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首页> 外文期刊>Cell cycle >Absence of an immediate G1/S checkpoint in primary MEFs following gamma-irradiation identifies a novel checkpoint switch.
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Absence of an immediate G1/S checkpoint in primary MEFs following gamma-irradiation identifies a novel checkpoint switch.

机译：伽马射线辐照后，主要MEF中不存在直接的G1 / S检查点，这标志着一种新颖的检查点开关。

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DNA double-strand breaks caused by ionizing radiation have been shown to induce G(1)/S, intra-S-phase, and G(2)/M cell cycle checkpoints. However, analysis of the immediate induction of G(1)/S checkpoint at a cellular level has been hampered by the inability to distinguish cells that were already replicating DNA at the time of damage from cells that entered S phase following the DNA damage. We have developed a novel strategy for assessing the initiation of the G(1)/S checkpoint following gamma-irradiation within asynchronous, low passage, primary mouse embryonic fibroblast cultures (MEFs) using a staggered CldU/IdU double-labeling protocol. Contrary to the current model of the G(1)/S checkpoint, we found that 65% of late- G(1) primary MEFs still proceeded into S phase after a gamma-irradiation dose of 5 Gy. The delayed p53-dependent G(1)/S checkpoint was intact in these cells, and a G2/M checkpoint that was over 90% effective was induced within one hour and maintained through six hours post-irradiation. Furthermore, these cells also exhibited an intra-S-phase replication slow-down, as there was a decrease in the S/ G(2) transition frequency of primary MEFs following gamma-irradiation. The absence of an immediate G(1)/S checkpoint in primary MEFs suggests that in late G(1) these cells may predominantly respond to DNA damage at the level of individual replication origins, rather than by inducing a complete shut-down of S-phase entry.
机译：由电离辐射引起的DNA双链断裂已显示可诱导G（1）/ S，S内和G（2）/ M细胞周期检查点。但是，无法在细胞水平上分析G（1）/ S检查点的即时诱导，因为无法区分在损伤时已经在复制DNA的细胞与在DNA损伤后进入S期的细胞。我们已经开发出一种新颖的策略，用于评估使用交错的CldU / IdU双标记方案在异步，低通量，原代小鼠胚胎成纤维细胞培养（MEF）中进行伽玛射线辐照后G（1）/ S检查点的启动。与当前的G（1）/ S检查点模型相反，我们发现，在5 Gy的伽马射线辐照剂量之后，有65％的晚期G（1）主要MEF仍进入S期。延迟的p53依赖的G（1）/ S检查点在这些细胞中是完整的，并且在1小时内诱导了90％以上的G2 / M检查点，并在照射后的6小时内一直保持该状态。此外，这些细胞还表现出S阶段内复制减慢，因为主要MEF的S / G（2）过渡频率在γ辐照后降低。在主要MEF中没有立即的G（1）/ S检查点表明，在G（1）晚期，这些细胞可能主要在单个复制起点的水平上对DNA损伤做出反应，而不是通过诱导S的完全关闭来进行。相输入。

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