Central role of PAFR signalling in ExoU-induced NF-κB activation.

Abstract

ExoU is an important virulence factor in acute Pseudomonas aeruginosa infections. Here, we unveiled the mechanisms of ExoU-driven NF-κB activation by using human airway cells and mice infected with P. aeruginosa strains. Several approaches showed that PAFR was crucially implicated in the activation of the canonical NF-κB pathway. Confocal microscopy of lungs from infected mice revealed that PAFR-dependent NF-κB activation occurred mainly in respiratory epithelial cells, and reduced p65 nuclear translocation was detected in mice PAFR-/- or treated with the PAFR antagonist WEB 2086. Several evidences showed that ExoU-induced NF-κB activation regulated PAFR expression. First, ExoU increased p65 occupation of PAFR promoter, as assessed by ChIP. Second, luciferase assays in cultures transfected with different plasmid constructs revealed that ExoU promoted p65 binding to the three κB sites in PAFR promoter. Third, treatment of cell cultures with the NF-κB inhibitor Bay 11-7082, or transfection with IκBα negative-dominant, significantly decreased PAFR mRNA. Finally, reduction in PAFR expression was observed in mice treated with Bay 11-7082 or WEB 2086 prior to infection. Together, our data demonstrate that ExoU activates NF-κB by PAFR signalling, which in turns enhances PAFR expression, highlighting an important mechanism of amplification of response to this P. aeruginosa toxin.

• Authors: Carolina Diettrich Mallet de Lima, Jessica da Conceição Costa, Sabrina Alves de Oliveira Lima Santos, Simone Carvalho, Laís de Carvalho, Rodolpho Mattos Albano, Mauro Martins Teixeira, Maria Cristina Maciel Plotkowski, Alessandra Mattos Saliba
• Journal: Cellular microbiology (Cell Microbiol) Vol. 16 Issue 8 Pg. 1244-54 (Aug 2014) ISSN: 1462-5822 [Electronic] India
• PMID: 24612488 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2014 John Wiley & Sons Ltd.

Chemical References

• Azepines
• Bacterial Proteins
• Bacterial Toxins
• Platelet Activating Factor
• Platelet Aggregation Inhibitors
• Platelet Membrane Glycoproteins
• Receptors, G-Protein-Coupled
• Rela protein, mouse
• Transcription Factor RelA
• Triazoles
• platelet activating factor receptor
• pseudomonas exoprotein A protein, Pseudomonas aeruginosa
• WEB 2086

Topics

• Animals
• Azepines (pharmacology)
• Bacterial Proteins (metabolism)
• Bacterial Toxins (metabolism)
• Cell Line
• Enzyme Activation
• Female
• Gene Expression Regulation
• Humans
• Lung (microbiology)
• Mice
• Mice, Inbred C57BL
• Mice, Knockout
• Microscopy, Confocal
• Platelet Activating Factor (metabolism, pharmacology)
• Platelet Aggregation Inhibitors (pharmacology)
• Platelet Membrane Glycoproteins (antagonists & inhibitors, biosynthesis, genetics)
• Promoter Regions, Genetic
• Protein Binding
• Pseudomonas Infections (pathology)
• Pseudomonas aeruginosa (pathogenicity)
• Receptors, G-Protein-Coupled (antagonists & inhibitors, biosynthesis, genetics)
• Signal Transduction (genetics)
• Transcription Factor RelA (metabolism)
• Triazoles (pharmacology)