Accumulation of Ku70 at DNA double-strand breaks in living epithelial cells
Accumulation of Ku70 at DNA double-strand breaks in living epithelial cells
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DOI：
10.1016/j.yexcr.2011.07.018
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发表时间：
2011-10-15
影响因子：
3.7
通讯作者：
Koike, Aki
中科院分区：
文献类型：
--
作者：
Koike, Manabu;Yutoku, Yasutomo;Koike, Aki
Ku70 和 Ku80 在 DNA 双链断裂 (DSB) 修复途径，即非同源 DNA 末端连接 (NHEJ) 中发挥重要作用。尽管 Ku70 在 DSB 的积累机制在调节 NHEJ 活性中发挥着关键作用，但目前尚未详细阐明 Ku70 在 DSB 的积累机制。在这里，我们展示了活肺上皮细胞中 DSB 上 Ku70 积累和功能的重要结构域。我们的结果表明，EGFP-Ku70 在 DSB 上的积累在照射后立即开始。我们的研究结果表明，Ku70 的三个结构域，即 α/β、DNA 结合结构域和 Ku80 结合结构域（而非 SAP 结构域）对于辐射后早期 DSB 的积累或识别是必需的。此外，我们的研究结果表明，Ku80 结合域中 Ku70 氨基酸 385 处的亮氨酸（而非 DNA 结合域中用于乙酰化的三个目标氨基酸）参与了 Ku70 在 DSB 的定位和积累。此外，XRCC4 和 XLF（而非 Artemis）在 DSB 的积累取决于 Ku70 的存在。这些发现表明，Artemis 不仅可以参与 Ku 依赖性修复过程，还可以参与活上皮细胞 DSB 的 Ku 独立修复过程。 (C) 2011 Elsevier Inc. 保留所有权利。
Ku70 and Ku80 play an essential role in the DNA double-strand break (DSB) repair pathway, i.e., nonhomologous DNA-end-joining (NHEJ). No accumulation mechanisms of Ku70 at DSBs have been clarified in detail, although the accumulation mechanism of Ku70 at DSBs plays key roles in regulating the NHEJ activity. Here, we show the essential domains for the accumulation and function of Ku70 at DSBs in living lung epithelial cells. Our results showed that EGFP-Ku70 accumulation at DSBs began immediately after irradiation. Our findings demonstrate that three domains of Ku70, i.e., the alpha/beta, DNA-binding, and Ku80-binding domains, but not the SAP domain, are necessary for the accumulation at or recognition of DSBs in the early stage after irradiation. Moreover, our findings demonstrate that the leucine at amino acid 385 of Ku70 in the Ku80-binding domain, but not the three target amino acids for acetylation in the DNA-binding domain, is involved in the localization and accumulation of Ku70 at DSBs. Furthermore, accumulations of XRCC4 and XLF, but not that of Artemis, at DSBs are dependent on the presence of Ku70. These findings suggest that Artemis can work in not only the Ku-dependent repair process, but also the Ku-independent process at DSBs in living epithelial cells. (C) 2011 Elsevier Inc. All rights reserved.