Structural analysis of the role of TPX2 in branching microtubule nucleation.
Structural analysis of the role of TPX2 in branching microtubule nucleation.
复制标题
DOI：
10.1083/jcb.201607060
复制
发表时间：
2017-04-03
期刊：
影响因子：
--
通讯作者：
Petry S
中科院分区：
文献类型：
--
作者：
Alfaro-Aco R;Thawani A;Petry S
TPX 2是有丝分裂中微管成核所必需的，但其功能机制尚不清楚。Alfaro-Aco等人分析了TPX 2活性所必需的结构域，并确定了分支微管成核所需的最小区域。有丝分裂纺锤体由微管（MT）组成，微管由γ-微管蛋白环复合物（γ-TuRC）成核。γ-TuRC如何在正确的时间和位置被激活仍然是一个谜。最近，它被发现，从有丝分裂纺锤体内的预先存在的MT成核，这需要蛋白TPX 2，但TPX 2的作用机制的基础是未知的。在这里，我们研究TPX 2在分支MT成核中的作用。我们建立了非洲爪蟾TPX 2的结构域组织，并定义了刺激分支MT成核的最小TPX 2版本，我们发现这与TPX 2在体外使MT成核的能力无关。TPX 2的几个结构域有助于其MT结合和捆绑活性。然而，TPX 2诱导分支MT成核所必需的性质包含在新鉴定的γ-TuRC成核活化剂基序内。功能分离突变使TPX 2与γ-TuRC的结合保持完整，而分支MT成核被消除，这表明TPX 2可能激活γ-TuRC以促进分支MT成核。
TPX2 is required for microtubule nucleation in mitosis, but the mechanism underlying its function is unclear. Alfaro-Aco et al. analyze the domains of TPX2 necessary for its activity and identify the minimal region required for branching microtubule nucleation. The mitotic spindle consists of microtubules (MTs), which are nucleated by the γ-tubulin ring complex (γ-TuRC). How the γ-TuRC gets activated at the right time and location remains elusive. Recently, it was uncovered that MTs nucleate from preexisting MTs within the mitotic spindle, which requires the protein TPX2, but the mechanism basis for TPX2 action is unknown. Here, we investigate the role of TPX2 in branching MT nucleation. We establish the domain organization of Xenopus laevis TPX2 and define the minimal TPX2 version that stimulates branching MT nucleation, which we find is unrelated to TPX2’s ability to nucleate MTs in vitro. Several domains of TPX2 contribute to its MT-binding and bundling activities. However, the property necessary for TPX2 to induce branching MT nucleation is contained within newly identified γ-TuRC nucleation activator motifs. Separation-of-function mutations leave the binding of TPX2 to γ-TuRC intact, whereas branching MT nucleation is abolished, suggesting that TPX2 may activate γ-TuRC to promote branching MT nucleation.