Laboratorio de Mutagenesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, Buenos Aires, Argentina.
Environ Mol Mutagen. 2012 Oct;53(8):608-18. doi: 10.1002/em.21729. Epub 2012 Sep 18.
The role of DNA double strand break (DSB) repair pathways, non-homologous end joining (NHEJ), and homologous recombination (HR) was evaluated to prevent the chromosome instability induced by the topoisomerase II (Top2) poisons, idarubicin, and etoposide in Chinese hamster cell lines. XR-C1 (DNA-PKcs deficient) and V-C8 (BRCA2 deficient) showed higher sensitivity to increased concentrations of Top2 poisons compared with their normal counterparts, CHO9 and V79. Both proficient and deficient cells exhibited a marked DSB induction in all phases of the cell cycle. Additionally, deficient cells showed persistent DNA damage 24 hr post-treatment. Chromosomal aberrations increased in the first mitosis following Top2 poison-treatments in G1 or G2 in proficient and deficient cells. CHO9 and V79 demonstrated chromosome and chromatid exchanges following treatments in G1 and G2 phases, respectively. Deficient cells showed high frequencies of chromatid exchanges following treatments in G1 and G2. Simultaneously, we analyzed the micronuclei (MN) induction in interphase cells after treatments in G1, S, or G2 of the previous cell cycle. Both Top2 poisons induced an important increase in MN in CHO9, V79, and V-C8 cells. XR-C1 exhibited an increased MN frequency when cells were treated in G1 phase but not in S or G2. This MN reduction was due to a cell accumulation at G2/M and death in G2-treated cells. Our data suggest that NHEJ and HR operate differentially throughout the cell cycle to protect from Top2 poison-induced chromosome instability, and that DNA-PKcs-dependent NHEJ pathway allows the survival of chromosome damaged cells during S/G2 to the next interphase.
我们评估了 DNA 双链断裂 (DSB) 修复途径、非同源末端连接 (NHEJ) 和同源重组 (HR) 的作用，以防止拓扑异构酶 II (Top2) 抑制剂伊达比星和依托泊苷在仓鼠细胞系中诱导染色体不稳定性。与正常对照 CHO9 和 V79 相比，XR-C1（DNA-PKcs 缺陷）和 V-C8（BRCA2 缺陷）对 Top2 抑制剂浓度的增加表现出更高的敏感性。所有细胞周期阶段， proficient 和 deficient 细胞均显示出明显的 DSB 诱导。此外，缺陷细胞在处理后 24 小时仍显示出持续的 DNA 损伤。在 proficient 和 deficient 细胞中，Top2 毒素处理后 G1 或 G2 期的第一次有丝分裂中染色体畸变增加。CHO9 和 V79 在 G1 和 G2 期分别显示染色体和染色单体交换。缺陷细胞在 G1 和 G2 期处理后显示出高频率的染色单体交换。同时，我们分析了前一个细胞周期 G1、S 或 G2 期处理后间期细胞中的微核 (MN) 诱导。两种 Top2 抑制剂均诱导 CHO9、V79 和 V-C8 细胞中 MN 显著增加。XR-C1 在 G1 期处理时 MN 频率增加，但在 S 期或 G2 期处理时则不增加。这种 MN 减少是由于 G2/M 期细胞积累和 G2 期处理细胞死亡所致。我们的数据表明，NHEJ 和 HR 在整个细胞周期中以不同的方式运作，以保护细胞免受 Top2 抑制剂诱导的染色体不稳定性，并且 DNA-PKcs 依赖性 NHEJ 途径允许染色体受损细胞在 S/G2 期存活到下一个间期。
