An ATM- and ATR-dependent checkpoint inactivates spindle assembly by targeting CEP63
An ATM- and ATR-dependent checkpoint inactivates spindle assembly by targeting CEP63
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DOI：
10.1038/ncb1835
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发表时间：
2009-03-01
影响因子：
21.3
通讯作者：
Costanzo, Vincenzo
中科院分区：
文献类型：
--
作者：
Smith, Eloise;Dejsuphong, Donniphat;Costanzo, Vincenzo
在染色体断裂后，蛋白激酶ATM和ATR的激活阻止了DNA复制的开始和进入有丝分裂。然而，ATM和ATR活化在细胞有丝分裂过程中的作用尚不清楚。本文报道了DNA双链断裂（DSBs）诱导的ATM和ATR激活抑制了非洲爪蟾有丝分裂卵提取物和体细胞中中心体驱动的纺锤体组装，延缓了有丝分裂进程。利用cDNA表达文库筛选ATM和ATR底物，我们发现中心体蛋白CEP63是正常纺锤体组装所需的ATM和ATR靶点。ATM和ATR使Xenopus CEP63 （XCEP63）的Ser 560磷酸化，并促进其从中心体脱位。抑制ATM和ATR活性或使XCEP63 Ser 560突变为Ala可防止纺锤体组装缺陷。在禽类DT40细胞中，CEP63基因的失活破坏了纺锤体组装，并阻止了ATM和atr对有丝分裂的依赖作用。这些数据表明，ATM和ATR通过靶向CEP63和中心体依赖的纺锤体组装来控制脊椎动物细胞的有丝分裂事件。
Activation of the protein kinases ATM and ATR following chromosomal breakage prevents initiation of DNA replication and entry into mitosis. However, the effects of ATM and ATR activation in cells already progressing through mitosis are poorly understood. Here we report that ATM and ATR activation induced by DNA double-strand breaks (DSBs) inhibits centrosome-driven spindle assembly in Xenopus laevis mitotic egg extract and somatic cells, delaying mitotic progression. Using a cDNA expression library to screen for ATM and ATR substrates, we identified centrosomal protein CEP63 as an ATM and ATR target required for normal spindle assembly. ATM and ATR phosphorylate Xenopus CEP63 (XCEP63) on Ser 560 and promote its delocalization from the centrosome. Suppression of ATM and ATR activity or mutation of XCEP63 Ser 560 to Ala prevented spindle assembly defects. Consistently, inactivation of the CEP63 gene in avian DT40 cells impaired spindle assembly and prevented ATM- and ATR-dependent effects on mitosis. These data indicate that ATM and ATR control mitotic events in vertebrate cells by targeting CEP63 and centrosome dependent spindle assembly.