In MCF-7 cells treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 12-O-tetradecanoylphorbol-13-acetate (TPA) causes a time- and concentration-dependent modulation of TCDD-induced CYP1A1 gene expression. Treatment of MCF-7 cells with 1 nM TCDD for 24 h resulted in the induction of ethoxyresorufin O-deethylase (EROD) activity and CYP1A1 mRNA levels. In cells treated with TCDD for 24 h and 100 ng/ml TPA for 26 and 30 h, the TCDD-induced CYP1A1 gene expression was detected. For example, TCDD-induced EROD activity decreased from 122 pmol/min/mg to 25.5 pmol/min/mg after treatment of MCF-7 cells with TPA for 26 h and this was also paralleled by a 44% decrease in CYP1A1 mRNA levels. There was also a decrease in nuclear Ah receptor levels and the binding of nuclear extracts to a 32P-labeled dioxin responsive element (DRE) in a gel mobility shift assay. In parallel studies which measured EROD activities, similar TCDD/TPA interactions were observed in wild-type Hepa 1c1c7 cells, whereas no interactive effects were observed in T47-D human breast cancer cells. In MCF-7 cells treated with TPA for 36 or 48 h, the TCDD-induced EROD activity and CYP1A1 mRNA levels were restored and in cells exposed to TPA for 72 or 96 h superinducibility of CYP1A1 gene expression was observed; there was a 2.8- and 2.2-fold increase in EROD activity and CYP1A1 mRNA levels, respectively, compared to MCF-7 cells treated with TCDD alone. The biphasic temporal effects of TPA on TCDD-induced CYP1A1 gene expression in MCF-7 cells were paralleled by comparable changes in nuclear Ah receptor levels and binding to a synthetic DRE. In contrast, prolonged exposure of the wild-type Hepa 1c1c7 or T47-D cells to both TCDD plus TPA gave results similar to those observed after 24 h. These data show that the effects of TPA on TCDD-induced expression of CYP1A1 are cell-specific and suggest that the proposed protein kinase C (PKC)-dependent activation of the nuclear Ah receptor complex may not be required in MCF-7 cells since TPA downregulates PKC activity within 11 h and this inactivation persists for at least 96 h.