Fig. 3. Akt underlies HER3 signal buffering and protection against lapatinib.
A) SkBr3/ myrAktΔER and SkBr3/ myr*AktΔER cells were treated with 4-hydroxy tamoxifen (4HT) to induce the activation of the engineered Akt construct and lapatinib at 0.2 or 5 μM for the indicated times. Cell lysates were immunoblotted as indicated. Arrows, endogenous cellular Akt; starred arrows, the larger engineered Akt-ΔER fusion proteins. B) SkBr3/ myr*AktΔER and SkBr3/ myrAktΔER cells were treated with 4HT to induce the activation of the engineered Akt construct and 200nM lapatinib for the indicated times (in hours). Cellular RNA was extracted and the relative expression of HER3 mRNA was assayed by real-time quantitative PCR with HER3 specific primers. GAPDH was used as normalization control. The results shown are the average of triplicates, and error bars represent SEM. Data are relative expression with values from untreated cells set as 1. * indicates significantly different from the adjacent induced data point (p=0.005), and significantly different from the 0 timepoint (p=0.01).