CryoET imaging reveals that mitotic spindles are locally organized by microtubule density and Eg5 activity

When eukaryotic cells divide, a mitotic spindle, composed of microtubule filaments interacting with molecular motors, self-assembles and is responsible for aligning and segregating chromosomes to the opposite daughter cells. Recent structural studies revealed that spindles are composed of short microtubules that do not span the full pole-to-chromosome distance. It remains unclear how these short, disconnected microtubules collectively assemble a bipolar spindle. Using cryo-electron tomography to map microtubule polarity in human spindles, we find that microtubules organize with consistent 8 nm wall-to-wall spacing. Spacing scales inversely with local microtubule density, consistent with organization by density-driven steric interactions, analogous to liquid crystal ordering. We find that the Eg5 motor establishes local antiparallel overlap independently of spindle bipolarity, while a balance of Eg5 and dynein regulates microtubule density to maintain spindle architecture. Together, these findings reveal the mesoscale principles by which cells build complex, self-healing cellular architectures from nanoscale building blocks.