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saeRS and sarA act synergistically to repress protease production and promote biofilm formation in Staphylococcus aureus.

Mrak LN, Zielinska AK, Beenken KE, Mrak IN, Atwood DN, Griffin LM, Lee CY, Smeltzer MS - PLoS ONE (2012)

Bottom Line: The reduced accumulation of Spa was reversed by mutation of the gene encoding aureolysin, while the reduced accumulation of FnbA was reversed by mutation of the sspABC operon.These results demonstrate that saeRS and sarA act synergistically to repress the production of extracellular proteases that would otherwise limit accumulation of critical proteins that contribute to biofilm formation, with constitutive activation of saeRS limiting protease production, even in a sarA mutant, to a degree that can be correlated with increased enhanced capacity to form a biofilm.Although it remains unclear whether these effects are mediated directly or indirectly, studies done with an sspA::lux reporter suggest they are mediated at a transcriptional level.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.

ABSTRACT
Mutation of the staphylococcal accessory regulator (sarA) limits biofilm formation in diverse strains of Staphylococcus aureus, but there are exceptions. One of these is the commonly studied strain Newman. This strain has two defects of potential relevance, the first being mutations that preclude anchoring of the fibronectin-binding proteins FnbA and FnbB to the cell wall, and the second being a point mutation in saeS that results in constitutive activation of the saePQRS regulatory system. We repaired these defects to determine whether either plays a role in biofilm formation and, if so, whether this could account for the reduced impact of sarA in Newman. Restoration of surface-anchored FnbA enhanced biofilm formation, but mutation of sarA in this fnbA-positive strain increased rather than decreased biofilm formation. Mutation of sarA in an saeS-repaired derivative of Newman (P18L) or a Newman saeRS mutant (ΔsaeRS) resulted in a biofilm-deficient phenotype like that observed in clinical isolates, even in the absence of surface-anchored FnbA. These phenotypes were correlated with increased production of extracellular proteases and decreased accumulation of FnbA and/or Spa in the P18L and ΔsaeRS sarA mutants by comparison to the Newman sarA mutant. The reduced accumulation of Spa was reversed by mutation of the gene encoding aureolysin, while the reduced accumulation of FnbA was reversed by mutation of the sspABC operon. These results demonstrate that saeRS and sarA act synergistically to repress the production of extracellular proteases that would otherwise limit accumulation of critical proteins that contribute to biofilm formation, with constitutive activation of saeRS limiting protease production, even in a sarA mutant, to a degree that can be correlated with increased enhanced capacity to form a biofilm. Although it remains unclear whether these effects are mediated directly or indirectly, studies done with an sspA::lux reporter suggest they are mediated at a transcriptional level.

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Related in: MedlinePlus

Impact of saeRS and sarA on the abundance of protein A (Spa).The abundance of surface associated (top) and extracellular Spa (bottom) was assessed by western blot using anti-Spa antibody. Strains include Newman (WT), its saeS-repaired derivative (P18L), its isogenic saeRS mutant, and derivatives of each in which sarA was mutated alone or in combination with aur.
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pone-0038453-g006: Impact of saeRS and sarA on the abundance of protein A (Spa).The abundance of surface associated (top) and extracellular Spa (bottom) was assessed by western blot using anti-Spa antibody. Strains include Newman (WT), its saeS-repaired derivative (P18L), its isogenic saeRS mutant, and derivatives of each in which sarA was mutated alone or in combination with aur.

Mentions: Mutation of saeRS or sarA has also been associated with reduced production of Spa, and this has been attributed to transcriptional changes [19]. However, like FnbA, the production of extracellular proteases has been shown to limit the accumulation of Spa [17]. The production of Spa in both its surface-associated and extracellular forms (eSpa) has also been correlated with an enhanced capacity to form a biofilm [18]. Based on these considerations, we examined the relative levels of surface-associated and eSpa in Newman and all of its saeRS and sarA derivatives. The amounts of both were comparable in Newman, its P18L derivative, and its isogenic saeRS mutant (Fig. 6). While indirect, this suggests that saeRS has relatively little impact on spa transcription. In contrast, the amount of both surface-associated and eSpa was decreased in a Newman sarA mutant, but decreased even further in the isogenic P18L sarA and saeRS/sarA mutants (Fig. 6), corresponding with biofilm formation (Fig. 1). The fact that this was protease mediated was confirmed by demonstrating that concomitant mutation of aur reversed this phenotype (Fig. 6).


saeRS and sarA act synergistically to repress protease production and promote biofilm formation in Staphylococcus aureus.

Mrak LN, Zielinska AK, Beenken KE, Mrak IN, Atwood DN, Griffin LM, Lee CY, Smeltzer MS - PLoS ONE (2012)

Impact of saeRS and sarA on the abundance of protein A (Spa).The abundance of surface associated (top) and extracellular Spa (bottom) was assessed by western blot using anti-Spa antibody. Strains include Newman (WT), its saeS-repaired derivative (P18L), its isogenic saeRS mutant, and derivatives of each in which sarA was mutated alone or in combination with aur.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3369899&req=5

pone-0038453-g006: Impact of saeRS and sarA on the abundance of protein A (Spa).The abundance of surface associated (top) and extracellular Spa (bottom) was assessed by western blot using anti-Spa antibody. Strains include Newman (WT), its saeS-repaired derivative (P18L), its isogenic saeRS mutant, and derivatives of each in which sarA was mutated alone or in combination with aur.
Mentions: Mutation of saeRS or sarA has also been associated with reduced production of Spa, and this has been attributed to transcriptional changes [19]. However, like FnbA, the production of extracellular proteases has been shown to limit the accumulation of Spa [17]. The production of Spa in both its surface-associated and extracellular forms (eSpa) has also been correlated with an enhanced capacity to form a biofilm [18]. Based on these considerations, we examined the relative levels of surface-associated and eSpa in Newman and all of its saeRS and sarA derivatives. The amounts of both were comparable in Newman, its P18L derivative, and its isogenic saeRS mutant (Fig. 6). While indirect, this suggests that saeRS has relatively little impact on spa transcription. In contrast, the amount of both surface-associated and eSpa was decreased in a Newman sarA mutant, but decreased even further in the isogenic P18L sarA and saeRS/sarA mutants (Fig. 6), corresponding with biofilm formation (Fig. 1). The fact that this was protease mediated was confirmed by demonstrating that concomitant mutation of aur reversed this phenotype (Fig. 6).

Bottom Line: The reduced accumulation of Spa was reversed by mutation of the gene encoding aureolysin, while the reduced accumulation of FnbA was reversed by mutation of the sspABC operon.These results demonstrate that saeRS and sarA act synergistically to repress the production of extracellular proteases that would otherwise limit accumulation of critical proteins that contribute to biofilm formation, with constitutive activation of saeRS limiting protease production, even in a sarA mutant, to a degree that can be correlated with increased enhanced capacity to form a biofilm.Although it remains unclear whether these effects are mediated directly or indirectly, studies done with an sspA::lux reporter suggest they are mediated at a transcriptional level.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States of America.

ABSTRACT
Mutation of the staphylococcal accessory regulator (sarA) limits biofilm formation in diverse strains of Staphylococcus aureus, but there are exceptions. One of these is the commonly studied strain Newman. This strain has two defects of potential relevance, the first being mutations that preclude anchoring of the fibronectin-binding proteins FnbA and FnbB to the cell wall, and the second being a point mutation in saeS that results in constitutive activation of the saePQRS regulatory system. We repaired these defects to determine whether either plays a role in biofilm formation and, if so, whether this could account for the reduced impact of sarA in Newman. Restoration of surface-anchored FnbA enhanced biofilm formation, but mutation of sarA in this fnbA-positive strain increased rather than decreased biofilm formation. Mutation of sarA in an saeS-repaired derivative of Newman (P18L) or a Newman saeRS mutant (ΔsaeRS) resulted in a biofilm-deficient phenotype like that observed in clinical isolates, even in the absence of surface-anchored FnbA. These phenotypes were correlated with increased production of extracellular proteases and decreased accumulation of FnbA and/or Spa in the P18L and ΔsaeRS sarA mutants by comparison to the Newman sarA mutant. The reduced accumulation of Spa was reversed by mutation of the gene encoding aureolysin, while the reduced accumulation of FnbA was reversed by mutation of the sspABC operon. These results demonstrate that saeRS and sarA act synergistically to repress the production of extracellular proteases that would otherwise limit accumulation of critical proteins that contribute to biofilm formation, with constitutive activation of saeRS limiting protease production, even in a sarA mutant, to a degree that can be correlated with increased enhanced capacity to form a biofilm. Although it remains unclear whether these effects are mediated directly or indirectly, studies done with an sspA::lux reporter suggest they are mediated at a transcriptional level.

Show MeSH
Related in: MedlinePlus