Induction of Cytochrome P-450 1A2 by Oxidized Tryptophan in Hepa lclc7 Cells1
Abstract
Recent studies from this laboratory have demonstrated thatl-tryptophan, after oxidation either by UV-irradiation or ozone, induces aryl hydrocarbon receptor (AhR) activation and binding of the liganded AhR complex to its specific DNA recognition site, thereby initiating transcription of the cytochrome P-450 1a1 (Cyp1a1) gene with concomitant increase of CYP1A1 protein and 7-ethoxyresorufin O-deethylase activity in wild-type mouse hepatoma cells, Hepa lclc7 (Hepa-1), in culture. Temporary inhibition of protein synthesis by cycloheximide resulted in superinduction of oxidized tryptophan-inducible CYP1A1 mRNA, protein, and 7-ethoxyresorufin O-deethylase activity in Hepa-1 cells. In the present communication, the results obtained by immunoblot analyses with monoclonal CYP1A1/1A2 antibody (NIH 1-7-1) demonstrate that both UV- or ozone-oxidized tryptophan also induce CYP1A2 protein in Hepa-1 cells. CYP1A2 mRNA, detected by reverse transcription-polymerase chain reaction, was markedly induced in the UV- or ozone-oxidized tryptophan-treated cells. Temporary inhibition of protein synthesis by cycloheximide further induced oxidized tryptophan-inducible CYP1A2 mRNA as well as the protein in Hepa-1 cells. This is the first report demonstrating the induction of CYP1A2 mRNA and protein in Hepa-1 cells.
Footnotes
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Send reprint requests to: Ram K. Sindhu, Ph.D., Department of Pathology, College of Medicine, University of California-Irvine, Irvine, CA 92697-4800. E-mail: rksindhu{at}uci.edu
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↵1 This work was supported by Grant HL-50450 from the National Heart, Lung and Blood Institute of the National Institutes of Health, U.S. Public Health Service.
- Abbreviations:
- CYP
- cytochrome P-450
- PAH
- polycyclic aromatic hydrocarbon
- Ah
- aryl hydrocarbon
- OT
- ozone-oxidized tryptophan
- UVT
- UV-oxidized tryptophan
- AhR
- aryl hydrocarbon receptor
- EROD
- 7-ethoxyresorufin O-deethylase
- TCDD
- 2,3,7,8-tetrachlorodibenzo-p-dioxin
- PAGE
- polyacrylamide gel electrophoresis
- RT-PCR
- reverse transcription-polymerase chain reaction
- bp
- base pair
- MC
- 3-methylcholanthrene
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- Received July 7, 1999.
- Accepted November 18, 1999.
- The American Society for Pharmacology and Experimental Therapeutics
