Fig. 5.

Effects of anillin depletion on epithelial junctions are not mediated by altered NM II activity. a, b DU145 cells were transfected with either control or anillin-specific siRNAs (duplex 1 and 3) and were harvested for total cell lysates on day 4 post-transfection. Expression of different NM II heavy chain isoforms, total regulatory myosin light chain (RMLC), as well as monophosphorylated (p) and diphosphorylated (pp) RMLC, was determined by immunoblotting. c Control and anillin-depleted DU145 cells were immunolabeled for p-RMLC and pp-RMLC. Arrows point out localization of phosphorylated RMLC at epithelial junctions in control DU145 cells, whereas arrowheads highlight loss of such localization following anillin depletion. d, e Control DU145 cells were treated for 24 h with either vehicle, or ROCK inhibitor, Y-27632 (25 µM), and examined for AJ/TJ integrity and RMLC phosphorylation. Arrows indicate intact AJ and TJ in either vehicle or Y-27632-treated epithelial cells. Scale bar 20 µm