Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe.
Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe.
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DOI：
10.1371/journal.pone.0130748
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Hilditch V
中科院分区：
文献类型：
--
作者：
Caspari T;Hilditch V
Cdc 2（CDK 1）激酶的活性，其协调细胞周期进程和DNA断裂修复，在存在DNA损伤的情况下通过Wee 1激酶在酪氨酸15（Y15）处磷酸化而被阻断。Cdc 2如何在被DNA损伤检查点灭活的同时支持DNA修复仍有待解释。人CDK 1在其T环中的苏氨酸161（T167 Sp）处被CDK活化激酶（CAK）修饰之前，在靠近其ATP结合位点的苏氨酸14（T14）处被Myt 1激酶磷酸化。虽然T161的修饰促进与细胞周期蛋白伴侣的结合，但T14的磷酸化抑制CDK 1-细胞周期蛋白复合物。在DNA损伤存在下，Wee 1对Y15的修饰进一步加强了这种抑制作用。在粟酒裂殖酵母中，Cdc 2的显性抑制由Y15的磷酸化提供，并且当细胞处于S期时，只有少量Cdc 2在T14处被修饰。与人类细胞不同，这两种抑制性修饰都是由Wee 1执行的。利用等电聚焦（IEF）和Phos-tag SDS电泳（PT）相结合的IEFPT技术，我们报道了粟酒裂殖酵母Cdc 2激酶的7种存在形式。虽然有五种形式是磷酸化的，但有两种不是。四种磷酸化形式与细胞周期蛋白B（Cdc 13）相关，其中只有两种在Y15被Wee 1修饰。有趣的是，只有一种Y15修饰的物种也携带T14修饰。第五磷酸化形式对细胞周期蛋白B具有低亲和力，并且既不是Y15也不是T14修饰的。这两种非磷酸化形式可能直接促进DNA损伤反应，因为只有它们与DNA损伤检查点激酶Chk 1相关。有趣的是，细胞周期蛋白B也存在于未磷酸化的池中。我们还表明，G146 D突变的Cdc2.1w，这使得Cdc 2不敏感的Wee 1抑制，是异常修改的Wee 1依赖的方式。总之，我们的工作增加了支持的想法，两个不同的Cdc 2池调节细胞周期的进程和DNA损伤的反应。
The activity of Cdc2 (CDK1) kinase, which coordinates cell cycle progression and DNA break repair, is blocked upon its phosphorylation at tyrosine 15 (Y15) by Wee1 kinase in the presence of DNA damage. How Cdc2 can support DNA repair whilst being inactivated by the DNA damage checkpoint remains to be explained. Human CDK1 is phosphorylated by Myt1 kinase at threonine 14 (T14) close to its ATP binding site before being modified at threonine 161 (T167Sp) in its T-loop by the CDK-activating kinase (CAK). While modification of T161 promotes association with the cyclin partner, phosphorylation of T14 inhibits the CDK1-cyclin complex. This inhibition is further enforced by the modification of Y15 by Wee1 in the presence of DNA lesions. In S.pombe, the dominant inhibition of Cdc2 is provided by the phosphorylation of Y15 and only a small amount of Cdc2 is modified at T14 when cells are in S phase. Unlike human cells, both inhibitory modifications are executed by Wee1. Using the novel IEFPT technology, which combines isoelectric focusing (IEF) with Phos-tag SDS electrophoresis (PT), we report here that S.pombe Cdc2 kinase exists in seven forms. While five forms are phosphorylated, two species are not. Four phospho-forms associate with cyclin B (Cdc13) of which only two are modified at Y15 by Wee1. Interestingly, only one Y15-modified species carries also the T14 modification. The fifth phospho-form has a low affinity for cyclin B and is neither Y15 nor T14 modified. The two unphosphorylated forms may contribute directly to the DNA damage response as only they associate with the DNA damage checkpoint kinase Chk1. Interestingly, cyclin B is also present in the unphosphorylated pool. We also show that the G146D mutation in Cdc2.1w, which renders Cdc2 insensitive to Wee1 inhibition, is aberrantly modified in a Wee1-dependent manner. In conclusion, our work adds support to the idea that two distinct Cdc2 pools regulate cell cycle progression and the response to DNA damage.
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影响因子：
56.9
作者：
Choudhary, Chunaram;Kumar, Chanchal;Mann, Matthias
通讯作者：
Mann, Matthias
影响因子：
64.5
作者：
FISHER, RP;MORGAN, DO
通讯作者：
MORGAN, DO
DOI：
10.1073/pnas.97.6.2579
发表时间：
2000-03-14
影响因子：
11.1
作者：
Christensen, PU;Bentley, NJ;Carr, AM
通讯作者：
Carr, AM
DOI：
10.1007/978-1-61779-821-4_5
发表时间：
2012-01-01
期刊：
Methods in molecular biology (Clifton, N.J.)
影响因子：
--
作者：
Arndt, Claudia;Koristka, Stefanie;Bachmann, Michael
通讯作者：
Bachmann, Michael
影响因子：
64.8
作者：
ALFA, CE;DUCOMMUN, B;HYAMS, JS
通讯作者：
HYAMS, JS