p27^(kip1)过表达对肝细胞癌细胞株细胞周期的影响  

Over-expression of p27^(kip1) affects cell cycle of hepatocellular carcinoma cell lines

作　　者：吴洁[1] 桑瑛颖[1] 陶静[1] 王英明[1] 吴超群[1] 邓可京[1] 乔守怡[1] 

机构地区：[1]复旦大学生命科学学院遗传学研究所遗传工程国家重点实验室,上海200433

出　　处：《复旦学报（医学版）》2007年第3期323-328,共6页Fudan University Journal of Medical Sciences

基　　金：复旦大学遗传工程国家重点实验室"211工程"支持项目

摘　　要：目的探讨肿瘤抑制基因p27kip1编码的P27蛋白过表达与HCC细胞株细胞周期的关系。方法本研究从野生型p27kip1基因真核表达质粒pEYFP-P27(WT)出发,采用定点诱变技术构建了突变型p27kip1基因表达载体pEYFP-P27(S10A)、pEYFP-P27(T157A)和pEYFP-P27(T187A),然后用脂质体法将上述质粒转染HCC细胞株HepG2和Hep3B,Western Blotting检测内源和转入的P27蛋白表达,流式细胞仪检测P27蛋白过表达对HCC细胞株细胞周期的影响。结果野生型和3种突变型的P27融合蛋白在HepG2和Hep3B中的过表达都能增强G0／G1期阻抑;同时,对HepG2细胞,核定位位点突变体P27(T157A)比野生型P27蛋白及其他两个突变体具有更强的细胞周期阻抑活性,而对Hep3B细胞,3个突变体和野生型p27蛋白对细胞周期的影响无明显差异。结论P27蛋白过表达能够显著增强HCC细胞株的G0／G1期阻抑。Purpose To study the effects of P27 protein,encoded by p27^kip1 , a crucial tumor suppression gene, on hepatocellular carcinoma（HCC） cell lines HepG2 and Hep3B, when it is over-expressed. Methods We constructed mutant P27-expressing plasmids, pEYFP-P27 （S10A）, pEYFP-P27 （T157A） and pEYFP-P27（T187A）, by site-mutation technique from pEYFP-P27（WT） which encodes wild type P27 protein. HCC cell lines, HepG2 and Hep3B, were transfected with pEYFP-P27（WT） as well as pEYFPK-P27（mutant）. Innate and over-expressed P27 protein were examined using Western blot, and their effects on cell cycle were analyzed by FACS,respectively. Results It was found that overexpression of all P27 fusion proteins could strengthen G1 phase arrest in HepG2 and Hep3B cells; T157A mutant beared stronger activity of G0/G1 arrest than wild type or other mutant in HepG2, while in Hep3B no remarkable differences were detected between 3 mutants and wild type P27. Conclusions Overexpression of P27 protein can remarkably strengthen G0/G1 phase arrest in HCC cell lines.

关 键 词：P27蛋白 细胞周期 肝细胞癌细胞株 

分 类 号：Q786[生物学—分子生物学]