title: Ribosomal Stalk Protein Silencing Partially Corrects the ΔF508-CFTR Functional Expression Defect.
abstract: The most common cystic fibrosis (CF) causing mutation, deletion of phenylalanine 508 (ΔF508 or Phe508del), results in functional expression defect of the CF transmembrane conductance regulator (CFTR) at the apical plasma membrane (PM) of secretory epithelia, which is attributed to the degradation of the misfolded channel at the endoplasmic reticulum (ER). Deletion of phenylalanine 670 (ΔF670) in the yeast oligomycin resistance 1 gene (YOR1, an ABC transporter) of Saccharomyces cerevisiae phenocopies the ΔF508-CFTR folding and trafficking defects. Genome-wide phenotypic (phenomic) analysis of the Yor1-ΔF670 biogenesis identified several modifier genes of mRNA processing and translation, which conferred oligomycin resistance to yeast. Silencing of orthologues of these candidate genes enhanced the ΔF508-CFTR functional expression at the apical PM in human CF bronchial epithelia. Although knockdown of RPL12, a component of the ribosomal stalk, attenuated the translational elongation rate, it increased the folding efficiency as well as the conformational stability of the ΔF508-CFTR, manifesting in 3-fold augmented PM density and function of the mutant. Combination of RPL12 knockdown with the corrector drug, VX-809 (lumacaftor) restored the mutant function to ~50% of the wild-type channel in primary CFTRΔF508/ΔF508 human bronchial epithelia. These results and the observation that silencing of other ribosomal stalk proteins partially rescue the loss-of-function phenotype of ΔF508-CFTR suggest that the ribosomal stalk modulates the folding efficiency of the mutant and is a potential therapeutic target for correction of the ΔF508-CFTR folding defect.
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.
authors.forename: Guido
authors.initials: G
authors.lastname: Veit
authors.affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.|Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: Kathryn
authors.initials: K
authors.lastname: Oliver
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.
authors.forename: Pirjo M
authors.initials: PM
authors.lastname: Apaja
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.
authors.forename: Doranda
authors.initials: D
authors.lastname: Perdomo
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.
authors.forename: Aurélien
authors.initials: A
authors.lastname: Bidaud-Meynard
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.
authors.forename: Sheng-Ting
authors.identifier: 0000-0002-8109-3557
authors.initials: ST
authors.lastname: Lin
authors.affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: Jingyu
authors.initials: J
authors.lastname: Guo
authors.affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: Mert
authors.initials: M
authors.lastname: Icyuz
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, United States of America.
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Sorscher
authors.affiliation: Department of Genetics, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.|Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.
authors.forename: John L
authors.initials: JL
authors.lastname: Hartman
authors.affiliation: Department of Physiology, McGill University, Montréal, Quebec, Canada.|Department of Biochemistry, McGill University, Montréal, Quebec, Canada.|Groupe de Recherche Axé sur la Structure des Protéines (GRASP), McGill University, Montréal, Quebec, Canada.
authors.forename: Gergely L
authors.initials: GL
authors.lastname: Lukacs
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000631:Aminopyridines / Q000494:pharmacology; D052117:Benzodioxoles / Q000494:pharmacology; D001980:Bronchi / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D003550:Cystic Fibrosis / Q000188:drug therapy / Q000235:genetics / Q000473:pathology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D057166:High-Throughput Screening Assays; D006801:Humans; D020652:Peptide Elongation Factor 2 / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D055550:Protein Stability; D034741:RNA, Small Interfering; D012269:Ribosomal Proteins / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D015003:Yeasts / Q000235:genetics
chemical_list: D018528:ATP-Binding Cassette Transporters; D000631:Aminopyridines; D052117:Benzodioxoles; C505032:CFTR protein, human; D020652:Peptide Elongation Factor 2; D034741:RNA, Small Interfering; C000620655:RPL12 protein, human; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; C097077:YOR1 protein, S cerevisiae; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; C569105:lumacaftor