FIG. 7.

Effect of cycloheximide treatment of Ras activation. (A) Quiescent Rb^−/− and Rb^+/+ 3T3 cells were restimulated with serum. At the indicated times, the activation state of Ras was measured. Cycloheximide (CHX) was added (25 μg/ml) to cultures 30 min before the indicated time point when levels of GTP-bound Ras were determined. Also shown are the levels of GTP-bound Ras determined 10 min after the addition of EGF alone (lane 17) or EGF with cycloheximide (lane 18) at time zero. In lane 11, cycloheximide was added at 3.5 h after serum stimulation and EGF was added 20 min later. Analysis of the activation state of Ras was performed at 4 h. The results shown are representative of at least 10 independent experiments. (B) Same as panel A, except primary MEFs were used, and the cycloheximide treatments at 30 min and 1 h were not included.