title: The human organic anion transporter 2 gene is transactivated by hepatocyte nuclear factor-4 alpha and suppressed by bile acids.
abstract: The human organic anion transporter 2 (hOAT2, SLC22A7) mediates the sodium-independent uptake of numerous drugs, including cephalosporins, salicylates, dicarboxylates, and prostaglandins, and is mainly expressed in hepatocytes. Because the regulation of hOAT2 expression is poorly understood, we characterized cis-acting elements in the 5'-flanking region that regulate hOAT2 transcription. A consensus binding motif for the hepatocyte nuclear factor-4 alpha (HNF-4 alpha), arranged as a direct repeat (DR)-1, is located at nucleotides -329/-317 relative to the transcription initiation site. This element specifically binds HNF-4 alpha in electrophoretic mobility shift assays. A luciferase-linked hOAT2 promoter fragment containing the HNF-4 alpha binding site was transactivated upon cotransfection of an HNF-4 alpha expression vector in Huh7 cells, whereas site-directed mutagenesis of the DR-1 element abolished activation by HNF-4 alpha. Short interfering RNAs inhibiting endogenous HNF-4 alpha expression markedly reduced endogenous expression of hOAT2 in Huh7 cells. Because HNF-4 alpha is a known target for bile acid-mediated repression of gene transcription, we studied whether chenodeoxycholic acid (CDCA) suppresses hOAT2 gene expression by inhibiting HNF-4 alpha-mediated transactivation. Treatment of Huh7 cells with CDCA or the synthetic farnesoid X receptor (FXR) agonist GW4064 decreased mRNA and protein levels and also nuclear binding activity of HNF-4 alpha. The FXR-inducible transcriptional repressor small heterodimer partner inhibited transactivation of hOAT2 promoter constructs and of endogenous hOAT2 expression by HNF-4 alpha. We conclude that the hOAT2 gene is critically dependent on HNF-4 alpha and that bile acids repress the hOAT2 gene by inhibiting HNF-4 alpha. Hepatic uptake of hOAT2 substrates may thus be decreased in disease conditions associated with elevated intracellular levels of bile acids.
authors.affiliation: Laboratory of Molecular Gastroenterology and Hepatology, University Hospital, Zurich, Switzerland.
authors.forename: Katrin
authors.initials: K
authors.lastname: Popowski
authors.forename: Jyrki J
authors.initials: JJ
authors.lastname: Eloranta
authors.forename: Michael
authors.initials: M
authors.lastname: Saborowski
authors.forename: Michael
authors.initials: M
authors.lastname: Fried
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Meier
authors.forename: Gerd A
authors.initials: GA
authors.lastname: Kullak-Ublick
mesh_terms: D001483:Base Sequence; D001647:Bile Acids and Salts / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D008969:Molecular Sequence Data; D029362:Organic Anion Transporters, Sodium-Independent / Q000235:genetics / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000502:physiology*; D015534:Trans-Activators / Q000502:physiology*; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D015533:Transcriptional Activation / Q000502:physiology*
chemical_list: D001647:Bile Acids and Salts; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; D029362:Organic Anion Transporters, Sodium-Independent; D010750:Phosphoproteins; C490280:SLC22A7 protein, human; D015534:Trans-Activators; D014157:Transcription Factors