Kinesin-5/Cut7 moves bidirectionally on fission-yeast spindles with activity that increases in anaphase

Abstract

ABSTRACTKinesin-5 motors are essential to separate mitotic spindle poles and assemble a bipolar spindle in many organisms. These tetrameric motors crosslink and slide apart antiparallel microtubules via microtubule plus-end-directed motility. However, kinesin-5s typically accumulate more at spindle poles than in the center of the spindle where antiparallel microtubule overlaps are most numerous. While the relevance of this localization has remained unclear, increasing evidence suggests that it occurs due to bidirectional motility or trafficking of kinesin-5 motors. The kinesin-5 motor Cut7p from fission yeast has been shown to move bidirectionally in reconstituted systems. However, bidirectional movement has not been shown in cells and the funtion of the minus-end-directed movement remains unclear. Here, we characterized the motility of kinesin-5/Cut7 on bipolar and monopolar spindles in fission yeast and observed movement both toward plus and minus ends of microtubules. Notably, we found that the activity of the motor increases at the onset on anaphase B. Perturbations to microtubule dynamics did not significantly change the observed Cut7p movement, while Cut7p mutation reduced or abolished observable movement. These results suggest that the directed movement of Cut7p was due to the motility of the motor itself. Mutations of Cut7p that decreased plus-end-directed motility enhanced its spindle-pole localization. In contrast, abolishing Cut7 motility or replacing it with plus-end-directed human Eg5 eliminates the pole localization. Our results suggest a new hypothesis for the function of minus-end-directed motility and spindle-pole localization of kinesin-5s in spindle assembly.