The Pseudomonas aeruginosa Exopolysaccharide Psl Facilitates Surface Adherence and NF-κB Activation in A549 Cells

Abstract

ABSTRACT In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P .  aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213–8221, 2006). Using an NF-κB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-κB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-κB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P .  aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-κB activation, likely as a result of increasing contact between bacterial cells and epithelial cells. IMPORTANCE Pseudomonas aeruginosa is the predominant airway pathogen causing morbidity and mortality in individuals affected by the genetic disease cystic fibrosis. P .  aeruginosa can also cause severe pneumonia, burn wound infections, and sepsis, making its overall impact on human health significant. The attachment of P .  aeruginosa to host tissues, often leading to recalcitrant biofilm infections, and inflammation induced by flagellin are both important mechanisms of virulence. We explored the role of the biofilm polysaccharide Psl in the pathogenesis of P .  aeruginosa and found that Psl is required for surface adherence to A549 epithelial cells, and as an adhesin, it facilitates flagellin-mediated NF-κB activation. This work was done to better understand the initial events of infection and revealed that a biofilm polysaccharide contributes to inflammation in a novel manner.