ACTIVATION OF P21RAS IS NOT SUFFICIENT TO ENSURE A COMPLETE G(1) PHASE OF THE CELL-DIVISION CYCLE IN MOUSE FIBROBLASTS

In the mouse BP-A31 fibroblasts, mRNAs coding the three isoforms (Ha, Ki, N) of ras are expressed, and there are no activating mutations in the codons 12, 13 or 61. We have produced a subline (Ras2) expressing an oestrogen-inducible v-Ha-ras gene. The contribution of v-Ha-ras to the overall p21ras-GTP content was evaluated by metabolic labelling wi th P-32. Surprisingly, p21ras-GTP complexes were predominant in the se rum-deprived BP-A31 cells as well as in the Ras2 cells. The excess of p21ras-GTP was not due to the lack of the GTPase activating protein. I n transient transfection experiments, the serum response element (SRE) -directed CAT was expressed in serum-deprived BP-A31 cells, and insuli n caused a further two- to threefold increase in CAT activity. A domin ant negative ras mutant (Ha-Ras Asn-17) cancelled both the basal and i nsulin-induced CAT expression in the BP-A31 but not in the Ras2 cells. Expression of v-Ha-ras in Ras2 cells did not relax their growth facto r-dependence and oestradiol had only a minor mitogenic effect. We conc lude that p21ras activation does not ensure a complete cell division c ycle in these cells, and does not entirely account for the transductio n of the mitogenic signal initiated by insulin.