title: 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.affiliation: Departamento de Microbiologia, Imunologia e Parasitologia, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, RJ, Brazil.
authors.forename: Carolina Diettrich
authors.initials: CD
authors.lastname: Mallet de Lima
authors.forename: Jessica
authors.initials: J
authors.lastname: da Conceição Costa
authors.forename: Sabrina Alves
authors.initials: SA
authors.lastname: de Oliveira Lima Santos
authors.forename: Simone
authors.initials: S
authors.lastname: Carvalho
authors.forename: Laís
authors.initials: L
authors.lastname: de Carvalho
authors.forename: Rodolpho Mattos
authors.initials: RM
authors.lastname: Albano
authors.forename: Mauro Martins
authors.initials: MM
authors.lastname: Teixeira
authors.forename: Maria Cristina Maciel
authors.initials: MC
authors.lastname: Plotkowski
authors.forename: Alessandra Mattos
authors.initials: AM
authors.lastname: Saliba
mesh_terms: D000818:Animals; D001381:Azepines / Q000494:pharmacology; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism; D002460:Cell Line; D004789:Enzyme Activation; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D008168:Lung / Q000382:microbiology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D010972:Platelet Activating Factor / Q000378:metabolism* / Q000494:pharmacology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011552:Pseudomonas Infections / Q000473:pathology; D011550:Pseudomonas aeruginosa / Q000472:pathogenicity*; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D015398:Signal Transduction / Q000235:genetics; D051996:Transcription Factor RelA / Q000378:metabolism*; D014230:Triazoles / Q000494:pharmacology
chemical_list: D001381:Azepines; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D010972:Platelet Activating Factor; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D043562:Receptors, G-Protein-Coupled; C498144:Rela protein, mouse; D051996:Transcription Factor RelA; D014230:Triazoles; C039351:platelet activating factor receptor; C111749:pseudomonas exoprotein A protein, Pseudomonas aeruginosa; C052518:WEB 2086