RNF8 promotes high linear energy transfer carbon-ion-induced DNA double-stranded break repair in serum-starved human cells
RNF8 promotes high linear energy transfer carbon-ion-induced DNA double-stranded break repair in serum-starved human cells
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DOI：
10.1016/j.dnarep.2020.102872
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发表时间：
2020-07-01
期刊：
影响因子：
3.8
通讯作者：
Shibata, Atsushi
中科院分区：
文献类型：
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作者：
Nakajima, Nakako Izumi;Yamauchi, Motohiro;Shibata, Atsushi
放射治疗的细胞杀伤效应主要依赖于未修复的DNA双链断裂（DSB）或DSB诱导的致死性染色体畸变。因此，DSB修复能力对于电离辐射的癌细胞杀伤作用至关重要。在这里，我们研究了参与的DNA损伤信号因子共济失调毛细血管扩张突变（ATM），环指蛋白8（RNF 8），RNF 168在静止的G 0/G1细胞，这是在大多数肿瘤细胞群中表达，高线性能量转移（LET）碳离子照射后。有趣的是，ATM抑制引起了大量的DSB修复缺陷后，高LET碳离子照射。类似地，RNF 8或RNF 168耗尽引起大量DSB修复缺陷。ATM抑制在RNF 8耗尽的细胞中没有产生累加效应，这表明ATM和RNF 8在相同的通路中起作用。重要的是，我们发现RNF 8 RING突变体表现出类似的DSB修复缺陷，这表明该修复途径需要遍在蛋白连接酶活性。RNF 8 FHA结构域也是该轴中DSB修复所需的。此外，p53结合蛋白1（53 BP 1）是RNF 8依赖性DSB修复的重要下游因子，也是这种修复所必需的。重要的是，ATM抑制或RNF 8耗尽增加了染色体断裂的频率，但减少了双着丝粒染色体的形成，表明ATM/RNF 8是形成双着丝粒染色体的DSB末端重新连接所必需的。最后，我们发现RNF 8耗竭增强了高LET碳离子照射后的放射敏感性。这项研究表明，抑制RNF 8活性或其下游途径可能会增加高LET碳离子治疗的疗效。
The cell-killing effect of radiotherapy largely depends on unrepaired DNA double-stranded breaks (DSBs) or lethal chromosome aberrations induced by DSBs. Thus, the capability of DSB repair is critically important for the cancer-cell-killing effect of ionizing radiation. Here, we investigated the involvement of the DNA damage signaling factors ataxia telangiectasia mutated (ATM), ring finger protein 8 (RNF8), and RNF168 in quiescent G0/G1 cells, which are expressed in the majority of cell populations in tumors, after high linear energy transfer (LET) carbon-ion irradiation. Interestingly, ATM inhibition caused a substantial DSB repair defect after high-LET carbon-ion irradiation. Similarly, RNF8 or RNF168 depletion caused a substantial DSB repair defect. ATM inhibition did not exert an additive effect in RNF8-depleted cells, suggesting that ATM and RNF8 function in the same pathway. Importantly, we found that the RNF8 RING mutant showed a similar DSB repair defect, suggesting the requirement of ubiquitin ligase activity in this repair pathway. The RNF8 FHA domain was also required for DSB repair in this axis. Furthermore, the p53-binding protein 1 (53BP1), which is an important downstream factor in RNF8-dependent DSB repair, was also required for this repair. Importantly, either ATM inhibition or RNF8 depletion increased the frequency of chromosomal breaks, but reduced dicentric chromosome formation, demonstrating that ATM/RNF8 is required for the rejoining of DSB ends for the formation of dicentric chromosomes. Finally, we showed that RNF8 depletion augmented radiosensitivity after high-LET carbon-ion irradiation. This study suggests that the inhibition of RNF8 activity or its downstream pathway may augment the efficacy of high-LET carbon-ion therapy.