Repo-Man and Sds22 stabilize anaphase chromosome segregation. (A and B) Depletion of Repo-Man or Sds22 increases the incidence of anaphase bridges (A) and lagging chromosomes (B). Segregation defects were detected in confocal videos of live HeLa Kyoto cells stably expressing H2B-mCherry (n > 500 cells; seven independent experiments; **, P < 0.001, using a two-sided binomial test). Bars, 10 µm. (C–I) 3D tracking of kinetochores in HeLa cells stably expressing EGFP–CENP-A (Jaqaman et al., 2010) reveals pauses in poleward segregation. (C and E) Kymographs of sister kinetochore pairs (yellow arrow highlights interkinetochore distance). Tracked sister kinetochores are highlighted by colored circles; green represents linear segregation, and red represents paused segregation (net movement of <0.1 µm in three subsequent frames). t = 0 min at chromosome segregation onset (dashed lines). Bars, 5 µm. (D, F, G, and H) 3D interkinetochore distances of sister kinetochore pairs from metaphase until full segregation. Paused segregation (net movement of <100 nm in three subsequent frames) is highlighted in red. t = 0 min at chromosome segregation onset (dashed lines). (I) Incidence of sister kinetochore pairs per cell with paused segregation after initially normal segregation during poleward movement 0–5 min after anaphase onset. n ≥ 9 cells (10 kinetochore pairs per cell) per condition. Mean ± SEM. **, P < 0.001, using a two-sided binomial test.