Fig. 1: PRC1-crosslinked microtubules undergo a transition from a loose array to bundles during spindle assembly.

a Spindles immunostained for α-tubulin and imaged at STED resolution in a HeLa-Kyoto BAC cell line stably expressing PRC1-GFP with DNA stained by DAPI (both imaged at a confocal resolution), in early prometaphase, late prometaphase, and metaphase. Spindles lying parallel (first row) or perpendicular (second row) to the imaging plane are shown. b Time-lapse images (single plane) of a cross-section of a vertically oriented prometaphase spindle in a HeLa-Kyoto BAC cell stably expressing PRC1-GFP, starting with the prometaphase rosette. c Number of PRC1 segments in the midplane over time for cells as in (b), obtained using Squassh segmentation. Gray lines represent individual cells, green line the mean and gray areas the standard deviation, n = 10 cells in 10 independent experiments. d Mean intensity of PRC1 segments from the same cells as in (c). e, f Intensity line plots of PRC1-GFP in the cell from (b) at t = 0 (e) and t = 7.2 min (f) along the periphery of the spindle cross-section (lines shown in Supplementary Fig. 1k). g Ratio of mean PRC1-GFP intensity in (f, e) (“Mean”) and ratio of the standard deviation in (f, e) (“Standard deviation”). The black line shows the mean; the light and dark gray areas mark 95% confidence interval on the mean and standard deviation, respectively; n = 10 cells in 10 independent experiments. Dots of the same color represent the same cells. All scale bars, 1 µm.