MUTATION OF THE ENDOGENOUS P53 GENE IN CELL TRANSFORMED BY HPV-16 E7 AND EJ C-RAS CONFERS A GROWTH ADVANTAGE INVOLVING AN AUTOCRINE MECHANISM

Rat embryo fibroblasts transformed with the HPV-16 E7 gene and the act ivated c-H-ras gene fall into two distinct phenotypic classes. At high cell density, clones of one class form colonies in methylcellulose su pplemented with low serum; at low cell density, these cells display re sponsiveness to mitogenic factors present in serum-free conditioned me dium from rat embryo fibroblasts. In contrast, clones of the second cl ass exhibit an absolute dependency on growth factors present in serum at all cell densities in the methylcellulose colony assay and fail to respond to conditioned medium. We find that the status of the endogeno us p53 gene is tightly correlated with these two classes of clones. Cl ones of the first class contain missense mutations in the p53 gene and have lost the wild-type allele. Clones of the second class express wi ld-type p53 protein. The importance of mutant p53 expression in reduci ng the growth factor dependency of transformed clones was confirmed in a separate series of experiments in which rat embryo fibroblasts were transformed with three genes, E7 + ras + mutant p53. The growth behav iour of these triply transfected clones was similar to that of the E7 + ras clones expressing endogenous mutant p53. We demonstrate that the enhanced proliferation of E7 + ras clones expressing mutant p53 prote in involves an autocrine mechanism.