Mapping the transcriptional machinery of the IL-8 gene in human bronchial epithelial cells.
IL-8 released from bronchial epithelial cells infected with Pseudomonas aeruginosa plays a crucial role in the chronic lung pathology of patients affected by cystic fibrosis. Novel anti-inflammatory approaches will benefit from a thorough understanding of the regulatory mechanisms involved in the transcription of this chemokine to identify potential pharmacological targets. We addressed this issue by investigating the role of phosphoproteins and transcription factors (TFs) on transcription of IL-8 gene in the human bronchial epithelial IB3-1, CuFi-1, and Calu-3 cells. P. aeruginosa increased the basal phosphorylation of the ERK1/2 pathway components 90-kDa ribosomal S6 kinase (RSK)1/2 and mitogen- and stress-activated kinase-2 and of the p38 MAPK pathway components p38α/δ/γ and heat shock protein 27 (HSP27). The involvement of these kinases in the expression of IL-8 gene was confirmed with pharmacological inhibitors of ERK1/2, RSK, p38, and HSP27 both at transcription and secretion levels. Transfection of TF decoy oligodeoxynucleotides, designed to interfere with the interaction of the TFs NF-κB, NF-IL6, AP-1, CREB, and CHOP with the corresponding consensus sequences identified in the IL-8 promoter, reduced the P. aeruginosa-dependent transcription of IL-8, suggesting their participation in the transcriptional machinery. Stimulation of IB3-1 cells with IL-1β led to a similar pattern of activation, whereas the pattern of phosphoproteins and of TFs modulated by TNF-α differentiated sharply. In conclusion, the results highlight a novel role for RSK1/2 and HSP27 phosphoproteins and of the cooperative role of the TFs NF-κB, NF-IL6, AP-1, CHOP, and CREB in P. aeruginosa-dependent induction of transcription of the IL-8 gene in human bronchial epithelial cells.
Laboratory of Molecular Pathology, Department of Pathology and Diagnostics, University Hospital of Verona, 37126 Verona, Italy.
Valentino
V
Bezzerri
Monica
M
Borgatti
Alessia
A
Finotti
Anna
A
Tamanini
Roberto
R
Gambari
Giulio
G
Cabrini
D001980:Bronchi / Q000276:immunology; D047369:Chromatin Immunoprecipitation; D024202:Electrophoretic Mobility Shift Assay; D004847:Epithelial Cells / Q000276:immunology; D005786:Gene Expression Regulation / Q000235:genetics*; D055551:HSP27 Heat-Shock Proteins / Q000235:genetics / Q000276:immunology; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis / Q000235:genetics*; D010750:Phosphoproteins / Q000235:genetics* / Q000276:immunology; D011552:Pseudomonas Infections / Q000235:genetics* / Q000276:immunology; D011550:Pseudomonas aeruginosa; D060888:Real-Time Polymerase Chain Reaction; D020545:Respiratory Mucosa / Q000276:immunology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000235:genetics / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology*; D014158:Transcription, Genetic; D014162:Transfection
D055551:HSP27 Heat-Shock Proteins; D016209:Interleukin-8; D010750:Phosphoproteins; C069329:RPS6KA1 protein, human; D038744:Ribosomal Protein S6 Kinases, 90-kDa; C069330:ribosomal protein S6 kinase, 90kDa, polypeptide 3