Nuclear factor erythroid 2-related factor 2 is a positive regulator of human bile salt export pump expression.
The bile salt export pump (BSEP) is the major determinant of bile salt-dependent bile secretion, and its deficiency leads to cholestatic liver injury. BSEP/Bsep gene expression is regulated by the nuclear farnesoid X receptor. However, BSEP expression, though reduced, is retained in the livers of Fxr(-/-) mice, indicating that additional transcriptional factors may regulate its expression. Nuclear factor erythroid 2-related factor 2 (Nrf2) plays a major role in response to oxidative stress by binding to antioxidant-responsive elements that regulate many hepatic phase I and II enzymes as well as hepatic efflux transporters. Computer software analysis of human BSEP reveals two musculo-aponeurotic fibrosacroma (Maf) recognition elements (MAREs) from the sequence in the proximal promoter region where Nrf2 may bind. In this study, we assessed whether Nrf2 plays a role in human BSEP expression and if this might be mediated by MAREs. Oltipraz, a potent activator of Nrf2, increased BSEP messenger RNA expression by approximately seven-fold in HepG2 cells and protein by approximately 70% in human hepatocytes. Small interfering RNAs lowered NRF2 expression in HepG2 cells and prevented the up-regulation of BSEP by oltipraz. Human BSEP promoter activity was stimulated by Nrf2 in a dose-dependent manner in luciferase reporter assays. Mutations of the predicted MARE1, but not MARE2, abolished this Nrf2 transcriptional activation. Chromatin immunoprecipitation assays also demonstrated that Nrf2 specifically bound to MARE1, but not MARE2 regions in the BSEP promoter in HepG2 cells. Electrophoretic mobility shift assays further demonstrated direct binding of MARE1 in the BSEP promoter.


CONCLUSIONS
Nrf2 is a positive transcriptional regulator of human BSEP expression. Pharmacological activation of Nrf2 may be beneficial for cholestatic liver injury.
Weerachayaphorn
Jittima
J
From the Department of Internal Medicine, Liver Center, Yale University, School of Medicine, New Haven, CT 06519-8019, USA.
Cai
Shi-Ying
SY
Soroka
Carol J
CJ
Boyer
James L
JL
D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D050977:Maf Transcription Factors / Q000378:metabolism; D008969:Molecular Sequence Data; D051267:NF-E2-Related Factor 2 / Q000235:genetics / Q000378:metabolism*; D011719:Pyrazines / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology; D013871:Thiones; D013876:Thiophenes
C093491:ABCB11 protein, human; D000074020:ATP Binding Cassette Transporter, Subfamily B, Member 11; D018528:ATP-Binding Cassette Transporters; D050977:Maf Transcription Factors; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D011719:Pyrazines; D012333:RNA, Messenger; D018894:Reverse Transcriptase Inhibitors; D013871:Thiones; D013876:Thiophenes; C026209:oltipraz