The Human Organic Anion Transporter 2 Gene Is Transactivated by Hepatocyte Nuclear Factor-4α and Suppressed by Bile Acids

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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-4α (HNF-4α), arranged as a direct repeat (DR)-1, is located at nucleotides –329/-317 relative to the transcription initiation site. This element specifically binds HNF-4α in electrophoretic mobility shift assays. A luciferase-linked hOAT2 promoter fragment containing the HNF-4α binding site was transactivated upon cotransfection of an HNF-4α expression vector in Huh7 cells, whereas site-directed mutagenesis of the DR-1 element abolished activation by HNF-4α. Short interfering RNAs inhibiting endogenous HNF-4α expression markedly reduced endogenous expression of hOAT2 in Huh7 cells. Because HNF-4α 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-4α-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-4α. The FXR-inducible transcriptional repressor small heterodimer partner inhibited transactivation of hOAT2 promoter constructs and of endogenous hOAT2 expression by HNF-4α. We conclude that the hOAT2 gene is critically dependent on HNF-4α and that bile acids repress the hOAT2 gene by inhibiting HNF-4α. Hepatic uptake of hOAT2 substrates may thus be decreased in disease conditions associated with elevated intracellular levels of bile acids.

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