Ⅰ型磷酸酶抑制亚基-1对人宫颈癌HeLa细胞增殖的抑制作用  

Effect of PPI1 on Proliferation of Human Cervix Carcinoma Cell Line HeLa

作　　者：曾麒燕[1] 黄毓[2] 曾麟杰 贺菽嘉[1] 张红[1] 

机构地区：[1]广西医科大学生物化学与分子生物学教研室,南宁5300212 [2]南方医院临床实验中心,广州510515 [3]广西贵港市骨科医院,广西贵港537100

出　　处：《基因组学与应用生物学》2009年第5期883-888,共6页Genomics and Applied Biology

基　　金：广西自然科学基金(No.0542077)资助

摘　　要：PPI1(Inhibitor-1of protein phosphatase 1)是Ⅰ型磷酸酶的抑制亚基之一,其活化依赖于35位苏氨酸蛋白激酶(PKA)的磷酸化而发挥抑制作用。本研究目的在于探讨PPI1持续活化型突变体的表达对人宫颈癌细胞株增殖的影响及其可能的作用机制。利用PPI1野生型和活化型突变体表达质粒分别转染HeLa细胞,首先通过H3TdR掺入实验、迁移实验观察PPI1基因对HeLa细胞增殖能力的影响,研究结果表明:在活化型突变体表达的HeLa细胞株中,细胞3H掺入量和迁移能力明显受抑。其次通过流式细胞术、Giemsa染色法分析PPI1对HeLa细胞的细胞周期的影响;FACS分析表明HeLa细胞G2/M期比例明显升高;经脱氧胸苷同步化后,该组细胞进入有丝分裂期明显滞后。最后利用Western blot分析PPI1对MAPK信号转导通路的影响,Western blot分析显示该组细胞的ERK磷酸化水平明显下降。研究表明PPI1活化型突变体的表达可抑制人宫颈癌细胞株的增殖,这与其诱导HeLa细胞G2/M期停滞、有丝分裂的进入延缓有关,其中涉及到MAPK信号转导通路的活化受抑制。Inhibitor-1 of protein phosphatase 1 （PPI1） is one of the regulatory subunits of protein phosphatase I. When phosphorylated by protein kinase A （PKA） at Thr-35, PPI1 inhibited PP 1 activity. In this research, we attempted to reveal the effects of expression of activated mutant ofPPI1 （PPI1/sI1） on cell proliferation and to explore its possible mechanism. Here, we employed the expression plasmids harbouring with wild type and activated mutant of PPI1 gene to transfecte into human cervix carcinoma cell line HeLa, respectively. Firstly, cell proliferation was evaluated by [3H]-TdR incorporation assay and migration assay. The result showed that the [3H]-thymidine incorporation and migration were impaired in HeLa-PPI1/sI1 cells. The analysis of cell cycle of HeLa cells was analyzed by flow cytometry and Giemsa staining presented that the proportion of cells in G2/M phase was higher than the references in transient transfected HeLa cells with activated mutant of PPI1. After released from thymidine arrest, in HeLa-PPI 1/sI 1 cells, the entry of mitosis was delayed, which was demonstrated by Giemsa staining, The further research on detecting the effects of PPI1 on MAPK signal transduction by immunoblotting. The result indicated that the level of ERK phosphorylation was declined sharply. To sum up, the expression of PPI1/sI1 suppresses proliferation of human cervix carcinoma cell line HeLa by inducing G2/M arrest and the delay in mitotic entry, which might involve in the inhibition of ERK activation.

关 键 词：Ⅰ型磷酸酶的抑制亚基-1 HELA细胞 增殖 细胞周期 信号转导 

分 类 号：R737.33[医药卫生—肿瘤]