Vas A, Mok W, Leatherwood J
Department of Molecular Genetics and Microbiology, State University of New York, Stony Brook, NY 11794-5222, USA.
Mol Cell Biol. 2001 Sep;21(17):5767-77. doi: 10.1128/MCB.21.17.5767-5777.2001.
Cdc2 kinase is a master regulator of cell cycle progression in the fission yeast Schizosaccharomyces pombe. Our data indicate that Cdc2 phosphorylates replication factor Orp2, a subunit of the origin recognition complex (ORC). Cdc2 phosphorylation of Orp2 appears to be one of multiple mechanisms by which Cdc2 prevents DNA rereplication in a single cell cycle. Cdc2 phosphorylation of Orp2 is not required for Cdc2 to activate DNA replication initiation. Phosphorylation of Orp2 appears first in S phase and becomes maximal in G(2) and M when Cdc2 kinase activity is required to prevent reinitiation of DNA replication. A mutant lacking Cdc2 phosphorylation sites in Orp2 (orp2-T4A) allowed greater rereplication of DNA than congenic orp2 wild-type strains when the limiting replication initiation factor Cdc18 was deregulated. Thus, Cdc2 phosphorylation of Orp2 may be redundant with regulation of Cdc18 for preventing reinitiation of DNA synthesis. Since Cdc2 phosphorylation sites are present in Orp2 (also known as Orc2) from yeasts to metazoans, we propose that cell cycle-regulated phosphorylation of the ORC provides a safety net to prevent DNA rereplication and resulting genetic instability.
Cdc2激酶是裂殖酵母粟酒裂殖酵母细胞周期进程的主要调节因子。我们的数据表明，Cdc2使复制因子Orp2磷酸化，Orp2是起始识别复合物（ORC）的一个亚基。Orp2的Cdc2磷酸化似乎是Cdc2在单个细胞周期中防止DNA重新复制的多种机制之一。Orp2的Cdc2磷酸化对于Cdc2激活DNA复制起始并非必需。Orp2的磷酸化首先出现在S期，并在需要Cdc2激酶活性以防止DNA复制重新起始的G2期和M期达到最大值。当限制复制起始因子Cdc18失调时，缺乏Orp2中Cdc2磷酸化位点的突变体（orp2-T4A）比同基因orp2野生型菌株允许更多的DNA重新复制。因此，Orp2的Cdc2磷酸化在防止DNA合成重新起始方面可能与Cdc18的调节冗余。由于从酵母到后生动物的Orp2（也称为Orc2）中都存在Cdc2磷酸化位点，我们提出ORC的细胞周期调节磷酸化为防止DNA重新复制和由此产生的遗传不稳定性提供了一个安全保障。
