Stimulation of matrix degradation by KIF5B correlates with increased ARF6-, JIP3/4-, and p150Glued-dependent endosome tubulogenesis. (A) Inverted still images from time-lapse sequences of MDA-MB-231 cells expressing KIF5B-YFP and MT1-MMPmCh treated with the indicated siRNAs (see Video 3). Bar, 10 µm. (B) Mean percentage distribution of MT1-MMPmCh–positive endosomes according to their cell center-to-cell periphery position ± SEM. (C) FITC-gelatin degradation by MDA-MB-231 cells overexpressing MT1-MMPmCh together with YFP or KIF5B-YFP. n, number of cells scored for each MDA-MB-231 cell population treated with the indicated siRNAs. (B and C) ***, P < 0.001 (as compared with siNT-treated cells). (D) High-magnification galleries of peripheral cell regions (boxed in A). Arrows point to tubulated MT1-MMP endosomes. Bar, 5 µm. (E) TIRF microscopy images of MDA-MB-231 cells expressing MT1-MMPmCh and KIF5B-YFP plated on gelatin. Gallery shows images with 0.2-s intervals corresponding to the boxed region. Arrows point to membrane tubulation events. Bars: 5 µm; (gallery) 2 µm. (F and G) Number of tubulated MT1-MMPmCh endosomes per frame normalized for the surface area. Mean ± SEM. n, number of peripheral cell regions scored for each cell population. *, P < 0.05; **, P < 0.01 (as compared with siNT condition).
