The mouse tyrosinase gene. Promoter modulation by positive and negative regulatory elements

Tyrosinase, the key enzyme in melanin synthesis, is expressed specifically in pigment-producing cells. Studies with transgenic mice and gene transfer experiments have demonstrated that the 270-base pair 5'-flanking sequence of the mouse tyrosinase gene leads to weak but cell type-specific and developmentally regulated expression. To elucidate the underlying transcriptional control, we focused on the identification of cis-acting elements within this 270-base pair minimal promoter. We also addressed the potential role of promoter elements in the control of cAMP regulation of the tyrosinase gene. Deletion and linker scanning mutagenesis revealed that promoter activity is modulated by two positive elements and one negative element. One of the positive elements includes the M-box, a sequence motif shared with the promoter of two other melanocyte-specific genes, trp-1 and trp-2. Cotransfection experiments provide evidence that a basic helix-loop-helix-zipper protein, encoded at the microphthalmia gene locus, transactivates the tyrosinase promoter, probably by binding to the M-box. Activating cis elements are bound by nuclear factors in vitro and confer increased expression to a reporter gene both in melanoma cells and in fibroblasts. We therefore suggest that the positive promoter elements modulate tyrosinase expression rather than determine cell specificity in vivo, whereas the negative element acts cell type specifically.