Changes in GTP-dependent fusion and ras-related proteins in membranes from rat hepatocellular carcinomas

The fusion capacity of rough endoplasmic reticulum membranes isolated from dissected liver tumor nodules of aflatoxin-treated rats was determined by cell free assay to be greater than that of homologous membranes from control liver. In a first attempt to understand the reason for this difference we compared the content of ras-related proteins in rough microsomal fractions and other cell fractions of both dissected tumor nodules and control liver. Using [alpha-32P]GTP blot overlay and densitometric analysis, homogenate, Golgi and rough endoplasmic reticulum fractions from dissected tumor nodules were observed to contain increased amounts of [alpha-32P]GTP binding to ras-related proteins when compared to homologous control fractions. Western blot analysis indicated that ras content was also increased in the tumor fractions. [alpha-32P]GTP-blot overlay using double-dimensional SDS-polyacrylamide gel electrophoresis confirmed quantitative differences in the amount of [alpha-32P]GTP binding to ras-related proteins between fractions from tumor and control tissues and indicated a surprising number of such proteins in each fraction. The data suggest that the changes in ras-related proteins could, in part, account for the enhanced GTP-dependent fusion capacity observed for the tumor-derived membranes.