Department of Genetics, University of Cambridge, Cambridge CB2 3EH , UK.
Open Biol. 2012 Feb;2(2):110032. doi: 10.1098/rsob.110032.
The formation of kinetochores shortly before each cell division is a prerequisite for proper chromosome segregation. The synchronous mitoses of Drosophila syncytial embryos have provided an ideal in vivo system to follow kinetochore assembly kinetics and so address the question of how kinetochore formation is regulated. We found that the nuclear exclusion of the Spc105/KNL1 protein during interphase prevents precocious assembly of the Mis12 complex. The nuclear import of Spc105 in early prophase and its immediate association with the Mis12 complex on centromeres are thus the first steps in kinetochore assembly. The cumulative kinetochore levels of Spc105 and Mis12 complex then determine the rate of Ndc80 complex recruitment commencing only after nuclear envelope breakdown. The carboxy-terminal part of Spc105 directs its nuclear import and is sufficient for the assembly of all core kinetochore components and CENP-C, when localized ectopically to centrosomes. Super-resolution microscopy shows that carboxy-terminus of Spc105 lies at the junction of the Mis12 and Ndc80 complexes on stretched kinetochores. Our study thus indicates that physical accessibility of kinetochore components plays a crucial role in the regulation of Drosophila kinetochore assembly and leads us to a model in which Spc105 is a licensing factor for its onset.
在每个细胞分裂之前，动粒的形成是正确染色体分离的前提条件。果蝇合胞胚胎的同步有丝分裂为跟踪动粒组装动力学提供了一个理想的体内系统，从而解决了动粒形成如何受到调节的问题。我们发现，Spc105/KNL1 蛋白在间期的核排斥阻止了 Mis12 复合物的过早组装。因此，Spc105 在早前期的核输入及其与着丝粒上的 Mis12 复合物的立即关联是动粒组装的第一步。随后，Spc105 和 Mis12 复合物的累积动粒水平决定了 Ndc80 复合物招募的速度，仅在核膜破裂后开始。Spc105 的羧基末端指导其核输入，并且当局部异位定位于中心体时，足以组装所有核心动粒成分和 CENP-C。超分辨率显微镜显示，Spc105 的羧基末端位于伸展的动粒上 Mis12 和 Ndc80 复合物的连接处。因此，我们的研究表明，动粒成分的物理可达性在果蝇动粒组装的调节中起着至关重要的作用，并使我们提出了一个模型，其中 Spc105 是其起始的许可因子。
