The proteolysis of key regulatory proteins is thought to control progress through mitosis. Here we analyse cyclin B1 degradation in real time and find that it begins as soon as the last chromosome aligns on the metaphase plate, just after the spindle-assembly checkpoint is inactivated. At this point, cyclin B1 staining disappears from the spindle poles and from the chromosomes. Cyclin B1 destruction can subsequently be inactivated throughout metaphase if the spindle checkpoint is reimposed, and this correlates with the reappearance of cyclin B1 on the spindle poles and the chromosomes. These results provide a temporal and spatial link between the spindle-assembly checkpoint and ubiquitin-mediated proteolysis.