Transcriptional activation and posttranscriptional modification of Cyp1a1 by arsenite, cadmium, and chromium.

Heavy metals alter the carcinogenicity of AhR ligands by modulating the induction of the Cyp1a1 enzyme, but the mechanism(s) remain unresolved. In this study, the effect of the heavy metals, As(3+), Cd(2+), and Cr(6+), on the transcriptional and posttranscriptional regulation of the Cyp1a1 gene was investigated in Hepa 1c1c7 cells. A time-dependent study showed that all metals significantly induced the basal Cyp1a1 mRNA, but only As(3+) and Cd(2+) further potentiated the inducible Cyp1a1 mRNA level. Alternately, Cr(6+) inhibited the TCDD-mediated induction of Cyp1a1 mRNA. As(3+) potentiated the induction of Cyp1a1 mRNA by TCDD only at 3h and 6h after treatment. Cd(2+) on the other hand further potentiated the induction up to 24h after treatment. The metal-mediated induction of Cyp1a1 mRNA was further potentiated by the protein synthesis inhibitor, cycloheximide and the 26S proteasome inhibitor, MG-132, but completely inhibited by the RNA transcription inhibitor, actinomycin-D, implying a transcriptional regulation of the Cyp1a1 gene by the heavy metals. Not surprisingly, Cd(2+) and Cr(6+) activated the DNA-binding capacity of the AhR for the xenobiotic responsive element, as measured by the electrophoretic-mobility shift assay while all three metals induced AhR-dependent luciferase reporter gene expression in transiently transfected Hepa 1c1c7 cells. On the other hand, only As(3+) increased the Cyp1a1 mRNA half-life while Cd(2+) and Cr(6+) increased the Cyp1a1 protein half-life, suggesting the involvement of posttranslational modifications. A significant decrease in TCDD-mediated induction of Cyp1a1 activity was associated with an increase in HO-1 mRNA levels and a concomitant decrease in cellular heme content after all metal treatments. We clearly demonstrated that As(3+), Cd(2+), and Cr(6+) increase Cyp1a1 mRNA levels at the transcriptional and posttranscriptional levels while decreasing Cyp1a1 activity at the posttranslational level.