Fig. 3

p38 kinase activity is required for stress-induced endocytosis of EGFR. A–C HeLa cells were cultured on glass cover slip-bottomed dishes (Matek) and serum-starved in DMEM plus 1 % FBS for 12 h. The Caspase-3 inhibitor, p38 kinase inhibitor SB203850 or MEK inhibitor U0126 was added into the medium 30 min before the treatment of EGF, anisomycin or UV irradiation. Then the cells were fixed and immunostained with anti-EGFR(1005) as described in “Methods” section. Bar 20 μm. D HeLa cells were biotin-labeled after treatment of anisomycin. The biotin-labeled EGFR was immunoprecipitated by anti-EGFR(Mab528) and detected by immunoblotting with anti-biotin (the top panel). The relative amount of lysates used for immunoprecipitation was controlled by immunoblotting of β-actin in the lysates (the bottom panel)