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 aflatoxintreated 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 [α-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 [α-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. [α-32P]GTP-blot overlay using double-dimensional SDS-polyacrylamide gel electrophoresis confirmed quantitative differences in the amount of [α-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.