Nasal Colonisation by Staphylococcus aureus Depends upon Clumping Factor B Binding to the Squamous Epithelial Cell Envelope Protein Loricrin

Abstract

Staphylococcus aureus asymptomatically colonises the anterior nares, but the host and bacterial factors that facilitate colonisation remain incompletely understood. The S. aureus surface protein ClfB has been shown to mediate adherence to squamous epithelial cells in vitro and to promote nasal colonisation in both mice and humans. Here, we demonstrate that the squamous epithelial cell envelope protein loricrin represents the major target ligand for ClfB during S. aureus nasal colonisation. In vitro adherence assays indicated that bacteria expressing ClfB bound loricrin most likely by the “dock, lock and latch” mechanism. Using surface plasmon resonance we showed that ClfB bound cytokeratin 10 (K10), a structural protein of squamous epithelial cells, and loricrin with similar affinities that were in the low µM range. Loricrin is composed of three separate regions comprising GS-rich omega loops. Each loop was expressed separately and found to bind ClfB, However region 2 bound with highest affinity. To investigate if the specific interaction between ClfB and loricrin was sufficient to facilitate S. aureus nasal colonisation, we compared the ability of ClfB+ S. aureus to colonise the nares of wild-type and loricrin-deficient (Lor−/−) mice. In the absence of loricrin, S. aureus nasal colonisation was significantly impaired. Furthermore a ClfB− mutant colonised wild-type mice less efficiently than the parental ClfB+ strain whereas a similar lower level of colonisation was observed with both the parental strain and the ClfB− mutant in the Lor−/− mice. The ability of ClfB to support nasal colonisation by binding loricrin in vivo was confirmed by the ability of Lactococcus lactis expressing ClfB to be retained in the nares of WT mice but not in the Lor−/− mice. By combining in vitro biochemical analysis with animal model studies we have identified the squamous epithelial cell envelope protein loricrin as the target ligand for ClfB during nasal colonisation by S. aureus.

Clumping Factor B Promotes Adherence of Staphylococcus aureus to Corneocytes in Atopic Dermatitis

“…ClfB is the only S. aureus factor that binds to L2v (Fig. 3) (14). Variation in the amino acid sequence of the ligand binding domains of ClfB has been reported previously (26).…”

“…2). The well-characterized S. aureus strain SH1000 was included as a control since the adherence of SH1000 to L2v is solely dependent on ClfB (14). The amount of adherence for each clinical strain was expressed as a percentage of the adherence value measured for SH1000.…”

“…Nevertheless, the control strains provided a useful comparison. Since ClfB is a major determinant of nasal colonization (14), it was surprising to find that ClfB ligand binding activity was lower among commensal control strains than among AD strains. However, nasal colonization is a multifactorial process involving several staphylococcal factors (30), and the relative contribution of each factor may differ from strain to strain.…”

“…For example, higher levels of fibronectin are found in the stratum corneum of patients with AD than in the stratum corneum of individuals with healthy skin, and the expression of S. aureus cell wall-anchored (CWA) fibronectin binding proteins (FnBPs) A and B enhances bacterial adherence to AD skin biopsy sections (13). The CWA protein clumping factor B (ClfB) binds to the cornified envelope proteins loricrin and cytokeratin 10 in the moist squamous epithelium, promoting colonization of the anterior nares (14,15). The binding of ClfB to its ligands is well understood at the molecular level.…”

Methicillin-resistant Staphylococcus aureus antibiotic resistance and virulence

“…Clumping factor B and Staphylococcus aureus surface proteins G and X (SasG, SasX) have been shown to combine to nasal epithelial cells (58)(59)(60). Among them, SasX lately has attracted more attention because it was found to play an important role in an MRSA epidemic (60).…”