转录因子KLF4与miR-106a的调控关系及其对人胃癌细胞迁移的影响

doi:10.3969/j.issn.1004-616x.2018.05.002

Abstract

Abstract: OBJECTIVE:To construct miR-106a promoter reporting plasmids and to explore their regulatory effects on the migration of human gastric cancer cells with transcription factor Krüppel-like factor 4. METHODS:Using UCSC genomic database to predict the promoter region of miR-106a,and to insert it into pGL3-basic reporter vector for construction of the wild-type reporter plasmid,pmiR-106a-WT-luc,and the point mutation reporter plasmid,pmiR-106a-MUT-luc. Using transcription factor database JASPAR to predict and to screen the transcription factor KLF4 that regulates the promoter region of miR-106a,and to clone it into pcDNA3.0 vector for construction of the over-expression plasmid KLF4-pcDNA3.0. KLF4^+/- -pcDNA3.0 and pmiR-106a-WT/MUT-luc were co-transferred into HEK293T cells and their fluorescence activities were detected by dual luciferase assay. 30 pairs of gastric cancer and adjacent non-tumor tissues were collected and qPCR was used to detect the expression level of miR-106a and KLF4. Transwell assay was used to detect the migratory capacity of human gastric cancer SGC-7901 cells. RESULTS:Both miR-106a reporter and KLF over-expression plasmids were constructed correctly through enzyme digestion,PCR amplification,sequencing and Blast comparison. KLF4-pcDNA3.0 significantly inhibited the luciferase activity of pmiR-106a-WT-luc (P=0.000);whereas, the effect on pmiR-106a-MUT-luc was little (P=0.553). The relative expression of KLF4 was 0.716±0.624 corresponding to 3.367±2.165 for miR-106a in gastric cancer tissues. KLF4-pcDNA3.0 partially blocked the stimulative effect of miR-106a on the migration of gastric cancer SGC-7901 cells (P=0.038). CONCLUSION:Our data show that there was a direct binding site between transcription factor KLF4 and the promoter region of miR-106a. In addition,KLF4 negatively regulated the expression of mature miR-106a at upstream transcription level. Thereby,it played a role in inhibiting the migration of gastric cancer cells induced by miR-106a.

Key words: miR-106a, KLF4, promoter, transcriptional factor, reporter plasmid

CLC Number:

R735.2

ZHU Meng, ZHAI Hualiang, ZHAO Wanying, ZHANG Ning, HE Shuixiang. Regulation of transcription factor KLF4 on miR-106a and their effects on gastric cancer cell migration[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2018, 30(5): 339-344.

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Regulation of transcription factor KLF4 on miR-106a and their effects on gastric cancer cell migration

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