Wang Hongyan, Wang Xiang, Zhang Piyan, Wang Ya
Department of Radiation Oncology and Kimmel Cancer Center of Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA.
DNA Repair (Amst). 2008 May 3;7(5):725-33. doi: 10.1016/j.dnarep.2008.01.010. Epub 2008 Mar 5.
Ionizing radiation (IR) induced DNA double strand breaks (DSBs) are repaired by both non-homologous end-joining (NHEJ) and homologous recombination repair (HRR) in mammalian cells. The NHEJ repair includes a Ku-dependent main pathway and a PARP-1-dependent complementary pathway. Compared with low linear energy transfer (LET) IR (X or gamma ray) at the same doses, high LET IR (high-charge particles) induces more cell death because of ineffective DNA repair. However, it remains unclear whether high LET IR inhibits all repair or specifically one repair pathway. By combining the assays of clonogenic survival, G2M checkpoint and gammaH2AX in the cell lines with deficiencies in different repair genes, we show here that high LET IR inhibits only the Ku-dependent main NHEJ pathway and does not inhibit either the HRR pathway or the PARP-1-dependent complementary NHEJ pathway. In addition, by developing an assay to detect small fragments of DSB (<400 bp) and by detecting the binding abilities of purified Ku and PARP to different sized dsDNA, we present a possible link for explaining the phenotypes. When compared with low LET IR at the same dose, high LET IR might induce similar yields of DNA DSBs in total but it might induce more small fragments of DNA DSBs (<40 base pairs) that prevent Ku binding efficiently to two ends of one DSB fragment at the same time, thus delaying Ku-dependent repair.
电离辐射（IR）诱导的DNA双链断裂（DSB）在哺乳动物细胞中通过非同源末端连接（NHEJ）和同源重组修复（HRR）进行修复。NHEJ修复包括一条Ku依赖的主要途径和一条PARP-1依赖的互补途径。与相同剂量的低传能线密度（LET）IR（X射线或γ射线）相比，高LET IR（高电荷粒子）由于DNA修复无效而诱导更多细胞死亡。然而，高LET IR是否抑制所有修复或特异性抑制某一种修复途径仍不清楚。通过将克隆存活分析、G2M检查点分析和γH2AX分析与不同修复基因缺陷的细胞系相结合，我们在此表明高LET IR仅抑制Ku依赖的主要NHEJ途径，而不抑制HRR途径或PARP-1依赖的互补NHEJ途径。此外，通过开发一种检测DSB小片段（<400 bp）的分析方法，并检测纯化的Ku和PARP与不同大小双链DNA的结合能力，我们提出了一个可能的联系来解释这些表型。与相同剂量的低LET IR相比，高LET IR可能在总体上诱导相似产量的DNA DSB，但它可能诱导更多的DNA DSB小片段（<40个碱基对），这些小片段会阻止Ku同时有效地结合到一个DSB片段的两端，从而延迟Ku依赖的修复。
