Study On The Interaction Between Transcription Factor TCF7l2and Ide

PART1DETECTION OF TCF7L2BINDING TO IDE GENEPROMOTERS BY CHROMATINIMMUNOPRECIPITATIONOBJECTIVE: To identify the region in IDE gene promoter where thetranscriptional factor TCF7L2can bind.METHODS: Human hepatoma HepG2cell were obtained as the studysubjects. Chromatin immunoprecipitation and PCR were performed usingthe antibody specific for TCF7L2to verify the binding of TCF7L2to IDEpromoter.RESULTS: IDE gene specific fragments were detected in the DNAfragments immunoprecipitated by antibody specific for TCF7L2.CONCLUSIONS: TCF7L2protein could bind to the promoterregions of IDE gene in HePG2cells, and may involved in regulation ofIDE gene expression．The result may lay a theoretical foundation forstudying the effect of TCF7L2RNA interference on expression of insulin degrading enzyme. PART2EFFECTS OF GENE SILENCE OF TCF7L2-TARGETINGON EXPRESSION OF INSULIN DEGRADING ENZYMEOBJECTIVE: To study the effect of TCF7L2on expression of insulindegrading enzyme, we constructed the shRNA lentiviral RNAi vectortargeting TCF7L2gene (LV-TCF7L2-shRNA), and detect the silencingeffect to TCF7L2gene in the Human hepatoma HepG2cells,then furtherstudied the effect of TCF7L2on expression of insulin degrading enzyme.METHODS：1. Specific shRNA lentiviral vetor targeting a sequenceof human TCF7L2mRNA coding region (LV-TCF7L2-shRNA) andnegative control vector (LV-NC-GFP-shRNA) were constructed.2. HePG2cells were divided into3groups: interference group，negative control group and blank control group. The groups wereadministrated with LV-TCF7L2-shRNA and LV-NC-GFP-shRNA andnothing respectively.3. The changes of mRNA of TCF7L2and IDE were analyzed usingFQ-PCR，the Protein level of TCF7L2and IDE were detected by Western blot.RESULTS:1.TCF7L2mRNA and protein expression inLV-TCF7L2-shRNA group cells were down regulated compared withLV-NC-GFP-shRNA group and blank Control group cells (P<0.05).2. The level of IDE mRNA and protein expression was significantlydecreased after the suppression of TCF7L2expression in HePG2cells(P<0.05).CONCLUSIONS: The lentivirus RNAi vector of LV-TCF7L2-shRNAwas constructed successfully. LV-TCF7L2-shRNA effectively inhibitedIDE expression. Thus, it suggested that TCF7L2may be an importanttranscription factor involving in regulation of IDE gene expression...