Novel Inhibitor of Geranylgeranyltransferase-I Figure 6. Stable expression of farnesylated RhoA prevents P61-E7-mediated increases in p21CIP/WAF1 and p27Kip1 protein levels. (A) Effects of GGTI-298 and FTI BMS-225975 on prenylation of wild-type 3xHA-RhoA (RhoA-GG, geranylgeranylated) and 3x-HA-RhoA-F (farnesylated) mutant. Panc-1 cells stably expressing 3xHA-RhoA-GG (right panels) or 3xHA-RhoA-F (left panels) were treated DMSO, BMS-225975, or GGTI-298 for 48 h. Whole cell lysates were collected and analyzed for RhoA, Rap1, and HDJ2 processing by Western blotting. (B) Expression levels of 3xHA-RhoA-F and 3xHA-RhoA-GG in Panc-1 stable cell lines used in this experiment were analyzed by immunoblotting using antibodies against the HA tag or actin (loading control). (C and D) Panc-1 cells and Panc-1 cells stably expressing 3xHA-RhoA-GG or 3xHA-RhoA-F were treated with DMSO or P61-E7 for 48 h. Whole cell lysates were collected and analyzed for protein levels of p21CIP/WAF1 (C, top panel), p27Kip1 (D, top panel), or Actin (C and D, bottom panels). Data shown are representative of three independent experiments. The p21CIP1/WAF1 (C, top panel), p27Kip1 (D, top panel), and Actin (C and D, bottom panels) bands were quantified using ImageJ. Protein Levels of p21CIP1/WAF1 and p27Kip1 are represented as relative level compared to the DMSO control. doi:10.1371/journal.pone.0026135.g006 PLoS ONE | www.plosone.org 9 October 2011 | Volume 6 | Issue 10 | e26135