Perez-Bertoldi Juan M, Zhao Yuanchang, Thawani Akanksha, Yildiz Ahmet, Nogales Eva
Biophysics Graduate Group, University of California, Berkeley, CA, USA.
Physics Department, University of California, Berkeley, CA, USA.
Res Sq. 2024 May 29:rs.3.rs-4249615. doi: 10.21203/rs.3.rs-4249615/v1.
During mitosis, microtubule dynamics are regulated to ensure proper alignment and segregation of chromosomes. The dynamics of kinetochore-attached microtubules are regulated by hepatoma-upregulated protein (HURP) and the mitotic kinesin-8 Kif18A, but the underlying mechanism remains elusive. Using single-molecule imaging , we demonstrate that Kif18A motility is regulated by HURP. While sparse decoration of HURP activates the motor, higher concentrations hinder processive motility. To shed light on this behavior, we determined the binding mode of HURP to microtubules using Cryo-EM. The structure reveals that one HURP motif spans laterally across β-tubulin, while a second motif binds between adjacent protofilaments. HURP partially overlaps with the microtubule-binding site of the Kif18A motor domain, indicating that excess HURP inhibits Kif18A motility by steric exclusion. We also observed that HURP and Kif18A function together to suppress dynamics of the microtubule plus-end, providing a mechanistic basis for how they collectively serve in spindle length control.
在有丝分裂过程中，微管动力学受到调控以确保染色体的正确排列和分离。着丝粒附着微管的动力学受肝癌上调蛋白（HURP）和有丝分裂驱动蛋白8 Kif18A调控，但其潜在机制仍不清楚。通过单分子成像，我们证明Kif18A的运动受HURP调控。当HURP稀疏修饰时会激活该驱动蛋白，而较高浓度则会阻碍其持续运动。为了解释这种行为，我们使用冷冻电镜确定了HURP与微管的结合模式。该结构显示，一个HURP基序横向跨越β-微管蛋白，而另一个基序则结合在相邻原纤维之间。HURP与Kif18A驱动结构域的微管结合位点部分重叠，这表明过量的HURP通过空间位阻抑制Kif18A的运动。我们还观察到，HURP和Kif18A共同作用以抑制微管正端的动力学，为它们如何共同参与纺锤体长度控制提供了一个机制基础。
