Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84112, USA.
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT 84112, USA.
Curr Biol. 2024 Jun 3;34(11):2294-2307.e4. doi: 10.1016/j.cub.2024.04.054. Epub 2024 May 21.
Accurate chromosome segregation relies on kinetochores carrying out multiple functions, including establishing and maintaining microtubule attachments, forming precise bi-oriented attachments between sister chromatids, and activating the spindle assembly checkpoint. Central to these processes is the highly conserved Ndc80 complex. This kinetochore subcomplex interacts directly with microtubules but also serves as a critical platform for recruiting kinetochore-associated factors and as a key substrate for error correction kinases. The precise manner in which these kinetochore factors interact and regulate each other's function remains unknown, considerably hindering our understanding of how Ndc80 complex-dependent processes function together to orchestrate accurate chromosome segregation. Here, we aimed to uncover the role of Nuf2's CH domain, a component of the Ndc80 complex, in ensuring these processes. Through extensive mutational analysis, we identified a conserved interaction domain composed of two segments in Nuf2's CH domain that form the binding site for Mps1 within the yeast Ndc80 complex. Interestingly, this site also associates with the Dam1 complex, suggesting Mps1 recruitment may be subject to regulation by competitive binding with other factors. Mutants disrupting this "interaction hub" exhibit defects in spindle assembly checkpoint function and severe chromosome segregation errors. Significantly, specifically restoring Mps1-Ndc80 complex association rescues these defects. Our findings shed light on the intricate regulation of Ndc80 complex-dependent functions and highlight the essential role of Mps1 in kinetochore bi-orientation and accurate chromosome segregation.
准确的染色体分离依赖于动粒执行多种功能，包括建立和维持微管连接、在姐妹染色单体之间形成精确的双取向连接，以及激活纺锤体组装检查点。这些过程的核心是高度保守的 Ndc80 复合物。这个动粒亚基复合物与微管直接相互作用，但也作为招募动粒相关因子的关键平台，并作为错误修正激酶的关键底物。这些动粒因子相互作用并调节彼此功能的确切方式仍然未知，这极大地阻碍了我们对 Ndc80 复合物依赖性过程如何协同作用以协调准确的染色体分离的理解。在这里，我们旨在揭示 Nuf2 的 CH 结构域（Ndc80 复合物的一个组成部分）在确保这些过程中的作用。通过广泛的突变分析，我们确定了一个保守的相互作用结构域，该结构域由 Nuf2 的 CH 结构域中的两个片段组成，形成了酵母 Ndc80 复合物中 Mps1 的结合位点。有趣的是，这个位点也与 Dam1 复合物相关联，表明 Mps1 的募集可能受到与其他因子竞争结合的调节。破坏这个“相互作用中心”的突变体表现出纺锤体组装检查点功能缺陷和严重的染色体分离错误。重要的是，特异性恢复 Mps1-Ndc80 复合物的关联可以挽救这些缺陷。我们的发现阐明了 Ndc80 复合物依赖性功能的复杂调节，并强调了 Mps1 在动粒双取向和准确染色体分离中的重要作用。
