Molecular and microscopical analysis of pathogenic streptococci - studies on surface proteins interacting with human cells and extracellular matrix

Abstract: Association to specific host tissue structures allows pathogenic bacteria to establish an infection and facilitates the spread within its host. Interactions between bacterial surface structures and human proteins might determine the outcome of the infection. Streptococcus pyogenes (S. pyogenes), is a human pathogen mostly causing localized infections of the skin and respiratory tract, but it is also capable of causing severe invasive disease such as necrotizing fasciitis, sepsis and toxic shock. Until recently, S. pyogenes has been considered as a strictly extracellular pathogen, but several studies has shown that it is capable of invading and surviving intracellularly in several human cell types. In this thesis I have investigated the interaction of S. pyogenes and Streptococcus pneumoniae (S. pneumoniae) with collagen type VI, the binding of S. pyogenes M1 protein and leucine rich (Slr) protein to collagen type I, and Slr’s interaction with human keratinocytes. The adherence of S. pyogenes and S. pneumoniae to collagen type VI in murine upper and lower airways was restricted to the proximity of the NH2 - and COOH-terminal globular domains of collagen type VI, and for S. pyogenes this interaction was mediated by the M1 protein. The M1 protein and Slr are co-expressed on the bacterial surface and both bind to collagen type I with high affinity. Slr exhibited multiple binding sites to collagen I, both to the monomeric and the fibrillar structure, with the most binding concentrated to the overlap region of the collagen I fibril. Slr is able to adhere to and internalize into human keratinocytes (HaCaT) cells in a time dependent manner and we were able to identify non-muscle myosin IIA as a potential cellular ligand for Slr. We could further confirm the presence of non-muscle myosin IIA in HaCat cell lysate and that Slr binds to non-muscle myosin IIA. In summary, we have characterized a novel adhesin of S. pyogenes, Slr, as a LRR containing lipoprotein that, in concert with the M1 protein, might utilize collagens as adhesive targets during the infection process. Slr and M1 bind to collagen type I, and M1 also binds to collagen type VI, a process that might play a role in the primary step of infection in the skin and respiratory tract. Furthermore, Slr binds to non-muscle myosin IIA possibly resulting in the bacterial adherence to and internalization into human keratinocytes. This could contribute to S. pyogenes avoidance of the immune system and/or bacterial dissemination into deeper tissues.