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Pseudomonas aeruginosa hemolytic phospholipase C suppresses neutrophil respiratory burst activity
L S Terada1, K A Johansen, S Nowbar
1University of Colorado Health Sciences Center, Denver, Colorado 80262, USA. lterada@aol.com
Abstract:
Pseudomonas aeruginosa is a persistent pathogen in the airways of patients with cystic fibrosis or bronchiectasis from other causes and appears to have evolved strategies to survive the inflammatory response of the host. We hypothesized that the secreted hemolytic phospholipase C (PLC) of P. aeruginosa (PlcHR) would decrease neutrophil respiratory burst activity. We found that while intact wild-type P. aeruginosa cells stimulated moderate respiratory burst activity from human neutrophils, an isogenic mutant pseudomonas (DeltaHR strain) containing a targeted deletion of the plcHR operon induced a much more robust oxidative burst from neutrophils. In contrast, a second pseudomonas mutant (DeltaN) containing a disruption in the gene encoding the nonhemolytic PLC (PlcN) was not different from the wild type in stimulating neutrophil O2.- production. Readdition of purified PlcHR to the DeltaHR strain suppressed neutrophil O2.- production to levels stimulated by wild-type bacteria. Interestingly, purified PlcHR decreased phorbol myristate acetate (PMA)- but not formyl methionyl-leucyl-proline (fMLP)-induced respiratory burst activity, suggesting interference by PlcHR with a protein kinase C (PKC)-specific signaling pathway. Accordingly, the PKC inhibitor bisindolylmaleimide inhibited the oxidative burst induced by either PMA or intact pseudomonas, but not by fMLP, whereas the p38 kinase inhibitor SB-203580 fully inhibited the respiratory burst induced by fMLP or the PlcHR-replete wild-type bacteria, but not PMA or the PlcHR-deficient DeltaHR bacterial mutant. We conclude that expression of PlcHR by P. aeruginosa suppresses bacterium-induced neutrophil respiratory burst by interfering with a PKC-dependent, non-p38 kinase-dependent pathway.
Insights
Pseudomonas aeruginosa uses hemolytic phospholipase C (PlcHR) to suppress neutrophil respiratory burst activity, a key immune response. This pathogen employs PlcHR to evade host defenses by interfering with specific signaling pathways.
Area of Science:
- Microbiology
- Immunology
- Pathogen-Host Interactions
Background:
- Pseudomonas aeruginosa is a significant opportunistic pathogen, particularly in chronic airway infections like cystic fibrosis.
- The host inflammatory response, including neutrophil respiratory burst, is crucial for combating bacterial infections.
- P. aeruginosa has evolved mechanisms to evade or suppress host immune defenses.
Purpose of the Study:
- To investigate the role of the hemolytic phospholipase C (PlcHR) from P. aeruginosa in modulating neutrophil respiratory burst activity.
- To elucidate the specific signaling pathways involved in PlcHR-mediated suppression of neutrophil oxidative burst.
Main Methods:
- Generation of isogenic P. aeruginosa mutants with targeted deletions in plcHR and nonhemolytic plcN genes.
- Assessment of neutrophil respiratory burst activity in response to wild-type and mutant bacterial strains.
- Evaluation of the effects of purified PlcHR and specific kinase inhibitors (PKC, p38) on neutrophil oxidative burst.
Main Results:
- Wild-type P. aeruginosa induced moderate neutrophil respiratory burst, while a plcHR deletion mutant induced a significantly more robust burst.
- Readdition of purified PlcHR to the mutant strain suppressed neutrophil oxidative burst.
- PlcHR interfered with phorbol myristate acetate (PMA)-induced, but not fMLP-induced, respiratory burst, implicating protein kinase C (PKC) signaling.
- Kinase inhibitors confirmed PlcHR's interference with a PKC-dependent, non-p38 kinase-dependent pathway.
Conclusions:
- The hemolytic phospholipase C (PlcHR) secreted by P. aeruginosa actively suppresses bacterium-induced neutrophil respiratory burst.
- PlcHR achieves this suppression by interfering with a host cell signaling pathway dependent on protein kinase C (PKC) but not p38 kinase.
- Understanding this mechanism provides insights into P. aeruginosa pathogenesis and potential therapeutic targets.
