scholarly journals Functional Characterization of the σB-Dependent yabJ-spoVG Operon in Staphylococcus aureus: Role in Methicillin and Glycopeptide Resistance

2009 ◽  
Vol 53 (5) ◽  
pp. 1832-1839 ◽  
Author(s):  
Bettina Schulthess ◽  
Stefan Meier ◽  
Dagmar Homerova ◽  
Christiane Goerke ◽  
Christiane Wolz ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Sigma Factor ◽  
Functional Characterization ◽  
Alternative Sigma Factor ◽  
Virulence Determinants ◽  
Glycopeptide Resistance ◽  
Global Regulators ◽  
Genetic Manipulations ◽  
Capsule Production

ABSTRACT The alternative sigma factor σB of Staphylococcus aureus controls the expression of multiple genes, including virulence determinants and global regulators; promotes capsule production; and increases the resistance levels of methicillin-resistant S. aureus (MRSA) and glycopeptide-intermediate-resistant S. aureus (GISA) strains. We show here that deletion of the σB-controlled yabJ-spoVG operon, which codes for potential downstream regulators of σB, abolished capsule synthesis and reduced resistance in MRSA and GISA to the same extent that σB inactivation did. Introduction of the yabJ-spoVG operon in trans restored the original phenotype. By genetic manipulations, we show that SpoVG but not YabJ is required for complementation. We therefore postulate that SpoVG is the major factor of the yabJ-spoVG operon required in S. aureus for capsule formation and antibiotic resistance.

scholarly journals Molecular Analysis and Organization of the σB Operon in Staphylococcus aureus

2005 ◽  
Vol 187 (23) ◽  
pp. 8006-8019 ◽  
Author(s):  
Maria Magdalena Senn ◽  
Philipp Giachino ◽  
Dagmar Homerova ◽  
Andrea Steinhuber ◽  
Jochen Strassner ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Hybrid Approach ◽  
Growth Cycle ◽  
Growth Stages ◽  
Virulence Determinants ◽  
Global Regulators ◽  
Genetic Manipulations ◽  
Cell Extracts ◽  
Activation Cascade

ABSTRACT The alternative sigma factor σB of Staphylococcus aureus controls the expression of a variety of genes, including virulence determinants and global regulators. Genetic manipulations and transcriptional start point (TSP) analyses showed that the sigB operon is transcribed from at least two differentially controlled promoters: a putative σA-dependent promoter, termed sigB p1, giving rise to a 3.6-kb transcript covering sa2059-sa2058-rsbU-rsbV-rsbW-sigB, and a σB-dependent promoter, sigB p3, initiating a 1.6-kb transcript covering rsbV-rsbW-sigB. TSP and promoter-reporter gene fusion experiments indicated that a third promoter, tentatively termed sigB p2 and proposed to lead to a 2.5-kb transcript, including rsbU-rsbV-rsbW-sigB, might govern the expression of the sigB operon. Environmental stresses, such as heat shock and salt stress, induced a rapid response within minutes from promoters sigB p1 and sigB p3. In vitro, the sigB p1 promoter was active in the early growth stages, while the sigB p2 and sigB p3 promoters produced transcripts throughout the growth cycle, with sigB p3 peaking around the transition state between exponential growth and stationary phase. The amount of sigB transcripts, however, did not reflect the concentration of σB measured in cell extracts, which remained constant over the entire growth cycle. In a guinea pig cage model of infection, sigB transcripts were as abundant 2 and 8 days postinoculation as values found in vitro, demonstrating that sigB is indeed transcribed during the course of infection. Physical interactions between staphylococcal RsbU-RsbV, RsbV-RsbW, and RsbW-σB were inferred from a yeast (Saccharomyces cerevisiae) two-hybrid approach, indicating the presence of a partner-switching mechanism in the σB activation cascade similar to that of Bacillus subtilis. The finding that overexpression of RsbU was sufficient to trigger an immediate and strong activation of σB, however, signals a relevant difference in the regulation of σB activation between B. subtilis and S. aureus in the cascade upstream of RsbU.

scholarly journals Functional Characterization of a Na+-Coupled Dicarboxylate Carrier Protein from Staphylococcus aureus

2005 ◽  
Vol 187 (15) ◽  
pp. 5189-5194 ◽  
Author(s):  
Jason A. Hall ◽  
Ana M. Pajor
Keyword(s):  
Staphylococcus Aureus ◽  
Structural Analysis ◽  
Transport Properties ◽  
Sequence Homology ◽  
Cytoplasmic Membrane ◽  
Functional Characterization ◽  
Carrier Protein ◽  
Dependent Manner ◽  
Reaction Sequence

ABSTRACT We have cloned and functionally characterized a Na+-coupled dicarboxylate transporter, SdcS, from Staphylococcus aureus. This carrier protein is a member of the divalent anion/Na+ symporter (DASS) family and shares significant sequence homology with the mammalian Na+/dicarboxylate cotransporters NaDC-1 and NaDC-3. Analysis of SdcS function indicates transport properties consistent with those of its eukaryotic counterparts. Thus, SdcS facilitates the transport of the dicarboxylates fumarate, malate, and succinate across the cytoplasmic membrane in a Na+-dependent manner. Furthermore, kinetic work predicts an ordered reaction sequence with Na+ (K 0.5 of 2.7 mM) binding before dicarboxylate (Km of 4.5 μM). Because this transporter and its mammalian homologs are functionally similar, we suggest that SdcS may serve as a useful model for DASS family structural analysis.

Activation of the alternative sigma factor SigB of Staphylococcus aureus following internalization by epithelial cells – An in vivo proteomics perspective

2014 ◽  
Vol 304 (2) ◽  
pp. 177-187 ◽  
Author(s):  
Henrike Pförtner ◽  
Marc S. Burian ◽  
Stephan Michalik ◽  
Maren Depke ◽  
Petra Hildebrandt ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Epithelial Cells ◽  
Sigma Factor ◽  
Alternative Sigma Factor

scholarly journals Characterization of sarR, a Modulator ofsar Expression in Staphylococcus aureus

2001 ◽  
Vol 69 (2) ◽  
pp. 885-896 ◽  
Author(s):  
Adhar Manna ◽  
Ambrose L. Cheung
Keyword(s):  
Staphylococcus Aureus ◽  
Fluorescent Protein ◽  
Regulatory Protein ◽  
Virulence Determinants ◽  
Protein Fusion ◽  
Putative Gene ◽  
P2 Promoter ◽  
Green Fluorescent ◽  
Overlapping Transcripts

ABSTRACT The expression of virulence determinants in Staphylococcus aureus is controlled by global regulatory loci (e.g.,sar and agr). The sar locus is composed of three overlapping transcripts (sar P1, P3, and P2 transcripts from P1, P3, and P2 promoters, respectively), all encoding the 372-bp sarA gene. The level of SarA, the major regulatory protein, is partially controlled by the differential activation of sar promoters. We previously partially purified a ∼12 kDa protein with a DNA-specific column containing asar P2 promoter fragment. In this study, the putative gene, designated sarR, was identified and found to encode a 13.6-kDa protein with homology to SarA. Transcriptional and immunoblot studies revealed the sarR gene to be expressed in other staphylococcal strains. Recombinant SarR protein bound sarP1, P2, and P3 promoter fragments in gel shift and footprinting assays. A sarR mutant expressed a higher level of P1 transcript than the parent, as confirmed by promoter green fluorescent protein fusion assays. As the P1 transcript is the predominant sartranscript, we confirmed that the sarR mutant expressed more SarA than the parental strain. We thus proposed that SarR is a regulatory protein that binds to the sar promoters to down-regulate P1 transcription and the ensuing SarA protein expression.

scholarly journals Reversion of the Glycopeptide Resistance Phenotype in Staphylococcus aureus Clinical Isolates

2000 ◽  
Vol 44 (2) ◽  
pp. 272-277 ◽  
Author(s):  
Susan Boyle-Vavra ◽  
Sarah K. Berke ◽  
Jean C. Lee ◽  
Robert S. Daum
Keyword(s):  
Staphylococcus Aureus ◽  
Clinical Laboratory ◽  
Serial Passage ◽  
The United States ◽  
Vancomycin Resistance ◽  
Clinical Isolates ◽  
Glycopeptide Resistance ◽  
Resistance Phenotype ◽  
Capsule Production ◽  
Cell Surface Changes

ABSTRACT The recent identification of glycopeptide intermediate-resistantStaphylococcus aureus (GISA) clinical isolates has provided an opportunity to assess the stability of the glycopeptide resistance phenotype by nonselective serial passage and to evaluate reversion-associated cell surface changes. Three GISA isolates from the United States (MIC of vancomycin = 8 μg/ml) and two from Japan (MICs of vancomycin = 8 and 2 μg/ml) were passaged daily on nutrient agar with or without vancomycin supplementation. After 15 days of passage on nonselective medium, vancomycin- and teicoplanin-susceptible revertants were obtained from each GISA isolate as determined by broth dilution MIC. Revertant isolates were compared with parent isolates for changes in vancomycin heteroresistance, capsule production, hemolysis phenotype, coagulase activity, and lysostaphin susceptibility. Several revertants lost the subpopulations with intermediate vancomycin resistance, whereas two revertants maintained them. Furthermore, although all of the parent GISA isolates produced capsule type 5 (CP5), all but one revertant tested no longer produced CP5. In contrast, passage on medium containing vancomycin yielded isolates that were still intermediately resistant to vancomycin, had no decrease in the MIC of teicoplanin, and produced detectable CP5. No consistent changes in the revertants in hemolysis phenotype, lysostaphin susceptibility, or coagulase activities were discerned. These data indicate that the vancomycin resistance phenotype is unstable in clinical GISA isolates. Reversion of the vancomycin resistance phenotype might explain the difficulty in isolating vancomycin-resistant clinical isolates from the blood of patients who fail vancomycin therapy and, possibly, may account for some of the difficulties in identifying GISA isolates in the clinical laboratory.

Functional characterization of lipase in the pathogenesis of Staphylococcus aureus

2012 ◽  
Vol 419 (4) ◽  
pp. 617-620 ◽  
Author(s):  
Chunyan Hu ◽  
Ning Xiong ◽  
Yong Zhang ◽  
Simon Rayner ◽  
Shiyun Chen
Keyword(s):  
Staphylococcus Aureus ◽  
Functional Characterization

scholarly journals CspA Regulates Pigment Production in Staphylococcus aureus through a SigB-Dependent Mechanism

2005 ◽  
Vol 187 (23) ◽  
pp. 8181-8184 ◽  
Author(s):  
Samuel Katzif ◽  
Eun-Hee Lee ◽  
Anthony B. Law ◽  
Yih-Ling Tzeng ◽  
William M. Shafer
Keyword(s):  
Staphylococcus Aureus ◽  
Sigma Factor ◽  
Cold Shock ◽  
Pigment Production ◽  
Cold Shock Protein ◽  
Alternative Sigma Factor ◽  
Expression Of Genes ◽  
Maximal Production ◽  
Dependent Mechanism

ABSTRACT We report that the cold shock protein CspA of Staphylococcus aureus is required for maximal production of pigment. Results from transcriptional studies revealed that loss of CspA resulted in decreased expression of genes needed for the biosynthesis of 4,4′-diaponeurosporene and the alternative sigma factor SigB.

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