title: Down-regulation of ATP-binding cassette C2 protein expression in HepG2 cells after rifampicin treatment is mediated by microRNA-379.
abstract: microRNAs (miRNAs), which contribute to the post-transcriptional processing through 3'-untranslated region-interference, have been shown to be involved in the regulation of ATP-binding cassette (ABC) membrane transporters. The aim of this study was to investigate whether ABCC2, an important efflux transporter for various endogenous and exogenous compounds at several compartment barriers, is subject to miRNA-mediated post-transcriptional gene regulation. We screened the expression of 377 human miRNAs in HepG2 cells after 48 h of treatment with 5 μM rifampicin [a pregnane X receptor (PXR) ligand] or vehicle using reverse transcription-polymerase chain reaction-based low-density arrays. Specific miRNA, ABCC2 mRNA, and protein expression were monitored in HepG2 cells undergoing rifampicin treatment for 72 h. Loss- and gain-of-function experiments and reporter gene assays were performed for further confirmation. Highly deregulated miRNAs compared with in silico data revealed miRNA (miR) 379 as candidate miRNA targeting ABCC2 mRNA. Under rifampicin treatment, ABCC2 mRNA increased significantly, with a maximal fold change of 1.56 ± 0.43 after 24 h. In addition, miR-379 increased (maximally 4.10 ± 1.33-fold after 48 h), whereas ABCC2 protein decreased with a maximal fold change of 0.47 ± 0.08 after 72 h. In contrast, transfection of miR-379 inhibitor led to an elevation of ABCC2 protein expression after rifampicin incubation for 48 h. We identify a miRNA negatively regulating ABCC2 on the post-transcriptional level and provide evidence that this miRNA impedes overexpression of ABCC2 protein after a PXR-mediated external transcriptional stimulus in HepG2 cells.
authors.affiliation: Institute for Experimental and Clinical Pharmacology, University Hospital Schleswig-Holstein, Bldg. 30, Arnold-Heller-Str. 3, D-24105 Kiel, Germany.
authors.forename: Sierk
authors.initials: S
authors.lastname: Haenisch
authors.forename: Sandra
authors.initials: S
authors.lastname: Laechelt
authors.forename: Henrike
authors.initials: H
authors.lastname: Bruckmueller
authors.forename: Anneke
authors.initials: A
authors.lastname: Werk
authors.forename: Andreas
authors.initials: A
authors.lastname: Noack
authors.forename: Oliver
authors.initials: O
authors.lastname: Bruhn
authors.forename: Cornelia
authors.initials: C
authors.lastname: Remmler
authors.forename: Ingolf
authors.initials: I
authors.lastname: Cascorbi
mesh_terms: D001665:Binding Sites / Q000235:genetics; D015536:Down-Regulation / Q000187:drug effects* / Q000235:genetics*; D056945:Hep G2 Cells; D006801:Humans; D035683:MicroRNAs / Q000502:physiology*; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000096:biosynthesis* / Q000235:genetics; D034622:RNA Interference / Q000502:physiology; D012293:Rifampin / Q000494:pharmacology*
chemical_list: D035683:MicroRNAs; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D012293:Rifampin; C000715087:ABCC2 protein, human; C561022:MIRN379 microRNA, human