Functional characterization of Bap subdomains required for biofilm development and host cell interaction in S. aureus

Abstract

Bap is the prototype of the proteinaceous biofilm matrix in bacteria. Members of this family confer the bacteria the capacity to form a biofilm and to interact with host cells. As common structural features, Bap-­‐related proteins: (i) show a high molecular weight; (ii) contain a core domain of tandem repeats and (iii) very often are harboured in mobile genetic elements. Most of our knowledge about the functionality of Bap proteins has been obtained with the Bap protein of S. aureus. This protein is 2276-­‐amino acid long and display the multidomain architecture characteristic of surface-­‐associated proteins from Gram-­‐positive bacteria. Bap protein promotes biofilm development on abiotic surfaces as well as intercellular adhesion in a process that it is inhibited by the presence of Ca+2. Bap also promotes the adhesion to epithelial cells but inhibits epithelial cell invasion through the interaction with host receptor Gp96 (Valle et al., Plos Pathog 2012). Several evidences suggest that Bap-­‐Gp96 interaction interferes with the fibronectin-­‐binding protein mediated invasion pathway. Although the role of Bap in biofilm development and host cell interaction is well established, our understanding of the Bap domains responsible for these phenotypes remains poorly characterized. In this communication we will present a functional characterization of the Bap subdomains necessary to promote multicellular behavior under in vitro conditions and bacterial adhesion to eukaryotic cells. By using chimeric proteins containing different regions of the Bap protein fused with the clumping factor subdomain R, we have determined the minimal region of Bap necessary and sufficient to mediate biofilm formation and cell-­‐to-­‐cell interactions. Interestingly, this subdomain contains the EF-­‐hand motifs and bacteria producing this subdomain are sensitive to the presence of Ca+2. We have also identified the region of Bap that it is responsible for the interaction with Gp96. We will discuss how the identification of these two subdomains will allow us to evaluate the relative contribution of biofilm development and host cell interaction during Bap-­‐mediated S. aureus chronic infection.