Local Load-Bearing by Kinetochore-Fibers in the Mammalian Spindle Provides Mechanical Isolation and Redundancy

Abstract

When cells divide, microtubule bundles called kinetochore-fibers (k-fibers) attach chromosomes to the mammalian spindle. Active forces generated at kinetochores move chromosomes, and the dynamic spindle must robustly anchor k-fibers to bear this load. Where and how anchorage occurs are not understood. To spatially map load-bearing by k-fibers and determine its molecular basis, we cut k-fibers at different spindle locations and quantitatively measure residual load-bearing in different molecular backgrounds. The relaxation response immediately after ablation indicates that k-fibers are anchored not only at their ends, but along their lengths. We find that the load of k-fiber anchorage is borne very locally: longitudinally in their first few microns from kinetochores, far from poles; and laterally within 1-2 μm, without neighboring k-fibers sharing load. Depleting the microtubule crosslinker NuMA reduces local load-bearing anchorage in the spindle body, while inhibiting or depleting microtubule motor Eg5 and crosslinker PRC1 do not. A simple viscoelastic model suggests that elastic connections of k-fibers to the spindle bear the load of active kinetochore forces moving chromosomes, while centromere viscosity determines the timescale of relaxation after ablation removes load-bearing connections. Together, the data indicate that the architecture of the dynamic mammalian spindle provides k-fibers with mechanical isolation and load-bearing redundancy well-suited for robust chromosome segregation.