Sanchez-Perez Isabel, Renwick Steven J, Crawley Karen, Karig Inga, Buck Vicky, Meadows John C, Franco-Sanchez Alejandro, Fleig Ursula, Toda Takashi, Millar Jonathan B A
Division of Yeast Genetics, National Institute for Medical Research, London, UK.
EMBO J. 2005 Aug 17;24(16):2931-43. doi: 10.1038/sj.emboj.7600761. Epub 2005 Aug 4.
We identified a truncated allele of dam1 as a multicopy suppressor of the sensitivity of cdc13-117 (cyclin B) and mal3-1 (EB-1) cells to thiabendazole, a microtubule poison. We find that Dam1 binds to the plus end of spindle microtubules and kinetochores as cells enter mitosis and this is dependent on other components of the fission yeast DASH complex, including Ask1, Duo1, Spc34 and Dad1. By contrast, Dad1 remains bound to kinetochores throughout the cell cycle and its association is dependent on the Mis6 and Mal2, but not Mis12, Nuf2 or Cnp1, kinetochore proteins. In cells lacking Dam1, or other components of the DASH complex, anaphase is delayed due to activation of the spindle assembly checkpoint and lagging sister chromatids are frequently observed and occasionally sister chromatid pairs segregate to the same spindle pole. We find that the mitotic centromere-associated Klp5/Klp6 kinesin complex is essential in cells lacking components of the DASH complex. Cells lacking both Dam1 and Klp5 undergo a first cell cycle arrest in mitosis due to a failure to establish bipolar chromosome attachment.
我们鉴定出dam1的一个截短等位基因，它是cdc13-117（细胞周期蛋白B）和mal3-1（EB-1）细胞对微管毒物噻苯咪唑敏感性的多拷贝抑制因子。我们发现，当细胞进入有丝分裂时，Dam1与纺锤体微管的正端和动粒结合，这依赖于裂殖酵母DASH复合体的其他组分，包括Ask1、Duo1、Spc34和Dad1。相比之下，Dad1在整个细胞周期中都与动粒结合，其结合依赖于动粒蛋白Mis6和Mal2，但不依赖于Mis12、Nuf2或Cnp1。在缺乏Dam1或DASH复合体其他组分的细胞中，由于纺锤体组装检查点的激活，后期延迟，经常观察到滞后的姐妹染色单体，偶尔姐妹染色单体对会分离到同一纺锤极。我们发现，有丝分裂着丝粒相关的Klp5/Klp6驱动蛋白复合体在缺乏DASH复合体组分的细胞中至关重要。同时缺乏Dam1和Klp5的细胞由于未能建立双极染色体附着而在有丝分裂中经历第一次细胞周期阻滞。
