Expression, localization, and function of ERM proteins. (A– E ) WT mice were left untreated (A–C) or treated for 30 min with 250 µM NSC (D) or vehicle (DMSO, E). Mice were injected with 0.01 mg/kg ISOP and after 10 min salivary glands were excised and processed for indirect immunofluorescence with Alexa488-phalloidin (green) and antibodies directed against Ezrin (A), Radixin (A and B), Moesin (A and B), pan phospho-ERM (C) or phospho-Ezrin (D and E). (A) Low mag of acini showing the localization of the various markers (magenta, arrows) at the APM. Only Radixin and Ezrin localize at the APM (arrows). Bar, 10 µm. (B) Radixin localizes on the secretory granules at early (E) but not late (L) stages (arrowheads). Moesin is not recruited on the fused granules. Bar, 1 µm. The graphs show the thickness of the actin coat measured as described in Material and Methods for granules labelled with Radixin or Ezrin. N = 100 granules, 20 cells, 3 animals. (C) pERM is detected on the secretory granules at both early (E) and late (L) stages (arrowheads). Bar, 10 µm. (D and E ) Treatment with NSC reduces the levels of p-Ezrin phosphorylation. Bar, 10 µm; 1 µm (high magnification).