Byrnes Amy E, Slep Kevin C
Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599.
Program in Molecular and Cellular Biophysics, University of North Carolina, Chapel Hill, NC 27599.
J Cell Biol. 2017 Jun 5;216(6):1641-1657. doi: 10.1083/jcb.201610090. Epub 2017 May 16.
XMAP215, CLASP, and Crescerin use arrayed tubulin-binding tumor overexpressed gene (TOG) domains to modulate microtubule dynamics. We hypothesized that TOGs have distinct architectures and tubulin-binding properties that underlie each family's ability to promote microtubule polymerization or pause. As a model, we investigated the pentameric TOG array of a XMAP215 member, Msps. We found that Msps TOGs have distinct architectures that bind either free or polymerized tubulin, and that a polarized array drives microtubule polymerization. An engineered TOG1-2-5 array fully supported Msps-dependent microtubule polymerase activity. Requisite for this activity was a TOG5-specific N-terminal HEAT repeat that engaged microtubule lattice-incorporated tubulin. TOG5-microtubule binding maintained mitotic spindle formation as deleting or mutating TOG5 compromised spindle architecture and increased the mitotic index. Mad2 knockdown released the spindle assembly checkpoint triggered when TOG5-microtubule binding was compromised, indicating that TOG5 is essential for spindle function. Our results reveal a TOG5-specific role in mitotic fidelity and support our hypothesis that architecturally distinct TOGs arranged in a sequence-specific order underlie TOG array microtubule regulator activity.
XMAP215、CLASP和Crescerin利用排列的微管蛋白结合肿瘤过表达基因（TOG）结构域来调节微管动力学。我们推测，TOG具有独特的结构和微管蛋白结合特性，这是每个家族促进微管聚合或暂停能力的基础。作为一个模型，我们研究了XMAP215成员Msps的五聚体TOG阵列。我们发现，Msps TOG具有独特的结构，可结合游离或聚合的微管蛋白，并且极化阵列驱动微管聚合。一个工程化的TOG1-2-5阵列完全支持依赖Msps的微管聚合酶活性。这种活性所必需的是一个TOG5特异性的N端HEAT重复序列，它与微管晶格结合的微管蛋白结合。TOG5与微管的结合维持了有丝分裂纺锤体的形成，因为删除或突变TOG5会损害纺锤体结构并增加有丝分裂指数。当TOG5与微管的结合受到损害时，Mad2基因敲低可释放触发的纺锤体组装检查点，这表明TOG5对纺锤体功能至关重要。我们的结果揭示了TOG5在有丝分裂保真度中的特定作用，并支持我们的假设，即按序列特异性顺序排列的结构上不同的TOG是TOG阵列微管调节活性的基础。
