title: Functional characterization of ABCB4 mutations found in progressive familial intrahepatic cholestasis type 3.
abstract: Multidrug resistance 3 (MDR3), encoded by the ATP-binding cassette, subfamily B, member 4 gene (ABCB4), localizes to the canalicular membrane of hepatocytes and translocates phosphatidylcholine from the inner leaflet to the outer leaflet of the canalicular membrane. Progressive familial intrahepatic cholestasis type 3 (PFIC3) is a rare hepatic disease caused by genetic mutations of ABCB4. In this study, we characterized 8 ABCB4 mutations found in PFIC3 patients, using in vitro molecular assays. First, we examined the transport activity of each mutant by measuring its ATPase activity using paclitaxel or phosphatidylcholine. Then, the pathogenic mechanisms by which these mutations affect MDR3 were examined through immunoblotting, cell surface biotinylation, and immunofluorescence. As a result, three ABCB4 mutants showed significantly reduced transport activity. Among these mutants, one mutation A364V, located in intracellular domains, markedly decreased MDR3 expression on the plasma membrane, while the others did not affect the expression. The expression of MDR3 on the plasma membrane and transport activity of A364V was rescued by a pharmacological chaperone, cyclosporin A. Our study provides the molecular mechanisms of ABCB4 mutations and may contribute to the understanding of PFIC3 pathogenesis and the development of a mutation-specific targeted treatment for PFIC3.
authors.affiliation: Department of Pharmacology, Tissue Injury Defense Research Center, School of Medicine, Ewha Womans University, Seoul, Korea.
authors.forename: Hyo Jin
authors.initials: HJ
authors.lastname: Park
authors.affiliation: Department of Pharmacology, Tissue Injury Defense Research Center, School of Medicine, Ewha Womans University, Seoul, Korea.
authors.forename: Tae Hee
authors.initials: TH
authors.lastname: Kim
authors.affiliation: Department of Pharmacology and the Institute for Clinical and Translational Research, Catholic Kwandong University College of Medicine, Gangneung, Korea.
authors.forename: So Won
authors.initials: SW
authors.lastname: Kim
authors.affiliation: Department of Pharmacology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Korea.
authors.forename: Shin Hye
authors.initials: SH
authors.lastname: Noh
authors.affiliation: Department of Pharmacology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Korea.
authors.forename: Kyeong Jee
authors.initials: KJ
authors.lastname: Cho
authors.affiliation: Department of Pharmacology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Korea.
authors.forename: Choe
authors.initials: C
authors.lastname: Choi
authors.affiliation: Department of Pharmacology, Tissue Injury Defense Research Center, School of Medicine, Ewha Womans University, Seoul, Korea.
authors.forename: Eun Young
authors.initials: EY
authors.lastname: Kwon
authors.affiliation: Department of Pharmacology, Tissue Injury Defense Research Center, School of Medicine, Ewha Womans University, Seoul, Korea.
authors.forename: Yang Ji
authors.initials: YJ
authors.lastname: Choi
authors.affiliation: Department of Pharmacology, Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Korea.
authors.forename: Heon Yung
authors.initials: HY
authors.lastname: Gee
authors.affiliation: Department of Pharmacology, Tissue Injury Defense Research Center, School of Medicine, Ewha Womans University, Seoul, Korea.
authors.forename: Ji Ha
authors.initials: JH
authors.lastname: Choi
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism; D002780:Cholestasis, Intrahepatic / Q000235:genetics*; D016572:Cyclosporine / Q000494:pharmacology; D015870:Gene Expression; D056726:Genetic Association Studies; D020022:Genetic Predisposition to Disease; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D007976:Leupeptins / Q000494:pharmacology; D018942:Macrolides / Q000494:pharmacology; D020125:Mutation, Missense; D017239:Paclitaxel / Q000378:metabolism; D021381:Protein Transport
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D007976:Leupeptins; D018942:Macrolides; D016572:Cyclosporine; C040929:bafilomycin A1; C095122:multidrug resistance protein 3; D017239:Paclitaxel; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde