人结肠癌细胞中TGF-β1基因-509 C>T单核苷酸多态性对启动转录活性的影响

Influence of-509 C>T single-nucleotide polymorphism on the transcriptional activity of transforming growth factor-beta 1 gene in human colorectal cancer cells

目的探讨在人结肠癌细胞中转化生长因子β1(TGF-β1)基因上游启动子-509 C>T位点多态性对其转录活性的影响。方法以特定-509 C>T基因型患者DNA为模板,用聚合酶链反应(PCR)扩增得到1对长度为2.14 kb(-1 328^+812)含有-509 C>T变异的TGF-β1上游基因片段,并将其与不含启动子的pCAT3-enhancer报告基因载体重组,构建重组体phTGF 2.14C和phTGF 2.14T。用脂质体转染法将2种重组体分别转染至人结肠癌细胞(SW480和LoVo细胞)中,酶联免疫吸附试验(ELISA)测定转染细胞的报告基因CAT活性。结果在SW480和LoVo细胞中,转染重组体phTGF 2.14C细胞的CAT活性均明显高于转染重组体phTGF2.14T细胞的CAT活性(P<0.05)。结论 -509位点C等位基因可明显增强TGF-β1基因上游调控序列的转录活性。

Objective To study the influence of -509 C 〉 T polymorphism on the transcriptional activity of transforming growth factor-beta 1 （TGF-131） gene in human colorectal cancer cells. Methods Sequence -1 328- ＋ 812 （2.14 kb） of TGF-β1 gene containing the -509 C 〉 T was selected as putative promoter by polymerase chain reaction （PCR） amplification. The -1 328-＋812 of TGF-β1 gene and pCAT3-enhaneer gene were recombined, and the phTGF 2.14C and phTGF 2.14T were constructed and transfected into human colorectal cancer cells （SW480 and LoVo） by liposomal transfection method. The transfected colorectal cancer cells were collected to analyze the reporter gene CAT activity by enzyme-linked immunosorbent assay （ELISA）. Results In SW480 and LoVo cells, reporter gene CAT activity in cells transfected with phTGF 2. 14C was significantly higher than that transfected with phTGF 2. 14T （P 〈 0.05）. Conclusions C allele at -509 C 〉 T can increase the promoter activity of TGF--β1 gene in colorectal cancer cells.