title: Changes in lipid raft proteome upon TNF-α stimulation of cystic fibrosis cells.
abstract: We have previously shown (i) that the cystic fibrosis transmembrane regulator (CFTR) locates to lipid raft-like microdomains of epithelial cells upon TNF-α proinflammatory stimulation; and (ii) that TNF-α increases the membrane localization and the channel function of F508del-mutated CFTR. In the present work, we hypothesized that CFTR mutations modify the proteome of lipid rafts in the same proinflammatory conditions. We prepared lipid rafts from HeLa cells transfected with either wild-type or F508del-CFTR and incubated for 10min with 100U/mL of TNF-α. Proteins were extracted, trypsin digested, and peptides analyzed by high resolution MS. Proteins were quantified by a stable isotope labeling with amino acids in cell culture approach. Out of the 22 proteins differentially recruited in lipid rafts after proinflammatory exposure, 17 were increased in F508del cells with respect to wild-type, including two G-protein coupled receptors, three anion transporters, and one cell surface mucin. In both HeLa and bronchial epithelial cells we confirmed that G-protein coupled receptor 5A relocates to lipid rafts along with F508del-CFTR after TNF-α treatment. These results could enlighten the cross-talk between CFTR and TNF-α and its impact on the cell response to proinflammatory challenge.
  
  
  
  CFTR mutations are at the origin of cystic fibrosis. The latter disease is characterized, among other symptoms, by a defective management of infection and inflammation in the airways. Short exposure to the proinflammatory cytokine TNF-α targets mutated CFTR to the plasma membrane and increases its chloride channel activity. The results hereby presented show a substantial modification of the lipid raft proteome in the same conditions, and may enlighten the effect of this cytokine and the role of CFTR in the cell response to inflammation.
authors.affiliation: Proteomic Platform Necker, PPN-3P5, Structure Fédérative de Recherche SFR Necker US24, 75015, Paris, France.
authors.forename: C
authors.initials: C
authors.lastname: Chhuon
authors.affiliation: Institut Necker Enfants Malades, INSERM, U1151, Paris, France.
authors.forename: I
authors.initials: I
authors.lastname: Pranke
authors.affiliation: Institut Necker Enfants Malades, INSERM, U1151, Paris, France.
authors.forename: F
authors.initials: F
authors.lastname: Borot
authors.affiliation: Institut Necker Enfants Malades, INSERM, U1151, Paris, France.
authors.forename: D
authors.initials: D
authors.lastname: Tondelier
authors.affiliation: The CPN Proteomics Facility - 3P5, Center of Psychiatry and Neuroscience, UMR INSERM 894, 75014, Paris, France.
authors.forename: J
authors.initials: J
authors.lastname: Lipecka
authors.affiliation: Institut Necker Enfants Malades, INSERM, U1151, Paris, France.
authors.forename: J
authors.initials: J
authors.lastname: Fritsch
authors.affiliation: Geneva University Hospitals and University of Geneva, 1211 Geneva, Switzerland.
authors.forename: M
authors.initials: M
authors.lastname: Chanson
authors.affiliation: Institut Necker Enfants Malades, INSERM, U1151, Paris, France.
authors.forename: A
authors.initials: A
authors.lastname: Edelman
authors.affiliation: Institut Mondor de Recherche Biomédicale, INSERM, U955, and Université Paris Est Créteil, 94010, Créteil, France. Electronic address: Mario.ollero@inserm.fr.
authors.forename: M
authors.initials: M
authors.lastname: Ollero
authors.affiliation: Proteomic Platform Necker, PPN-3P5, Structure Fédérative de Recherche SFR Necker US24, 75015, Paris, France. Electronic address: chiara.guerrera@inserm.fr.
authors.forename: I C
authors.initials: IC
authors.lastname: Guerrera
mesh_terms: D001980:Bronchi; D002478:Cells, Cultured; D003550:Cystic Fibrosis / Q000473:pathology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics; D004847:Epithelial Cells; D006367:HeLa Cells; D006801:Humans; D007249:Inflammation; D021962:Membrane Microdomains / Q000737:chemistry*; D009154:Mutation; D020543:Proteome / Q000032:analysis / Q000187:drug effects*; D040901:Proteomics / Q000379:methods; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D020543:Proteome; D014409:Tumor Necrosis Factor-alpha; D019005:Cystic Fibrosis Transmembrane Conductance Regulator
keywords: CFTR; Cystic fibrosis; GPCR; Inflammation; Lipid rafts; SILAC