FIGURE 4.

Cleavage-resistant and truncated HER-2 confer greater protection against apoptosis than WT HER-2. A, immunoblot analysis of MDA-MB-231 pools stably expressing vector, WT, caspase-truncated (amino acids 1-1016, designated Trun), or caspase cleavage-resistant 4× HER-2. B, FACS analysis of cell surface expression of HER-2 proteins in MDA-MB-231 pools. C, MDA-MB-231 pools were treated with 0-5 μg/ml TRAIL for 24 h, and apoptotic nuclei were scored (mean ± S.E., n = 3). ^**, p < 0.01; ^***, p < 0.001 for the indicated comparisons by two-way ANOVA with Bonferroni post-test. D, immunoblot analysis of MDA-MB-231 pools treated with TRAIL (0 or 5 μg/ml) for 24 h. E, MDA-MB-231 pools were serum-starved for 24 h, treated with 100 ng/ml EGF for 10 min, and examined for phospho-Akt (serine 473) and total Akt levels by immunoblotting. F, MDA-MB-231 pools were treated with 5 μg/ml TRAIL for 24 h. Phospho-Akt (serine 473), total Akt levels, phospho-p42/p44 MAPK (Thr²⁰²/Tyr²⁰⁴), and total p42/p44 MAPK were determined by immunoblotting. A positive control lysate for phospho-Akt (serine 473; Cell Signaling) was also immunoblotted.