title: Functional Rescue of Trafficking-Impaired ABCB4 Mutants by Chemical Chaperones.
abstract: Multidrug resistance protein 3 (MDR3, ABCB4) is a hepatocellular membrane protein that mediates biliary secretion of phosphatidylcholine. Null mutations in ABCB4 gene give rise to severe early-onset cholestatic liver disease. We have previously shown that the disease-associated mutations p.G68R, p.G228R, p.D459H, and p.A934T resulted in retention of ABCB4 in the endoplasmic reticulum, thus failing to target the plasma membrane. In the present study, we tested the ability of two compounds with chaperone-like activity, 4-phenylbutyrate and curcumin, to rescue these ABCB4 mutants by assessing their effects on subcellular localization, protein maturation, and phospholipid efflux capability. Incubation of transfected cells at a reduced temperature (30°C) or exposure to pharmacological doses of either 4-PBA or curcumin restored cell surface expression of mutants G228R and A934T. The delivery of these mutants to the plasma membrane was accompanied by a switch in the ratio of mature to inmature protein forms, leading to a predominant expression of the mature protein. This effect was due to an improvement in the maturation rate and not to the stabilization of the mature forms. Both mutants were also functionally rescued, displaying bile salt-dependent phospholipid efflux activity after addition of 4-PBA or curcumin. Drug-induced rescue was mutant specific, given neither 4-PBA nor curcumin had an effect on the ABCB4 mutants G68R and A934T. Collectively, these data indicate that the functionality of selected trafficking-defective ABCB4 mutants can be recovered by chemical chaperones through restoration of membrane localization, suggesting a potential treatment for patients carrying such mutations.
authors.affiliation: La Paz University Hospital Health Research Institute-IdiPAZ, Madrid, Spain.
authors.forename: Raquel
authors.initials: R
authors.lastname: Gordo-Gilart
authors.affiliation: La Paz University Hospital Health Research Institute-IdiPAZ, Madrid, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: Andueza
authors.affiliation: La Paz University Hospital Health Research Institute-IdiPAZ, Madrid, Spain.|Pediatric Liver Service, La Paz Children's University Hospital, Madrid, Spain.
authors.forename: Loreto
authors.initials: L
authors.lastname: Hierro
authors.affiliation: La Paz University Hospital Health Research Institute-IdiPAZ, Madrid, Spain.|Pediatric Liver Service, La Paz Children's University Hospital, Madrid, Spain.
authors.forename: Paloma
authors.initials: P
authors.lastname: Jara
authors.affiliation: La Paz University Hospital Health Research Institute-IdiPAZ, Madrid, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Alvarez
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D003474:Curcumin / Q000494:pharmacology; D004285:Dogs; D018832:Molecular Chaperones / Q000494:pharmacology*; D010654:Phenylbutyrates / Q000494:pharmacology; D010743:Phospholipids / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects*
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D018832:Molecular Chaperones; D010654:Phenylbutyrates; D010743:Phospholipids; C075773:4-phenylbutyric acid; C095122:multidrug resistance protein 3; D003474:Curcumin