title: Virulence Factors of Pseudomonas aeruginosa Induce Both the Unfolded Protein and Integrated Stress Responses in Airway Epithelial Cells.
abstract: Pseudomonas aeruginosa infection can be disastrous in chronic lung diseases such as cystic fibrosis and chronic obstructive pulmonary disease. Its toxic effects are largely mediated by secreted virulence factors including pyocyanin, elastase and alkaline protease (AprA). Efficient functioning of the endoplasmic reticulum (ER) is crucial for cell survival and appropriate immune responses, while an excess of unfolded proteins within the ER leads to "ER stress" and activation of the "unfolded protein response" (UPR). Bacterial infection and Toll-like receptor activation trigger the UPR most likely due to the increased demand for protein folding of inflammatory mediators. In this study, we show that cell-free conditioned medium of the PAO1 strain of P. aeruginosa, containing secreted virulence factors, induces ER stress in primary bronchial epithelial cells as evidenced by splicing of XBP1 mRNA and induction of CHOP, GRP78 and GADD34 expression. Most aspects of the ER stress response were dependent on TAK1 and p38 MAPK, except for the induction of GADD34 mRNA. Using various mutant strains and purified virulence factors, we identified pyocyanin and AprA as inducers of ER stress. However, the induction of GADD34 was mediated by an ER stress-independent integrated stress response (ISR) which was at least partly dependent on the iron-sensing eIF2α kinase HRI. Our data strongly suggest that this increased GADD34 expression served to protect against Pseudomonas-induced, iron-sensitive cell cytotoxicity. In summary, virulence factors from P. aeruginosa induce ER stress in airway epithelial cells and also trigger the ISR to improve cell survival of the host.
authors.affiliation: Department of Pulmonology, Leiden University Medical Centre, Leiden, the Netherlands; Cambridge Institute for Medical Research (CIMR), University of Cambridge, Cambridge, United Kingdom.
authors.forename: Emily F A
authors.initials: EF
authors.lastname: van 't Wout
authors.affiliation: Department of Pulmonology, Leiden University Medical Centre, Leiden, the Netherlands.
authors.forename: Annemarie
authors.initials: A
authors.lastname: van Schadewijk
authors.affiliation: Department of Molecular Microbiology, Utrecht University, Utrecht, the Netherlands.
authors.forename: Ria
authors.initials: R
authors.lastname: van Boxtel
authors.affiliation: Cambridge Institute for Medical Research (CIMR), University of Cambridge, Cambridge, United Kingdom.
authors.forename: Lucy E
authors.initials: LE
authors.lastname: Dalton
authors.affiliation: Cambridge Institute for Medical Research (CIMR), University of Cambridge, Cambridge, United Kingdom.
authors.forename: Hanna J
authors.initials: HJ
authors.lastname: Clarke
authors.affiliation: Department of Molecular Microbiology, Utrecht University, Utrecht, the Netherlands.
authors.forename: Jan
authors.initials: J
authors.lastname: Tommassen
authors.affiliation: Cambridge Institute for Medical Research (CIMR), University of Cambridge, Cambridge, United Kingdom.
authors.forename: Stefan J
authors.initials: SJ
authors.lastname: Marciniak
authors.affiliation: Department of Pulmonology, Leiden University Medical Centre, Leiden, the Netherlands.
authors.forename: Pieter S
authors.initials: PS
authors.lastname: Hiemstra
mesh_terms: D015153:Blotting, Western; D002478:Cells, Cultured; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D006801:Humans; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D011552:Pseudomonas Infections / Q000378:metabolism* / Q000473:pathology; D011550:Pseudomonas aeruginosa; D020545:Respiratory Mucosa / Q000378:metabolism / Q000382:microbiology / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D056811:Unfolded Protein Response / Q000502:physiology*; D037521:Virulence Factors / Q000378:metabolism*
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D037521:Virulence Factors; C087519:PPP1R15A protein, human; D054645:Protein Phosphatase 1