Rac1 promotes pioneer MTs in U2-OS cells.

<p>(A) Immunolocalization of microtubules (MTs) and fluorescent phalloidin staining of F-actin in mock-transfected cells (control) or cells expressing CA-Rac1 or DN-Rac1. Contrast inverted, left, center. Zoom of boxed region, second column. Insets, BFP-Rac1. Bar, 10 µm. (B) Workflow of plusTipTracker software for detecting mKO-EB3 comets, tracking them, and classifying MT growth excursions. (C) Top: Proportion of MT growth excursions in each subpopulation in non-targeting control vector, CA-Rac1 or DN-Rac1 expressing cells. Bottom: Color key showing MT growth speed and growth excursion lifetime ranges for subpopulations. Box-plots of speed (D) and lifetime (E) of MT growth excursions, conditions as in C. (*, p<0.001, Kolmogorov-Smirnov, *, p<0.05, Students). (F) Top: mKO-EB3 tracks from 2 min time-lapse movies (frame rate = 3 s) colored according to the key in C overlaid on images of mKO-EB3 (inverted contrast), conditions described in A. Bottom: zoom of boxed region. Bars, 10 µm. (G) Percentage of MT growth tracks within 5 µm from the leading edge whose angle relative to the edge is between 0–45° (blue) or between 45–90° (green), conditions as in C.