Characterization of the 5'-flanking region of the human multidrug resistance protein 2 (MRP2) gene and its regulation in comparison withthe multidrug resistance protein 3 (MRP3) gene.
The multidrug resistance proteins MRP2 (symbol ABCC2) and MRP3 (symbol ABCC3) are conjugate export pumps expressed in hepatocytes. MRP2 is localized exclusively to the apical membrane and MRP3 to the basolateral membrane. MRP2 mRNA is expressed at a high level under normal conditions, whereas MRP3 mRNA expression is low and increases only when secretion across the apical membrane by MRP2 is impaired. We studied some of the regulatory properties of the two human genes using transient transfection assays with promoter-luciferase constructs in HepG2 cells and cloned fragments of 1229 nucleotides and 1287 nucleotides of the MRP2 and MRP3 5'-flanking regions, respectively. The sequence between nucleotides -517 and -197 was decisive for basal MRP2 expression. Basal promoter activity of MRP3 was only 4% of that measured for MRP2. At submicromolar concentrations, the histone deacetylase inhibitor trichostatin A reduced the MRP2 reporter gene activity and expression of the protein. Disruption of microtubules with nocodazole decreased gene and protein expression of MRP2 and increased MRP3 reporter gene activity. The genotoxic 2-acetylaminofluorene decreased the activity of the human MRP2 reporter gene construct, but increased MRP3 gene activity and enhanced the amounts of mRNA and protein of MRP2 and MRP3. Thus, regulation of the expression of these ATP-dependent conjugate export pumps is not co-ordinate, but in part inverse. The inverse regulation of the two MRP isoforms is consistent with their distinct localization, their different mRNA expression under normal and pathophysiological conditions, and their different directions of substrate transport in polarized cells.
Stöckel
B
B
Division of Tumor Biochemistry, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
König
J
J
Nies
A T
AT
Cui
Y
Y
Brom
M
M
Keppler
D
D
D015073:2-Acetylaminofluorene / Q000494:pharmacology; D018528:ATP-Binding Cassette Transporters / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D015151:Immunoblotting; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D015739:Nocodazole / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D012269:Ribosomal Proteins / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D014162:Transfection; D014407:Tumor Cells, Cultured
D015073:2-Acetylaminofluorene; D018528:ATP-Binding Cassette Transporters; D018076:DNA, Complementary; D006877:Hydroxamic Acids; D024101:Mitochondrial Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D015739:Nocodazole; D012333:RNA, Messenger; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; C000715087:ABCC2 protein, human; C093947:MRP2 protein, S cerevisiae; C114462:multidrug resistance-associated protein 3; C012589:trichostatin A