Regulation of Chk1 includes chromatin association and 14-3-3 binding following phosphorylation on Ser-345
Regulation of Chk1 includes chromatin association and 14-3-3 binding following phosphorylation on Ser-345
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DOI：
10.1074/jbc.m300070200
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发表时间：
2003-07-04
影响因子：
4.8
通讯作者：
Sanchez, Y
中科院分区：
文献类型：
--
作者：
Jiang, KC;Pereira, E;Sanchez, Y
检查点是生化途径，为细胞提供检测DNA损伤的机制，并通过阻止细胞周期以允许DNA修复来做出反应。保守的检查点激酶Chk1通过阻断Cdk1/cyclin b的激活来调节DNA损伤和复制干扰的有丝分裂进程。Chk1在接受检查点信号后在Ser-317和Ser-345上磷酸化，这一过程由Atr和含有Rad17和Hus1的传感器复合物调节。我们发现Chk1与循环细胞中的染色质相关，并且染色质相关的Chk1在没有外源DNA损伤的情况下被磷酸化。紫外线诱导的ser -345磷酸化形式的Chk1在治疗后几分钟出现，主要与染色质相关。Ser-345位点位于14-3-3一致结合基序中，是在羟基脲诱导的检查点信号后Chk1的核保留所必需的；然而，Ser-345或Ser-317并不是Chk1染色质关联所必需的。Hus1是增殖细胞核抗原样损伤识别复合体的一个成员，在Ser-345上的Chk1磷酸化中起作用，然而，在Ser-317上的磷酸化或Chk1定位到染色质上并不需要Hus1。这些结果表明，Chk1的激活不止一个步骤，并且对这个检查点信号的调节至少部分是通过Ser-345的磷酸化实现的，Ser-345可能通过阻断依赖crm1的核输出来定位Chk1在细胞核中的位置。
Checkpoints are biochemical pathways that provide the cell with mechanisms to detect DNA damage and respond by arresting the cell cycle to allow DNA repair. The conserved checkpoint kinase Chk1 regulates mitotic progression in response to DNA damage and replication interference by blocking the activation of Cdk1/cyclin B. Chk1 is phosphorylated on Ser-317 and Ser-345 following a checkpoint signal, a process that is regulated by Atr, and by the sensor complexes containing Rad17 and Hus1. We show that Chk1 is associated with chromatin in cycling cells and that the chromatin-associated Chk1 is phosphorylated in the absence of exogenous DNA damage. The UV-induced Ser-345-phosphorylated forms of Chk1 that appear minutes after treatment are predominantly associated with chromatin. The Ser-345 site is in a 14-3-3 consensus binding motif and is required for nuclear retention of Chk1 following an hydroxyurea-induced checkpoint signal; nonetheless, Ser-345 or Ser-317 are not required for the chromatin association of Chk1. Hus1, a member of the proliferating cell nuclear antigen-like damage recognition complex plays a role in the phosphorylation of Chk1 on Ser-345, however, Hus1 is not required for phosphorylation on Ser-317 or for Chk1 localization to chromatin. These results indicate that there is more than one step in Chk1 activation and that the regulation of this checkpoint signaling is achieved at least in part through phosphorylation of Ser-345, which serves to localize Chk1 in the nucleus presumably by blocking Crm1-dependent nuclear export.