Identification of RKIP Ligands Figure 7. Compound 48 reduces EGF-induced MAPK activity dose dependently. HeLa cells (0.56106 cells/ml) expressing wild-type (A) or depleted RKIP (B) were serum starved overnight and then pre-treated for 30 min with increasing doses of DMSO (control) or compound 48. EGF (10 ng/ml) was added in the final 5 minutes of incubation. Whole cell lysates (30 mg) were separated on SDS-PAGE gels (12.5%), transferred to nitrocellulose and analysed by immunoblotting with anti-phospho ERK and anti-total ERK antibodies. ERK phosphorylation was assessed by normalizing phospho-ERK levels to total ERK levels as depicted in line graphs. The results shown are mean 6 range of two independent experiments. doi:10.1371/journal.pone.0010479.g007 PLoS ONE | www.plosone.org 11 May 2010 | Volume 5 | Issue 5 | e10479