Tuberculosis ◽

Surface Stress ◽

Sigma Factor ◽

Clustering Analysis ◽

Transcriptional Response ◽

Alternative Sigma Factor ◽

Hierarchical Clustering Analysis ◽

Physiological Mechanisms ◽

Basal Resistance ◽

Antibacterial Drugs

In order to gain additional understanding of the physiological mechanisms used by bacteria to maintain surface homeostasis and to identify potential targets for new antibacterial drugs, we analysed the variation of the Mycobacterium tuberculosis transcriptional profile in response to inhibitory and subinhibitory concentrations of vancomycin. Our analysis identified 153 genes differentially regulated after exposing bacteria to a concentration of the drug ten times higher than the MIC, and 141 genes differentially expressed when bacteria were growing in a concentration of the drug eightfold lower than the MIC. Hierarchical clustering analysis indicated that the response to these different conditions is different, although with some overlap. This approach allowed us to identify several genes whose products could be involved in the protection from antibiotic stress targeting the envelope and help to confer the basal level of M. tuberculosis resistance to antibacterial drugs, such as Rv2623 (UspA-like), Rv0116c, PE20-PPE31, PspA and proteins related to toxin–antitoxin systems. Moreover, we also demonstrated that the alternative sigma factor σ E confers basal resistance to vancomycin, once again underlining its importance in the physiology of the mycobacterial surface stress response.

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Roles of rpoN, fliA,and flgR in Expression of Flagella inCampylobacter jejuni

Journal of Bacteriology ◽

10.1128/jb.183.9.2937-2942.2001 ◽

2001 ◽

Vol 183 (9) ◽

pp. 2937-2942 ◽

Cited By ~ 59

Author(s):

Aparna Jagannathan ◽

Chrystala Constantinidou ◽

Charles W. Penn

Keyword(s):

Campylobacter Jejuni ◽

Genome Sequence ◽

Sigma Factor ◽

Transcriptional Activator ◽

Alternative Sigma Factor ◽

Wild Type ◽

Electron Microscopic ◽

Global Regulation ◽

Mutant Strains ◽

Microscopic Studies

ABSTRACT Three potential regulators of flagellar expression present in the genome sequence of Campylobacter jejuni NCTC 11168, the genes rpoN, flgR, andfliA, which encode the alternative sigma factor ς54, the ς54-associated transcriptional activator FlgR, and the flagellar sigma factor ς28, respectively, were investigated for their role in global regulation of flagellar expression. The three genes were insertionally inactivated inC. jejuni strains NCTC 11168 and NCTC 11828. Electron microscopic studies of the wild-type and mutant strains showed that therpoN and flgR mutants were nonflagellate and that the fliA mutant had truncated flagella. Immunoblotting experiments with the three mutants confirmed the roles of rpoN, flgR, and fliA in the expression of flagellin.

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