The phorbol ester, 12-0-tetradecanoylphorbol-13-acetate (TPA), was used as a reversible inhibitor of melanogenesis. Chick-melanocyte cultures of the black genotype, EIE, were grown in conditioned medium plus TPA. After growth in TPA and after its removal, the cells were pulse labeled with 3H-leucine. The membrane fraction, which included all tyrosinase activity as well as both mature and immature melanosomes, was solubilized with Triton X-100. The proteins were separated using two-dimensional electrophoresis and visualized by fluorography. One defined melanogenic protein, tyrosinase, was isolated, and its location was determined in the two-dimensional protein pattern. The protein patterns for both the TPA-inhibited cells and the cells in which the TPA effects were reversed after removal were compared. in addition to tyrosinase, at least nine TPA-sensitive proteins were found. These were designated as being putative melanogenic proteins which, along with tyrosinase, may be responsible for melanin-granule synthesis.
|Original language||English (US)|
|Number of pages||7|
|State||Published - 1985|
Bibliographical noteFunding Information:
Acknowledgements. This work was supported by a grant (GM 18969) from the National Institutes of Health and an NIH Biomedical Research Support Grant (RR 07055). The authors wish to thank Tom Bargar, Dana Scheele, and Roxanne Martin for their assistance in the preparation of this manuscript.