title: Fibronectin Binding Proteins SpsD and SpsL Both Support Invasion of Canine Epithelial Cells by Staphylococcus pseudintermedius.
abstract: In this study, we investigated the cell wall-anchored fibronectin-binding proteins SpsD and SpsL from the canine commensal and pathogen Staphylococcus pseudintermedius for their role in promoting bacterial invasion of canine progenitor epidermal keratinocytes (CPEK). Invasion was examined by the gentamicin protection assay and fluorescence microscopy. An ΔspsD ΔspsL mutant of strain ED99 had a dramatically reduced capacity to invade CPEK monolayers, while no difference in the invasion level was observed with single mutants. Lactococcus lactis transformed with plasmids expressing SpsD and SpsL promoted invasion, showing that both proteins are important. Soluble fibronectin was required for invasion, and an RGD-containing peptide or antibodies recognizing the integrin α5β1 markedly reduced invasion, suggesting an important role for the integrin in this process. Src kinase inhibitors effectively blocked internalization, suggesting a functional role for the kinase in invasion. In order to identify the minimal fibronectin-binding region of SpsD and SpsL involved in the internalization process, recombinant fragments of both proteins were produced. The SpsD520-846 and SpsL538-823 regions harboring the major fibronectin-binding sites inhibited S. pseudintermedius internalization. Finally, the effects of staphylococcal invasion on the integrity of different cell lines were examined. Because SpsD and SpsL are critical factors for adhesion and invasion, blocking these processes could provide a strategy for future approaches to treating infections.
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, Pavia, Italy.
authors.forename: Giampiero
authors.initials: G
authors.lastname: Pietrocola
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, Pavia, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Gianotti
authors.affiliation: The Roslin Institute and Edinburgh Infectious Diseases, University of Edinburgh, Easter Bush, Midlothian, Scotland, United Kingdom.
authors.forename: Amy
authors.initials: A
authors.lastname: Richards
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, Pavia, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Nobile
authors.affiliation: Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College Dublin, Dublin, Ireland.
authors.forename: Joan A
authors.initials: JA
authors.lastname: Geoghegan
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, Pavia, Italy.
authors.forename: Simonetta
authors.initials: S
authors.lastname: Rindi
authors.affiliation: Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College Dublin, Dublin, Ireland.
authors.forename: Ian R
authors.initials: IR
authors.lastname: Monk
authors.affiliation: Center for Infectious and Inflammatory Diseases, Texas A&M Health Science Center, Houston, Texas, USA.
authors.forename: Andrea S
authors.initials: AS
authors.lastname: Bordt
authors.affiliation: Department of Microbiology, Moyne Institute of Preventive Medicine, Trinity College Dublin, Dublin, Ireland.
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Foster
authors.affiliation: The Roslin Institute and Edinburgh Infectious Diseases, University of Edinburgh, Easter Bush, Midlothian, Scotland, United Kingdom.
authors.forename: J Ross
authors.initials: JR
authors.lastname: Fitzgerald
authors.affiliation: Department of Molecular Medicine, Unit of Biochemistry, Pavia, Italy pspeziale@unipv.it.
authors.forename: Pietro
authors.initials: P
authors.lastname: Speziale
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002473:Cell Wall / Q000235:genetics / Q000378:metabolism; D004283:Dog Diseases / Q000378:metabolism / Q000382:microbiology*; D004285:Dogs; D004847:Epithelial Cells / Q000382:microbiology*; D005353:Fibronectins / Q000378:metabolism*; D011485:Protein Binding; D013203:Staphylococcal Infections / Q000378:metabolism / Q000382:microbiology / Q000662:veterinary*; D013210:Staphylococcus / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; D005353:Fibronectins