Figure 3.

(A) Functional characterization of the mutant R379C P63 protein using the yeast-based transactivation assay. ΔN-P63α or TA-P63α isoforms of the wt or mutant R379C P63 protein were expressed under the GAL1-10 inducible promoter (8 h at 0.128% galactose) in two isogenic yeast reporter strains (yLFM-P21-5′; yLFM-PERP) having the luciferase reporter gene under the control of P53 responsive element present in the promoter region of the P21 and the PERP gene. Presented in the graph is the fold-induction over the empty expression vector (pRS314) of mean luciferase activities normalized to optical density (OD) at 600 nm. Average and standard errors of three biological replicates are plotted. (B) Western blot analysis of different isoforms (TA and ΔN) of P63α proteins revealed that wt and R379C are expressed at similar level in yeast cells. A specific antibody directed against P63 was used to quantify the P63 isoforms. PGK1 was used as a loading control. The P63/PGK1 ratio is similar within a specific situation (e.g., strain and isoform) for the wt and R379C protein. PGK1, phosphoglycerate kinase 1.