2.
Bielecka, Ewa, et al.
(författare)
Peptidyl arginine deiminase from Porphyromonas gingivalis abolishes C5a activity.
2014
Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 289:47, s. 32481-32487
Tidskriftsartikel (refereegranskat) abstract
Evasion of killing by the complement system, a crucial part of innate immunity, is a key evolutionary strategy of many human pathogens. A major etiological agent of chronic periodontitis, the Gram-negative bacterium Porphyromonas gingivalis produces a vast arsenal of virulence factors that compromise human defense mechanisms. One of these is peptidylarginine deiminase (PPAD), an enzyme unique to P. gingivalis among bacteria, which converts Arg residues in polypeptide chains into citrulline. Here, we report that PPAD citrullination of a critical C-terminal arginine of the anaphylatoxin C5a disabled the protein function. Treatment of C5a with PPAD in vitro resulted in decreased chemotaxis of human neutrophils and diminished calcium signaling in monocytic cell line U937 transfected with the C5a receptor (C5aR) and loaded with a fluorescent intracellular calcium probe: Fura 2-AM. Moreover, a low degree of citrullination of internal arginine residues by PPAD was also detected using mass spectrometry. Further, after treatment of C5 with outer membrane vesicles (OMVs) naturally shed by P. gingivalis we observed generation of C5a totally citrullinated at the C-terminal Arg74 residue (Arg74Cit). In stark contrast only native C5a was detected after treatment with PPAD-null OMVs. Our study suggests reduced antibacterial and proinflammatory capacity of citrullinated C5a, achieved via lower level of chemotactic potential of the modified molecule, and weaker cell activation. In the context of previous studies, which showed crosstalk between C5aR and toll-like receptors, as well as enhanced arthritis development in mice infected with PPAD expressing P. gingivalis, our findings support a crucial role of PPAD in the virulence of P. gingivalis.
3.
Jusko, Monika
(författare)
Complement evasion strategies of human pathogens - the evolutionary arms race
2014
Doktorsavhandling (övrigt vetenskapligt/konstnärligt) abstract
Although the complement system, a pivotal component of innate immunty, is centrally involved in host defense against pathogens, its overactivation or deregulation may excessively amplify inflammation and contribute to immunopathology. Periodontitis, an oral infection-driven chronic inflammatory disease, has been linked with complement disruption by periodontal bacteria, resulting in inflammation and pathogenesis. The mechanisms of this disruption have partially been revealed, yet in this thesis we investigated novel periodontal pathogens and/or complement evasion pathways in periodontitis. Furthermore, complement inhibition by S. aureus is a key step in its infection, but the impact of its proteases on complement, highlighted in our studies, has not been well characterized before. We focused on a major but relatively poorly studied periodontal bacterium T. forsythia. We showed that its two novel peptidases contribute to its complement resistance by cleaving several key complement components. Interestingly, both of the proteases were able to cleave C5 to release biologically active anaphylatoxins C5a, activity of which has been largely implicated in pathogenesis of periodontitis. Further, we showed that another important periopathogen, Prevotella intermedia, acquires resistance towards complement by binding complement inhibitor found in human serum, factor I, and its two major cofactors C4b-binding protein (C4BP) and Factor H. This mechanism contributes to complement resistance of this bacterium. We also found that major periopathogens have the ability to bind to the membrane-bound complement inhibitor CD46. Strikingly, even though this molecule improved initial attachment of bacteria to the epithelial cells, the final outcome was not beneficial for the bacteria – in cells without CD46 they persisted much longer without getting cleared, indicating activation of certain killing mechanisms upon CD46 stimulation. The phenomenon may be related to autophagy, which may be affected by CD46 as shown previously. Recent studies implicated an involvement of a novel gram-positive Filifactor alocis in the pathogenesis of the periodontal disease. Taking into account the key role of complement deregulation by periodontal bacteria, we focused on F. alocis capacity to manipulate this system. We pinpointed different strategies, such as production of proteases or non-productive binding of C3, employed by this bacterium. Finally, Staphylococcus aureus, is known for its impressive repertoire of complement inhibitors. We tested a panel of major proteases of S. aureus and identified their substrates within complement cascades. We also showed that two of the proteases are able to release C5a, similarly to proteases of periodontal bacteria. Taken together, we explored the knowledge about complement manipulation by various common human pathogens. We identified both unique as wells as common strategies of bacterial complement evasion. Exploring virulence mechanisms shared by different pathogens can give rationales for developing effective therapies for infectious diseases.
