A short G1 phase imposes constitutive replication stress and fork remodelling in mouse embryonic stem cells.
A short G1 phase imposes constitutive replication stress and fork remodelling in mouse embryonic stem cells.
复制标题
DOI：
10.1038/ncomms10660
复制
发表时间：
2016-02-15
影响因子：
16.6
通讯作者：
Lopes M
中科院分区：
文献类型：
--
作者：
Ahuja AK;Jodkowska K;Teloni F;Bizard AH;Zellweger R;Herrador R;Ortega S;Hickson ID;Altmeyer M;Mendez J;Lopes M
胚胎干细胞（ESC）代表一种短暂的生物学状态，其中多能性与快速增殖相结合。ESCs表现出组成型活性DNA损伤反应（DDR），但其分子决定因素仍然难以捉摸。在这里，我们表明在培养的胚胎干细胞和小鼠胚胎H2 AX磷酸化是依赖于共济失调毛细血管扩张和Rad 3相关（ATR），并与染色质加载的ssDNA结合蛋白RPA和RAD 51。复制中间体的单分子分析揭示了大量的ssDNA缺口积累，减少叉速度和频繁的叉逆转。所有这些复制应激的标记不损害有丝分裂过程，并且在分化开始时迅速丢失。延迟ESCs中的G1/S转换允许53 BP 1核体的形成并抑制ssDNA积累，在随后的S期中叉慢化和逆转。基因失活的叉减慢和逆转导致染色体断裂在未受干扰的胚胎干细胞。我们认为，快速的细胞周期进程使ESCs依赖于有效的复制偶联机制来保护基因组的完整性。 在快速增殖的胚胎干细胞（ESC）中，DNA损伤反应是由尚未阐明的机制激活的。在这里，Ahuja等人将DNA损伤反应与小鼠ESCs中由短G1期引起的复制应激联系起来，并提出叉重塑作为维持胚胎中基因组稳定性的方法。
Embryonic stem cells (ESCs) represent a transient biological state, where pluripotency is coupled with fast proliferation. ESCs display a constitutively active DNA damage response (DDR), but its molecular determinants have remained elusive. Here we show in cultured ESCs and mouse embryos that H2AX phosphorylation is dependent on Ataxia telangiectasia and Rad3 related (ATR) and is associated with chromatin loading of the ssDNA-binding proteins RPA and RAD51. Single-molecule analysis of replication intermediates reveals massive ssDNA gap accumulation, reduced fork speed and frequent fork reversal. All these marks of replication stress do not impair the mitotic process and are rapidly lost at differentiation onset. Delaying the G1/S transition in ESCs allows formation of 53BP1 nuclear bodies and suppresses ssDNA accumulation, fork slowing and reversal in the following S-phase. Genetic inactivation of fork slowing and reversal leads to chromosomal breakage in unperturbed ESCs. We propose that rapid cell cycle progression makes ESCs dependent on effective replication-coupled mechanisms to protect genome integrity. In fast proliferating embryonic stem cells (ESC) the DNA damage response is activated by mechanisms that are as yet elusive. Here, Ahuja et al. link the DNA damage response to replication stress in mouse ESCs, caused by a short G1 phase, and propose fork remodelling as maintaining genome stability in embryos.