Guo B, Lee H
Northeastern Ontario Regional Cancer Centre, 41 Ramsey Lake Road, Sudbury, Ontario, Canada.
Gene. 2001 Feb 21;264(2):249-56. doi: 10.1016/s0378-1119(01)00332-8.
The Dbf4 protein is the regulatory subunit of Cdc7 serine/threonine kinase, which is essential for entry into S phase. We report here the cloning and initial characterization of the Chinese hamster homologue of yeast DBF4. The deduced ChDbf4 protein contains 676 amino acids with a predicted molecular mass of 75.8 kDa, and shares extensive identity overall with those of human (68%) and mouse (73%). The ChDBF4 mRNA level was barely detectable in the cells arrested in the quiescent stage (G(0)) by isoleucine starvation. When cells in G(0) were released into the cell cycle, the ChDBF4 mRNA level did not significantly change until the cells reached the G(1)/S boundary, when the level rapidly increased and reached approximately 70% of the maximum level that was observed in mid to late S phase. Interestingly, gamma-irradiation rapidly and transiently downregulated the level of ChDBF4 mRNA in asynchronous cell populations. Since Dbf4-Cdc7 kinase is involved in the regulation of replication initiation, which can be transiently downregulated by irradiation (Larner et al., 1994. Mol. Cell. Biol. 14, 1901, our data raise the possibility that the downregulation of DBF4 (and, thus, the Cdc7 kinase activity) by irradiation may play a role in the cell-cycle checkpoint that functions at the G(1)/S transition and in S phase (Lee et al., 1997. Proc. Natl. Acad. Sci. USA 94, 526).
Dbf4蛋白是Cdc7丝氨酸/苏氨酸激酶的调节亚基，对进入S期至关重要。我们在此报告了酵母DBF4的中国仓鼠同源物的克隆及初步特性分析。推导的ChDbf4蛋白含有676个氨基酸，预测分子量为75.8 kDa，与人类（68%）和小鼠（73%）的蛋白在整体上有广泛的同源性。在通过异亮氨酸饥饿阻滞于静止期（G(0)）的细胞中，几乎检测不到ChDBF4 mRNA水平。当处于G(0)期的细胞进入细胞周期时，ChDBF4 mRNA水平直到细胞到达G(1)/S边界时才发生显著变化，此时该水平迅速升高并达到在S期中期至后期观察到的最大水平的约70%。有趣的是，γ射线照射可使异步细胞群体中ChDBF4 mRNA水平迅速且短暂地下调。由于Dbf4 - Cdc7激酶参与复制起始的调控，而复制起始可被照射短暂下调（Larner等人，1994。《分子与细胞生物学》14，1901，我们的数据提示，照射导致的DBF4下调（进而Cdc7激酶活性下调）可能在G(1)/S期转换和S期发挥作用的细胞周期检查点中起作用（Lee等人，1997。《美国国家科学院院刊》94，526）。
