抗酶1基因转染对HeLa细胞增殖及细胞周期的抑制作用  

Inhibitory Effect of Antizyme 1 Transfection on Cell Proliferation and Cell Cycle in HeLa Cells

作　　者：姜立[1] 马文丽[1] 李晋[2] 彭翼飞[1] 徐兵[3] 郑文岭[1] 

机构地区：[1]南方医科大学基因工程研究所,广州510515 [2]南方医科大学人体解剖学教研室,广州510515 [3]南方医科大学附属南方医院,广州510515

出　　处：《中国生物化学与分子生物学报》2007年第12期1006-1011,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：广东省自然科学基金(No.5004737)资助~~

摘　　要：研究抗酶(antizyme)1对人宫颈癌HeLa细胞增殖与细胞周期的影响,并分析抗酶1对细胞周期蛋白D1(cyclin D1)的表达影响.采用定点突变技术,将抗酶1的frameshift位点缺失,随后将突变基因重组至真核表达载体pEGFP-N1中,鉴定后转染HeLa细胞.通过MTT法检测细胞增殖变化,流式细胞术分析抗酶1对细胞周期的影响.RT-PCR和Western印迹检测抗酶1转染对细胞周期蛋白D1基因表达的影响.酶切结果显示,抗酶1突变基因成功克隆至pEGFP-N1中.成功转染HeLa细胞后,检测结果显示,抗酶1能够减慢HeLa细胞增殖速度,并使细胞停滞于G0/G1期,细胞周期蛋白D1基因的表达同时受到抑制.实验说明,抗酶1基因能够抑制HeLa细胞增殖,通过降低细胞周期蛋白D1的表达阻滞细胞周期.To investigate the effect of antizyme 1 on the cell proliferation and cell cycle in connection with cyclin D1 expression in HeLa cells, a frame-shift mutant antizyme 1 was created by site-directed mutagenesis, then cloned into a pEGFP-N1 vector. After transfection into HeLa cells, the cell proliferation was determined by MTT assay and the cell cycle was characterized by flow cytometry. The level of cyclin D1 in response to antizyme 1 was determined by RT-PCR and Western blotting. The results showed that the mutant antizyme 1 inhibited cell proliferation and the cell cycle was arrested at G0/G1 phase. Meanwhile, the expression of cyclin D1 was down-regulated. The data suggested that the reduced expression of cyclin D1 might be important for the anti-proliferation effect of antizymel in HeLa cells.

关 键 词：抗酶 细胞增殖 细胞周期 细胞周期素D1 

分 类 号：Q78[生物学—分子生物学]