title: Na+/H+ exchanger regulatory factor isoform 1 overexpression modulates cystic fibrosis transmembrane conductance regulator (CFTR) expression and activity in human airway 16HBE14o- cells and rescues DeltaF508 CFTR functional expression in cystic fibrosis cells.
abstract: There is evidence that cystic fibrosis transmembrane conductance regulator (CFTR) interacting proteins play critical roles in the proper expression and function of CFTR. The Na(+)/H(+) exchanger regulatory factor isoform 1 (NHERF1) was the first identified CFTR-binding protein. Here we further clarify the role of NHERF1 in the regulation of CFTR activity in two human bronchial epithelial cell lines: the normal, 16HBE14o-, and the homozygous DeltaF508 CFTR, CFBE41o-. Confocal analysis in polarized cell monolayers demonstrated that NHERF1 distribution was associated with the apical membrane in 16HBE14o- cells while being primarily cytoplasmic in CFBE41o- cells. Transfection of 16HBE14o- monolayers with vectors encoding for wild-type (wt) NHERF1 increased both apical CFTR expression and apical protein kinase A (PKA)-dependent CFTR-mediated chloride efflux, whereas transfection with NHERF1 mutated in the binding groove of the PDZ domains or truncated for the ERM domain inhibited both the apical CFTR expression and the CFTR-dependent chloride efflux. These data led us to hypothesize an important role for NHERF1 in regulating CFTR localization and stability on the apical membrane of 16HBE14o- cell monolayers. Importantly, wt NHERF1 overexpression in confluent DeltaF508 CFBE41o- and DeltaF508 CFT1-C2 cell monolayers induced both a significant redistribution of CFTR from the cytoplasm to the apical membrane and a PKA-dependent activation of CFTR-dependent chloride secretion.
authors.affiliation: Department of General and Environmental Physiology, University of Bari, Via Amendola 165/A, Bari 70126, Italy.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Guerra
authors.forename: Teresa
authors.initials: T
authors.lastname: Fanelli
authors.forename: Maria
authors.initials: M
authors.lastname: Favia
authors.forename: Stefania M
authors.initials: SM
authors.lastname: Riccardi
authors.forename: Giovanni
authors.initials: G
authors.lastname: Busco
authors.forename: Rosa Angela
authors.initials: RA
authors.lastname: Cardone
authors.forename: Salvatore
authors.initials: S
authors.lastname: Carrabino
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Weinman
authors.forename: Stephan Joel
authors.initials: SJ
authors.lastname: Reshkin
authors.forename: Massimo
authors.initials: M
authors.lastname: Conese
authors.forename: Valeria
authors.initials: V
authors.lastname: Casavola
mesh_terms: D001665:Binding Sites / Q000235:genetics; D001980:Bronchi / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D002712:Chlorides / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003550:Cystic Fibrosis / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D003593:Cytoplasm / Q000737:chemistry; D004355:Drug Stability; D004847:Epithelial Cells; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D005786:Gene Expression Regulation; D006706:Homeostasis; D006801:Humans; D018613:Microscopy, Confocal; D009154:Mutation; D010750:Phosphoproteins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D017923:Sodium-Hydrogen Exchangers; D014162:Transfection
chemical_list: D002712:Chlorides; D010750:Phosphoproteins; D017923:Sodium-Hydrogen Exchangers; C093349:cystic fibrosis transmembrane conductance regulator delta F508; C093376:sodium-hydrogen exchanger regulatory factor; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D017868:Cyclic AMP-Dependent Protein Kinases