Tyrosine phosphorylation of cdc2 is required for the replication checkpoint in Schizosaccharomyces pombe.
Tyrosine phosphorylation of cdc2 is required for the replication checkpoint in Schizosaccharomyces pombe.
复制标题
裂殖酵母中的复制检查点需要 cdc2 的酪氨酸磷酸化。
DOI：
10.1128/mcb.18.7.3782
复制
发表时间：
1998
影响因子：
5.3
通讯作者：
Russell,P
中科院分区：
文献类型：
--
作者：
Rhind,N;Russell,P
DNA 复制检查点会抑制无法复制 DNA 的细胞的有丝分裂，就像羟基脲抑制核苷酸生物合成一样。在裂殖酵母粟酒裂殖酵母中，遗传证据表明该检查点涉及通过 tyrosine-15 磷酸化来抑制 Cdc2 活性。相反，最近的一项生化研究表明，Cdc2 在复制检查点期间处于激活状态，这表明 Cdc2 在 tyrosine-15 上的磷酸化并不是复制检查点机制的一部分。我们进行了生化和遗传学研究来解决这一争议。我们报告 DNA 复制检查点位于 S。庞贝病毒在携带等位基因 ecdc2-Y15F 的细胞中被消除，表达非磷酸化形式的 Cdc2。此外，从复制检查点停滞的细胞中分离的Cdc2可以在体外被Cdc25激活，Cdc25是负责体内Cdc2去磷酸化的酪氨酸磷酸酶，其程度与从cdc25ts阻断的细胞中分离的Cdc2相同，表明羟基脲处理导致Cdc2活性维持在不足以诱导有丝分裂的低水平。这些研究表明，Cdc2 的抑制性酪氨酸 15 磷酸化对于 DNA 复制检查点至关重要，并表明 Cdc25 和/或 Wee1 和 Mik1 之一或两者（磷酸化 Cdc2 的酪氨酸激酶）受到复制检查点的调节。
The DNA replication checkpoint inhibits mitosis in cells that are unable to replicate their DNA, as when nucleotide biosynthesis is inhibited by hydroxyurea. In the fission yeastSchizosaccharomyces pombe, genetic evidence suggests that this checkpoint involves the inhibition of Cdc2 activity through the phosphorylation of tyrosine-15. On the contrary, a recent biochemical study indicated that Cdc2 is in an activated state during a replication checkpoint, suggesting that phosphorylation of Cdc2 on tyrosine-15 is not part of the replication checkpoint mechanism. We have undertaken biochemical and genetic studies to resolve this controversy. We report that the DNA replication checkpoint inS. pombeis abrogated in cells that carry the allelecdc2-Y15F, expressing an unphosphorylatable form of Cdc2. Furthermore, Cdc2 isolated from replication checkpoint-arrested cells can be activated in vitro by Cdc25, the tyrosine phosphatase responsible for dephosphorylating Cdc2 in vivo, to the same extent as Cdc2 isolated fromcdc25ts-blocked cells, indicating that hydroxyurea treatment causes Cdc2 activity to be maintained at a low level that is insufficient to induce mitosis. These studies show that inhibitory tyrosine-15 phosphorylation of Cdc2 is essential for the DNA replication checkpoint and suggests that Cdc25, and/or one or both of Wee1 and Mik1, the tyrosine kinases that phosphorylate Cdc2, are regulated by the replication checkpoint.