The pregnane X receptor binds to response elements in a genomic context-dependent manner, and PXR activator rifampicin selectively alters the binding among target genes.
The pregnane X receptor (PXR) is a key regulator of genes encoding several major types of cytochrome P450 enzymes and transporters (e.g., multidrug resistance-1, MDR1); therefore, PXR contributes significantly to drug-drug interactions. PXR binds to response elements and confers transactivation. Several target genes such as CYP3A4 and 3A7 contain two PXR elements (distant and proximal) that are separated by more than 7000 nucleotides in the genome. Disruption of the distant element causes a 73% decrease of the reporter activity, whereas inactivation of the proximal element decreases by only 53%. This study was undertaken to test the hypothesis that PXR differentially binds to the elements with the distant enhancer being bound to a higher extent. To test this hypothesis, a stable transfected line (hPXR-HRE) was prepared to constitutively express human PXR and harbor a chromatinized CYP3A4-ER6 reporter. This line responded to rifampicin and dexamethasone similarly as hepatocytes based on the relative potency and activation kinetics. Contrary to the hypothesis, chromatin immunoprecipitation experiments showed that the genomic fragment harboring the proximal element was preferably precipitated over the fragment containing the distant element in the CYP3A4 gene, but the opposite was true with the CYP3A7 gene. In addition, the promoters from the MDR1 and CYP2B6 genes were abundantly present in the PXR immunocomplexes from the vehicle-treated cells. However, such abundant interactions were markedly diminished when cells were treated with PXR activator rifampicin. These findings suggest that PXR binding is dependent on the genomic context and PXR activators modulate such bindings.
Song
Xiulong
X
Department of Biomedical Sciences, University of Rhode Island, 41 Lower College Road, Kingston, RI 02881, USA.
Xie
Mingxing
M
Zhang
He
H
Li
Yuxin
Y
Sachdeva
Karuna
K
Yan
Bingfang
B
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000378:metabolism; D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D065702:Cytochrome P-450 CYP2B6; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D003907:Dexamethasone / Q000494:pharmacology; D005557:Formaldehyde / Q000737:chemistry; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D010089:Oxidoreductases, N-Demethylating / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011233:Precipitin Tests; D000077297:Pregnane X Receptor; D011401:Promoter Regions, Genetic / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D011987:Receptors, Steroid / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics*; D012293:Rifampin / Q000494:pharmacology*; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D003432:Cross-Linking Reagents; D017931:DNA Primers; D018076:DNA, Complementary; D000077297:Pregnane X Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D005557:Formaldehyde; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; C104464:CYP3A protein, human; D065702:Cytochrome P-450 CYP2B6; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D010089:Oxidoreductases, N-Demethylating; D012293:Rifampin