Radiation Effect Mechanisms Research Group, Research Center for Radiation Protection, International Open laboratory, National Institute of Radiological Sciences, Anagawa, Inage-ku, Chiba-shi, Japan.
J Radiat Res. 2011;52(4):415-24. doi: 10.1269/jrr.10168.
Non-homologous end-joining (NHEJ) is the predominant pathway for the repair of DNA double-strand breaks (DSBs) in human cells. XRCC4 is indispensable to NHEJ and functions together with DNA ligase IV in the rejoining of broken DNA ends. Artemis is a nuclease required for trimming of some, but not all, types of broken DNA ends prior to rejoining by the DNA ligase IV/XRCC4 complex. To better understand the roles of these factors, we generated XRCC4- and Artemis-deficient cells from the human colon adenocarcinoma cell line HCT116 by gene targeting and examined their cellular responses to several DNA-damaging agents including X-rays. As anticipated, kinetic analyses of γ-H2AX foci and chromosomal aberrations after ionizing radiation (IR) demonstrated a serious incompetence of DSB repair in the XRCC4-deficient cells, and relatively moderate impairment in the Artemis-deficient cells. The XRCC4-deficient cells were highly sensitive to etoposide and 5-fluoro-2'-deoxyuridine as well as IR, and moderately sensitive to camptothecin, methyl methanesulfonate, cisplatin, mitomycin C, aphidicolin and hydroxyurea, compared to the parental HCT116 cells. The Artemis-deficient cells were not as sensitive as the XRCC4-deficient cells, except to cisplatin and mitomycin C. By contrast, the Artemis-deficient cells were significantly more resistant to hydroxyurea than the parental cells. These observations suggest that Artemis also functions in some DNA damage response pathways other than NHEJ in human cells.
非同源末端连接（NHEJ）是人类细胞修复 DNA 双链断裂（DSB）的主要途径。XRCC4 对 NHEJ 是不可或缺的，它与 DNA 连接酶 IV 一起在断裂 DNA 末端的重新连接中发挥作用。Artemis 是一种核酸内切酶，需要在 DNA 连接酶 IV/XRCC4 复合物重新连接之前，对一些但不是所有类型的断裂 DNA 末端进行修剪。为了更好地理解这些因素的作用，我们通过基因靶向从人结肠腺癌细胞系 HCT116 中生成了 XRCC4 和 Artemis 缺陷细胞，并研究了它们对几种 DNA 损伤剂（包括 X 射线）的细胞反应。正如预期的那样，对 γ-H2AX 焦点和染色体畸变的动力学分析表明，XRCC4 缺陷细胞的 DSB 修复严重无能，Artemis 缺陷细胞的修复能力相对中等受损。与亲本 HCT116 细胞相比，XRCC4 缺陷细胞对依托泊苷和 5-氟-2'-脱氧尿苷以及 IR 非常敏感，对卡铂、甲磺酸甲酯、顺铂、丝裂霉素 C、阿非迪霉素和羟基脲中度敏感。Artemis 缺陷细胞不像 XRCC4 缺陷细胞那样敏感，除了对顺铂和丝裂霉素 C 敏感。相比之下，Artemis 缺陷细胞对羟基脲的耐药性明显高于亲本细胞。这些观察结果表明，Artemis 也在人类细胞中的某些除 NHEJ 以外的 DNA 损伤反应途径中发挥作用。
