Cep57是一个中心体蛋白，我们前期的结果显示该蛋白与中心体结构的稳定性相关，并且是胞质分裂过程所必需。此外，该蛋白还定位于动粒，与动粒蛋白Mis12结合，敲低Cep57导致染色体分离异常；我们还发现一个与Cep57结合的未知功能蛋白,命名为AK57，也定位于动粒，进一步的结果显示Cep57，AK57和Mis12这三个动粒定位蛋白质都能被Nek2A磷酸化。在本项目中，一方面将研究Cep57在动粒上是否通过介导Mad1和Mad2的动粒定位从而促进纺锤体装配检验点激活；另一方面,将研究AK57这一新蛋白是否在动粒上参与微管的捕捉、动粒微管的连接及染色体的运动；此外还将探究Nek2A磷酸化Cep57、AK57和Mis12对这三个蛋白的功能发挥和整个有丝分裂过程的调控机制。希望通过本项目的研究对有丝分裂的过程机制有更深入全面的了解，并为探讨多种染色体非整倍性疾病和恶性肿瘤的发病机理提供分子基础。

Cep57 is a centrosome-localized protein. Our previous results have shown that Cep57 is required for centrosome integrity and completion of cytokinesis. Our further study indicated that Cep57, as a kinetochore component, localized to kinetochores and interacted with kinetochore protein Mis12. Depletion of Cep57 led to chromosome missegregation. We also found that a function-unknown protein, we named it as AK57, not only interacted with Cep57, but also localized to kinetochores. Cep57, AK57 and Mis12, as kinetochore localized proteins, could be phosphorylated by mitotic kinase Nek2A. In this project, we would like to investigate whether Cep57 recruits Mad1 and Mad2 to the kinetochores to promote the activation of spindle assembly checkpoint. On the other hand, we will study whether AK57, as a novel kinesin-like motor domain containing protein, is involved in the microtubule capture of kinetochore, kinetochore-microtubule attachment and chromosome movement. Finally, we will determine the mechanism of Nek2A in regulating the function of Cep57, AK57 and Mis12, and even the process of mitosis. We hope this project will help us to further understand the mechanism of mitosis, and provide new clues for investigation of the pathogenesis of chromosome aneuploidy diseases and cancer.

我们在前期工作的基础上分析了Cep57 在染色体动粒结构域的定位，及其激活纺锤体组装检验点的机制，发现在有丝分裂前期和中期，Cep57通过其N端结构域与动粒蛋白Mis12相互作用。在动粒定位的Cep57进一步招募纺锤体组装检验点蛋白Mad1/Mad2，从而激活纺锤体组装检验点，保障染色体的正确列队。随后，Cep57通过在动粒微管与Mad1之间的竞争性结合调控Mad1从结合微管的动粒解离，从而保证纺锤体组装检验点的及时失活，使有丝分裂能正常进行。磷酸化是调控蛋白质之间相互作用的重要方式，我们分析了Cep57、Mis12 和一个功能未知的与Cep57相互作用的动粒定位蛋白AK57的磷酸化状态，并证明这三个动粒蛋白都是Nek2A的底物。进一步的结果显示在有丝分裂的前中期Nek2A使Mis12的S177位点磷酸化。而在有丝分裂中期，Nek2A与AK57相互作用，并将其S203和T371位点磷酸化，推测该蛋白的磷酸化修饰与其从动粒部位解离，以及细胞周期向后期推进相关。