摘要
目的:研究转化生长因子β(TGF-β)在翻译水平、转录水平对肝癌细胞表达高尔基蛋白73(GP73)的影响。方法:用不同时间(0、1、2、4、8 h),不同浓度(0、1、2 ng/mL)的TGF-β1刺激转染了pcDNA3-Flag-GP73、pcDNA3-Flag-V质粒的HepG2细胞,分别用免疫印迹检测细胞内GP73蛋白水平变化,荧光定量PCR检测细胞内GP73 mRNA表达水平变化;双萤光素酶报告基因检测TGF-β1能否激活GP73基因启动子。结果:在TGF-β1的刺激下,细胞GP73蛋白及mRNA水平均明显上调;TGF-β能激活GP73启动子且使其活性增强6倍左右,但对GP73启动子突变体无作用。结论:TGF-β能在蛋白水平、mRNA水平影响GP73的表达,GP73是TGF-β作用的靶分子。
Objective： To study the effects of transforming growth factor β（TGF-β） on the expression of Golgi protein 73（GP73） in hepatocellular carcinoma cells at the translation level and transcription level. Methods： Human hepatocellular carcinoma cells HepG2 were transfected with pcDNA3-Flag-GP73, pcDNA3-Flag-V plasmid, then we detected the changes of GP73 protein and mRNA in different times（0, 1, 2, 4, 8 h） and different concentrations （0, 1, 2 ng/mL） of using external TGF-β1 stimulation, GP73 protein were detected by Western blotting, the relative expression of GP73 mRNA in cells was detected by real time fluorescence quantitative PCR（qPCR） and the effect of TGF-β1 on GP73 promoter was detected by luciferase reporter gene method. Results： TGF-β1 stimulation led to increased expression of GP73, moreover, TGF-β stimulation significantly enhanced GP73 promoter luciferase activity to about 6 fold, but with no effect on the GP73 promoter mutant. Conclusion： TGF-β can affect the expression of GP73 at protein level and mRNA level in conclusion GP73 is a novel TGF-β target gene.
作者
刘宁
孙鹤鸣
赵怡雯
马雪萍
陈健康
魏从文
钟辉
杨晓莉
LIU Ning;SUN He-Ming;ZHAO Yi-Wen;MA Xue-Ping;CHEN Jian-Kang;WEI Cong-Wen;ZHONG Hui;YANG Xiao-Li(General Hospital of Chinese People＇s Armed Police Forces,Beijing 100039;Xijing Hospital,Fourth Military Medical University,Xi＇an 710032;Beijing Institute of Biotechnology,Beijing 100850,China)
出处
《生物技术通讯》
CAS
2018年第5期636-640,共5页
Letters in Biotechnology
基金
武警部队课题(WZ2010016
WZ2011021)
