细胞周期蛋白依赖性激酶1在S期细胞检测点的激活  

Activity of CDK1 in S phase cell checkpoint

作　　者：高霞 龙浩成 潘志坚 龙志雄 龚建平[3] 

机构地区：[1]武汉市第五医院肿瘤科,430050 [2]武汉市第五医院普外科,430050 [3]华中科技大学同济医学院肿瘤研究所

出　　处：《肿瘤研究与临床》2012年第3期165-168,共4页Cancer Research and Clinic

摘　　要：目的以细胞周期特异性T淋巴细胞型白血病Molt-4细胞S期凋亡为模式，检测细胞周期蛋白依赖性激酶1（CDK1）在s期细胞周期检测点蛋白磷酸化及蛋白去磷酸化，探讨细胞周期时相特异性细胞凋亡的CDK1的活性改变。方法对数生长期的人类急性淋巴细胞白血病细胞系Molt-4经喜树碱（CPT）诱导细胞凋亡，经不同的作用时间、剂量处理后，以API法检测细胞凋亡的细胞周期特异性，以分选后激光共聚焦荧光显微镜技术（PSC）对API结果进一步验证，从而获取最佳条件下细胞周期特异性细胞凋亡，蛋白电泳分析法检测CDK1的磷酸化、去磷酸化。结果对数生长期的Molt-4细胞经CTP诱导后发生细胞周期特异性细胞凋亡，CPT0．2μg／ml处理后6h出现S期特异性细胞凋亡。CTP诱导出现S期特异性细胞凋亡时CDK1-第15位酪氨酸（Tyr15）与对照Westernblot结果条带相近，而CDK1-第161位苏氨酸（Thr161）条带较窄。结论细胞凋亡往往伴有细胞周期特异性；经PSC形态学证实的API流式细胞术是有效、快捷的细胞周期特异性细胞凋亡分析方法；CDK1在S期特异性细胞凋亡时磷酸化活性降低。Objective To investigate the phosphorylation and dephosphorylation of CDK1 based on the specific cell cycle apoptosis in Moh-4 cells and active variety of CDK1 in cell cycle specific apoptosis. Methods The exponential phase of growth Molt-4 cells （the human acute lymphoblastic leukemia cell line） were induced with dose response and time course of Camptothecin （CPT）. The specific cell cycle apoptosis was detected with API method, then cell apoptosis was verified with post sorting confocal method. Meanwhile, the phosphorylation and dephosphorylation of CDK1 were detected by the protein electrophoretic analysis （Western blot）. Results The specific cell cycle apoptosis occurred on exponential phase of growth Moh-4 cells after CPT treatment.When Molt-4 cells occured S-phase apoptosis, the apoptosis cell phosphorylation of CDK1-Thr161 band was more narrow than that of control cells, the apoptosis cell phosphorylation of CDK1-Thr15 hand almost had the same band with control cells. Conclusion Cell apoptosis frequently developed in different cell cycle phase. API assay is quick and efficient method to analyze specific cell cycle apoptosis. When cell apoptosis take place in S-phase ,the phosphorylation activity of CDK1 is inhibited.

关 键 词：细胞周期 细胞凋亡 CDK1活性 

分 类 号：R363[医药卫生—病理学]