Differential radiosensitivity phenotypes of DNA-PKcs mutations affecting NHEJ and HRR systems following irradiation with gamma-rays or very low fluences of alpha particles.
Differential radiosensitivity phenotypes of DNA-PKcs mutations affecting NHEJ and HRR systems following irradiation with gamma-rays or very low fluences of alpha particles.
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DOI：
10.1371/journal.pone.0093579
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Chen BP
中科院分区：
文献类型：
--
作者：
Lin YF;Nagasawa H;Little JB;Kato TA;Shih HY;Xie XJ;Wilson PF Jr;Brogan JR;Kurimasa A;Chen DJ;Bedford JS;Chen BP
我们研究了在S2056簇、T2609簇或激酶结构域中携带DNA-PKcs突变的细胞中，高或低剂量率γ射线照射后染色体畸变诱导和细胞杀伤的细胞周期依赖性。我们还比较了DNA-PKcs突变细胞和同源重组修复（HRR）突变细胞（包括Rad 51 C、Rad 51 D和Fancg/xrcc 9）中极低通量的α颗粒产生的姐妹染色单体交换（SCE）。一般来说，染色体畸变和γ射线对细胞的杀伤作用与DNA-PKcs突变的影响相似，这些突变细胞在G1期比S/G2期更敏感。在G1-照射DNA-PKcs突变细胞，染色体和染色单体型断裂和交换超过野生型细胞。对于在后期S/G2期照射的细胞，突变体细胞表现出非常高的产量的染色单体断裂相比，野生型细胞。在DNA-PKcs-null，Ku 80-null或DNA-PKcs激酶死亡突变体中观察到很少的交换，但在S2506或T2609簇突变体中检测到过量的交换。极低剂量的α粒子诱发SCE是由于未被α粒子穿过的细胞中的旁观者效应。在野生型细胞中观察到的SCE在Rad 51 C-或Rad 51 D-缺陷细胞中完全消除，但在Fancg/xrcc 9细胞中接近正常。与此形成鲜明对比的是，在DNA-PKcs-null、DNA-PKcs激酶-dead和Ku 80-null突变体中观察到非常高水平的SCE。SCE诱导在T2609簇突变细胞中也被取消，但在S2056簇突变细胞中仅略有减少。由于非同源末端连接（NHEJ）和HRR系统都利用初始DNA损伤作为底物，这些结果表明NHEJ和至少HRR的Rad 51 C/D组分之间存在竞争性干扰现象的可能性;受损DNA和特定DNA-PK组分之间的相互作用水平可能决定此类DNA与相关HRR组分的相互作用水平。
We have examined cell-cycle dependence of chromosomal aberration induction and cell killing after high or low dose-rate γ irradiation in cells bearing DNA-PKcs mutations in the S2056 cluster, the T2609 cluster, or the kinase domain. We also compared sister chromatid exchanges (SCE) production by very low fluences of α-particles in DNA-PKcs mutant cells, and in homologous recombination repair (HRR) mutant cells including Rad51C, Rad51D, and Fancg/xrcc9. Generally, chromosomal aberrations and cell killing by γ-rays were similarly affected by mutations in DNA-PKcs, and these mutant cells were more sensitive in G1 than in S/G2 phase. In G1-irradiated DNA-PKcs mutant cells, both chromosome- and chromatid-type breaks and exchanges were in excess than wild-type cells. For cells irradiated in late S/G2 phase, mutant cells showed very high yields of chromatid breaks compared to wild-type cells. Few exchanges were seen in DNA-PKcs-null, Ku80-null, or DNA-PKcs kinase dead mutants, but exchanges in excess were detected in the S2506 or T2609 cluster mutants. SCE induction by very low doses of α-particles is resulted from bystander effects in cells not traversed by α-particles. SCE seen in wild-type cells was completely abolished in Rad51C- or Rad51D-deficient cells, but near normal in Fancg/xrcc9 cells. In marked contrast, very high levels of SCEs were observed in DNA-PKcs-null, DNA-PKcs kinase-dead and Ku80-null mutants. SCE induction was also abolished in T2609 cluster mutant cells, but was only slightly reduced in the S2056 cluster mutant cells. Since both non-homologous end-joining (NHEJ) and HRR systems utilize initial DNA lesions as a substrate, these results suggest the possibility of a competitive interference phenomenon operating between NHEJ and at least the Rad51C/D components of HRR; the level of interaction between damaged DNA and a particular DNA-PK component may determine the level of interaction of such DNA with a relevant HRR component.
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