AN ACTIVATED MUTANT OF R-RAS INHIBITS CELL-DEATH CAUSED BY CYTOKINE DEPRIVATION IN BAF3 CELLS IN THE PRESENCE OF IGF-I

R-Ras belongs to a family of low molecular weight GTP-binding proteins and exhibits 55% amino acid identity to H-Ras. It has been demonstrat ed that H-Ras inhibits cell death caused by interleukin-3 (IL-3) withd rawal in BaF3 cells (Kinoshita rt al. (1995b); Terada et al. (1995)). In the present study, we examined whether R-Ras also rescues BaF3 cell s from the factor-deprived cell death, To do this, several BaF3 transf ectants were established, in which expression of wild-type as well as mutant R-Ras was regulated by an inducible promoter. Using these trans fectants, we found that expression of an activated R-Ras mutant, R-Ras (Q87L), suppressed the death of IL-3-deprived BaF3 cells. On the othe r hand, expression of the wild-type and the dominant-negative mutant o f R-Ras showed no inhibitory effect on cell death, indicating that R-R as.GTP abrogated cell death caused by deprivation of IL-3, Furthermore , it was found that IGF-I in serum was required for the anti-apoptotic activity of R-Ras, Suppression of cell death by R-Ras(Q87L) was inhib ited by wortmannin, LY294002 (phosphatidylinositol 3-kinase (PI3K) inh ibitors), or PD98059 (inhibitor for MEK, a specific activator of mitog en-activated protein kinase (MAPK)), In addition, we have shown that, in HEK293 cells, R-Ras and IGF-I could activate MAPK synergistically. Also, PI3K activity was co-immunoprecipitated with an activated mutant of R-Ras. These results suggest that R-Ras in collaboration with IGF- I suppressed apoptotic cell death of BaF3 caused by IL-3 deprivation, presumably by modulating the activitites of MAPK and PI3K.