Fig. 4

Proteasomal activity is required for stress-induced degradation of EGFR and cell death, but not endocytosis of EGFR. HeLa cells were cultured and serum-starved the same as described in Fig. 1. The proteasome inhibitor MG-132 (10 μM) was added to the cells 30 min prior to the treatment of EGF, anisomycin, UV irradiation or 0.7 M NaCl. A The cells were lysed and EGFR was immunoprecipitated and detected by immunoblotting with anti-EGFR(1005) (the top panel). B The immunoblotting data of EGFR from three repetitions of experiments was quantified by Kodak EDAS290 image system. ***p < 0.001. C Examination of effect of MG-132 on stress-induced endocytosis of EGFR by immunofluorescent staining of EGFR. The experimental procedures were the same as described in Fig. 3A–C Bar 20 μm. D MG-132 protects stress-induced cell death. The cells were incubated in MG-132 (10 μM)-contained medium 30 min prior to the treatment of anisomycin or UV irradiation. The viable cell numbers were counted under a microscope with a hemacytometer. The relative cell survival was plotted as percentage of the control cell numbers. In statistical analysis, the samples treated with MG-132 alone are used as the control data set. *p < 0.05, **p < 0.01, ***p < 0.001