Figure 4

Microtubule detachment from centrosomes. (A) Paclitaxel dependent mutants Tax 11-6 and Tax 18 were grown for 2 d without paclitaxel, transfected with EGFP-MAP4 and viewed by live cell imaging. Fluorescent images taken 5 s apart were deconvolved to improve contrast. Note detachment and shortening from the minus end of a long microtubule in Tax 11-6 (arrows, upper panels); and detachment and translocation of a newly nucleated microtubule in Tax 18 (arrows, lower panels). Asterisks mark the centrosomal area of the cells. Scale bar equals 5 μm. (B) Cells treated with the indicated concentrations of paclitaxel for 2 d were viewed by time-lapse fluorescence microscopy. Microtubules that detached from the centrosome were counted from 25-40 min of time-lapse video recordings representing 5 cells in 2 separate experiments, and the rate was calculated as the number of detachments/cell/min. Error bars indicate the standard deviation from the mean.