Autocatalytic microtubule nucleation determines the size and mass of Xenopus laevis egg extract spindles.
Autocatalytic microtubule nucleation determines the size and mass of Xenopus laevis egg extract spindles.
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DOI：
10.7554/elife.31149
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发表时间：
2018-01-11
期刊：
影响因子：
7.7
通讯作者：
Brugués J
中科院分区：
文献类型：
--
作者：
Decker F;Oriola D;Dalton B;Brugués J
大小和生长的调节是生物学中的一个基本问题。一个突出的例子是有丝分裂纺锤体的形成，其中染色体周围的蛋白质浓度梯度被认为通过控制微管成核来调节纺锤体的生长。以前的证据表明，微管成核整个纺锤体结构。然而，微管成核及其空间调节的机制仍不清楚。在这里，我们开发了一种基于激光烧蚀直接探测非洲爪蟾卵提取物中微管成核事件的测定方法。将这种方法与理论和定量显微镜相结合，我们表明，纺锤体的大小是由微管的自催化生长控制的，由微管刺激的微管成核驱动。该成核系统的自催化活性在空间上受活性微管成核剂的限制量的调节，所述活性微管成核剂随着与染色体的距离而减少。即使资源不受限制，这种机制也为主轴大小提供了上限。当细胞分裂时，它们首先需要创建一个遗传物质的副本，然后将其均匀分布在子细胞之间。这是由一种称为有丝分裂纺锤体的蛋白质复合体完成的，纺锤体将携带基因形式遗传物质的染色体分开。有丝分裂纺锤体主要由微管蛋白组成，微管蛋白排列成中空的索状细丝，称为微管。微管是动态结构，可以通过添加或去除微管蛋白来生长或收缩。与纺锤体不同，纺锤体可以“存活”长达数小时，微管只能存活约20秒，需要不断更新以维持结构。为了成功地分配遗传物质，纺锤体需要有合适的长度。先前的研究表明，纺锤体的长度适应细胞的大小-细胞越大，纺锤体越大。然而，在非常大的细胞中，例如胚胎细胞第一次分裂时，纺锤体有一个上限。据认为，染色体产生的特定蛋白质有助于调节新微管的形成，从而也影响纺锤体的大小。然而，到目前为止，尚不清楚他们是如何做到这一点的，以及这是否也设定了上限。为了进一步研究微管更新及其与纺锤体大小的关系，德克尔等人使用了非洲爪蛙卵细胞提取物中组装的纺锤体。结果表明，新的微管从现有的微管上生长出来，就像树的新分支一样。当建立的微管与染色体发射的特定分子相互作用时，就会发生分支，这些分子的浓度随着与染色体的距离而降低。这种浓度梯度调节了在离染色体不同距离处生长的微管数量，从而确定了纺锤体的大小。这些发现有助于我们理解生物结构是如何由动态和短暂的成分组成的。此外，更好地了解有丝分裂纺锤体如何生长可能最终有助于开发癌症和其他疾病的新疗法。
Regulation of size and growth is a fundamental problem in biology. A prominent example is the formation of the mitotic spindle, where protein concentration gradients around chromosomes are thought to regulate spindle growth by controlling microtubule nucleation. Previous evidence suggests that microtubules nucleate throughout the spindle structure. However, the mechanisms underlying microtubule nucleation and its spatial regulation are still unclear. Here, we developed an assay based on laser ablation to directly probe microtubule nucleation events in Xenopus laevis egg extracts. Combining this method with theory and quantitative microscopy, we show that the size of a spindle is controlled by autocatalytic growth of microtubules, driven by microtubule-stimulated microtubule nucleation. The autocatalytic activity of this nucleation system is spatially regulated by the limiting amounts of active microtubule nucleators, which decrease with distance from the chromosomes. This mechanism provides an upper limit to spindle size even when resources are not limiting. When cells divide, they first need to create a copy of their genetic material, which they then evenly distribute between their daughter cells. This is done by a complex of proteins known as the mitotic spindle, which divides the chromosomes that carry the genetic material in the form of genes. The mitotic spindle is mainly made of tubulin proteins that are arranged to form hollow cable-like filaments, called the microtubules. Microtubules are dynamic structures that can grow or shrink by adding or removing tubulin proteins. Unlike the spindle, which can ‘live’ up to hours, the microtubules only live for about 20 seconds and need to be constantly renewed to maintain the structure. To successfully distribute the genetic material, spindles need to have the right length. Previous research has shown that the length of a spindle adapts to the size of a cell – the larger the cells, the larger the spindles. However, in very large cells, such as the cells of an embryo when they first divide, spindles have an upper size limit. It is thought that specific proteins produced by the chromosomes help to regulate the formation of new microtubules and thereby also influence the size of the spindle. However, until now it was not clear how exactly they do so and if this also sets the upper size limit. To further investigate microtubule renewal and its relation to spindle size, Decker et al. used spindles assembled in cell extracts from the eggs of the African clawed frog. The results showed that the new microtubules grow off the existing ones, like new branches of a tree. The branching happens when the established microtubules interact with specific molecules emitted by the chromosomes, and the concentration of these molecules decreases with distance from the chromosomes. This concentration gradient regulates how many microtubules grow at different distances from the chromosomes and so sets the size of spindles. These findings help us to understand how biological structures are built out of dynamic and short-lived components. Moreover, a better understanding of how mitotic spindles grow might eventually help to develop new treatments for cancer and other diseases.