Bao S, Shen X, Shen K, Liu Y, Wang X F
Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Cell Growth Differ. 1998 Dec;9(12):961-7.
Cell cycle checkpoint proteins play critical roles in maintaining genomic stability and integrity to prevent the development of cancer and hereditary diseases. Here we report the isolation of a novel mouse gene encoding the protein MmRad24 [MmRad24 is the mouse homologue of HRad17, which was described recently by A. E. Parker et al. (J. Biol. Chem., 273: 18340-18346, 1998)], which shares significant sequence and structural homology with the budding yeast Rad24 and its fission yeast counterpart Rad17, both of which are required for DNA damage checkpoints. Confocal microscopy revealed that the green fluorescent protein-tagged MmRad24 protein is localized to the nucleus in living cells. Fluorescence-activated cell-sorting analysis showed that overexpression of the wild-type MmRad24 in diploid fibroblast WI-38 cells caused a significant G2 arrest of the cell cycle, whereas overexpression of a mutant MmRad24 (mutated on the nucleotide-binding site) that likely functions as a dominant-negative protein resulted in a defect in cell cycle arrest after DNA damage treatment as measured by bromodeoxyuridine pulse-chase labeling experiments. Taken together, these results suggest that the mammalian Rad24 protein may function as a critical gatekeeper in DNA damage checkpoint control.
细胞周期检查点蛋白在维持基因组稳定性和完整性以预防癌症和遗传性疾病的发生中起着关键作用。在此，我们报告分离出一个编码MmRad24蛋白的新型小鼠基因[MmRad24是HRad17的小鼠同源物，A. E. Parker等人最近对其进行了描述（《生物化学杂志》，273: 18340 - 18346，1998年）]，它与芽殖酵母的Rad24及其裂殖酵母对应物Rad17具有显著的序列和结构同源性，这两者都是DNA损伤检查点所必需的。共聚焦显微镜显示，绿色荧光蛋白标记的MmRad24蛋白定位于活细胞的细胞核中。荧光激活细胞分选分析表明，在二倍体成纤维细胞WI - 38中过表达野生型MmRad24会导致细胞周期显著停滞于G2期，而过表达一种可能作为显性负性蛋白起作用的突变型MmRad24（在核苷酸结合位点发生突变），通过溴脱氧尿苷脉冲追踪标记实验测量，会导致DNA损伤处理后细胞周期停滞出现缺陷。综上所述，这些结果表明哺乳动物的Rad24蛋白可能在DNA损伤检查点控制中作为关键的守门人发挥作用。
