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Secretome analysis defines the major role of SecDF in Staphylococcus aureus virulence.

Quiblier C, Seidl K, Roschitzki B, Zinkernagel AS, Berger-Bächi B, Senn MM - PLoS ONE (2013)

Bottom Line: Numerous Sec signal containing proteins involved in virulence were found to be decreased in the supernatant of the secDF mutant.Adhesion, invasion, and cytotoxicity of the secDF mutant were reduced in human umbilical vein endothelial cells.Virulence was significantly reduced using a Galleria mellonella insect model.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland.

ABSTRACT
The Sec pathway plays a prominent role in protein export and membrane insertion, including the secretion of major bacterial virulence determinants. The accessory Sec constituent SecDF has been proposed to contribute to protein export. Deletion of Staphylococcus aureus secDF has previously been shown to reduce resistance, to alter cell separation, and to change the expression of certain virulence factors. To analyse the impact of the secDF deletion in S. aureus on protein secretion, a quantitative secretome analysis was performed. Numerous Sec signal containing proteins involved in virulence were found to be decreased in the supernatant of the secDF mutant. However, two Sec-dependent hydrolases were increased in comparison to the wild type, suggesting additional indirect, regulatory effects to occur upon deletion of secDF. Adhesion, invasion, and cytotoxicity of the secDF mutant were reduced in human umbilical vein endothelial cells. Virulence was significantly reduced using a Galleria mellonella insect model. Altogether, SecDF is a promising therapeutic target for controlling S. aureus infections.

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Related in: MedlinePlus

Impact of SecDF on fibrinogen and fibronectin binding.Binding properties of Newman and CHE482 strain sets to immobilized human fibrinogen and fibronectin was assessed as described in materials and methods. (A) Newman pEmpty, Newman ΔsecDF pEmpty and the complemented mutant Newman ΔsecDF pSecDF. FnBPs in the Newman background are truncated due to a point mutation leading to a stop codon before the sortase motif, which is required for cell wall anchoring, and therefore are secreted [56]. Hence, Cowan I was used as a functional control strain in the fibronectin binding assay. (B) CHE482 harbouring empty plasmid pEmpty (pCN34), the mutant strain CHE482 ΔsecDF harbouring empty plasmid pEmpty (pCN34) and the complemented mutant CHE482 ΔsecDF pSecDF. Mean of three independent experiments are shown with their standard deviation.
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pone-0063513-g003: Impact of SecDF on fibrinogen and fibronectin binding.Binding properties of Newman and CHE482 strain sets to immobilized human fibrinogen and fibronectin was assessed as described in materials and methods. (A) Newman pEmpty, Newman ΔsecDF pEmpty and the complemented mutant Newman ΔsecDF pSecDF. FnBPs in the Newman background are truncated due to a point mutation leading to a stop codon before the sortase motif, which is required for cell wall anchoring, and therefore are secreted [56]. Hence, Cowan I was used as a functional control strain in the fibronectin binding assay. (B) CHE482 harbouring empty plasmid pEmpty (pCN34), the mutant strain CHE482 ΔsecDF harbouring empty plasmid pEmpty (pCN34) and the complemented mutant CHE482 ΔsecDF pSecDF. Mean of three independent experiments are shown with their standard deviation.

Mentions: Binding of Newman ΔsecDF pEmpty to fibrinogen was reduced by up to 50% in comparison to the wild type strain Newman pEmpty and the complemented mutant Newman ΔsecDF pSecDF (Figure 3A). In CHE482, binding of CHE482 ΔsecDF pEmpty was reduced at low concentrations of fibrinogen, but not at concentrations above 2 µg/ml (Figure 3B). This phenotype was restored in both strains by the complementing plasmid pSecDF.


Secretome analysis defines the major role of SecDF in Staphylococcus aureus virulence.

Quiblier C, Seidl K, Roschitzki B, Zinkernagel AS, Berger-Bächi B, Senn MM - PLoS ONE (2013)

Impact of SecDF on fibrinogen and fibronectin binding.Binding properties of Newman and CHE482 strain sets to immobilized human fibrinogen and fibronectin was assessed as described in materials and methods. (A) Newman pEmpty, Newman ΔsecDF pEmpty and the complemented mutant Newman ΔsecDF pSecDF. FnBPs in the Newman background are truncated due to a point mutation leading to a stop codon before the sortase motif, which is required for cell wall anchoring, and therefore are secreted [56]. Hence, Cowan I was used as a functional control strain in the fibronectin binding assay. (B) CHE482 harbouring empty plasmid pEmpty (pCN34), the mutant strain CHE482 ΔsecDF harbouring empty plasmid pEmpty (pCN34) and the complemented mutant CHE482 ΔsecDF pSecDF. Mean of three independent experiments are shown with their standard deviation.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3643904&req=5

pone-0063513-g003: Impact of SecDF on fibrinogen and fibronectin binding.Binding properties of Newman and CHE482 strain sets to immobilized human fibrinogen and fibronectin was assessed as described in materials and methods. (A) Newman pEmpty, Newman ΔsecDF pEmpty and the complemented mutant Newman ΔsecDF pSecDF. FnBPs in the Newman background are truncated due to a point mutation leading to a stop codon before the sortase motif, which is required for cell wall anchoring, and therefore are secreted [56]. Hence, Cowan I was used as a functional control strain in the fibronectin binding assay. (B) CHE482 harbouring empty plasmid pEmpty (pCN34), the mutant strain CHE482 ΔsecDF harbouring empty plasmid pEmpty (pCN34) and the complemented mutant CHE482 ΔsecDF pSecDF. Mean of three independent experiments are shown with their standard deviation.
Mentions: Binding of Newman ΔsecDF pEmpty to fibrinogen was reduced by up to 50% in comparison to the wild type strain Newman pEmpty and the complemented mutant Newman ΔsecDF pSecDF (Figure 3A). In CHE482, binding of CHE482 ΔsecDF pEmpty was reduced at low concentrations of fibrinogen, but not at concentrations above 2 µg/ml (Figure 3B). This phenotype was restored in both strains by the complementing plasmid pSecDF.

Bottom Line: Numerous Sec signal containing proteins involved in virulence were found to be decreased in the supernatant of the secDF mutant.Adhesion, invasion, and cytotoxicity of the secDF mutant were reduced in human umbilical vein endothelial cells.Virulence was significantly reduced using a Galleria mellonella insect model.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medical Microbiology, University of Zurich, Zurich, Switzerland.

ABSTRACT
The Sec pathway plays a prominent role in protein export and membrane insertion, including the secretion of major bacterial virulence determinants. The accessory Sec constituent SecDF has been proposed to contribute to protein export. Deletion of Staphylococcus aureus secDF has previously been shown to reduce resistance, to alter cell separation, and to change the expression of certain virulence factors. To analyse the impact of the secDF deletion in S. aureus on protein secretion, a quantitative secretome analysis was performed. Numerous Sec signal containing proteins involved in virulence were found to be decreased in the supernatant of the secDF mutant. However, two Sec-dependent hydrolases were increased in comparison to the wild type, suggesting additional indirect, regulatory effects to occur upon deletion of secDF. Adhesion, invasion, and cytotoxicity of the secDF mutant were reduced in human umbilical vein endothelial cells. Virulence was significantly reduced using a Galleria mellonella insect model. Altogether, SecDF is a promising therapeutic target for controlling S. aureus infections.

Show MeSH
Related in: MedlinePlus