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为了探究A33核心启动子结肠癌特异性及SV40增强子对其转录水平的影响,该研究通过构建A33核心启动子和带SV40增强子的A33核心启动子(eA33)的荧光素酶报告基因载体pGL3-A33和pGL3-eA33,与内参照pRL-SV40质粒共转染至不同的细胞系中,利用双荧光素酶检测系统检测分析了A33和eA33启动子在不同细胞系中的转录活性。结果显示,A33核心启动子在结肠癌细胞系中具有转录活性低,但结肠癌特异性好的特点,而在其他类型癌细胞中基本没有活性。同时发现,eA33在各类癌细胞中的转录水平与A33相比,均呈大幅度提高,有显著性差异(P<0.01),但SV40增强子能显著增强A33启动子转录活性的同时减低了其结肠癌特异性。这为靶向癌症基因—病毒治疗策略在结肠癌的生物治疗应用中寻找结肠癌特异性的启动子奠定了研究基础。
In order to explore the effect of A33 core promoter colon cancer specificity and SV40 enhancer on its transcriptional level, we constructed the A33 core promoter and the luciferase reporter gene vector pGL3 of the A33 core promoter (eA33) with SV40 enhancer -A33 and pGL3-eA33 were co-transfected into different cell lines with the internal reference pRL-SV40 plasmid. The transcriptional activity of A33 and eA33 promoters in different cell lines was analyzed by dual luciferase detection system. The results showed that the A33 core promoter has the characteristics of low transcriptional activity in colorectal cancer cell lines but good specificity of colon cancer and little activity in other types of cancer cells. At the same time, it was found that the transcriptional level of eA33 in all kinds of cancer cells was significantly increased compared with that of A33 (P <0.01), but SV40 enhancer significantly enhanced the transcription activity of A33 promoter Its colon cancer specificity. This laid the foundation for a targeted cancer gene-viral therapy strategy to find colon-specific promoters in biotherapeutic applications of colon cancer.