Johnson Victoria L, Scott Maria I F, Holt Sarah V, Hussein Deema, Taylor Stephen S
School of Biological Sciences, University of Manchester, 2.205 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
J Cell Sci. 2004 Mar 15;117(Pt 8):1577-89. doi: 10.1242/jcs.01006.
During mitosis, the recruitment of spindle-checkpoint-associated proteins to the kinetochore occurs in a defined order. The protein kinase Bub1 localizes to the kinetochore very early during mitosis, followed by Cenp-F, BubR1, Cenp-E and finally Mad2. Using RNA interference, we have investigated whether this order of binding reflects a level of dependency in human somatic cells. Specifically, we show that Bub1 plays a key role in the assembly of checkpoint proteins at the kinetochore, being required for the subsequent localization of Cenp-F, BubR1, Cenp-E and Mad2. In contrast to studies in Xenopus, we also show that BubR1 is not required for kinetochore localization of Bub1. Repression of Bub1 increases the number of cells with lagging chromosomes at metaphase, suggesting that Bub1 plays a role in chromosome congression. However, repression of Bub1 does not appear to compromise spindle checkpoint function either during normal mitosis or in response to spindle damage. This raises the possibility that, in the absence of Bub1, other mechanisms contribute to spindle checkpoint function.
在有丝分裂过程中，纺锤体检查点相关蛋白向动粒的募集按特定顺序发生。蛋白激酶Bub1在有丝分裂早期就定位于动粒，随后是Cenp - F、BubR1、Cenp - E，最后是Mad2。我们利用RNA干扰研究了这种结合顺序在人类体细胞中是否反映了一种依赖水平。具体而言，我们发现Bub1在动粒处检查点蛋白的组装中起关键作用，是Cenp - F、BubR1、Cenp - E和Mad2随后定位所必需的。与非洲爪蟾中的研究不同，我们还发现BubR1对于Bub1在动粒处的定位不是必需的。抑制Bub1会增加中期有落后染色体的细胞数量，这表明Bub1在染色体排列中起作用。然而，在正常有丝分裂期间或对纺锤体损伤作出反应时，抑制Bub1似乎并未损害纺锤体检查点功能。这就提出了一种可能性，即在没有Bub1的情况下，其他机制有助于纺锤体检查点功能。
