The DNA damage-dependent intra-S phase checkpoint is regulated by parallel pathways
The DNA damage-dependent intra-S phase checkpoint is regulated by parallel pathways
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DOI：
10.1038/ng845
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发表时间：
2002-03-01
期刊：
影响因子：
30.8
通讯作者：
Bartek, J
中科院分区：
文献类型：
--
作者：
Falck, J;Petrini, JHJ;Bartek, J
为了保持遗传完整性，暴露于电离辐射下的哺乳动物细胞激活ATM激酶，从而启动包括s期检查点途径在内的复杂反应，以延迟DNA复制（1,2）。ATM或其底物Nbs1或Chk2中的缺陷（参考文献3），与Nbs1相互作用的Mre11蛋白(4)，或Chk2调控的Cdc25A-Cdk2级联都会导致耐辐射DNA合成（RDS）（5,6）。然而，尚不清楚这些蛋白是否在一个共同的信号级联中起作用。本研究表明，通过实验阻断Nbs1-Mre11功能或chk2触发的事件可导致人类细胞出现部分RDS表型。相反，同时干扰Nbs1-Mre11和Chk2-Cdc25A-Cdk2通路完全消除电离辐射诱导的DNA合成抑制，导致完全RDS，类似于有缺陷的ATM引起的RDS。此外，Cdc45在复制起点上的cdk2依赖性装载是DNA聚合酶募集的先决条件（7,8），在正常或Nbs1/ mre11缺陷细胞而非ATM缺陷细胞的照射下被阻止。我们得出结论，在电离辐射的作用下，ATM对Nbs1和Chk2的磷酸化触发了DNA损伤依赖性s期检查点的两个平行分支，它们通过抑制DNA复制的不同步骤进行合作。
To preserve genetic integrity, mammalian cells exposed to ionizing radiation activate the ATM kinase, which initiates a complex response including the S-phase checkpoint pathways to delay DNA replication(1,2). Defects in ATM or its substrates Nbs1 or Chk2 (ref. 3), the Nbs1-interacting Mre11 protein(4), or the Chk2-regulated Cdc25A-Cdk2 cascade all cause radio-resistant DNA synthesis (RDS)(5,6). It is unknown, however, whether these proteins operate in a common signaling cascade. Here we show that experimental blockade of either the Nbs1-Mre11 function or the Chk2-triggered events leads to a partial RDS phenotype in human cells. In contrast, concomitant interference with Nbs1-Mre11 and the Chk2-Cdc25A-Cdk2 pathways entirely abolishes inhibition of DNA synthesis induced by ionizing radiation, resulting in complete RDS analogous to that caused by defective ATM. In addition, Cdk2-dependent loading of Cdc45 onto replication origins, a prerequisite for recruitment of DNA polymerase(7,8), was prevented upon irradiation of normal or Nbs1/Mre11-defective cells but not cells with defective ATM. We conclude that in response to ionizing radiation, phosphorylations of Nbs1 and Chk2 by ATM trigger two parallel branches of the DNA damage-dependent S-phase checkpoint that cooperate by inhibiting distinct steps of DNA replication.