A yeast GSK-3 kinase Mck1 promotes Cdc6 degradation to inhibit DNA re-replication.
A yeast GSK-3 kinase Mck1 promotes Cdc6 degradation to inhibit DNA re-replication.
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DOI：
10.1371/journal.pgen.1003099
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发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Yoshida S
中科院分区：
文献类型：
--
作者：
Ikui AE;Rossio V;Schroeder L;Yoshida S
CDC6P是复制前复合体(Pre-RC)的重要组成部分，它与DNA复制起点结合，促进DNA复制的启动。DNA复制在每个细胞周期中只发生一次。在启动之后，拆卸预RC组件以防止重新复制。已有研究表明，CDC6p的N-末端区域被细胞周期蛋白依赖性激酶(CDK)磷酸化后被靶向降解。在这里，我们表明，Mck1p，一种酵母GSK-3激酶的同源物，也是通过一个不同的机制降解CDC6所必需的。CDC6是一种不稳定的蛋白质，在野生型细胞中仅在G1期和S期早期积累在细胞核中。在MCK1缺失细胞中，CDC6p稳定并积聚在细胞核中，甚至在S晚期和有丝分裂时也是如此。Mck1p的过表达诱导了CDC6p的快速降解，其方式依赖于苏氨酸-368(GSK-3的共同磷酸化位点)和SCFCDC4。我们的证据表明，依赖Mck1p的CDC6的降解是防止DNA重新复制所必需的。Mck1活性的丧失会导致与其他先前参与再复制控制的前RC突变体的合成致死性，并且这些双突变株在单个细胞周期内过度复制DNA。这些结果表明，GSK3家族蛋白在防止酿酒酵母中通过降解CDC6而防止DNA过度复制方面发挥了意想不到的作用。我们认为CDK和Mck1激酶都是CDC6降解所必需的，以确保对DNA复制的严格控制。DNA复制是发生在所有生物体内的一个基本的细胞过程。这一细胞事件必须受到严格的监管，以确保准确的基因组完整性，以便每个细胞周期只进行一次DNA复制。在这里，我们展示了在酿酒酵母中，细胞周期蛋白依赖性激酶(CDK)和酵母GSK-3激酶(Mck1p)控制DNA再复制的机制。我们发现Mck1p促进了CDC6蛋白的降解。Mck1p通过GSK-3共有位点(T368)靶向CDC6p，CDC6p蛋白的降解也通过同一T368位点介导。GSK-3在高等真核生物中具有多种细胞功能，包括在肿瘤发生中的作用。这一发现尤其重要，因为这是第一个证明GSK-3家族的激酶调节DNA复制的证据。
Cdc6p is an essential component of the pre-replicative complex (pre-RC), which binds to DNA replication origins to promote initiation of DNA replication. Only once per cell cycle does DNA replication take place. After initiation, the pre-RC components are disassembled in order to prevent re-replication. It has been shown that the N-terminal region of Cdc6p is targeted for degradation after phosphorylation by Cyclin Dependent Kinase (CDK). Here we show that Mck1p, a yeast homologue of GSK-3 kinase, is also required for Cdc6 degradation through a distinct mechanism. Cdc6 is an unstable protein and is accumulated in the nucleus only during G1 and early S-phase in wild-type cells. In mck1 deletion cells, CDC6p is stabilized and accumulates in the nucleus even in late S phase and mitosis. Overexpression of Mck1p induces rapid Cdc6p degradation in a manner dependent on Threonine-368, a GSK-3 phosphorylation consensus site, and SCFCDC4. We show evidence that Mck1p-dependent degradation of Cdc6 is required for prevention of DNA re-replication. Loss of Mck1 activity results in synthetic lethality with other pre-RC mutants previously implicated in re-replication control, and these double mutant strains over-replicate DNA within a single cell cycle. These results suggest that a GSK3 family protein plays an unexpected role in preventing DNA over-replication through Cdc6 degradation in Saccharomyces cerevisiae. We propose that both CDK and Mck1 kinases are required for Cdc6 degradation to ensure a tight control of DNA replication. DNA replication is a fundamental cellular process that takes place in all living organisms. This cellular event has to be tightly regulated to ensure an accurate genome integrity such that DNA replication takes place only once per cell cycle. Here we show a mechanism by which DNA re-replication is controlled by Cyclin Dependent Kinase (CDK) and a yeast GSK-3 kinase (Mck1p) in S. cerevisiae. We found that Mck1p promoted Cdc6 protein degradation. Mck1p targets Cdc6p through a GSK-3 consensus site (T368), and Cdc6p protein degradation was also mediated through the same T368 site. The GSK-3 kinase has diverse cellular functions in higher eukaryotes including roles in tumorigenesis. This finding is particularly important, since this is the first evidence to show that a GSK-3 family kinase regulates DNA replication.
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DOI：
10.1016/s0006-291x(03)01082-9
发表时间：
2003-07-11
影响因子：
3.1
作者：
Luo, KQ;Elsasser, S;Campbell, JL
通讯作者：
Campbell, JL
影响因子：
10.5
作者：
EPSTEIN, CB;CROSS, FR
通讯作者：
CROSS, FR
影响因子：
16
作者：
Kõivomägi M;Valk E;Venta R;Iofik A;Lepiku M;Morgan DO;Loog M
通讯作者：
Loog M
影响因子：
3.3
作者：
Archambault, V;Li, CHX;Cross, FR
通讯作者：
Cross, FR
影响因子：
3.3
作者：
Honey, Sangeet;Futcher, Bruce
通讯作者：
Futcher, Bruce