Kinetochores form the link between chromosomes and microtubules of the mitotic spindle. The heterodecameric Dam1 complex (Dam1c) is a major component of the Saccharomyces cerevisiae outer kinetochore, assembling into 3 MDa-sized microtubule-embracing rings, but how ring assembly is specifically initiated in vivo remains to be understood. Here, we describe a molecular pathway that provides local control of ring assembly during the establishment of sister kinetochore bi-orientation. We show that Dam1c and the general microtubule plus end-associated protein (+TIP) Bim1/EB1 form a stable complex depending on a conserved motif in the Duo1 subunit of Dam1c. EM analyses reveal that Bim1 crosslinks protrusion domains of adjacent Dam1c heterodecamers and promotes the formation of oligomers with defined curvature. Disruption of the Dam1c-Bim1 interaction impairs kinetochore localization of Dam1c in metaphase and delays mitosis. Phosphorylation promotes Dam1c-Bim1 binding by relieving an intramolecular inhibition of the Dam1 C-terminus. In addition, Bim1 recruits Bik1/CLIP-170 to Dam1c and induces formation of full rings even in the absence of microtubules. Our data help to explain how new kinetochore end-on attachments are formed during the process of attachment error correction.

中文翻译：

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磷调节的 Bim1/EB1 相互作用在出芽的酵母外动粒处触发 Dam1c 环组装

动粒形成染色体和有丝分裂纺锤体微管之间的联系。异十二聚体 Dam1 复合物 (Dam1c) 是酿酒酵母外动粒的主要成分，组装成 3 个 MDa 大小的微管环，但环组装是如何在体内特异性启动的仍有待了解。在这里，我们描述了一种分子通路，该通路在建立姐妹动粒双向定向期间提供对环组装的局部控制。我们显示 Dam1c 和一般微管加末端相关蛋白 (+TIP) Bim1/EB1 形成稳定的复合物，这取决于 Dam1c 的 Duo1 亚基中的保守基序。EM 分析表明 Bim1 交联了相邻 Dam1c 异十二聚体的突起域，并促进了具有确定曲率的低聚物的形成。Dam1c-Bim1 相互作用的中断会损害 Dam1c 在中期的动粒定位并延迟有丝分裂。磷酸化通过解除 Dam1 C 端的分子内抑制来促进 Dam1c-Bim1 结合。此外，即使在没有微管的情况下，Bim1 也会将 Bik1/CLIP-170 招募到 Dam1c 并诱导完整环的形成。