Melanosome formation in cultured amelanotic melanophores of Rana brevipoda by a frog tyrosinase cDNA Transfection.

A cDNA encoding tyrosinase of Rana nigromaculata was introduced into cultured, tyrosinase-negative amelanotic melanophores of R. brevipoda by a calcium phosphate precipitation method. Within a few days following transfection, dark pigmentation became visible in a small number of cells. Light microscopic observation revealed that the morphology of these transformed cells was comparable to that of normal melanophores in culture, and their proliferative activity was lower than that of amelanotic cells. Ultrastructural examination verified that amelanotic melanophores possessed a relatively small number of premelanosomes while the transformants contained numerous melanosomes at various stages of pigment deposition. The result indicated that tyrosinase cDNA of R. nigromaculata was expressed in amelanotic melanophores of R. brevipoda including the maturation of premelanosomes. It was also suggested that the expression of transfected tyrosinase cDNA had promoted differentiation of the amelanotic cells into fully developed melanophores.