IDENTIFICATION AND CHARACTERIZATION OF A NOVEL PHORBOL ESTER-RESPONSIVE DNA-SEQUENCE IN THE 5'-FLANKING REGION OF THE HUMAN DOPAMINE-BETA-HYDROXYLASE GENE

The phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), enhance s transcription of many eukaryotic genes, including that for dopamine P-hydroxylase (DBH). In the present study, we report identification an d characterization of a novel sequence motif residing in the 5'-flanki ng region of the human DBH gene, which mediates transcriptional induct ion by TPA. Deletional analyses indicated the promoter region between -223 and -187 base pairs to be critical. Whereas this region does not contain any putative regulatory motifs with significant sequence homol ogy to the AP-1 motif, extensive deletional and site-directed mutation al analyses indicated that a sequence between -210 and -199 base pairs , B'-ATCCGCCTGTCT-3', may represent a novel TPA-response element (TRE) , In addition, alteration of the YY1-binding site decreased TPA-mediat ed induction of the DBH promoter activity, suggesting that contiguous cis-regulatory element(s) cooperate with this novel sequence motif, Fu rthermore, insertional mutation analyses between the YY1-binding site and the cyclic AMP-responsive element indicated that the stereospecifi city of these motifs is important for intact transcriptional induction by TPA, Taken together, these data suggest that transcriptional up-re gulation of the human DBH gene in response to TPA requires coordinatio n of a novel TRE (human DBH TRE, hDTRE), cyclic AMP-responsive element , and the YY1-binding site.