Fig 1: Modulating PKC alters cell morphology and signalling. (A) Western blot demonstrates PKC activity in both total cell lysates and integrin adhesion complexes from podocytes spread onto collagen IV or laminin for 210 min. (B) The colour-coded shape outlines indicate representative protrusive activities at 5-min intervals recorded over 100-min period. For measurements of protrusive activity live cell imaging was performed between 180 and 280 min of cell spreading. Podocytes were treated with either G?6976 or PMA to suppress or induce PKC activity respectively during cell spreading on matrix ligand. (C,D) PKC modulation impacts podocyte morphology and pseudopodial projection formation. (E,F) PKCa localization to adhesions is controlled through ligand specificity. Podocytes were spread on circles and lines coated with collagen IV or laminin for 3.5 h in serum-free media and stained for PKCa and paxillin and ratiometric imaging was performed. Scale bar represents 20 µm. Data are pooled from three independent experiments. G?6976 induced inhibition of PKCa localization to FA was significant for collagen circles, lines and laminin lines. For ratiometric imaging 20–40 cells were analysed per experiment and each experiment was performed four times. All bar graph measurements are shown as mean ± standard deviation. Box plots indicate 25th and 75th percentiles (lower and upper bounds, respectively), 1.5 × interquartile range (whiskers) and median (black line). Scale bar in (e) represents 5 µm; ****, p < 0.0001; NS, not significant; LAM511, laminin-511; COL4, collagen IV. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)