FIGURE 5:

Analysis of spindle pole displacements in living cells. Spindle dynamical behavior was measured using 3D spindle-pole tracking and extended projection kymographs. (A) Both the kymograph and tracking data show that a wild-type spindle does not increase in net length, whereas pole tracking demonstrates that the amplitude of length fluctuations decreases over time. (B) Both the kymograph and tracking data show that a γ-tubS360A undergoes steady increase in length. The kymograph reveals that net increase in length correlates with outward displacement of both old and new spindle poles. (C) γ-tubS360D spindles are unstable, exhibiting very large fluctuations in length. Frequent tilting of the γ-tubS360D spindle through more than a single focal plane results in lack of correspondence between kymograph and pole tracking data and illustrates the need for pole tracking for accurate measurement of spindle dynamics. (D) Initial pole separation is delayed as a consequence of a cin8∆ loss-of-function mutation but is unaffected by γ-tubS360A and γ-tubS360D mutations.