title: Sodium 4-phenylbutyrate downregulates Hsc70: implications for intracellular trafficking of DeltaF508-CFTR.
abstract: The most common mutation of the cystic fibrosis transmembrane conductance regulator (CFTR), DeltaF508, is a trafficking mutant that has prolonged associations with molecular chaperones and is rapidly degraded, at least in part by the ubiquitin-proteasome system. Sodium 4-phenylbutyrate (4PBA) improves DeltaF508-CFTR trafficking and function in vitro in cystic fibrosis epithelial cells and in vivo. To further understand the mechanism of action of 4PBA, we tested the hypothesis that 4PBA modulates the targeting of DeltaF508-CFTR for ubiquitination and degradation by reducing the expression of Hsc70 in cystic fibrosis epithelial cells. IB3-1 cells (genotype DeltaF508/W1282X) that were treated with 0.05-5 mM 4PBA for 2 days in culture demonstrated a dose-dependent reduction in Hsc70 protein immunoreactivity and mRNA levels. Immunoprecipitation with Hsc70-specific antiserum demonstrated that Hsc70 and CFTR associated under control conditions and that treatment with 4PBA reduced these complexes. Levels of immunoreactive Hsp40, Hdj2, Hsp70, Hsp90, and calnexin were unaffected by 4PBA treatment. These data suggest that 4PBA may improve DeltaF508-CFTR trafficking by allowing a greater proportion of mutant CFTR to escape association with Hsc70.
authors.affiliation: Division of Pulmonary Medicine, Children's Hospital of Philadelphia and Department of Pediatrics, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA. rrubenst@mail.med.upenn.edu
authors.forename: R C
authors.initials: RC
authors.lastname: Rubenstein
authors.forename: P L
authors.initials: PL
authors.lastname: Zeitlin
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001692:Biological Transport / Q000187:drug effects; D002087:Butyrates / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000235:genetics; D037281:Calnexin; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003550:Cystic Fibrosis / Q000235:genetics / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation / Q000187:drug effects; D015870:Gene Expression / Q000187:drug effects; D005990:Glycerol / Q000494:pharmacology; D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics; D006360:Heat-Shock Proteins / Q000235:genetics; D006801:Humans; D009097:Multienzyme Complexes / Q000378:metabolism; D009154:Mutation; D010654:Phenylbutyrates / Q000494:pharmacology*; D046988:Proteasome Endopeptidase Complex; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D014452:Ubiquitins / Q000378:metabolism
chemical_list: D000970:Antineoplastic Agents; D002087:Butyrates; C505032:CFTR protein, human; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; C107481:DNAJA1 protein, human; C494647:DNAJB1 protein, human; D050883:HSC70 Heat-Shock Proteins; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; C494371:HSPA8 protein, human; D006360:Heat-Shock Proteins; D009097:Multienzyme Complexes; D010654:Phenylbutyrates; D012333:RNA, Messenger; D014452:Ubiquitins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D037281:Calnexin; C075773:4-phenylbutyric acid; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D005990:Glycerol