<i>Staphylococcus aureus</i>Staphopain A inhibits CXCR2-dependent neutrophil activation and chemotaxis

Laarman, Alexander J.; Mijnheer, Gerdien; Mootz, Joe M.; Rooijen, Willemien J. M. van; Ruyken, Maartje; Malone, Cheryl L.; Heezius, Erik; Ward, Richard J.; Milligan, Graeme; Strijp, Jos A. G. van; Haas, Carla J. C. de; Horswill, Alexander R.; Kessel, Kok P. M. van; Rooijakkers, Suzan H.M.

doi:10.1038/emboj.2012.212

Cited by 97 publications

(84 citation statements)

References 59 publications

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“…As a control, a ⌬sigB ⌬sspB ⌬scpA triple mutant was tested and ScpA was not detectable under any of the tested conditions. In LAC-WT, the fact that ScpA was not detectable at 24 h even under the TSB-grown conditions matched previous reports (54). To confirm that the anti-ScpA antibodies were functioning properly, an immunoblot time course experiment was performed (see Fig.…”

Section: Resultsmentioning

confidence: 73%

Staphopains Modulate Staphylococcus aureus Biofilm Integrity

et al. 2013

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Staphylococcus aureus is a known cause of chronic biofilm infections that can reside on medical implants or host tissue. Recent studies have demonstrated an important role for proteinaceous material in the biofilm structure. The S. aureus genome encodes many secreted proteases, and there is growing evidence that these enzymes have self-cleavage properties that alter biofilm integrity. However, the specific contribution of each protease and mechanism of biofilm modulation is not clear. To address this issue, we utilized a sigma factor B (⌬sigB) mutant where protease activity results in a biofilm-negative phenotype, thereby creating a condition where the protease(s) responsible for the phenotype could be identified. Using a plasma-coated microtiter assay, biofilm formation was restored to the ⌬sigB mutant through the addition of the cysteine protease inhibitor E-64 or by using Staphostatin inhibitors that specifically target the extracellular cysteine proteases SspB and ScpA (called Staphopains). Through construction of gene deletion mutants, we determined that an sspB scpA double mutant restored ⌬sigB biofilm formation, and this recovery could be replicated in plasma-coated flow cell biofilms. Staphopain levels were also found to be decreased under biofilm-forming conditions, possibly allowing biofilm establishment. The treatment of S. aureus biofilms with purified SspB or ScpA enzyme inhibited their formation, and ScpA was also able to disperse an established biofilm. The antibiofilm properties of ScpA were conserved across S. aureus strain lineages. These findings suggest an underappreciated role of the SspB and ScpA cysteine proteases in modulating S. aureus biofilm architecture.

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Section: Resultsmentioning

confidence: 73%

Staphopains Modulate Staphylococcus aureus Biofilm Integrity

et al. 2013

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“…Based on our ecp regulatory findings, we analyzed Ecp activity to assess the impact of the agr system on the extracellular proteome. We recently developed assays for S. aureus staphopain A based on the CXCR2 receptor (33,39), and we tested whether these assays could translate to S. epidermidis. 1457 spent medium containing Ecp cleaved the CXCR2 substrate (data not shown), and this activity was inhibited with E-64, as expected for this protease (40).…”

Section: Resultsmentioning

confidence: 99%

Staphylococcus epidermidis agr Quorum-Sensing System: Signal Identification, Cross Talk, and Importance in Colonization

et al. 2014

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e Staphylococcus epidermidis is an opportunistic pathogen that is one of the leading causes of medical device infections. Global regulators like the agr quorum-sensing system in this pathogen have received a limited amount of attention, leaving important questions unanswered. There are three agr types in S. epidermidis strains, but only one of the autoinducing peptide (AIP) signals has been identified (AIP-I), and cross talk between agr systems has not been tested. We structurally characterized all three AIP types using mass spectrometry and discovered that the AIP-II and AIP-III signals are 12 residues in length, making them the largest staphylococcal AIPs identified to date. S. epidermidis agr reporter strains were developed for each system, and we determined that cross-inhibitory interactions occur between the agr type I and II systems and between the agr type I and III systems. In contrast, no cross talk was observed between the type II and III systems. To further understand the outputs of the S. epidermidis agr system, an RNAIII mutant was constructed, and microarray studies revealed that exoenzymes (Ecp protease and Geh lipase) and low-molecular-weight toxins were downregulated in the mutant. Follow-up analysis of Ecp confirmed the RNAIII is required to induce protease activity and that agr cross talk modulates Ecp activity in a manner that mirrors the agr reporter results. Finally, we demonstrated that the agr system enhances skin colonization by S. epidermidis using a porcine model. This work expands our knowledge of S. epidermidis agr system function and will aid future studies on cell-cell communication in this important opportunistic pathogen.

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“…VfrB impacts virulence in an SSTI model. Alpha-hemolysin has been shown previously to be important in a variety of infection models (5)(6)(7)(8)(9)(10)(11)(12), and proteases are thought to contribute to pathogenesis by modulating secreted virulence factors and degrading host proteins (21,(28)(29)(30)(31)(32)(33)(34). To assess the contribution of VfrBregulated virulence factors in vivo, we chose a mouse model of dermonecrosis due to the following considerations: (i) S. aureus is the dominant cause of all skin and soft tissue infections (SSTI) in patients presenting to emergency departments in the United States (49); (ii) more than 90% of all S. aureus infections are SSTI (50, 51); (iii) our wild-type strain is derived from an SSTI isolate (39,52); and (iv) both alpha-hemolysin (8) and proteases have been demonstrated to contribute to skin infections (21).…”

Section: Figmentioning

confidence: 99%

“…These proteases belong to several classes, including the Aur metalloprotease, the cysteine proteases ScpA and SspB, the serine protease SspA (V8), and the serine-like proteases SplABCDEF. These proteases have been shown to contribute to the stability of virulence factors (21), including alphahemolysin, as well as to cleave host proteins (28)(29)(30)(31)(32)(33)(34). Importantly, a mutant lacking all of these proteases was found to have an increased abundance of virulence factors and to cause increased mortality in an animal model of infection (21).…”

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confidence: 99%

Identification of the Staphylococcus aureus vfrAB Operon, a Novel Virulence Factor Regulatory Locus

et al. 2014

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b During a screen of the Nebraska Transposon Mutant Library, we identified 71 mutations in the Staphylococcus aureus genome that altered hemolysis on blood agar medium. Although many of these mutations disrupted genes known to affect the production of alpha-hemolysin, two of them were associated with an apparent operon, designated vfrAB, that had not been characterized previously. Interestingly, a ⌬vfrB mutant exhibited only minor effects on the transcription of the hla gene, encoding alphahemolysin, when grown in broth, as well as on RNAIII, a posttranscriptional regulatory RNA important for alpha-hemolysin translation, suggesting that VfrB may function at the posttranscriptional level. Indeed, a ⌬vfrB mutant had increased aur and sspAB protease expression under these conditions. However, disruption of the known secreted proteases in the ⌬vfrB mutant did not restore hemolytic activity in the ⌬vfrB mutant on blood agar. Further analysis revealed that, in contrast to the minor effects of VfrB on hla transcription when strains were cultured in liquid media, the level of hla transcription was decreased 50-fold in the absence of VfrB on solid media. These results demonstrate that while VfrB represses protease expression when strains are grown in broth, hla regulation is highly responsive to factors associated with growth on solid media. Intriguingly, the ⌬vfrB mutant displayed increased pathogenesis in a model of S. aureus dermonecrosis, further highlighting the complexity of VfrB-dependent virulence regulation. The results of this study describe a phenotype associated with a class of highly conserved yet uncharacterized proteins found in Gram-positive bacteria, and they shed new light on the regulation of virulence factors necessary for S. aureus pathogenesis.

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Staphylococcus aureusStaphopain A inhibits CXCR2-dependent neutrophil activation and chemotaxis

Abstract: Neutrophil activation and recruitment to the site of infection are critical for host immunity. In humans, the cysteine protease Staphopain A of the pathogen S. aureus blocks this process by cleaving the chemokine receptor CXCR2.

Cited by 97 publications

References 59 publications

Staphopains Modulate Staphylococcus aureus Biofilm Integrity

Staphopains Modulate Staphylococcus aureus Biofilm Integrity

Staphylococcus epidermidis agr Quorum-Sensing System: Signal Identification, Cross Talk, and Importance in Colonization

Identification of the Staphylococcus aureus vfrAB Operon, a Novel Virulence Factor Regulatory Locus

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