小鼠白蛋白增强子在肝癌细胞系中无增强功能

Mouse albumin enhancer lost its enhancement function in hepatoma cell line

目的 :鉴定小鼠白蛋白增强子序列在肝癌细胞系中对白蛋白启动子转录活性的影响。方法 :以绿色荧光蛋白 (GFP)基因作为报告基因 ,将小鼠白蛋白增强子序列 (_10 .2 1～_8.4 3kb)之间的不同区段与白蛋白启动子相融合 ,转染不同组织来源细胞系 ,用荧光显微镜和流式细胞仪分别对GFP的表达情况进行分析。结果 :瞬时转染小鼠肝癌细胞系Hepa 1_6 ,上游的不同增强子区段均不能增强白蛋白启动子的转录活性 ,相反 ,它们都不同程度地下调了启动子的转录活性 ;稳定转染期 ,E1E3能极弱地增强启动子的转录活性。稳定转染人肝癌细胞系HepG2 ,上游的不同增强子区段均能抑制白蛋白启动子的转录活性 ,导致报告基因不能表达。在非肝脏组织的细胞系中 ,小鼠白蛋白启动子无转录活性。结论 :只使用白蛋白启动子在肝癌细胞系中表达目的基因较同时使用白蛋白启动子和增强子效果会更好。

Objective:To identify the enhancement function of mouse albumin enhancer in the transcription activity of albumin promoter in hepatoma cell lines. Methods:Green fluorescent protein (GFP) gene was used as a reporter gene to study the enhancement function of mouse albumin enhancer in the cells derived from the different tissues. Mouse albumin enhancer fragments of -10.21?kb to -8.43?kb and albumin promoter were fused with the 5′-end of GFP gene in vector pEGFP-C1. The identified recombinant vector and its control vector only containing albumin promoter were transfected into Hepa 1-6 (mouse hepatoma cell line), HepG2 (human hepatoma cell line), CHO (Chinese hamster ovary cell line) and PLA801(human lung cancer cell line), respectively. Results:Albumin promoter could drive GFP transient and stable expression in Hepa 1-6 and HepG2, but could not drive GFP stable expression not in CHO and PLA801. These enhancer fragments could not enhance the transcription of albumin promoter when transiently transfected in Hepa 1-6 and stably transfected in HepG2. On the contrary, these enhancer fragments suppressed the transcription activity of mouse albumin promoter. Albumin promoter can not transcribe the interest gene in non-liver derived cell lines. Conclusion:It is better to use only albumin promoter to express a gene in hepatoma cell lines than to use albumin promoter and its enhancer.