Abstract The role of myosin II in mitosis is generally thought to be restricted to cytokinesis. We present surprising new evidence that cortical myosin II is also required for spindle assembly in cells. Drug- or RNAi-mediated disruption of myosin II in cells interferes with normal spindle assembly and positioning. Time-lapse movies reveal that these treatments block the separation and positioning of duplicated centrosomes after nuclear envelope breakdown (NEBD), thereby preventing the migration of the microtubule asters to opposite sides of chromosomes. Immobilization of cortical movement with tetravalent lectins produces similar spindle defects to myosin II disruption and suggests that myosin II activity is required within the cortex. Latex beads bound to the cell surface move in a myosin II-dependent manner in the direction of the separating asters. We propose that after NEBD, completion of centrosome separation and positioning around chromosomes depends on astral microtubule connections to a moving cell cortex.