title: Enhanced Ca2+ entry due to Orai1 plasma membrane insertion increases IL-8 secretion by cystic fibrosis airways.
abstract: Cystic fibrosis (CF) is caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR). The most common mutation, ΔF508, causes retention of CFTR in the endoplasmic reticulum (ER). Some CF abnormalities can be explained by altered Ca(2+) homeostasis, although it remains unknown how CFTR influences calcium signaling. This study examined the novel hypothesis that store-operated calcium entry (SOCE) through Orai1 is abnormal in CF. The significance of Orai1-mediated SOCE for increased interleukin-8 (IL-8) expression in CF was also investigated. CF and non-CF human airway epithelial cell line and primary cells (obtained at lung transplantation) were used in Ca(2+) imaging, electrophysiology, and fluorescence imaging experiments to explore differences in Orai1 function in CF vs. non-CF cells. Protein expression and localization was assessed by Western blots, cell surface biotinylation, ELISA, and image correlation spectroscopy (ICS). We show here that store-operated Ca(2+) entry (SOCE) is elevated in CF human airway epithelial cells (hAECs; ≈ 1.8- and ≈ 2.5-fold for total Ca(2+)(i) increase and Ca(2+) influx rate, respectively, and ≈ 2-fold increase in the I(CRAC) current) and is caused by increased exocytotic insertion (≈ 2-fold) of Orai1 channels into the plasma membrane, which is normalized by rescue of ΔF508-CFTR trafficking to the cell surface. Augmented SOCE in CF cells is a major factor leading to increased IL-8 secretion (≈ 2-fold). CFTR normally down-regulates the Orai1/stromal interaction molecule 1 (STIM1) complex, and loss of this inhibition due to the absence of CFTR at the plasma membrane helps to explain the potentiated inflammatory response in CF cells.
authors.affiliation: Department of Physiology, McGill University, Montréal, Québec, Canada. haouaria.balghi@mail.mcgill.ca
authors.forename: Haouaria
authors.initials: H
authors.lastname: Balghi
authors.forename: Renaud
authors.initials: R
authors.lastname: Robert
authors.forename: Benjamin
authors.initials: B
authors.lastname: Rappaz
authors.forename: Xuexin
authors.initials: X
authors.lastname: Zhang
authors.forename: Adeline
authors.initials: A
authors.lastname: Wohlhuter-Haddad
authors.forename: Alexandra
authors.initials: A
authors.lastname: Evagelidis
authors.forename: Yishan
authors.initials: Y
authors.lastname: Luo
authors.forename: Julie
authors.initials: J
authors.lastname: Goepp
authors.forename: Pasquale
authors.initials: P
authors.lastname: Ferraro
authors.forename: Philippe
authors.initials: P
authors.lastname: Roméo
authors.forename: Mohamed
authors.initials: M
authors.lastname: Trebak
authors.forename: Paul W
authors.initials: PW
authors.lastname: Wiseman
authors.forename: David Y
authors.initials: DY
authors.lastname: Thomas
authors.forename: John W
authors.initials: JW
authors.lastname: Hanrahan
mesh_terms: D001483:Base Sequence; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003550:Cystic Fibrosis / Q000235:genetics / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D055785:Gene Knockdown Techniques; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis*; D008564:Membrane Potentials; D008565:Membrane Proteins / Q000378:metabolism; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D000071740:ORAI1 Protein; D034741:RNA, Small Interfering / Q000235:genetics; D020545:Respiratory Mucosa / Q000378:metabolism; D015398:Signal Transduction; D000071737:Stromal Interaction Molecule 1
chemical_list: C505032:CFTR protein, human; C508606:CXCL8 protein, human; D015220:Calcium Channels; D017931:DNA Primers; D016209:Interleukin-8; D008565:Membrane Proteins; D050505:Mutant Proteins; D009363:Neoplasm Proteins; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D034741:RNA, Small Interfering; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1; D019005:Cystic Fibrosis Transmembrane Conductance Regulator