DNA Damage-Induced Accumulation of Centrosomal Chk1 Contributes to its Checkpoint Function
DNA Damage-Induced Accumulation of Centrosomal Chk1 Contributes to its Checkpoint Function
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DOI：
10.4161/cc.6.20.4810
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发表时间：
2007-10
期刊：
影响因子：
4.3
通讯作者：
H. Löffler;T. Bochtler;B. Fritz;B. Tews;A. Ho;J. Lukas;J. Bartek;A. Krämer
中科院分区：
文献类型：
--
作者：
H. Löffler;T. Bochtler;B. Fritz;B. Tews;A. Ho;J. Lukas;J. Bartek;A. Krämer
检查点激酶Chk 1是响应DNA损伤的ATR和ATM依赖性信号传导的既定转导子。除了它的核定位，Chk 1定位于间期中心体，从而负调控进入有丝分裂，防止过早激活细胞周期蛋白B-Cdk 1在未受干扰的细胞周期。在这里，我们表明，紫外线照射或羟基脲治疗引起的DNA损伤导致中心体积累的内源性Chk 1在正常人BJ成纤维细胞和ATR或ATM缺陷的成纤维细胞。咖啡因对ATR/ATM的化学抑制导致中心体Chk 1沉积增强，与核Chk 1耗竭相关。与正常或ATM缺陷型成纤维细胞相反，遗传ATR缺陷型Seckel成纤维细胞显示出可检测的Chk 1的组成性中心体积累，即使在没有外源性损伤的情况下。DNA损伤后，Chk 1的中心体部分被发现在ATR/ATM磷酸化位点磷酸化。强迫固定激酶失活，但不是野生型Chk 1的中心体导致G2/M检查点缺陷。最后，DNA损伤和强迫Chk 1的中心体表达在没有遗传毒性处理的情况下，诱导中心体扩增的细胞亚群，这一现象可以抑制ATM/ATR介导的信号转导的抑制。两者合计，我们的研究结果表明，磷酸化Chk 1在中心体的积累构成了一个额外的元素在DNA损伤反应。中心体Chk 1诱导G2/M细胞周期停滞，并可能引起中心体扩增，后者可能提供一个备份机制，用于消除细胞与受损的DNA损伤检查点在细胞周期的早期操作。
The checkpoint kinase Chk1 is an established transducer of ATR- and ATM-dependent signalling in response to DNA damage. In addition to its nuclear localization, Chk1 localizes to interphase centrosomes and thereby negatively regulates entry into mitosis by preventing premature activation of cyclin B-Cdk1 during unperturbed cell cycles. Here, we demonstrate that DNA damage caused by ultraviolet irradiation or hydroxyurea treatment leads to centrosomal accumulation of endogenous Chk1 in normal human BJ fibroblasts and in ATR- or ATM-deficient fibroblasts. Chemical inhibition of ATR/ATM by caffeine led to enhanced centrosomal Chk1 deposition associated with nuclear Chk1 depletion. In contrast to normal or ATM-deficient fibroblasts, genetically ATR-deficient Seckel-fibroblasts showed detectable constitutive centrosomal accumulation of Chk1 even in the absence of exogenous insults. After DNA damage, the centrosomal fraction of Chk1 was found to be phosphorylated at ATR/ATM phosphorylation sites. Forced immobilization of kinase-inactive but not wild-type Chk1 to centrosomes resulted in a G2/M checkpoint defect. Finally, both DNA damage, and forced centrosomal expression of Chk1 in the absence of genotoxic treatments, induced centrosome amplification in a subset of cells, a phenomenon which could be suppressed by inhibition of ATM/ATR-mediated signaling. Taken together, our results suggest that accumulation of phosphorylated Chk1 at centrosomes constitutes an additional element in the DNA damage response. Centrosomal Chk1 induces G2/M cell cycle arrest and may evoke centrosome amplification, the latter possibly providing a backup mechanism for elimination of cells with impaired DNA damage checkpoints operating earlier during the cell cycle.