Degradation of ATM-Independent Decatenation Checkpoint Function in Human Cells is Secondary to Inactivation of p53 and Correlated with Chromosomal Destabilization
Degradation of ATM-Independent Decatenation Checkpoint Function in Human Cells is Secondary to Inactivation of p53 and Correlated with Chromosomal Destabilization
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DOI：
10.4161/cc.1.3.127
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发表时间：
2002-05-01
期刊：
影响因子：
4.3
通讯作者：
Downes, C. Stephen
中科院分区：
文献类型：
--
作者：
Kaufmann, William K.;Campbell, Christine B.;Downes, C. Stephen
DNA拓扑异构酶II在细胞周期中是必需的，以在有丝分裂之前使缠结的子染色单体解链。为了研究细胞用于实现及时染色单体脱连环的机制，在DNA损伤G2检查点中具有遗传性或获得性缺陷的人皮肤成纤维细胞中监测了连锁反应检查点的活性。G2延迟在与ICRF-193短暂孵育后不久定量，ICRF-193阻断拓扑异构酶II使染色单体十链化的能力，或用电离辐射（IR）处理后不久定量，IR损伤DNA。两种处理均诱导正常人成纤维细胞中的G2延迟。具有对IR的G2检查点反应缺陷的共济失调毛细血管扩张成纤维细胞在用ICRF-193处理后显示正常的G2延迟，表明当染色单体脱链被阻断时，ATM激酶不是信号传导所必需的。ICRF-193诱导的G2延迟被咖啡因逆转，表明参与了主动检查点信号传导。ICRF-193诱导的G2期延迟也不依赖于p53功能，这在表达HPV 16 E6的细胞中很明显可以灭活p53。然而，随着表达HPV 16 E6的成纤维细胞在培养中老化，它们在DNA损伤后和去连环化被阻断时都失去了延迟进入有丝分裂的能力。这种年龄相关的损失G2延迟响应ICRF-193和IR在E6表达细胞被阻断诱导端粒酶。端粒酶的表达也防止了衰老E6表达细胞中的染色体不稳定。这些观察结果导致了一种新的遗传不稳定性模型，其中G2脱连环检查点功能的衰减允许细胞进入具有不充分脱连环染色单体的有丝分裂，导致非整倍性和多倍性。
DNA topoisomerase II is required in the cell cycle to decatenate intertwined daughter chromatids prior to mitosis. To study the mechanisms that cells use to accomplish timely chromatid decatenation, the activity of a catenation-responsive checkpoint was monitored in human skin fibroblasts with inherited or acquired defects in the DNA damage G2 checkpoint. G2 delay was quantified shortly after a brief incubation with ICRF-193, which blocks the ability of topoisomerase II to decatenate chromatids, or treatment with ionizing radiation (IR), which damages DNA. Both treatments induced G2 delay in normal human fibroblasts. Ataxia telangiectasia fibroblasts with defective G2 checkpoint response to IR displayed normal G2 delay after treatment with ICRF-193, demonstrating that ATM kinase was not required for signaling when chromatid decatenation was blocked. The G2 delay induced by ICRF-193 was reversed by caffeine, indicating that active checkpoint signaling was involved. ICRF-193-induced G2 delay also was independent of p53 function, being evident in cells expressing HPV16E6 to inactivate p53. However, as fibroblasts expressing HPV16E6 aged in culture, they lost the ability to delay entry to mitosis, both after DNA damage and when decatenation was blocked. This age-related loss of G2 delay in response to ICRF-193 and IR in E6-expressing cells was blocked by induction of telomerase. Expression of telomerase also prevented chromosomal destabilization in aging E6-expressing cells. These observations lead to a new model of genetic instability, in which attenuation of G2 decatenatory checkpoint function permits cells to enter mitosis with insufficiently decatenated chromatids, leading to aneuploidy and polyploidy.