Farnesoid X receptor, hepatocyte nuclear factors 1alpha and 3beta are essential for transcriptional activation of the liver-specific organic anion transporter-2 gene.
BACKGROUND
We isolated the human liver-specific organic anion transporter gene, LST-2 (OATP8/SLCO1B3), which is exclusively expressed in the basolateral membrane of the hepatocytes. In this study, we analyzed the transcriptional regulation of the LST-2 gene in hepatocyte-derived cells and the effect of bile acid.


METHODS
Transcriptional activity of the LST-2 gene was measured using a human LST-2 promoter-luciferase reporter plasmid under various concentrations of bile acids. Electrophoresis mobility shift assays of farnesoid X receptor (FXR), hepatocyte nuclear factor (HNF) 1alpha, and HNF3beta were performed.


RESULTS
Luciferase analysis showed that the 5'-flanking region from -180 to -20 bp is responsible for LST-2 transcriptional activity. By site-directed mutation analysis, it was revealed that the consensus binding sites for FXR, HNF1alpha, and HNF3beta play important roles in the transcriptional activity of the LST-2 gene. By electrophoresis mobility shift assay, we observed specific protein-DNA complexes of FXR, HNF1alpha, and HNF-3beta. Luciferase activity was increased fivefold when chenodeoxycholate or deoxycholate were added. Northern blot analyses revealed that the expression of LST-2 was increased by addition of chenodeoxycholate or deoxycholate in a dose-dependent manner.


CONCLUSIONS
This study demonstrated that the transcription of the LST-2 gene is regulated by three transcription factors, FXR, HNF1alpha, and HNF3beta. HNF1alpha and HNF3beta might contribute to its liver-specific expression, and FXR might play a role in its transcriptional activation by bile acids.
Ohtsuka
Hideo
H
Division of Gastroenterological Surgery, Department of Surgery, Tohoku University Graduate School of Medical Science, Sendai 980-8574, Japan.
Abe
Takaaki
T
Onogawa
Tohru
T
Kondo
Noriko
N
Sato
Takeaki
T
Oshio
Hiroshi
H
Mizutamari
Hiroya
H
Mikkaichi
Tsuyoshi
T
Oikawa
Masaya
M
Rikiyama
Toshiki
T
Katayose
Yu
Y
Unno
Michiaki
M
D001647:Bile Acids and Salts / Q000494:pharmacology; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000378:metabolism*; D051542:Hepatocyte Nuclear Factor 3-beta / Q000378:metabolism*; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D016296:Mutagenesis; D029362:Organic Anion Transporters, Sodium-Independent / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation; D000099150:Receptor, Farnesoid X-Activated
D001647:Bile Acids and Salts; D004247:DNA; D004268:DNA-Binding Proteins; D051538:Hepatocyte Nuclear Factor 1-alpha; D051542:Hepatocyte Nuclear Factor 3-beta; D029362:Organic Anion Transporters, Sodium-Independent; D018160:Receptors, Cytoplasmic and Nuclear; D000074542:Solute Carrier Organic Anion Transporter Family Member 1B3; D014157:Transcription Factors; D000099150:Receptor, Farnesoid X-Activated; C421209:SLCO1B3 protein, human