蛋白激酶Cα对人正常肝和肝癌细胞中Ha-ras基因表达的影响  

Effect of PKCα on Ha-ras Gene Promoter Activity in Human Normal Liver and Hepatoma Cell Lines

作　　者：冯怡[1] 柳惠图[1] 高萍[1] 

机构地区：[1]北京师范大学生命科学学院,细胞增殖及调控生物学教育部重点实验室,北京100875

出　　处：《生物化学与生物物理进展》2002年第5期781-785,共5页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金资助项目 (3 9870 3 66)~~

摘　　要：运用基因转染技术 ,将蛋白激酶C(PKCα)cDNA正向插入的真核表达重组质粒pXJ4 1 PKCα ,导入人正常肝细胞 (L 0 2 ) ,经蛋白质免疫印迹等检验 ,表明成功构建了稳定过表达PKCα的人正常肝细胞模型 (LT3) ,用LT3和表达反义PKCα的人肝癌细胞HT6为细胞模型 ,通过RT PCR ,蛋白质印迹 (Westernblot)等分析和进一步运用自行构建的真核表达质粒prasGL3,进行荧光素酶活性检测表明 :过表达PKCα可以促进L 0 2细胞的增殖速率 ,并引起Ha ras基因转录水平上升和Ha ras启动子活性升高 ;反之 ,表达反义PKCα的BEL 74 0 2细胞增殖被抑制 ,Ha ras基因转录水平下降 ,Ha ras启动子活性降低 .通过实验我们首次观察到 ,PKCα亚类对Ha ras癌基因表达的影响与其对Ha ras启动子活性的作用有关 ,PKCα可能参与了对HaThe human normal liver cells (L 02) were transfected with plasmid pXJ41 neo and pXJ41 PKCα respectively by lipofectamine agent and were selected positive clones by using G418. The analysis of RT PCR and Western blot showed that the cell model overexpressing PKCα was constructed successfully. In contrast of control cells (LTC) transfected with pXJ41 neo, the PKCα overexpressing cells (LT3) have enhanced growth rate. The RT PCR analysis showed that the LT3 cells have elevated transciption level of Ha ras gene and the luciferase assay showed that Ha ras gene promoter activity increased, in which the prasGL3 plasmid containing Ha ras promoter was constructed. On the contrary, the BEL 7402 cells (HT6) transfected with antisense PKCα displayed the decrease of growth rate, Ha ras gene transciption level and Ha ras promoter activity compared with the control cells (HTC). The results suggested that the effect of PKCα isoform on Ha ras oncogene expressing was related with the Ha ras promoter activity. It seems that PKCα play a positive role in Ha ras gene expressing regulation.

关 键 词：蛋白激酶CΑ 人正常肝 肝癌细胞 HA-RAS 基因表达 

分 类 号：Q26[生物学—细胞生物学] R735.7[医药卫生—肿瘤]