High-resolution imaging reveals how the spindle midzone impacts chromosome movement

Abstract

In the spindle midzone, microtubules from opposite half-spindles form bundles between segregating chromosomes. Microtubule bundles can either push or restrict chromosome movement during anaphase in different cellular contexts, but how these activities are achieved remains poorly understood. Here, we use high-resolution live-cell imaging to analyze individual microtubule bundles, growing filaments, and chromosome movement in dividing human cells. Within bundles, filament overlap length marked by the cross-linking protein PRC1 decreases during anaphase as chromosome segregation slows. Filament ends within microtubule bundles appear capped despite dynamic PRC1 turnover and submicrometer proximity to growing microtubules. Chromosome segregation distance and rate are increased in two human cell lines when microtubule bundle assembly is prevented via PRC1 knockdown. Upon expressing a mutant PRC1 with reduced microtubule affinity, bundles assemble but chromosome hypersegregation is still observed. We propose that microtubule overlap length reduction, typically linked to pushing forces generated within filament bundles, is needed to properly restrict spindle elongation and position chromosomes within daughter cells.

Aurora A promotes chromosome congression by activating the condensin-dependent pool of KIF4A

“…3, B-F). Consistent with other work (Pamula et al, 2019), we observed that PRC1 localized to bundles centered in the middle of the cell away from the cortex early in anaphase before furrowing ( Fig. 4 A, WT 6-8 min), and this organization persists once furrowing commences ( Fig.…”

“…PRC1-binding mutants of KIF4A are defective for central spindle length control Two key hallmarks are associated with of loss of KIF4A and PRC1 function in anaphase. First, the integrity of the central spindle is lost and microtubule bundles become disorganized and fail to form an ordered centered array, and second, chromosomes show enhanced separation and movement in anaphase (Nunes Pamula et al, 2019). KIF4A is crucial for anaphase spindle length control and the formation of organized bundles of anaphase spindle microtubules ( Fig.…”

“…In anaphase, a second pool of KIF4A interacts with PRC1, a highly conserved, homodimeric microtubule-binding protein required for central spindle formation (Mollinari et al, 2002(Mollinari et al, , 2005. PRC1 and KIF4A form a minimal self-organizing microtubule bundling system in anaphase (Hannabuss et al, 2019) and are mutually dependent on each other for localization to the anaphase spindle midzone, where they together determine central spindle length and contribute to anaphase chromosome position (Bieling et al, 2010;Hu et al, 2011;Pamula et al, 2019;Subramanian et al, 2010). This is achieved by molecular tagging of the ends of microtubules in the central spindle zone by KIF4A and PRC1 in a microtubule-length-dependent manner (Subramanian et al, 2013).…”

Chromosome segregation is driven by joint microtubule sliding action of kinesins KIF4A and EG5

“…3, D and E) demonstrating that the effect of modulating kinesin and PRC1 does not depend on the MT turnover. Spindle elongation during anaphase is thus, contrary to earlier suggestions (20), independent of midzone MT stability provided primarily by MT bundling protein PRC1.…”

“…To test this, we depleted the main crosslinker of antiparallel MTs and scaffold for recruitment of multiple motors, protein regulator of cytokinesis 1 (PRC1) (19), with a siRNA. PRC1 depletion often led to excessive spindle and cell elongation, as reported previously (20), while not affecting rates of spindle elongation in early anaphase ( fig. S3, A to E).…”