Kuhn Jonathan, Dumont Sophie
Tetrad Graduate Program, University of California, San Francisco, San Francisco, CA.
Department of Cell and Tissue Biology, University of California, San Francisco, San Francisco, CA.
J Cell Biol. 2017 Jun 5;216(6):1533-1542. doi: 10.1083/jcb.201611104. Epub 2017 May 23.
To ensure accurate chromosome segregation, the spindle assembly checkpoint (SAC) prevents anaphase until all kinetochores attach to the spindle. What signals the SAC monitors remains unclear. We do not know the contributions of different microtubule attachment features or tension from biorientation to SAC satisfaction nor how these possible cues change during attachment. In this study, we quantify concurrent Mad1 intensity and report on SAC silencing, real-time attachment geometry, occupancy, and tension at individual mammalian kinetochores. We show that Mad1 loss from the kinetochore is switch-like with robust kinetics and that tension across sister kinetochores is established just before Mad1 loss events at the first sister. We demonstrate that CenpE-mediated lateral attachment of the second sister can persistently generate this metaphase-like tension before biorientation, likely stabilizing sister end-on attachment, yet cannot induce Mad1 loss from that kinetochore. Instead, Mad1 loss begins after several end-on microtubules attach. Thus, end-on attachment provides geometry-specific molecular cues or force on specific kinetochore linkages that other attachment geometries cannot provide.
为确保染色体准确分离，纺锤体组装检查点（SAC）会阻止细胞进入后期，直到所有动粒都附着到纺锤体上。SAC监测的信号尚不清楚。我们不知道不同微管附着特征或双定向产生的张力对SAC满足的贡献，也不知道这些可能的线索在附着过程中是如何变化的。在这项研究中，我们量化了同时存在的Mad1强度，并报告了单个哺乳动物动粒处SAC沉默、实时附着几何形状、占有率和张力。我们表明，动粒上Mad1的丧失具有类似开关的特性，动力学稳健，并且在第一个姐妹动粒处Mad1丧失事件之前，姐妹动粒之间的张力就已建立。我们证明，CenpE介导的第二个姐妹动粒的侧向附着可以在双定向之前持续产生这种类似中期的张力，这可能稳定了姐妹端对端附着，但不能诱导该动粒上的Mad1丧失。相反，在几个端对端微管附着后，Mad1才开始丧失。因此，端对端附着在特定动粒连接上提供了其他附着几何形状无法提供的特定几何形状的分子线索或力。
