Vimentin interplay with actin filaments and microtubules. (A–C) Fluorescence images taken from FRAP experiments on vimentin-EGFP–expressing astrocytoma cells treated with DMSO (A), 10 µM latrunculin A (B), and 2 µM blebbistatin (C). The images were acquired 0 and 9 min after photobleaching. Kymographs are boxed in yellow and show the fluorescence intensity profile along the yellow arrows in the corresponding images over time for each condition. The higher-magnification images of regions indicated by a blue dotted box are shown in the corresponding dotted line boxes. (D) Fluorescence image of an astrocytoma cell expressing vimentin-EGFP (green) and mCherry-tubulin (magenta). Time-lapse images of the region indicated by a dotted blue box are shown on the right. Arrowheads highlight a microtubule tip, and asterisks show the tip of a vimentin IF moving along this microtubule (Video 5; total time, 12 min). (E) Fluorescence images of an astrocytoma cell expressing vimentin-EGFP and mCherry-tubulin showing EGFP fluorescence 2 min after photobleaching and mCherry fluorescence before photobleaching. The merged image and the higher-magnification image of the boxed region are shown on the right. (F) STORM image of the edge of a migrating astrocytoma cell stained for vimentin (green) merged with the corresponding epifluorescence (epi) image of the microtubule network (magenta). A higher magnification of the region indicated by a dotted line rectangle on the vimentin network is shown on the right. (G) FRAP experiment on a vimentin-EGFP–expressing astrocytoma cell treated with 10 µM nocodazole. White dashed outline indicates the cell contour. A higher magnification of the boxed region (cyan) shows single vimentin filaments moving into the bleached area from the adjacent regions. The kymograph shows the fluorescence intensity profile along the yellow arrow over time. Bars: (main images) 10 µm; (insets) 2 µm; unless otherwise indicated.