Pseudomonas aeruginosa Alkaline Protease Blocks Complement Activation via the Classical and Lectin Pathways

Abstract

The complement system rapidly detects and kills Gram-negative bacteria and supports bacterial killing by phagocytes. However, bacterial pathogens exploit several strategies to evade detection by the complement system. The alkaline protease (AprA) of Pseudomonas aeruginosa has been associated with bacterial virulence and is known to interfere with complement-mediated lysis of erythrocytes, but its exact role in bacterial complement escape is unknown. In this study, we analyzed how AprA interferes with complement activation and whether it could block complement-dependent neutrophil functions. We found that AprA potently blocked phagocytosis and killing of Pseudomonas by human neutrophils. Furthermore, AprA inhibited opsonization of bacteria with C3b and the formation of the chemotactic agent C5a. AprA specifically blocked C3b deposition via the classical and lectin pathways, whereas the alternative pathway was not affected. Serum degradation assays revealed that AprA degrades both human C1s and C2. However, repletion assays demonstrated that the mechanism of action for complement inhibition is cleavage of C2. In summary, we showed that P. aeruginosa AprA interferes with classical and lectin pathway-mediated complement activation via cleavage of C2.

Quorum sensing enhancement of the stress response promotes resistance to quorum quenching and prevents social cheating

“…QS also allows P. aeruginosa biofilms to inhibit phagocytosis and oxygen radical bursts by neutrophils (Bjarnsholt et al, 2005). Furthermore, the activities of QS-controlled virulence factors such as elastase, alkaline protease and rhamnolipids interfere with the activation of phagocytosis and cell signaling (Leid et al, 2005;Kuang et al, 2011;Dö ssel et al, 2012;Laarman et al, 2013). The relationship between QS and resistance of biofilms to several antibiotics like tobramycin is also well documented (Davies et al, 1998;Hentzer et al, 2003).…”

Pouring Salt on a Wound: Pseudomonas aeruginosa Virulence Factors Alter Na+ and Cl- Flux in the Lung

“…AprA has been shown to aid in P. aeruginosa survival in the lung by cleaving transferrin to facilitate iron acquisition by siderophores (83), as well as inhibiting immune recognition by cleaving flagellin monomers to prevent TLR5 recognition (84). Furthermore, AprA degrades complement proteins C1q, C2, and C3, as well as gamma interferon (IFN-␥) (63,85,86), and loss of complement proteins has been shown to block phagocytosis and killing by neutrophils (63). Thus, it appears that AprA degrades many extracellular proteins that may limit the life span of P. aeruginosa in the lung.…”

“…Another pseudomonal protein shown to be important for phagocytic evasion is alkaline protease (63). Alkaline protease (AprA, Fig.…”

The LasB Elastase of Pseudomonas aeruginosa Acts in Concert with Alkaline Protease AprA To Prevent Flagellin-Mediated Immune Recognition

“…For instance, LasB has been reported to degrade mucins and surfactant proteins which collectively function to promote bacterial clearance (9)(10)(11). Similarly, AprA also has been shown to impede bacterial clearance by degrading the C2 component of the complement system, thereby preventing complement-mediated phagocytosis (12). In addition to acting against responding host factors, the immune-modulating capabilities of AprA also include the ability to degrade flagellin, a known activator of proinflammatory responses via toll-like receptor 5 (TLR5) recognition (13,14).…”