Figure 4

DI1 Peptide Inhibits Raf Dimerization and Signaling (A) B-Raf structure is shown with the amino acids of DI1 highlighted in red. (B-D) Cos cells were transfected to express GFP or GFP-DI1 as indicated. (B) Endogenous Raf proteins were immunoprecipitated and probed for the binding of GFP-DI1. (C,D) Endogenous C-Raf proteins were immunoprecipitated from serum-starved or EGF-treated cells and probed for the binding of GFPDI1 or B-Raf. Lysates were probed for pMEK levels in (C). (E) Cos cells stably expressing the indicated B-Raf or C-Raf mutants were transfected to express GFP or GFP-DI1. Lysates were probed for levels of pMEK. (F-H) Cancer cell lines were treated with the indicated concentrations of TAT-DI1 or TAT-Scram. (F) Cell viability was determined by cell counting. Error bars represent the standard deviation of at least three independent experiments. (G) Lysates were probed for pMEK levels. (H) Lysates were examined for PARP cleavage.