Fig. 8. Dominant-negative effects of N19RhoA and of N-terminal domain of ezrin (N-ezrin) in podoplanin-induced EMT. (A) Phase-contrast micrographs and confocal fluorescence detection of podoplanin in MDCK cells cotransfected with EGFP-tagged PWT and N-ezrin or N19RhoA. Control cells expressing EGFP alone, EGFP/N-ezrin and EGFP/N19RhoA are also shown. Confocal images are maximum projections of horizontal optical sections through the whole depth of the transfectants. (B) Western blot analysis of N-ezrin (VSVG), N19RhoA (HA) and differentiation-related proteins in MDCK cell transfectants. N-ezrin and N19RhoA expression was determined by using antibodies against their respective tags. (C) N-ezrin and N19RhoA inhibits podoplanin-mediated RhoA activation and ERM phosphorylation. The levels of RhoA-GTP relative to the levels of total RhoA and the levels of phospho-ERM relative to the total expression levels of ezrin and moesin are shown. The expression of -tubulin was determined as a control for protein loading.