RAS SIGNALING IS REQUIRED FOR INACTIVATION OF THE TUMOR-SUPPRESSOR PRB CELL-CYCLE CONTROL PROTEIN

Ras proteins act as molecular switches, responding to signals by enter ing the active GTP-bound, rather than the inactive GDP-bound, state, T he inhibition of normal Ras proteins by microinjection of neutralizing antibody or expression of dominant-negative mutants has shown that Ra s signalling is required for growth factors to stimulate DNA synthesis [1,2], but the link between Ras and the cell cycle machinery is not c lear, Regulation of the phosphorylation state of the retinoblastoma pr otein (pRb), the product of the tumour suppressor gene Rb, is a key ev ent in the progression of cells from G1 phase into S phase, In growth- arrested or early G1 cells, pRb is hypophosphorylated and binds to tra nscription factors of the E2F family [3]. These pRb-E2F complexes act to suppress gene transcription required for entry into DNA synthesis e ither by preventing E2F from stimulating transcription or by actively repressing transcription [4]. During G1, cyclin-dependent kinases (CDK s) become activated and phosphorylate pRb at multiple sites, leading t o the dissolution of pRb-E2F complexes and gene transcription [5]. Her e, we have tested the hypothesis that Ras signalling is required for t he inactivation of pRb, A neutralizing antibody directed against p21Ra s was microinjected into cells derived from mutant mouse embryos that lack Rb or CDK inhibitors (CDKIs), Cells without pRb or the pie CDKI w ere more resistant to the inhibitory effects of the anti-Ras antibody, DNA synthesis in some tumour cell lines was completely resistant to t he anti-Ras injection, indicating that p21Ras is required for pRb inac tivation but also has other functions in cell cycle progression.