Cell cycle checkpoint control

在减数分裂过程中，DNA双链断裂(DSB)引发的高水平重组只有在DNA复制之后才会发生。然而，DSB的形成如何与DNA复制相耦合尚不清楚。我们研究了几种DNA复制蛋白在裂殖酵母中这种偶联过程中的作用，现在发现脱氧核糖核苷酸合成的限速酶和DNA合成抑制剂羟基脲(HU)的靶标--核糖核苷酸还原酶(RNR)间接地参与了与DNA复制相关的DSB的形成。在DNA复制检查点蛋白Rad1p、Rad3p、Rad9p、Rad17p、Rad26p、Huslp或CDslp功能受损的细胞中，在没有或存在Hu的情况下，DSB的形成频率相似。经HU处理的突变细胞中的双链断裂发生在正常位置，并与重组相关。我们认为，在真核细胞的有丝分裂和减数分裂过程中，激酶CDC2p是进入和通过核分裂进入和进行的中心调节因子。在有丝分裂开始时，CDc2p通过酪氨酸-15的去磷酸化而被激活，酪氨酸-15的磷酸化状态主要由激酶Weel p和磷酸酶CDc25p决定。在分裂酵母中，在减数分裂过程中，许多基因的表达都需要叉头型转录因子Mei4p，其中mei4突变细胞在减数分裂I之前就被阻止了。然而，mei4细胞中细胞周期停滞的机制尚不清楚。我们现在发现CDC25+是Mei4p控制进入减数分裂I的一个重要靶点。强迫酪氨酸-15上的CDc2p去磷酸化从而在没有形成孢子的情况下立即诱导mei4突变细胞的减数分裂I。我们认为，Mei4p通过与其他减数分裂事件协同激活CDC25+转录，发挥减数分裂I的限速调节作用。

During meiosis, high levels of recombination initiated by DNA double-strand breaks (DSBs) occur only after DNA replication. How DSB formation is coupled to DNA replication is unknown, however. We examined several DNA replication proteins for a role in this coupling in Schizosaccharomyces pombe and now show that ribonucleotide reductase (RNR), the rate-limiting enzyme of deoxyribonucleotide synthesis and the target of the DNA synthesis inhibitor hydroxyurea (HU), is indirectly required for DSB formation linked to DNA replication. In cells in which the function of the DNA replication checkpoint proteins Rad1p, Rad3p, Rad9p, Radl7p, Rad26p, Huslp, or Cdslp was compromised, however, DSB formation occurred at similar frequencies in the absence or presence of HU. The DSBs in the HU-treated mutant cells occurred at normal sites and were associated with recombination. We propose that the sequence of meiotic S phase and initiation of recombination is coordinated by DNA replication checkpoint proteins.The kinase Cdc2p is a central regulator of entry into and progression through nuclear division during mitosis and meiosis in eukaryotes. Cdc2p is activated at the onset of mitosis by dephosphorylation on tyrosine-15, the phosphorylation status of which is determined mainly by the kinase Weel p and the phosphatase Cdc25p. In fission yeast, the forkhead-type transcription factor Mei4p is required for expression of many genes during meiosis, with mei4 mutant cells arresting before meiosis I. The mechanism of cell cycle arrest in mei4 cells has remained unknown, however. We now show that cdc25+ is an important target of Mei4p in control of entry into meiosis I. Forced dephosphorylation of Cdc2p on tyrosine-15 thus induced meiosis I in mei4 mutant cells without a delay, although no spores were formed. We propose that Mei4p acts as a rate-limiting regulator of meiosis I by activating cdc25+ transcription in coordination with other meiotic events.