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Stable maintenance of the Mre11-Rad50-Nbs1 complex is sufficient to restore the DNA double-strand break response in cells lacking RecQL4 helicase activity.
Journal of Biological Chemistry ( IF 4.0 ) Pub Date : 2021-08-30 , DOI: 10.1016/j.jbc.2021.101148
Hyunsup Kim 1 , Hyemin Choi 2 , Jun-Sub Im 2 , Soon-Young Park 2 , Gwangsu Shin 1 , Jung-Ho Yoo 2 , Gyungmin Kim 2 , Joon-Kyu Lee 3
Affiliation  

The proper cellular response to DNA double-strand breaks (DSBs) is critical for maintaining the integrity of the genome. RecQL4, a DNA helicase of which mutations are associated with Rothmund-Thomson syndrome (RTS), is required for the DNA DSB response. However, the mechanism by which RecQL4 performs these essential roles in the DSB response remains unknown. Here, we show that RecQL4 and its helicase activity are required for maintaining the stability of the Mre11-Rad50-Nbs1 (MRN) complex on DSB sites during a DSB response. We found using immunocytochemistry and live-cell imaging that the MRN complex is prematurely disassembled from DSB sites in a manner dependent upon Skp2-mediated ubiquitination of Nbs1 in RecQL4-defective cells. This early disassembly of the MRN complex could be prevented by altering the ubiquitination site of Nbs1 or by expressing a deubiquitinase, Usp28, which sufficiently restored homologous recombination repair and ATM, a major checkpoint kinase against DNA DSBs, activation abilities in RTS, and RecQL4-depleted cells. These results suggest that the essential role of RecQL4 in the DSB response is to maintain the stability of the MRN complex on DSB sites and that defects in the DSB response in cells of patients with RTS can be recovered by controlling the stability of the MRN complex.

中文翻译:


Mre11-Rad50-Nbs1 复合物的稳定维持足以恢复缺乏 RecQL4 解旋酶活性的细胞中的 DNA 双链断裂反应。



细胞对 DNA 双链断裂 (DSB) 的适当反应对于维持基因组的完整性至关重要。 RecQL4 是一种 DNA 解旋酶,其突变与 Rothmund-Thomson 综合征 (RTS) 相关,是 DNA DSB 反应所必需的。然而,RecQL4 在 DSB 响应中发挥这些重要作用的机制仍然未知。在这里,我们表明 RecQL4 及其解旋酶活性对于在 DSB 响应期间维持 DSB 位点上的 Mre11-Rad50-Nbs1 (MRN) 复合物的稳定性是必需的。我们使用免疫细胞化学和活细胞成像发现,MRN 复合物以依赖于 RecQL4 缺陷细胞中 Skp2 介导的 Nbs1 泛素化的方式过早地从 DSB 位点分解。这种 MRN 复合物的早期分解可以通过改变 Nbs1 的泛素化位点或表达去泛素酶 Usp28 来防止,Usp28 可以充分恢复同源重组修复和 ATM(一种针对 DNA DSB 的主要检查点激酶)、RTS 和 RecQL4 的激活能力。耗尽的细胞。这些结果表明,RecQL4在DSB反应中的重要作用是维持DSB位点上MRN复合物的稳定性,并且RTS患者细胞中DSB反应的缺陷可以通过控制MRN复合物的稳定性来恢复。
更新日期:2021-08-30
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