Figure 3. Loss of cohesin leads to unstable kinetochore-microtubule attachments.

Rad21^TEV embryos were injected with TEV protease to trigger sister centromere disjunction in Mad2^L13Q-induced metaphase-arrested embryos. a) Selected stills from a movie showing His2Av-mRFP1 (red) and Cid-EFGP (green); arrows follow the trajectory of a single centromere after the initial pole-ward movement; times (min:sec) are relative to TEV injection; scale bar is 5 μm; b) representative kymograph showing centromere behaviour for 15 minutes after TEV-induced segregation. Vertical scale bars are 5 μm and horizontal scale bars correspond to 1 minute; c) example of a single centromere trajectory for 15 minutes after TEV-mediated centromere disjunction; pauses are in red and runs in blue (away from starting point) and green (towards starting point); grey line represents the segregation axis; d) directions of centromere movement (measured by the angle of trajectory) in Mad2-TEV experiments plotted on rose diagrams to show overall bias in run directions towards the axis of segregation (0°-180°); e) frequency (%) of maximum speed per run in control and Mad2-TEV embryos; top two panels show the maximum speeds per run observed during the initial segregation phase (until 2 minutes after the initial separation of sister centromeres); bottom panel shows later chromosome movements observed in Mad2-TEV experiments (from 2 minutes after separation onwards); insets show speeds >10 μm/min enlarged (5x) on the y axis; single centromere trajectories were analysed from 80 individual centromeres (out of 5 independent experiments); note that while during the segregation phase the chromosome movements in TEV-induced pseudo-anaphases are consistently slower than in controls, later chromosome movements are still overall slower but exhibit infrequent very fast movements (>12 μm/min).