title: NHE-RF1 protein rescues DeltaF508-CFTR function.
abstract: In cystic fibrosis (CF), the DeltaF508-CFTR anterograde trafficking from the endoplasmic reticulum to the plasma membrane is inefficient. New strategies for increasing the delivery of DeltaF508-CFTR to the apical membranes are thus pathophysiologically relevant targets to study for CF treatment. Recent studies have demonstrated that PDZ-containing proteins play an essential role in determining polarized plasma membrane expression of ionic transporters. In the present study we have hypothesized that the PDZ-containing protein NHE-RF1, which binds to the carboxy terminus of CFTR, rescues DeltaF508-CFTR expression in the apical membrane of epithelial cells. The plasmids encoding DeltaF508-CFTR and NHE-RF1 were intranuclearly injected in A549 or Madin-Darby canine kidney (MDCK) cells, and DeltaF508-CFTR channel activity was functionally assayed using SPQ fluorescent probe. Cells injected with DeltaF508-CFTR alone presented a low chloride channel activity, whereas its coexpression with NHE-RF1 significantly increased both the basal and forskolin-activated chloride conductances. This last effect was lost with DeltaF508-CFTR deleted of its 13 last amino acids or by injection of a specific NHE-RF1 antisense oligonucleotide, but not by NHE-RF1 sense oligonucleotide. Immunocytochemical analysis performed in MDCK cells transiently transfected with DeltaF508-CFTR further revealed that NHE-RF1 specifically determined the apical plasma membrane expression of DeltaF508-CFTR but not that of a trafficking defective mutant potassium channel (KCNQ1). These data demonstrate that the modulation of the expression level of CFTR protein partners, like NHE-RF1, can rescue DeltaF508-CFTR activity.
authors.affiliation: Institut National de la Santé et de la Recherche Médicale Unité 533, l' Institut du Thorax, Faculté de Médecine, Nantes, France.
authors.forename: Florian
authors.initials: F
authors.lastname: Bossard
authors.forename: Amal
authors.initials: A
authors.lastname: Robay
authors.forename: Gilles
authors.initials: G
authors.lastname: Toumaniantz
authors.forename: Shehrazade
authors.initials: S
authors.lastname: Dahimene
authors.forename: Frédéric
authors.initials: F
authors.lastname: Becq
authors.forename: Jean
authors.initials: J
authors.lastname: Merot
authors.forename: Chantal
authors.initials: C
authors.lastname: Gauthier
mesh_terms: D000818:Animals; D002460:Cell Line; D016764:Cell Polarity; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000502:physiology*; D017931:DNA Primers; D004285:Dogs; D005786:Gene Expression Regulation; D006801:Humans; D007668:Kidney; D010750:Phosphoproteins / Q000235:genetics / Q000502:physiology*; D016133:Polymerase Chain Reaction; D020545:Respiratory Mucosa / Q000502:physiology*; D017384:Sequence Deletion; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000502:physiology*
chemical_list: C505032:CFTR protein, human; D017931:DNA Primers; D010750:Phosphoproteins; D017923:Sodium-Hydrogen Exchangers; C093376:sodium-hydrogen exchanger regulatory factor; D019005:Cystic Fibrosis Transmembrane Conductance Regulator