Inducer-dependent transcriptional activation of the P4501A2 gene in vivo and in isolated hepatocytes.
In vitro nuclear run-on transcription analysis using probes directed against different regions of CYP1A2 revealed that the 70-100-fold induction of CYP1A2 mRNA by polycyclic aromatic compounds is associated with a corresponding increase in the transcriptional activation of this gene in rat liver. Probes from regions of the 1st, 2nd, and 4th introns detected approximately 50-100-fold higher CYP1A2 run-on transcription in liver nuclei from inducer-treated animals than in nuclei from untreated animals. The run-on signals from untreated rats were 3-5-fold above background signals. Additional experiments using single-stranded DNA probes and a probe from a region 5' to the CYP1A2 transcription start site revealed that the inducer-dependent transcripts were colinear with the CYP1A2 mRNA and that they did not result from read through of an initiation event 5' to CYP1A2. Run-on transcription analyses were also carried out with nuclei from isolated hepatocytes using the same series of probes spanning CYP1A2. These analyses indicated that the inducer-dependent accumulation of CYP1A2 mRNA in hepatocytes is associated with at least a 20-fold increase in CYP1A2 transcription. In contrast to liver and hepatocytes, these probes failed to detect run-on transcripts from kidney nuclei, indicating that the lack of CYP1A2 mRNA in this tissue is due to the lack of transcriptional activation of this gene by polycyclic aromatic compounds.
Pasco
D S
DS
Molecular Biology Laboratory, Maharishi International University, Fairfield, Iowa 52557.
Boyum
K W
KW
Elbi
C
C
Siu
C S
CS
Fagan
J B
JB
D000818:Animals; D001571:Benzoflavones / Q000494:pharmacology; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics*; D015342:DNA Probes; D015971:Gene Expression Regulation, Enzymologic; D007438:Introns; D007668:Kidney / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D051381:Rats; D014158:Transcription, Genetic; D019324:beta-Naphthoflavone
D001571:Benzoflavones; D015342:DNA Probes; D000072317:Polychlorinated Dibenzodioxins; D019324:beta-Naphthoflavone; D003577:Cytochrome P-450 Enzyme System