McAlear M A, Tuffo K M, Holm C
Department of Pharmacology, University of California at San Diego, La Jolla 92093-0651, USA.
Genetics. 1996 Jan;142(1):65-78. doi: 10.1093/genetics/142.1.65.
We used genetic and biochemical techniques to characterize the phenotypes associated with mutations affecting the large subunit of replication factor C (Cdc44p or Rfc1p) in Saccharomyces cerevisiae. We demonstrate that Cdc44p is required for both DNA replication and DNA repair in vivo. Cold-sensitive cdc44 mutants experience a delay in traversing S phase at the restrictive temperature following alpha factor arrest; although mutant cells eventually accumulate with a G2/M DNA content, they undergo a cell cycle arrest and initiate neither mitosis nor a new round of DNA synthesis. cdc44 mutants also exhibit an elevated level of spontaneous mutation, and they are sensitive both to the DNA damaging agent methylmethane sulfonate and to exposure to UV radiation. After exposure to UV radiation, cdc44 mutants at the restrictive temperature contain higher levels of single-stranded DNA breaks than do wild-type cells. This observation is consistent with the hypothesis that Cdc44p is involved in repairing gaps in the DNA after the excision of damaged bases. Thus, Cdc44p plays an important role in both DNA replication and DNA repair in vivo.
我们运用遗传学和生物化学技术，对酿酒酵母中与影响复制因子C（Cdc44p或Rfc1p）大亚基的突变相关的表型进行了表征。我们证明，Cdc44p在体内对于DNA复制和DNA修复均是必需的。温度敏感型的cdc44突变体在α因子阻断后于限制温度下经历S期进程延迟；尽管突变细胞最终积累到G2/M期的DNA含量，但它们会发生细胞周期停滞，既不进行有丝分裂也不启动新一轮的DNA合成。cdc44突变体还表现出自发突变水平升高，并且它们对DNA损伤剂甲磺酸甲酯以及紫外线照射均敏感。在紫外线照射后，处于限制温度的cdc44突变体比野生型细胞含有更高水平的单链DNA断裂。这一观察结果与Cdc44p参与在切除受损碱基后修复DNA缺口的假说相一致。因此，Cdc44p在体内的DNA复制和DNA修复过程中均发挥着重要作用。
