Degradation of human exonuclease 1b upon DNA synthesis inhibition
Degradation of human exonuclease 1b upon DNA synthesis inhibition
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DOI：
10.1158/0008-5472.can-04-4069
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发表时间：
2005-05-01
期刊：
影响因子：
11.2
通讯作者：
Ferrari, S
中科院分区：
文献类型：
--
作者：
El-Shemerly, M;Janscak, P;Ferrari, S
作为对DNA损伤的响应，信号通路被触发，要么阻止细胞在特定的过渡期(G(1)-S和G(2)-M)的分裂周期，要么减缓S期的进展。核酸酶在DNA合成、重组、修复和细胞凋亡中起着重要作用。在这项研究中，我们研究了人外切酶I(HEXO1b)的调节。内源性hEXO1b蛋白只有在免疫沉淀富集后才能检测到。我们发现hEXO1b在整个细胞周期中持续表达。然而，用导致DNA复制停止的试剂处理细胞会导致hEXO1b的快速降解。这一效果在移除封闭块后完全逆转。对同步化细胞的分析表明，hEXO1b在S期的降解严格依赖于DNA合成的抑制。UV-C辐射、电离辐射、顺铂或烷基化试剂N-甲基-N‘-硝基-N-亚硝基不影响hEXO1b的稳定性。我们发现hEXO1b被磷酸化以响应DNA合成的抑制，并且磷酸化与蛋白质通过泛素-蛋白酶体途径的快速降解相一致。我们的数据支持这样的证据，即控制外切核酸酶I的活性可能对维持停滞的复制分叉至关重要。
In response to DNA damage, signaling pathways are triggered that either block the cell division cycle at defined transitions (G(1)-S and G(2)-M) or slow down progression through the S phase. Nucleases play important roles in DNA synthesis, recombination, repair, and apoptosis. In this study, we have examined the regulation of human exonuclease I (hEXO1b). The endogenous hEXO1b protein was only detected upon enrichment by immunoprecipitation. We found that hEXO1b was constantly expressed throughout the cell cycle. However, treatment of cells with agents that cause arrest of DNA replication led to rapid degradation of hEXO1b. This effect was fully reversed upon removal of the block. Analysis of synchronized cells showed that degradation of hEXO1b during the S phase was strictly dependent on DNA synthesis inhibition. DNA damage caused by UV-C radiation, ionizing radiation, cisplatin, or the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine did not affect hEXO1b stability. We show that hEXO1b was phosphorylated in response to inhibition of DNA synthesis and that phosphorylation coincided with rapid protein degradation through ubiquitin-proteasome pathways. Our data support the evidence that control of exonuclease I activity may be critical for the maintenance of stalled replication forks.