TY - JOUR A1 - Okafornta, Chukwuebuka William A1 - Farhadifar, Reza A1 - Fabig, Gunar A1 - Wu, Hai-Yin A1 - Köckert, Maria A1 - Vogel, Martin A1 - Baum, Daniel A1 - Haase, Robert A1 - Shelley, Michael J. A1 - Needleman, Daniel J. A1 - Müller-Reichert, Thomas T1 - Cell size reduction scales spindle elongation but not chromosome segregation in C. elegans JF - bioRxiv N2 - How embryos adapt their internal cellular machinery to reductions in cell size during development remains a fundamental question in cell biology. Here, we use high-resolution lattice light-sheet fluorescence microscopy and automated image analysis to quantify lineage-resolved mitotic spindle and chromosome segregation dynamics from the 2– to 64–cell stages in Caenorhabditis elegans embryos. While spindle length scales with cell size across both wild-type and size-perturbed embryos, chromosome segregation dynamics remain largely invariant, suggesting that distinct mechanisms govern these mitotic processes. Combining femtosecond laser ablation with large-scale electron tomography, we find that central spindle microtubules mediate chromosome segregation dynamics and remain uncoupled from cell size across all stages of early development. In contrast, spindle elongation is driven by cortically anchored motor proteins and astral microtubules, rendering it sensitive to cell size. Incorporating these experimental results into an extended stoichiometric model for both the spindle and chromosomes, we find that allowing only cell size and microtubule catastrophe rates to vary reproduces elongation dynamics across development. The same model also accounts for centrosome separation and pronuclear positioning in the one-cell C. elegans embryo, spindle-length scaling across nematode species spanning ~100 million years of divergence, and spindle rotation in human cells. Thus, a unified stoichiometric framework provides a predictive, mechanistic account of spindle and nuclear dynamics across scales and species. Y1 - 2025 U6 - https://doi.org/10.1101/2025.10.13.681585 ER - TY - JOUR A1 - Kiewisz, Robert A1 - Fabig, Gunar A1 - Conway, William A1 - Baum, Daniel A1 - Needleman, Daniel A1 - Müller-Reichert, Thomas T1 - Three-dimensional structure of kinetochore-fibers in human mitotic spindles JF - eLife N2 - During cell division, kinetochore microtubules (KMTs) provide a physical linkage between the chromosomes and the rest of the spindle. KMTs in mammalian cells are organized into bundles, so-called kinetochore-fibers (k-fibers), but the ultrastructure of these fibers is currently not well characterized. Here we show by large-scale electron tomography that each k-fiber in HeLa cells in metaphase is composed of approximately nine KMTs, only half of which reach the spindle pole. Our comprehensive reconstructions allowed us to analyze the three-dimensional (3D) morphology of k-fibers and their surrounding MTs in detail. We found that k-fibers exhibit remarkable variation in circumference and KMT density along their length, with the pole-proximal side showing a broadening. Extending our structural analysis then to other MTs in the spindle, we further observed that the association of KMTs with non-KMTs predominantly occurs in the spindle pole regions. Our 3D reconstructions have implications for KMT growth and k-fiber self-organization models as covered in a parallel publication applying complementary live-cell imaging in combination with biophysical modeling (Conway et al., 2022). Finally, we also introduce a new visualization tool allowing an interactive display of our 3D spindle data that will serve as a resource for further structural studies on mitosis in human cells. Y1 - 2022 U6 - https://doi.org/10.7554/eLife.75459 VL - 11 SP - e75459 ER -