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Investigation of A33 Promoter’s Specifically Transcriptional Activity in Colorectal Cancer and the Influence When Combined with SV40 Enhancer
Zhong Dan1, Zheng Shuidi1, Shenghe Kuanzi1, Yuan Sujing1, Zhang Kangjian2*
1Xin Yuan Institute of Medicine and Biotechnology, College of Life Sciences, Zhejiang Sci-Tech University, Hangzhou 310018, China;2Institute of Biochemistry and Cell Biology, Shanghai Institute of Biological Sciences, Chinese Academy
Abstract: In order to investigate A33 core promoter’s specifically transcriptional activity in colorectal cancer (CRC) and to identify its transcriptional activity combined with SV40 enhancer， the transcriptional regulatory elements were constructed by appending a SV40 enhancer 5’ to the A33 core promoter (eA33)， and then we constructed two luciferase reporter gene vectors which containing A33 promoter and eA33 promoter， pGL3-A33 and pGL3-eA33， and made them co-transfected with transfection-efficiency normalization vector pRL-SV40 into different cell lines by liposome transfection. The transcriptional activity of A33 and eA33 promoter was analyzed by Dual-Luciferase Assay system and relative luciferase unit (RLU) was used to evaluate the expression efficiency. The results showed that A33 promoter performs high CRC specific expression but low transcriptional activity， and showed barely any activity in other types of cancer cells. Meanwhile， the eA33 promoter’s transcriptional activity is slightly stronger than the A33 promoter’s in all kinds of cell lines. Therefore， the SV40 enhancer is able to greatly up-regulate A33 promoter transcriptional activity， and by the meantime， attenuate the CRC specific of A33 promoter. It will definitely provide useful information for targeted gene-viro-therapy of CRC.
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