Induction of cytochrome (CYP) P4501A2 by such polycyclic aromatic hydrocarbons as 3-methylcholanthrene (3MC) can lead to the bioactivation of carcinogenic aromatic amines and heterocyclic amines. A 3MC response element was recently identified approximately 2.2 kb upstream of the transcription start site of the human CYP1A2 gene. Sequence analysis of this enhancer identified, in addition to a binding site for the aryl hydrocarbon receptor, two other sequences, referred to as 5'AP1 and 3'AP1, each with complete homology to the phorbol 12-O-tetradecanoate 13-acetate (TPA) response element consensus sequence. Nuclear extracts from TPA-treated HepG2 cells protected both the 5'AP1 and 3'AP1 sequences against digestion with DNase I. Gel mobility shift and supershift assays revealed that TPA treatment of HepG2 results in increased binding activity of the AP-1 proteins, c-Jun, JunD, and c-Fos, to both sites. We transiently expressed, in HepG2, either a fragment containing both the 5'AP1 and 3'AP1 sites (-2.3pT81Luc) or only the 3'AP1 site (-2.2pT81Luc) cloned into a plasmid containing the luciferase gene under transcriptional control of the thymidine kinase promoter. TPA treatment of cells transfected with -2.3pT81Luc resulted in an approximately threefold induction of luciferase activity over untreated control cells, while the -2.2pT81Luc construction containing only the 3'AP1 site displayed an approximately sixfold induction. These studies suggest that the human CYP1A2 gene may be regulated by tumor promoters in addition to polycyclic aromatic hydrocarbons.