Fig. 3.

Effect of androgen treatment on Skp2 mRNA expression. A: LNCaP cells were treated with different doses of R1881 as indicated for 48 h. Total RNA (15 μg) was applied for Northern blot analysis and hybridized with Skp2 and GAPDH cDNAs as probes. B: Time-course study on the androgenic effect on Skp2 mRNA expression. LNCaP Cells were treated with 5 nM of R1881 for varying lengths of time, from 6–48 h, and Skp2 mRNA expression was examined by Northern blot analysis. C: Skp2 repression by androgens is blocked by the protein synthesis inhibition. LNCaP cells were pretreated with CHX (20 μg/ml) for 30 min and then treated with or without 5 nM of R1881. At the time points indicated, cells were harvested and RNAs were isolated and subject to Northern blot analysis. D: Effect of ActD on Skp2 repression by androgens. LNCaP cells were pretreated with 4 μM ActD for 30 min and then treated with or without 5 nM of R1881. At the time points indicated, cells were harvested and RNAs were isolated and subject to Northern blot analysis. GAPDH cDNA was used as a control for the normalization of RNA loaded in these experiments.