A molecular basis for the differential roles of Bub1 and BubR1 in the spindle assembly checkpoint.
A molecular basis for the differential roles of Bub1 and BubR1 in the spindle assembly checkpoint.
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DOI：
10.7554/elife.05269
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发表时间：
2015-01-22
期刊：
影响因子：
7.7
通讯作者：
Musacchio A
中科院分区：
文献类型：
--
作者：
Overlack K;Primorac I;Vleugel M;Krenn V;Maffini S;Hoffmann I;Kops GJ;Musacchio A
纺锤体组装检查点（SAC）在有丝分裂过程中监测和促进着丝粒微管附着。Bub 1和BubR 1是SAC组分，起源于祖先基因的复制。随后的子功能化建立了从属关系：Bub 1，首先招募到动粒，促进连续BubR 1招聘。由于Bub 1和BubR 1均与Bub 3（磷酸化动粒的靶向衔接子）异源二聚化，因此这种亚功能化的分子基础尚不清楚。我们证明，Bub 1，而不是BubR 1，增强Bub 3磷酸化kinetochores的结合。将Bub 1的短基序嫁接到BubR 1上促进BubR 1的Bub 1-独立动粒募集。这种功能获得性BubR 1突变体不能维持功能检查点。我们表明，动粒本地化的BubR 1依赖于直接异源二聚体与Bub 1在一个伪对称接口。这种假对称的相互作用支持了一个模板复制的关系，对着丝粒微管附着和SAC信号传导至关重要。我们的研究结果说明了基因复制和亚功能化如何塑造一个重要的分子网络的运作。DOI：http://dx.doi.org/10.7554/eLife.05269.001我们细胞内的遗传物质排列在称为染色体的结构中。在细胞分裂之前，它会精确复制所有的DNA。然后，遗传物质需要平均分裂，以便两个子细胞都有一套完整的染色体。当细胞准备分裂时，每个染色体由两个相同的姐妹染色单体组成，排列在一个被称为纺锤体的结构上，纺锤体是由称为微管的细丝组成的。细胞有一个复杂的安全机制，称为纺锤体组装检查点，以确保染色体在细胞分裂之前有时间在纺锤体上正确排列。一旦这个检查点得到满足，微管就会把姐妹染色单体拉开，这样每个子细胞就从每对姐妹染色单体中得到一个染色单体。微管通过一个称为动粒的蛋白质复合体附着在染色体上，该复合体聚集在每个姐妹染色单体上。纺锤体组装检查点监测着丝粒与微管的附着;两种称为Bub 1和BubR 1的蛋白质在这一过程中起着至关重要的作用。这些蛋白质与另一种称为Bub 3的蛋白质结合，Bub 3也是纺锤体组装检查点的一部分。虽然Bub 1和BubR 1非常相似，但它们似乎并没有发挥相同的作用，但它们差异的精确分子细节仍不清楚。在这项研究中，Overlack，Primorac等人研究了人类细胞中的Bub 1和BubR 1。实验表明，Bub 1可以在BubR 1不存在的情况下被招募到动粒，但BubR 1只有在Bub 1存在时才会移动到动粒。此外，BubR 1需要直接绑定到Bub 1才能移动到动粒。Overlack，Primorac等人还鉴定了Bub 1中与Bub 3结合的区域，并且该区域在BubR 1中显著不同。当Bub 1的这一区域被移植到BubR 1中时，即使在Bub 1不存在的情况下，所得的蛋白质也能够结合动粒。编码Bub 1和BubR 1蛋白质的基因源自在进化过程中复制的单个祖先基因。因此，Overlack，Primorac等人的研究结果表明，基因的复制如何通过创造在细胞中具有不同作用的产物而对细胞有益。DOI：http://dx.doi.org/10.7554/eLife.05269.002网站
The spindle assembly checkpoint (SAC) monitors and promotes kinetochore–microtubule attachment during mitosis. Bub1 and BubR1, SAC components, originated from duplication of an ancestor gene. Subsequent sub-functionalization established subordination: Bub1, recruited first to kinetochores, promotes successive BubR1 recruitment. Because both Bub1 and BubR1 hetero-dimerize with Bub3, a targeting adaptor for phosphorylated kinetochores, the molecular basis for such sub-functionalization is unclear. We demonstrate that Bub1, but not BubR1, enhances binding of Bub3 to phosphorylated kinetochores. Grafting a short motif of Bub1 onto BubR1 promotes Bub1-independent kinetochore recruitment of BubR1. This gain-of-function BubR1 mutant cannot sustain a functional checkpoint. We demonstrate that kinetochore localization of BubR1 relies on direct hetero-dimerization with Bub1 at a pseudo-symmetric interface. This pseudo-symmetric interaction underpins a template–copy relationship crucial for kinetochore–microtubule attachment and SAC signaling. Our results illustrate how gene duplication and sub-functionalization shape the workings of an essential molecular network. DOI: http://dx.doi.org/10.7554/eLife.05269.001 The genetic material within our cells is arranged in structures called chromosomes. Before a cell divides it makes an accurate copy of all of its DNA. The genetic material then needs to be equally split so that both daughter cells have a complete set of chromosomes. As the cell prepares to divide, each chromosome—consisting of two identical sister chromatids—lines up on a structure known as the spindle, which is made of filaments called microtubules. Cells have a sophisticated safety mechanism known as the spindle assembly checkpoint to ensure that chromosomes have time to correctly line up on the spindle before the cell can divide. Once this checkpoint is satisfied, the microtubules pull the sister chromatids apart so that each daughter cell receives one chromatid from each pair. The microtubules attach to the chromosomes through a large protein complex known as the kinetochore that assembles on each sister chromatid. The spindle assembly checkpoint monitors the attachment of the kinetochores to the microtubules; and two proteins, called Bub1 and BubR1, play an essential role in this process. These proteins bind to another protein called Bub3 that is also part of the spindle assembly checkpoint. Although Bub1 and BubR1 are very similar, they do not appear to perform the same roles, but the precise molecular details of their differences remain unclear. In this study, Overlack, Primorac et al. studied Bub1 and BubR1 in human cells. The experiments show that Bub1 can be recruited to kinetochores in the absence of BubR1, but BubR1 will only move to kinetochores when Bub1 is present. Furthermore, BubR1 needs to bind to Bub1 directly to move to the kinetochores. Overlack, Primorac et al. also identified a region in Bub1 that binds to Bub3, and which is considerably different in BubR1. When this region of Bub1 was grafted into BubR1, the resulting protein was able to bind kinetochores even in the absence of Bub1. The genes that encode the Bub1 and BubR1 proteins originate from a single ancestor gene that was duplicated during evolution. Therefore, the findings of Overlack, Primorac et al. show how the duplication of a gene can be beneficial for cells by creating products that have different roles in cells. DOI: http://dx.doi.org/10.7554/eLife.05269.002