Akkari Y M, Bateman R L, Reifsteck C A, Olson S B, Grompe M
Department of Molecular and Medical Genetics, Oregon Health Sciences University, Portland, Oregon 97201, USA.
Mol Cell Biol. 2000 Nov;20(21):8283-9. doi: 10.1128/MCB.20.21.8283-8289.2000.
Following introduction of DNA interstrand cross-links (ICLs), mammalian cells display chromosome breakage or cell cycle delay with a 4N DNA content. To further understand the nature of the delay, previously described as a G(2)/M arrest, we developed a protocol to generate ICLs during specific intervals of the cell cycle. Synchronous populations of G(1), S, and G(2) cells were treated with photoactivated 4'-hydroxymethyl-4,5',8-trimethylpsoralen (HMT) and scored for normal passage into mitosis. In contrast to what was found for ionizing radiation, ICLs introduced during G(2) did not result in a G(2)/M arrest, mitotic arrest, or chromosome breakage. Rather, subsequent passage through S phase was required to trigger both chromosome breakage and arrest in the next cell cycle. Similarly, ICLs introduced during G(1) did not cause a G(1)/S arrest. We conclude that DNA replication is required to elicit the cellular responses of cell cycle arrest and genomic instability after psoralen-induced ICLs. In primary human fibroblasts, the 4N DNA content cell cycle arrest triggered by ICLs was long lasting but reversible. Kinetic analysis suggested that these cells could remove up to approximately 2,500 ICLs/genome at an average rate of 11 ICLs/genome/h.
在引入DNA链间交联（ICL）后，哺乳动物细胞会出现染色体断裂或细胞周期延迟，并伴有4N DNA含量。为了进一步了解这种延迟的本质，此前将其描述为G(2)/M期阻滞，我们开发了一种在细胞周期的特定间隔内产生ICL的方案。用光活化的4'-羟甲基-4,5',8-三甲基补骨脂素（HMT）处理同步化的G(1)、S和G(2)期细胞群体，并对进入有丝分裂的正常进程进行评分。与电离辐射的情况不同，在G(2)期引入的ICL不会导致G(2)/M期阻滞、有丝分裂阻滞或染色体断裂。相反，需要随后经过S期才能在下一个细胞周期触发染色体断裂和阻滞。同样，在G(1)期引入的ICL不会导致G(1)/S期阻滞。我们得出结论，在补骨脂素诱导ICL后，需要DNA复制来引发细胞周期阻滞和基因组不稳定的细胞反应。在原代人成纤维细胞中，由ICL触发的4N DNA含量细胞周期阻滞持续时间长但可逆。动力学分析表明，这些细胞平均每小时可去除约2500个ICL/基因组，去除速率为11个ICL/基因组/小时。
