Early events in midzone formation stabilize nascent bipolar spindles

The formation of a stable mitotic spindle is critical to the accurate partitioning of the chromosomes during mitosis. Interpolar microtubules of the spindle midzone consist of antiparallel microtubules crosslinked by kinesin-5 and stabilize the spindle by opposing forces produced when sister chromatids are attached to microtubules and under tension. Despite the importance of the interpolar microtubules, how and when they form and what determines their number remain unknown. In this study, we report that a γ-tubulin mutation (γtub-Y445D) disrupts the localization of kinesin-5 and the formation of the interpolar microtubules, resulting in spindle instability. We find that kinesin-5 crosslinking is intact in this mutant, but that it is incapable of the subsequent kinesin-5 microtubule sliding needed to stabilize the nascent spindle. Early activation of the PRC1 homolog Ase1 restores nascent spindle stability to the γtub-Y445D mutant but cannot stabilize spindles during centromere attachment. Our work shows that midzone assembly begins with the formation of interpolar microtubule precursors in monopolar spindles that persist until early metaphase and limit the formation of kinetochore attachments in new spindles.