Dual mechanisms regulate the recruitment of spindle assembly checkpoint proteins to the budding yeast kinetochore.
Dual mechanisms regulate the recruitment of spindle assembly checkpoint proteins to the budding yeast kinetochore.
复制标题
双重机制调节纺锤体组装检查点蛋白募集到萌芽的酵母菌。
DOI：
10.1091/mbc.e16-01-0007
复制
发表时间：
2016-11-07
影响因子：
3.3
通讯作者：
Joglekar AP
中科院分区：
文献类型：
--
作者：
Aravamudhan P;Chen R;Roy B;Sim J;Joglekar AP
纺锤体组装检查点（SAC）蛋白质的招募由动粒的定量知识是必不可少的了解调节蛋白质招募的机制，因此SAC的强度。在这里，这种招聘是定量的，并确定了新的机制，强烈调节SAC蛋白招聘的动粒。纺锤体组装检查点（SAC）蛋白被未附着的动粒募集导致SAC活化。这种募集是由Mps 1激酶许可的，Mps 1激酶在其六个MELT重复中的一个或多个处磷酸化动粒蛋白Spc 105。Spc 105然后招募Bub 3-Bub 1和Mad 1-Mad 2复合物，其产生阻止细胞分裂的抑制信号。这种信号的强度部分取决于Spc 105招募的Bub 3-Bub 1和Mad 1-Mad 2分子的数量。因此，这种募集的调节将影响SAC信号传导。为了理解这种调节，我们建立了描述Bub 3-Bub 1和Mad 1-Mad 2与芽殖酵母动粒结合的生理结合曲线。我们发现两者的结合遵循质量作用定律。Mps 1可能磷酸化Spc 105的所有六个MELT重复。然而，有两种机制阻止Spc 105招募6个Bub 3-Bub 1分子：Bub 1丰度低和阻碍一个以上的Bub 3-Bub 1分子与同一Spc 105结合。令人惊讶的是，动粒为每个Bub 3-Bub 1分子招募两个Mad 1-Mad 2异四聚体。最后，每个Spc 105至少需要三个MELT重复序列才能进行准确的染色体分离。这些数据表明，kinetochore的内在和外在机制影响SAC信号的生理操作，有可能最大限度地提高染色体分离的准确性。
Quantitative knowledge of the recruitment of spindle assembly checkpoint (SAC) proteins by the kinetochore is essential to understanding the mechanisms that regulate protein recruitment and hence the strength of the SAC. Here this recruitment is quantified, and novel mechanisms are identified that strongly modulate SAC protein recruitment by the kinetochore. Recruitment of spindle assembly checkpoint (SAC) proteins by an unattached kinetochore leads to SAC activation. This recruitment is licensed by the Mps1 kinase, which phosphorylates the kinetochore protein Spc105 at one or more of its six MELT repeats. Spc105 then recruits the Bub3-Bub1 and Mad1-Mad2 complexes, which produce the inhibitory signal that arrests cell division. The strength of this signal depends, in part, on the number of Bub3-Bub1 and Mad1-Mad2 molecules that Spc105 recruits. Therefore regulation of this recruitment will influence SAC signaling. To understand this regulation, we established the physiological binding curves that describe the binding of Bub3-Bub1 and Mad1-Mad2 to the budding yeast kinetochore. We find that the binding of both follows the mass action law. Mps1 likely phosphorylates all six MELT repeats of Spc105. However, two mechanisms prevent Spc105 from recruiting six Bub3-Bub1 molecules: low Bub1 abundance and hindrance in the binding of more than one Bub3-Bub1 molecule to the same Spc105. Surprisingly, the kinetochore recruits two Mad1-Mad2 heterotetramers for every Bub3-Bub1 molecule. Finally, at least three MELT repeats per Spc105 are needed for accurate chromosome segregation. These data reveal that kinetochore-intrinsic and -extrinsic mechanisms influence the physiological operation of SAC signaling, potentially to maximize chromosome segregation accuracy.