FIG. 1.

Forskolin treatment increases tyrosinase, TRP1, and TRP2 gene expression in B16 mouse melanoma cells. (A) Thirty micrograms of proteins from control cells and cells treated for 24 or 48 h with 20 μM forskolin (FSK) was subjected to Western blot analysis using antibodies PEP7 for tyrosinase detection, B8G3 for TRP1, and PEP8 for TRP2 (upper panel). The detection of the 44-kDa ERK1 protein ensured even loading of lanes (lower panel). (B) Ten micrograms of total RNA from control B16 cells or cells treated for 48 h with 20 μM forskolin was reverse transcribed. The resulting cDNAs were subjected to 28-cycle PCRs using specific primers that gave amplified products of 1,191 bp for tyrosinase (TYRO), 784 bp for TRP1, and 518 bp for TRP2. A control of PCR amplification with specific primers of GAPDH transcript showed a 983-bp fragment. PCR products were electrophoresed on a 1% agarose gel and stained with ethidium bromide before UV light visualization. DNA molecular weight markers (lane M), from bottom to top: 510, 1,018, 1,636, 2,036, 3,054, and 4,072 bp.