Apoptosis induced by replication inhibitors in Chk1-depleted cells is dependent upon the helicase cofactor Cdc45
Apoptosis induced by replication inhibitors in Chk1-depleted cells is dependent upon the helicase cofactor Cdc45
复制标题
DOI：
10.1038/cdd.2008.4
复制
发表时间：
2008-05-01
影响因子：
12.4
通讯作者：
Meuth, M.
中科院分区：
文献类型：
--
作者：
Rodriguez, R.;Gagou, M. E.;Meuth, M.
检查点激酶1（Chk 1）响应DNA复制的破坏以维持停滞叉的完整性，促进复制叉损伤的同源重组介导的修复，并控制复制起点的不适当发射。这种反应对于生存力是必不可少的，因为复制抑制剂在Chk 1耗尽的S期细胞中触发凋亡。考虑到Chk 1控制的细胞反应的复杂网络，我们的目的是确定这些保护细胞在复制应激后免于凋亡。无细胞系统的研究表明，RPA-ssDNA复合物在复制抑制后通过复制和解旋酶复合物的解偶联形成。在这里，我们表明，复制蛋白A（RPA）焦点的形式与复制抑制剂处理的细胞和焦点的数量显着增加，连同高度磷酸化的RPA 34在Chk 1耗竭细胞诱导凋亡的提前。RPA焦点，RPA 34过度磷酸化，和细胞凋亡抑制siRNA介导的敲低Cdc 45，一个必要的复制解旋酶辅因子所需的DNA复制的起始和延伸步骤。与此相反，p21的损失，一个负效应源发射，刺激RPA灶和细胞凋亡的积累。综上所述，这些结果表明，Chk 1耗竭后复制起点放电控制的丧失引发了RPA-ssDNA复合物的积累和复制被阻断时的细胞凋亡。
Checkpoint kinase 1 (Chk1) responds to disruption of DNA replication to maintain the integrity of stalled forks, promote homologous recombination-mediated repair of replication fork lesions, and control inappropriate firing of replication origins. This response is essential for viability as replication inhibitors trigger apoptosis in S-phase cells depleted of Chk1. Given the complex network of cellular responses controlled by Chk1, our aim was to determine which of these protect cells from apoptosis following replication stress. Work with cell-free systems has shown that RPA-ssDNA complex forms following replication inhibition through the uncoupling of replication and helicase complexes. Here we show that replication protein A (RPA) foci form in cells treated with replication inhibitors and that the number of foci dramatically increases together with hyperphosphorylation of RPA34 in Chk1-depleted cells in advance of the induction of apoptosis. RPA foci, RPA34 hyperphosphorylation, and apoptosis were suppressed by siRNA-mediated knockdown of Cdc45, an essential replication helicase cofactor required for both the initiation and elongation steps of DNA replication. In contrast, loss of p21, a negative effector of origin firing, stimulates both the accumulation of RPA foci and apoptosis. Taken together, these results suggest that the loss of control of replication origin firing following Chk1 depletion triggers the accumulation of the RPA-ssDNA complex and apoptosis when replication is blocked.