Polak Bruno, Risteski Patrik, Lesjak Sonja, Tolić Iva M
Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia.
Division of Molecular Biology, Ruđer Bošković Institute, Zagreb, Croatia
EMBO Rep. 2017 Feb;18(2):217-230. doi: 10.15252/embr.201642650. Epub 2016 Dec 27.
In the mitotic spindle, kinetochore microtubules form k-fibers, whereas overlap or interpolar microtubules form antiparallel arrays containing the cross-linker protein regulator of cytokinesis 1 (PRC1). We have recently shown that an overlap bundle, termed bridging fiber, links outermost sister k-fibers. However, the relationship between overlap bundles and k-fibers throughout the spindle remained unknown. Here, we show that in a metaphase spindle more than 90% of overlap bundles act as a bridge between sister k-fibers. We found that the number of PRC1-GFP-labeled bundles per spindle is nearly the same as the number of kinetochore pairs. Live-cell imaging revealed that kinetochore movement in the equatorial plane of the spindle is highly correlated with the movement of the coupled PRC1-GFP-labeled fiber, whereas the correlation with other fibers decreases with increasing distance. Analysis of endogenous PRC1 localization confirmed the results obtained with PRC1-GFP PRC1 knockdown reduced the bridging fiber thickness and interkinetochore distance throughout the spindle, suggesting a function of PRC1 in bridging microtubule organization and force balance in the metaphase spindle.
在有丝分裂纺锤体中，动粒微管形成 k 纤维，而重叠微管或极间微管形成包含细胞分裂调节蛋白 1（PRC1）的反平行阵列。我们最近发现，一种称为桥纤维的重叠束连接着最外层的姐妹 k 纤维。然而，整个纺锤体中重叠束与 k 纤维之间的关系仍然未知。在这里，我们表明，在中期纺锤体中，超过 90%的重叠束充当姐妹 k 纤维之间的桥梁。我们发现每个纺锤体中 PRC1-GFP 标记的束的数量与动粒对的数量几乎相同。活细胞成像显示，纺锤体赤道平面上的动粒运动与耦合的 PRC1-GFP 标记纤维的运动高度相关，而与其他纤维的相关性随着距离增加而降低。对内源性 PRC1 定位的分析证实了用 PRC1-GFP 获得的结果，PRC1 敲低降低了整个纺锤体中的桥纤维厚度和动粒间距离，表明 PRC1 在中期纺锤体中桥接微管组织和力平衡方面具有功能。
