FIG. 1.

ERK and PI3-kinase-Akt signaling are required for cyclin D1 expression in C10 cells. C10 cells were arrested in G₀ by serum deprivation for 72 h and then pretreated with either PD98059 (50 μM), LY-294,002 (25 μM), wortmannin (300 nM), or an equal volume of DMSO as a vehicle control for 30 min. Media containing 10% FBS and inhibitors or DMSO at the same concentrations were added. Total-cell lysates were prepared 1 h (A) or 8 h (B) after serum stimulation and analyzed for total and phosphorylated forms of Akt and ERK, cyclin D1 expression, and phosphorylation of retinoblastoma protein by immunoblotting. CDK4 was used as a loading control.