Melanosomal and lysosomal alterations in murine melanocytes following transfection with the v-rasHa oncogene

Abstract

Melanomas exhibiting mutated ras genes are frequently invasive and amelanotic. Transfecting melanocytes with ras oncogenes causes transformation and a loss of visible pigmentation. We analyzed murine melanocytes rendered amelanotic by transfection with the v‐ras^Ha oncogene. Consistent with previous reports, tyrosinase and tyrosinase‐related protein‐1 (TRP‐1) were not expressed by transformed cells. In addition, lack of expression of TRP‐2 and the product of the silver locus was documented. Levels of melanosomal matrix antigens, the pink‐eyed dilution locus protein and lysosome‐associated membrane protein‐I were markedly reduced. Residual matrix antigens were localized by immunofluorescence to large vacuoles distributed peri‐nuclearly in transfected cells. Electron microscopy demonstrated the absence of typical melanosomes and the presence of large vacuolar structures, also in a peri‐nuclear distribution. Although levels of lysosomal hydrolases, such as beta‐glucuronidase and cathepsin D, were diminished, marked elevations were observed in the expression of cathepsins B and L, 2 thiol proteases implicated in the acquisition of invasiveness. Our data demonstrate that transfection of melanocytes with v‐ras^Ha is sufficient to disrupt the biogenesis of melanosomes and to up‐regulate thiol protease synthesis, providing insights into the amelanotic and invasive nature of melanomas exhibiting mutations in ras genes.