SUPPRESSION OF TYROSINASE GENE-EXPRESSION BY BROMODEOXYURIDINE IN SYRIAN-HAMSTER MELANOMA-CELLS IS NOT DUE TO ITS INCORPORATION INTO UPSTREAM OR CODING SEQUENCES OF THE TYROSINASE GENE

5-Bromodeoxyuridine (BrdU), a thymidine analog, suppresses melanogenes is in Syrian hamster melanoma cells. Tyrosinase, which is the key enzy me for the synthesis of melanin, is suppressed by exposure to BrdU, an d the drop in enzyme activity is correlated with a drop in tyrosinase mRNA level. In order to investigate whether suppression of tyrosinase mRNA by BrdU is due to BrdU substitution into coding sequences or upst ream sequences of the tyrosinase gene, we carried out stable and trans ient transfection assays with constructs containing either the human t yrosinase cDNA sequence under the control of a nontyrosinase promoter or a chloramphenicol acetyltransferase (CAT) reporter gene under the c ontrol of 5' flanking sequences of the mouse tyrosinase gene. When the plasmid containing the tyrosinase cDNA was stably transfected into mo use fibroblasts, tyrosinase activity in the transfectants was not supp ressed by BrdU. Since BrdU would be incorporated into the tyrosinase c DNA integrated in these transfectants, the results suggest that BrdU s uppression of tyrosinase gene expression is not due to its incorporati on into coding sequences of the tyrosinase gene. When plasmids with ty rosinase regulatory sequences were transfected into melanoma cells for transient expression assays, CAT gene expression was suppressed by Br dU. Because the CAT plasmids do not contain a mammalian origin of repl ication and should not replicate under the conditions of transient tra nsfection, BrdU would not be incorporated into the DNA of those plasmi ds. Therefore, these results suggest that the suppression of tyrosinas e gene expression by BrdU also is not due to the incorporation of BrdU into upstream sequences of the tyrosinase gene.