The Chk1 protein kinase and the Cdc25C regulatory pathways are targets of the anticancer agent UCN-01
The Chk1 protein kinase and the Cdc25C regulatory pathways are targets of the anticancer agent UCN-01
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DOI：
10.1074/jbc.275.8.5600
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发表时间：
2000-02-25
影响因子：
4.8
通讯作者：
Piwnica-Worms, H
中科院分区：
文献类型：
--
作者：
Graves, PR;Yu, LJ;Piwnica-Worms, H
在细胞周期的G(2)阶段运行的检查点防止在存在DNA损伤的情况下进入有丝分裂，目前正在进行癌症治疗临床试验的蛋白激酶抑制剂UCN-01取消了G(2)检查点的功能，并使p53缺陷的癌细胞对DNA损伤剂敏感。在大多数物种中，G(2)检查点阻止CDC25磷酸酶从促进有丝分裂的激酶CDC2中移除抑制磷酸基团。这是通过维持CDC25的磷酸化形式与14-3-3蛋白结合来实现的。Chk1和CDs1是通过磷酸化丝氨酸216上的CDc25C来调节人CDc25C和14-3-3蛋白之间相互作用的检查点蛋白。而14-3-3蛋白则起到阻止CDC25C进入细胞核的作用。在这里，我们报告了UCN-01在DNA损伤细胞中导致丝氨酸216磷酸化和14-3-3与CDC25C结合的丢失。此外，UCN-01在体外可有效抑制Chk1对CDc25C的磷酸化作用。相反，CDS1在体外对UCN-01的抑制作用不明显，经UCN-01处理的照射细胞中，CDS1仍被磷酸化。因此，无论是cds1还是cds1上游的激酶，例如突变的共济失调毛细血管扩张症，都不是UCN-01体内作用的靶点。综上所述，我们的结果确定Chk1激酶和CDc25C通路是UCN-01消除G(2)检查点的潜在靶点。
A checkpoint operating in the G(2) phase of the cell cycle prevents entry into mitosis in the presence of DNA damage, UCN-01, a protein kinase inhibitor currently undergoing clinical trials for cancer treatment, abrogates G(2) checkpoint function and sensitizes p53-defective cancer cells to DNA damaging agents. In most species, the G(2) checkpoint prevents the Cdc25 phosphatase from removing inhibitory phosphate groups from the mitosis-promoting kinase Cdc2. This is accomplished by maintaining Cdc25 in a phosphorylated form that binds 14-3-3 proteins. The checkpoint kinases, Chk1 and Cds1, are proposed to regulate the interactions between human Cdc25C and 14-3-3 proteins by phosphorylating Cdc25C on serine 216. 14-3-3 proteins, in turn, function to keep Cdc25C out of the nucleus. Here we report that UCN-01 caused loss of both serine 216 phosphorylation and 14-3-3 binding to Cdc25C in DNA-damaged cells. In addition, UCN-01 potently inhibited the ability of Chk1 to phosphorylate Cdc25C in vitro. In contrast, Cds1 was refractory to inhibition by UCN-01 in vitro, and Cds1 was still phosphorylated in irradiated cells treated with UCN-01. Thus, neither Cds1 nor kinases upstream of Cds1, such as ataxia telangiectasia-mutated, are targets of UCN-01 action in vivo. Taken together our results identify the Chk1 kinase and the Cdc25C pathway as potential targets of G(2) checkpoint abrogation by UCN-01.