Figure 3. Drug-stabilized microtubule... | Rockefeller University Press

$Drug-stabilized microtubule dynamics dramatically reduce the frequency of detachment events in low-tension cells. (A) Time series depicting sister centromeres (green) and spindle poles (red) in metaphase yeast cells (all images have been band-pass filtered in MATLAB). Top: Untreated Cin8-degron cells. Bottom: Cin8-degron cells treated with a low dose of benomyl (30 µg/ml). (B) Sister centromere spacing is measured as the spacing between two green lacO-lacI-GFP signals. Sister centromere spacing was plotted against time for each single cell. Left: Sister centromere spacing history for an untreated Cin8-degron cell with a detachment event. Right: Sister centromere spacing history for a Cin8-degron benomyl-treated cell with no detachment event. (C) Blue: Combined tension history for untreated Cin8-degron cells with detachment events (n = 20 cells; binned at 1 min intervals; mean ± SEM). Magenta: Combined tension history for Cin8-degron benomyl-treated cells without detachment events (n = 30 cells; binned at 1 min intervals; mean ± SEM). (D) Tension history data was fit to a linear model for untreated Cin8-degron cells with detachment events (blue), and Cin8-degron benomyl-treated cells without detachment events (magenta). The variability in tension from the mean fit line appeared larger for untreated cells (blue open circles) as compared to benomyl-treated cells (magenta open circles). (E) Cumulative squared error vs. time, with cumulative squared error calculated by squaring the difference between the linear predicted tension value and the actual tension value, at each time point. (F) Top: Representative image of an untreated Cin8-degron cell with attached kinetochores. Bottom: Representative image of a Cin8-degron benomyl-treated cell with attached kinetochores. (G) Tension distribution of untreated (n = 105 cells) and benomyl-treated cells (n = 93 cells), measured in population sampling experiments. (H) Left-Top: Representative image of an untreated Cin8-degron cell with a detached kinetochore (white arrow; dotted white line denotes spindle axis). Left-Bottom: Representative image of a benomyl-treated Cin8-degron cell with a detached kinetochore (white arrow; dotted white line denotes spindle axis). Right: Fraction of cells with detached kinetochore microtubules, measured in population sampling experiments. Benomyl-treated cells (n = 261 cells) have a nearly twofold lower fraction of cells with detached kinetochore microtubules relative to untreated cells (n = 265 cells; P = 0.002, Z test).$

Figure 3.

Drug-stabilized microtubule dynamics dramatically reduce the frequency of detachment events in low-tension cells. (A) Time series depicting sister centromeres (green) and spindle poles (red) in metaphase yeast cells (all images have been band-pass filtered in MATLAB). Top: Untreated Cin8-degron cells. Bottom: Cin8-degron cells treated with a low dose of benomyl (30 µg/ml). (B) Sister centromere spacing is measured as the spacing between two green lacO-lacI-GFP signals. Sister centromere spacing was plotted against time for each single cell. Left: Sister centromere spacing history for an untreated Cin8-degron cell with a detachment event. Right: Sister centromere spacing history for a Cin8-degron benomyl-treated cell with no detachment event. (C) Blue: Combined tension history for untreated Cin8-degron cells with detachment events (n = 20 cells; binned at 1 min intervals; mean ± SEM). Magenta: Combined tension history for Cin8-degron benomyl-treated cells without detachment events (n = 30 cells; binned at 1 min intervals; mean ± SEM). (D) Tension history data was fit to a linear model for untreated Cin8-degron cells with detachment events (blue), and Cin8-degron benomyl-treated cells without detachment events (magenta). The variability in tension from the mean fit line appeared larger for untreated cells (blue open circles) as compared to benomyl-treated cells (magenta open circles). (E) Cumulative squared error vs. time, with cumulative squared error calculated by squaring the difference between the linear predicted tension value and the actual tension value, at each time point. (F) Top: Representative image of an untreated Cin8-degron cell with attached kinetochores. Bottom: Representative image of a Cin8-degron benomyl-treated cell with attached kinetochores. (G) Tension distribution of untreated (n = 105 cells) and benomyl-treated cells (n = 93 cells), measured in population sampling experiments. (H) Left-Top: Representative image of an untreated Cin8-degron cell with a detached kinetochore (white arrow; dotted white line denotes spindle axis). Left-Bottom: Representative image of a benomyl-treated Cin8-degron cell with a detached kinetochore (white arrow; dotted white line denotes spindle axis). Right: Fraction of cells with detached kinetochore microtubules, measured in population sampling experiments. Benomyl-treated cells (n = 261 cells) have a nearly twofold lower fraction of cells with detached kinetochore microtubules relative to untreated cells (n = 265 cells; P = 0.002, Z test).

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