Fig. 6: CathD regulates cell mitosis and cytokinesis via its cofilin phosphatase activity.
From: The noncanonical role of the protease cathepsin D as a cofilin phosphatase
a, b CathD-specific (cathDRNAi) or scrambled shRNA (scramble) was co-expressed with GFP in asynchronous HeLa cells. After cultured in 15 μM Hsp inhibitor I or vehicle for 48 h, cells were fixed and stained for F-actin with rhodamine-conjugated phalloidin (red). Nuclei were counter-stained with DAPI. (a) Representative results showing that downregulation of cathD by transfection of cathD-targeting shRNA generates giant multinuclear cells. Arrows indicate nuclei in control cells, while arrowheads indicate nuclei in a tetraploidy cell induced by cathD shRNA. Scale bar, 30 μm. (b) Quantifications of multinuclear HeLa cells. Compared with controls (scramble), downregulation of cathD (cathDRNAi) increases giant multinuclear cells. c A significant increase of giant multinuclear cells was found in cathDD33G, cathDD75G, and cathDE117G mutants. Scale bar, 30 μm. d Quantifications of Ki67-positive A549 cells with indicated cathD mutants. Data are means ± SEM. One-way ANOVA followed with the Tukey’s test, **P < 0.01 and ***P < 0.001 compared with cathDwt (n = 6). e CCK-8 assay of A549 cells at indicated time points. Relative proliferation ratios were normalized to the values at 0 h. f Representative images showing A549 xenografts carrying indicated cathD mutations in nude mice after subcutaneous transplantation. g The change in tumor volume of A549 xenografts (relative to the initial volume) carrying indicated cathD mutations (n = 9). Data are means ± SEM, *P < 0.05, **P < 0.01, and ***P < 0.001 the by Mann–Whitney U test, Kruskal–Wallis test (with Dunn’s test), or ANOVA (with Tukey’s test), respectively.
