Oncode Institute, University Medical Center Utrecht, Utrecht University, 3584, Utrecht, the Netherlands; Center for Molecular Medicine, University Medical Center Utrecht, Utrecht University, 3584 Utrecht, the Netherlands.
Cell Biology, Neurobiology and Biophysics, Department of Biology, Faculty of Science, Utrecht University, 3584 Utrecht, the Netherlands.
Curr Biol. 2020 Jul 6;30(13):2628-2637.e9. doi: 10.1016/j.cub.2020.04.081. Epub 2020 Jun 4.
During cytokinesis, signals from the anaphase spindle direct the formation and position of a contractile ring at the cell cortex [1]. The chromosomal passenger complex (CPC) participates in cytokinesis initiation by signaling from the spindle midzone and equatorial cortex [2], but the mechanisms underlying the anaphase-specific CPC localization are currently unresolved. Accumulation of the CPC at these sites requires the presence of microtubules and the mitotic kinesin-like protein 2, MKLP2 (KIF20A), a member of the kinesin-6 family [2-7], and this has led to the hypothesis that the CPC is transported along microtubules by MKLP2 [3-5, 7]. However, the structure of the MKLP2 motor domain with its extended neck-linker region suggests that this kinesin might not be able to drive processive transport [8, 9]. Furthermore, experiments in Xenopus egg extracts indicated that the CPC might be transported by kinesin-4, KIF4A [10]. Finally, CPC-MKLP2 complexes might be directly recruited to the equatorial cortex via association with actin and myosin II, independent of kinesin activity [4, 8]. Using microscopy-based assays with purified proteins, we demonstrate that MKLP2 is a processive plus-end directed motor that can transport the CPC along microtubules in vitro. In cells, strong suppression of MKLP2-dependent CPC motility by expression of an MKLP2 P-loop mutant perturbs CPC accumulation at both the spindle midzone and equatorial cortex, whereas a weaker inhibition of MKLP2 motor using Paprotrain mainly affects CPC localization to the equatorial cortex. Our data indicate that control of cytokinesis initiation by the CPC requires its directional MKLP2-dependent transport.
在胞质分裂过程中，来自后期纺锤体的信号指导收缩环在细胞皮层的形成和定位[1]。染色体乘客复合物（CPC）通过纺锤体中间区和赤道皮层的信号参与胞质分裂的起始[2]，但目前尚不清楚 CPC 在后期特异性定位的机制。CPC 在这些部位的积累需要微管的存在和有丝分裂驱动蛋白样蛋白 2（MKLP2），MKLP2（KIF20A）是驱动蛋白-6 家族的成员[2-7]，这导致了 CPC 沿着微管由 MKLP2 运输的假说[3-5,7]。然而，MKLP2 马达结构域及其延伸的颈连接区的结构表明，这种驱动蛋白可能无法进行连续的运输[8,9]。此外，在非洲爪蟾卵提取物中的实验表明，CPC 可能由驱动蛋白-4（KIF4A）运输[10]。最后，CPC-MKLP2 复合物可能通过与肌动蛋白和肌球蛋白 II 的关联，而无需依赖驱动蛋白的活性，被直接募集到赤道皮层[4,8]。通过使用纯化蛋白的基于显微镜的测定，我们证明 MKLP2 是一种具有沿微管进行连续运动的正极定向的马达，能够在体外沿微管运输 CPC。在细胞中，通过表达 MKLP2 P 环突变体强烈抑制 MKLP2 依赖性 CPC 运动，会扰乱 CPC 在纺锤体中间区和赤道皮层的积累，而使用 Paprotrain 较弱地抑制 MKLP2 马达主要影响 CPC 定位到赤道皮层。我们的数据表明，CPC 对胞质分裂起始的控制需要其定向的 MKLP2 依赖性运输。
