In vivo activation of the human CYP3A4 promoter in mouse liver and regulation by pregnane X receptors.
Human cytochrome P450 3A4 (CYP3A4) is responsible for the metabolism of numerous xenobiotics in the human liver. We have examined the activation of the human CYP3A4 promoter in mouse liver by using in vivo bioluminescent imaging (BLI). Transcription of the CYP3A4 promoter occurs as a result of a ligand binding to a nuclear orphan receptor, pregnane X receptor (PXR), followed by dimerization with another nuclear receptor, retinoid X receptor (RXR). Since this heterodimer then binds to xenobiotic response elements to activate transcription of CYP3A4, we examined a 13kb promoter region of CYP3A4 for responsiveness to dexamethasone and rifampicin. A reporter vector CYP3A4-luc was constructed consisting of the CYP3A4 promoter driving the firefly luciferase gene. This DNA was injected into the tail veins of mice, and reporter gene expression was monitored in the liver region using BLI. Treatment of transfected mice with dexamethasone resulted in a 188-fold induction of luciferase, whereas treatment with rifampicin resulted in a 68-fold induction. Co-injection with a human PXR expression vector resulted in a dramatic increase in rifampicin-induced activity and a smaller increase of dexamethasone-induced activity. Co-injection of an antisense murine PXR construct with the CYP3A4-luc reduced both the dexamethasone- and rifampicin-induced responses, thus demonstrating that the murine PXR receptor can participate in the regulation of the human CYP3A4 promoter in mice. The approach described here will be of general use in studying the regulation of nuclear receptors in vivo.
Zhang
Weisheng
W
Xenogen Corporation, 860 Atlantic Avenue, Alameda, CA 94501, USA. wzhang@xenogen.com
Purchio
Anthony
A
Chen
Kevin
K
Burns
Stacy M
SM
Contag
Christopher H
CH
Contag
Pamela R
PR
D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics*; D003907:Dexamethasone / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D017930:Genes, Reporter; D006801:Humans; D008099:Liver / Q000502:physiology*; D008156:Luciferases / Q000096:biosynthesis; D051379:Mice; D000077297:Pregnane X Receptor; D011401:Promoter Regions, Genetic; D018160:Receptors, Cytoplasmic and Nuclear / Q000502:physiology*; D011987:Receptors, Steroid / Q000502:physiology*; D012293:Rifampin / Q000494:pharmacology
D000893:Anti-Inflammatory Agents; D004791:Enzyme Inhibitors; D000077297:Pregnane X Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; D008156:Luciferases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D012293:Rifampin