FIG. 4.
The effect of activated MEK1 on CDK4 activity and cyclin D1 levels in cells transiently expressing RasN17. (A) NIH 3T3 cells expressing ectopic cyclin D1 or MEK-EE or cells infected with control virus were transiently transfected with control plasmid (pMT-ΔBam), pMT-RasN17, or pcDNA3-p16. The transfected population, cultured in DMEM–5% BCS, was magnetically sorted, and CDK4 kinase assays were performed as described for Fig. 3B. Kinase activity for vector control transfections was set to 100% as indicated. The bar graph represents the relative kinase activity with the mean standard error from at least three independent experiments. A representative autoradiogram is shown. GST, glutathione S-transferase. (B) Same as panel A except that cell lysates were used to assess the relative amounts of cyclin D1 protein present as a function of RasN17 expression by Western blot analysis.