title: Cystic fibrosis transmembrane conductance regulator trafficking is mediated by the COPI coat in epithelial cells.
abstract: Cystic fibrosis (CF) is caused by defects in the CF transmembrane conductance regulator (CFTR) that functions as a chloride channel in epithelial cells. The most common cause of CF is the abnormal trafficking of CFTR mutants. Therefore, understanding the cellular machineries that transit CFTR from the endoplasmic reticulum to the plasma membrane (PM) is important. The coat protein complex I (COPI) has been implicated in the anterograde and retrograde transport of proteins and lipids between the endoplasmic reticulum and the Golgi. Here, we investigated the role of COPI in CFTR trafficking. Blocking COPI recruitment to membranes by expressing an inactive form of the GBF1 guanine nucleotide exchange factor for ADP-ribosylation factor inhibits CFTR trafficking to the PM. Similarly, inhibiting COPI dissociation from membranes by expressing a constitutively active ADP-ribosylation factor 1 mutant arrests CFTR within disrupted Golgi elements. To definitively explore the relationship between COPI and CFTR in epithelial cells, we depleted beta-COP from the human colonic epithelial cell HT-29Cl.19A using small interfering RNA. Beta-COP depletion did not affect CFTR synthesis but impaired its trafficking to the PM. The arrest occurred pre-Golgi as shown by reduced level of glycosylation. Importantly, decreased trafficking of CFTR had a functional consequence as cells depleted of beta-COP showed decreased cAMP-activated chloride currents. To explore the mechanism of COPI action in CFTR traffic we tested whether CFTR was COPI cargo. We discovered that the alpha-, beta-, and gamma-subunits of COPI co-immunoprecipitated with CFTR. Our results indicate that the COPI complex plays a critical role in CFTR trafficking to the PM.
authors.affiliation: Division of Pulmonary, Critical Care, and Sleep Medicine, The Ohio State University, Columbus, Ohio 43210, USA.
authors.forename: Jessica
authors.initials: J
authors.lastname: Rennolds
authors.forename: Cristy
authors.initials: C
authors.lastname: Tower
authors.forename: Lois
authors.initials: L
authors.lastname: Musgrove
authors.forename: Lijuan
authors.initials: L
authors.lastname: Fan
authors.forename: Kevin
authors.initials: K
authors.lastname: Maloney
authors.forename: John Paul
authors.initials: JP
authors.lastname: Clancy
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Kirk
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Sztul
authors.forename: Estelle
authors.initials: E
authors.lastname: Cormet-Boyaka
mesh_terms: D000230:Adenocarcinoma; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000502:physiology; D020755:Coat Protein Complex I / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000502:physiology*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004847:Epithelial Cells / Q000502:physiology*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005905:Glyburide / Q000494:pharmacology; D006056:Golgi Apparatus / Q000502:physiology; D006801:Humans; D012334:RNA, Neoplasm / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D020755:Coat Protein Complex I; D012334:RNA, Neoplasm; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D005905:Glyburide