Experimental Center, Faculty of Medicine Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany.
Department of Physics, Harvard University, Cambridge, United States.
Elife. 2022 Jul 27;11:e75459. doi: 10.7554/eLife.75459.
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.
在细胞分裂过程中，动粒微管（KMTs）为染色体和纺锤体的其余部分提供物理连接。哺乳动物细胞中的 KMT 组织成束，称为动粒纤维（k-fibers），但这些纤维的超微结构目前尚未很好地描述。在这里，我们通过大规模电子断层扫描表明，在有丝分裂中期的 HeLa 细胞中，每条 k 纤维由大约 9 个 KMT 组成，只有一半到达纺锤极。我们的全面重建使我们能够详细分析 k 纤维及其周围 MT 的三维（3D）形态。我们发现，k 纤维在其长度上的周长和 KMT 密度表现出显著的变化，其极近侧显示出变宽。然后，我们将结构分析扩展到纺锤体中的其他 MT，进一步观察到 KMT 与非 KMT 的结合主要发生在纺锤极区域。我们的 3D 重建对 KMT 生长和 k 纤维自组织模型具有影响，这在应用互补活细胞成像结合生物物理建模的平行出版物中得到了报道（Conway 等人，2022 年）。最后，我们还引入了一种新的可视化工具，允许交互式显示我们的 3D 纺锤体数据，这将作为进一步研究人类细胞有丝分裂结构的资源。
