Fig. 4

Spatiotemporal control over MT dynamics in 2D-cultured HeLa cells. (a, b) MT inhibition in SBTubA4P-treated cells is initiated only upon 405 nm illumination pulses and only in ROI-targeted cells (data related to Movie S1; live-cell EB3-tdTomato comets quantify polymerizing MTs). (a) Comet count statistics are similar to cosolvent-only baseline in both ROI-pulsed-cosolvent and non-ROI-SBTubA4P conditions; ROI-SBTubA4P statistics show inhibition spikes. (b) Stills from Movie S1 at the times indicated in (a), initially during the untreated timecourse, then during the SBTubA4P-treated timecourse on the same cells. Purple arrowhead indicates the ROI cell; purple dotted circle indicates where the 405 nm ROI is applied at times 26, 88, and 148 s; and white arrowhead indicates the non-ROI cell quantified as the internal control (scale bar 15 μm). (c) EB3 comet counts of cells imaged at 561 nm only (dark, gray), with 47 frames at 487 nm applied to full field of view during the time span indicated with dashed lines (“487”, cyan) and SBTubA4P (6 μM), or with single-frame 405 nm pulses, SBTubA4P (0.6 μM), applied to full field of view at times indicated with dashed lines (“405”, violet) (n = 3 cells). Temporally precise onset and full-field diffusional reversibility are shown (data related to Movies S2 and S3). [(a, c) Mean ± standard error of the mean (SEM) EB3 comet counts as normalized to the means of the first five time points; 405 nm ROIs applied at indicated times; for further details, see the Supporting Information].