Fig. 8. Inactivation of ERM proteins by neomycin. (A) L cells were microinjected with 10 mM neomycin/F-dextran and incubated for 10 (a-c) or 30 (d) minutes. Cells were double stained with anti-ERM mAb, CR22 (b) and anti-CPERM mAb (c). In neomycin-microinjected cells (a), ERM proteins were dephosphorylated (c) and translocated to the cytoplasm (b). Scanning electron microscopy showed that microvilli disappeared from the cell surface (d). (B) L cells were homogenized in the presence of 1 mM neomycin, incubated for 30 minutes on ice and centrifuged. The supernatant (S) and pellet (P) were resolved by SDS-PAGE followed by immunobloting with anti-ERM pAb, TK89. Neomycin induced translocation of ERM proteins from the insoluble to soluble cell fraction. (C) MDCK II cells were microinjected with 10 mM neomycin/F-dextran and incubated for 10 minutes. Cells were doubly stained with anti-CPERM mAb (a) and anti-ERM mAb, CR22 (b). In neomycin-microinjected cells (arrows), ERM proteins were dephosphorylated (a) and translocated to the cytoplasm (b). Bars, Aa-c, 10 µm; Ad, 2 µm; C, 10 µm.