SEPT9 promotes the motility of renal epithelia during EMT. (A) Western blot of HGF-treated MDCK cell lysates probed for SEPT9, actin, and GAPDH. (B) Graphs show the median, lowest, and highest (error bars) ratio values of the SEPT9, actin, and GAPDH band intensities from three independent experiments. Values were normalized to the protein band intensity ratios at 0 h of HGF treatment. (C) Maximum intensity projections from 3D confocal microscopy images of HGF-treated renal cysts of stable MDCK cells expressing SEPT9-mCherry (green) at low and high levels of expression. 3D renal cysts were grown in collagen and stained for nuclei (DAPI; blue) and F-actin (phalloidin; red). (D) Graphs show the number of extensions per MDCK cyst (n = 23) and the number of cells per extension from untransfected (n = 18), low-expressing (n = 35), and high-expressing (n = 51) MDCK cysts (n = 15). (E) Mean number of extensions per cyst after treatment with DMSO (n = 23) and FCF (n = 19). (F and G) Graphs show the mean percentage of transmigrated 786-O cells expressing GFP and SEPT9-GFP (F), and control and SEPT9 shRNAs from 20 different areas of a transwell filter. (H and I) Graphs show the mean velocity (H; n = 16) and axial ratio of the long to short axes (I; n = 31) of MDCK cells migrating in a 3D matrix in the presence of HGF after transfection with control or SEPT9 shRNAs. Error bars indicate SEM. **, P < 0.01; ***, P < 0.001.