The THCRE2 site in the rat tyrosine hydroxylase gene promoter is responsive to phorbol ester

Recently, our laboratory has identified a tyrosine hydroxylase (TH) gene promoter element, THCRE2 (located at -97 to -90), that is required for maximal response to cyclic AMP. In this study we test whether the THCRE2 is responsive to phorbol ester. Rat PC12 cells were transfected with a reporter gene construct, TH(-272/+27)-CAT, which is driven by the first 272 bp of the rat TH gene 5' flanking region. Treatment of transfected cells with 0.1 microM 12-O-tetradecanoylphorbol 13-acetate (TPA) elicited a 5-6-fold increase in TH gene proximal promoter activity. Mutagenesis of the THCRE2 sequence diminished TPA-responsiveness of the TH gene promoter by approximately 50%. Minimal promoter constructs driven by a single copy of the region of the TH gene that encodes the THCRE2 (from -117 to -59) were also responsive to TPA. Our results suggest that the THCRE2 is a phorbol ester-responsive element, as well as a cyclic AMP-responsive element.