过表达核苷二磷酸X水解酶21(NUDT21)通过阻断P53/CDK2/Rb通路抑制HCT-116结肠癌细胞的增殖

Over-expression of NUDT21 inhibits the proliferation of HCT-116 colon cancer cells via blocking P53/CDK2/Rb pathway

目的探讨过表达核苷二磷酸X水解酶21(NUDT21)对HCT-116结肠癌细胞增殖的影响及可能的机制。方法Gibson组装法构建NUDT21过表达质粒。克隆形成实验、CCK-8法检测细胞增殖活性,流式细胞术检测细胞周期变化,Western blot法检测P53、细胞周期蛋白依赖激酶2(CDK2)的蛋白水平,以及第780位丝氨酸磷酸化的视网膜母细胞瘤蛋白(p-Rb-Ser780)和p-Rb-Ser608的蛋白水平。结果测序结果提示NUDT21过表达载体构建成功,HCT-116细胞转染NUDT21过表达质粒后,其mRNA和蛋白水平表达量明显升高。过表达NUDT21抑制结肠癌HCT-116细胞增殖,细胞周期阻滞于G0/G1期;过表达NUDT21后,CDK2、p-Rb-Ser608与p-Rb-Ser780蛋白表达量降低,P53蛋白水平增高。结论过表达NUDT21通过阻断P53/CDK2/Rb信号通路抑制结肠癌HCT-116细胞的增殖。

Objective To investigate the effect of over-expression of nudix hydrolase 21(NUDT21)on the proliferation of colon cancer HCT-116 cells and its mechanism.Methods The NUDT21 over-expression plasmid was constructed by Gibson assembly.The colony formation assay and CCK-8 assay were used to detect cell proliferation.The cell cycle of HCT-116 cells was detected by flow cytometry.Western blot was performed to detect the expressions of P53,cyclin-dependent kinase 2(CDK2),phosphorylated retinoblastoma protein at serine 780(p-Rb-Ser780),and p-Rb-Ser608.Results The sequencing results showed that the NUDT21 over-expression plasmid was successfully constructed.After the NUDT21 over-expression plasmid was transfected into HCT-116 cells,the expressions of NUDT21 mRNA and protein in the cells were significantly increased.The over-expression of NUDT21 inhibited the proliferation of HCT-116 cells and arrested the cell cycle in G0/G1 phase.The expressions of CDK2,p-Rb-Ser608,and p-Rb-Ser780 proteins decreased while the expression of P53 protein increased.Conclusion Over-expression of NUDT21 inhibits the proliferation of HCT-116 cells by blocking P53/CDK2/Rb signal pathway.