Cleavage of claspin by caspase-7 during apoptosis inhibits the Chk1 pathway
Cleavage of claspin by caspase-7 during apoptosis inhibits the Chk1 pathway
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DOI：
10.1074/jbc.m506460200
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发表时间：
2005-10-21
影响因子：
4.8
通讯作者：
Clarke, PR
中科院分区：
文献类型：
--
作者：
Clarke, CAL;Bennett, LN;Clarke, PR
Claspin在DNA复制期间和响应DNA损伤时被ATR磷酸化和激活Chk 1蛋白激酶所需。该检查点途径在细胞对遗传毒性应激的抗性中起关键作用。在这里，我们表明，人类Claspin是裂解的caspase- 7在启动细胞凋亡。在细胞中，依托泊苷诱导的DNA损伤首先在ATR靶向的位点产生Chk 1的快速磷酸化。随后，依托泊苷引起caspase- 7的活化，Claspin的裂解和Chk 1的去磷酸化。在凋亡细胞提取物中，Claspin在单个天冬氨酸残基处被胱天蛋白酶-7切割成含有不同功能结构域的大的N-末端片段和较小的C-末端片段。大的N-末端片段在人无细胞系统中响应于双链DNA寡核苷酸而被严重磷酸化，并且该片段保留Chk 1结合活性。相反，较小的C-末端片段不结合Chk 1，但确实与DNA结合并抑制与其活化相关的Chk 1的DNA依赖性磷酸化。这些结果表明，由胱天蛋白酶-7切割Claspin使Chk 1信号传导途径失活。这一机制可能在基因毒性应激反应中调节细胞周期阻滞和诱导凋亡之间的平衡。
Claspin is required for the phosphorylation and activation of the Chk1 protein kinase by ATR during DNA replication and in response to DNA damage. This checkpoint pathway plays a critical role in the resistance of cells to genotoxic stress. Here, we show that human Claspin is cleaved by caspase- 7 during the initiation of apoptosis. In cells, induction of DNA damage by etoposide at first produced rapid phosphorylation of Chk1 at a site targeted by ATR. Subsequently, etoposide caused activation of caspase- 7, cleavage of Claspin, and dephosphorylation of Chk1. In apoptotic cell extracts, Claspin was cleaved by caspase- 7 at a single aspartate residue into a large N- terminal fragment and a smaller C- terminal fragment that contain different functional domains. The large N- terminal fragment was heavily phosphorylated in a human cell- free system in response to double- stranded DNA oligonucleotides, and this fragment retained Chk1 binding activity. In contrast, the smaller C- terminal fragment did not bind Chk1, but did associate with DNA and inhibited the DNA- dependent phosphorylation of Chk1 associated with its activation. These results indicate that cleavage of Claspin by caspase- 7 inactivates the Chk1 signaling pathway. This mechanism may regulate the balance between cell cycle arrest and induction of apoptosis during the response to genotoxic stress.