In vitro and in vivo regulation of liver epithelial cells carrying a metallothionein-rasT24 fusion gene.

We inserted a zinc-responsive metallothionein-rasT24 fusion gene in both orientations into a retroviral vector (SVX) and infected Fisher rat liver epithelial cells. Only the construction in which the viral long terminal repeat and the metallothionein promoters were in opposite orientations resulted in cell lines with biologically altered behavior. Two lines (MTR-1 and MTR-6) showed altered morphology in vitro when ZnSO4 was added to the medium. These cell lines grew in soft agarose in a dose-dependent manner. Immunoprecipitation experiments revealed dose-dependent increases in the rate of synthesis of the mutant p21Ha-ras protein in these lines in response to ZnSO4. Both lines produced poorly differentiated metastatic adenocarcinomas when injected subcutaneously in Fisher rats, and tumors derived from MTR-6 cells grew more rapidly in animals on a zinc-supplemented diet than on a zinc-deficient diet. Uninfected liver epithelial cells showed no change in morphology in vitro after ZnSO4 addition and did not grow in soft agarose or after subcutaneous transplantation during the 14-wk experimental period. These results indicate that altered levels of expression of a single gene (rasT24) can have profound effect on the biologic behavior of tumor cells both in vitro and in vivo.