The Fibrinogen- and Fibronectin-Binding Domains of Staphylococcus aureus Fibronectin-Binding Protein A Synergistically Promote Endothelial Invasion and Experimental Endocarditis

ABSTRACT Staphylococcus aureus experimental endocarditis relies on sequential fibrinogen binding (for valve colonization) and fibronectin binding (for endothelial invasion) conferred by peptidoglycan-attached adhesins. Fibronectin-binding protein A (FnBPA) reconciles these two properties—as well as elastin binding—and promotes experimental endocarditis by itself. Here we attempted to delineate the minimal subdomain of FnBPA responsible for fibrinogen and fibronectin binding, cell invasion, and in vivo endocarditis. A large library of truncated constructs of FnBPA was expressed in Lactococcus lactis and tested in vitro and in animals. A 127-amino-acid subdomain spanning the hinge of the FnBPA fibrinogen-binding and fibronectin-binding regions appeared necessary and sufficient to confer the sum of these properties. Competition with synthetic peptides could not delineate specific fibrinogen- and fibronectin-binding sites, suggesting that dual binding arose from protein folding, irrespective of clearly defined binding domains. Moreover, coexpressing the 127-amino-acid subdomain with remote domains of FnBPA further increased fibrinogen binding by ≥10 times, confirming the importance of domain interactions for binding efficacy. In animals, fibrinogen binding (but not fibronectin binding) was significantly associated with endocarditis induction, whereas both fibrinogen binding and fibronectin binding were associated with disease severity. Moreover, fibrinogen binding also combined with fibronectin binding to synergize the invasion of cultured cell lines significantly, a feature correlating with endocarditis severity. Thus, while fibrinogen binding and fibronectin binding were believed to act sequentially in colonization and invasion, they appeared unexpectedly intertwined in terms of both functional anatomy and pathogenicity (in endocarditis). This unforeseen FnBPA subtlety might bear importance for the development of antiadhesin strategies.

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Correlation between fibronectin binding protein A expression level at the surface of recombinant lactococcus lactis and plasmid transfer in vitro and in vivo

BMC Microbiology ◽

10.1186/s12866-014-0248-9 ◽

2014 ◽

Vol 14 (1) ◽

Cited By ~ 12

Author(s):

Juliana F Almeida ◽

Denis Mariat ◽

Vasco Azevedo ◽

Anderson Miyoshi ◽

Alejandra de Moreno de LeBlanc ◽

...

Keyword(s):

Lactococcus Lactis ◽

Binding Protein ◽

Protein A ◽

Plasmid Transfer ◽

Expression Level ◽

Fibronectin Binding Protein ◽

Fibronectin Binding ◽

Recombinant Lactococcus Lactis

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Impacts of sarA and agr in Staphylococcus aureus Strain Newman on Fibronectin-Binding Protein A Gene Expression and Fibronectin Adherence Capacity In Vitro and in Experimental Infective Endocarditis

Infection and Immunity ◽

10.1128/iai.72.3.1832-1836.2004 ◽

2004 ◽

Vol 72 (3) ◽

pp. 1832-1836 ◽

Cited By ~ 45

Author(s):

Yan-Qiong Xiong ◽

Arnold S. Bayer ◽

Michael R. Yeaman ◽

Willem van Wamel ◽

Adhar C. Manna ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Binding Capacity ◽

Protein A ◽

Aureus Strain ◽

Global Regulators ◽

Fibronectin Binding Protein ◽

Regulatory Loci ◽

Fibronectin Binding

ABSTRACT We investigated the impacts of sarA and agr on fnbA expression and fibronectin-binding capacity in Staphylococcus aureus in vitro and in experimental endocarditis. Although sarA up-regulated and agr down-regulated both fnbA expression and fibronectin binding in vitro and in vivo, fnbA expression was positively regulated in the absence of both global regulators. Thus, additional regulatory loci contribute to fnbA regulation and fibronectin-binding capacities in S. aureus.

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Reassessing the Role of Staphylococcus aureus Clumping Factor and Fibronectin-Binding Protein by Expression in Lactococcus lactis

Infection and Immunity ◽

10.1128/iai.69.10.6296-6302.2001 ◽

2001 ◽

Vol 69 (10) ◽

pp. 6296-6302 ◽

Cited By ~ 115

Author(s):

Yok-Ai Que ◽

Patrice François ◽

Jacques-Antoine Haefliger ◽

José-Manuel Entenza ◽

Pierre Vaudaux ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Lactococcus Lactis ◽

Gene Knockout ◽

Binding Protein ◽

Single Gene ◽

Fibronectin Binding Protein ◽

Fibronectin Binding

ABSTRACT Since Staphylococcus aureus expresses multiple pathogenic factors, studying their individual roles in single-gene-knockout mutants is difficult. To circumvent this problem,S. aureus clumping factor A (clfA) and fibronectin-binding protein A (fnbA) genes were constitutively expressed in poorly pathogenic Lactococcus lactis using the recently described pOri23 vector. The recombinant organisms were tested in vitro for their adherence to immobilized fibrinogen and fibronectin and in vivo for their ability to infect rats with catheter-induced aortic vegetations. In vitro, bothclfA and fnbA increased the adherence of lactococci to their specific ligands to a similar extent as theS. aureus gene donor. In vivo, the minimum inoculum size producing endocarditis in ≥80% of the rats (80% infective dose [ID80]) with the parent lactococcus was ≥107CFU. In contrast, clfA-expressing andfnbA-expressing lactococci required only 105CFU to infect the majority of the animals (P < 0.00005). This was comparable to the infectivities of classical endocarditis pathogens such as S. aureus and streptococci (ID80 = 104 to 105 CFU) in this model. The results confirmed the role ofclfA in endovascular infection, but with a much higher degree of confidence than with single-gene-inactivated staphylococci. Moreover, they identified fnbA as a critical virulence factor of equivalent importance. This was in contrast to previous studies that produced controversial results regarding this very determinant. Taken together, the present observations suggest that if antiadhesin therapy were to be developed, at least both of theclfA and fnbA products should be blocked for the therapy to be effective.

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Role of σB in the Expression of Staphylococcus aureus Cell Wall Adhesins ClfA and FnbA and Contribution to Infectivity in a Rat Model of Experimental Endocarditis

Infection and Immunity ◽

10.1128/iai.73.2.990-998.2005 ◽

2005 ◽

Vol 73 (2) ◽

pp. 990-998 ◽

Cited By ~ 47

Author(s):

Jose-Manuel Entenza ◽

Philippe Moreillon ◽

Maria Magdalena Senn ◽

Jan Kormanec ◽

Paul M. Dunman ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Binding Protein ◽

Protein A ◽

Northern Blotting ◽

Cell Matrix ◽

Fibrinogen Binding ◽

Fibronectin Binding Protein ◽

Coated Surfaces ◽

Fibrin Clots

ABSTRACT Isogenic Staphylococcus aureus strains with different capacities to produce σB activity were analyzed for their ability to attach to fibrinogen- or fibronectin-coated surfaces or platelet-fibrin clots and to cause endocarditis in rats. In comparison to the σB-deficient strain, BB255, which harbors an rsbU mutation, both rsbU-complemented and σB-overproducing derivatives exhibited at least five times greater attachment to fibrinogen- and fibronectin-coated surfaces and showed increased adherence to platelet-fibrin clots. No differences in adherence were seen between BB255 and a ΔrsbUVWsigB isogen. Northern blotting analyses revealed that transcription of clfA, encoding fibrinogen-binding protein clumping factor A, and fnbA, encoding fibronectin-binding protein A, were positively influenced by σB. σB overproduction resulted in a statistically significant increase in positive spleen cultures and enhanced bacterial densities in both the aortic vegetations and spleens at 16 h postinoculation. In contrast, at 72 h postinoculation, tissues infected with the σB overproducer had lower bacterial densities than did those infected with BB255. These results suggest that although σB appears to increase the adhesion of S. aureus to various host cell-matrix proteins in vitro, it has limited effect on pathogenesis in the rat endocarditis model. σB appears to have a transient enhancing effect on bacterial density in the early stages of infection that is lost during progression.

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Amino acid alterations in fibronectin binding protein A (FnBPA) and bacterial genotype are associated with cardiac device related infection in Staphylococcus aureus bacteraemia

Journal of Infection ◽

10.1016/j.jinf.2014.09.005 ◽

2015 ◽

Vol 70 (2) ◽

pp. 153-159 ◽

Cited By ~ 12

Author(s):

Nina J. Hos ◽

Siegbert Rieg ◽

Winfried V. Kern ◽

Daniel Jonas ◽

Vance G. Fowler ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Amino Acid ◽

Binding Protein ◽

Protein A ◽

Cardiac Device ◽

Fibronectin Binding Protein ◽

Fibronectin Binding

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Fibrinogen binding is affected by amino acid substitutions in C-terminal repeat region of fibronectin binding protein A

Scientific Reports ◽

10.1038/s41598-019-48031-5 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 5

Author(s):

Nadia N. Casillas-Ituarte ◽

Alex C. DiBartola ◽

Megan J. Broughton ◽

Lumarie Pérez-Guzmán ◽

Robert M. Wheeler ◽

...

Keyword(s):

Amino Acid ◽

Binding Protein ◽

Protein A ◽

Terminal Repeat ◽

Amino Acid Substitutions ◽

Repeat Region ◽

Fibrinogen Binding ◽

Fibronectin Binding Protein ◽

Fibronectin Binding ◽

Terminal Repeat Region

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Production of Fibronectin Binding Protein A at the Surface of Lactococcus lactis Increases Plasmid Transfer In Vitro and In Vivo

PLoS ONE ◽

10.1371/journal.pone.0044892 ◽

2012 ◽

Vol 7 (9) ◽

pp. e44892 ◽

Cited By ~ 23

Author(s):

Daniela Pontes ◽

Silvia Innocentin ◽

Silvina del Carmen ◽

Juliana Franco Almeida ◽

Jean-Guy LeBlanc ◽

...

Keyword(s):

Lactococcus Lactis ◽

Binding Protein ◽

Protein A ◽

Plasmid Transfer ◽

Fibronectin Binding Protein ◽

Fibronectin Binding

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Increased Virulence of a Fibronectin-Binding Protein Mutant of Staphylococcus aureus in a Rat Model of Pneumonia

Infection and Immunity ◽

10.1128/iai.70.7.3865-3873.2002 ◽

2002 ◽

Vol 70 (7) ◽

pp. 3865-3873 ◽

Cited By ~ 42

Author(s):

Mary C. McElroy ◽

David J. Cain ◽

Christine Tyrrell ◽

Timothy J. Foster ◽

Christopher Haslett

Keyword(s):

Staphylococcus Aureus ◽

Epithelial Cells ◽

Rat Model ◽

Binding Proteins ◽

Binding Protein ◽

Alveolar Epithelial Cells ◽

Alveolar Epithelial ◽

Fibronectin Binding Protein ◽

Fibronectin Binding

ABSTRACT Fibronectin-binding proteins mediate Staphylococcus aureus internalization into nonphagocytic cells in vitro. We have investigated whether fibronectin-binding proteins are virulence factors in the pathogenesis of pneumonia by using S. aureus strain 8325-4 and isogenic mutants in which fibronectin-binding proteins were either deleted (DU5883) or overexpressed [DU5883(pFnBPA4)]. We first demonstrated that fibronectin-binding proteins mediate S. aureus internalization into alveolar epithelial cells in vitro and that S. aureus internalization into alveolar epithelial cells requires actin rearrangement and protein kinase activity. Second, we established a rat model of S. aureus-induced pneumonia and measured lung injury and bacterial survival at 24 and 96 h postinoculation. S. aureus growth and the extent of lung injury were both increased in rats inoculated with the deletion mutant (DU5883) in comparison with rats inoculated with the wild-type (8325-4) and the fibronectin-binding protein-overexpressing strain DU5883(pFnBPA4) at 24 h postinfection. Morphological evaluation of infected lungs at the light and electron microscopic levels demonstrated that S. aureus was present within neutrophils from both 8325-4- and DU5883-inoculated lungs. Our data suggest that fibronectin-binding protein-mediated internalization into alveolar epithelial cells is not a virulence mechanism in a rat model of pneumonia. Instead, our data suggest that fibronectin-binding proteins decrease the virulence of S. aureus in pneumonia.

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