Beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation.
The Wnt/beta-catenin/TCF4 pathway plays critical roles in the maintenance of small intestinal epithelium; however, downstream targets of the beta-catenin/TCF4 complex are not extensively characterized. We identified miR-30e as an immediate target activated by the beta-catenin/TCF4 complex. miR-30e was detected in the peri-nuclear region of the intestinal crypt IEC-6 cells. Bioinformatics analysis revealed clustered beta-catenin/TCF4 binding sites within the miR-30e promoter region. This promoter region was cloned into pGL3-control luciferase reporter vector, with the enhancer region removed. Transfection of pCMV-SPORT6-beta-catenin expression vector dose-dependently increased luciferase activity, and co-transfection of pCMV-SPORT6-TCF4 expression vector further enhanced the promoter activity. Dexamethasone-induced IEC-6 cells differentiation caused a 2.5-fold increase in miR-30e expression, and upon beta-catenin siRNA transfection, miR-30e increased 1.3-fold. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed the binding between beta-catenin/TCF4 complexes from IEC-6 nuclear extracts and the putative sequences in the miR-30e promoter. These results demonstrate that beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation.
Liao
Y
Y
Department of Nutrition, University of California, One Shields Ave., Davis, CA, 95616, USA.
Lönnerdal
B
B
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D003907:Dexamethasone; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D015151:Immunoblotting; D007413:Intestinal Mucosa / Q000166:cytology* / Q000502:physiology; D035683:MicroRNAs / Q000378:metabolism*; D018613:Microscopy, Confocal; D016133:Polymerase Chain Reaction; D000073940:Transcription Factor 4; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology*; D051176:beta Catenin / Q000378:metabolism*
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D017931:DNA Primers; C546086:MIRN30b microRNA, human; D035683:MicroRNAs; C532096:TCF4 protein, human; D000073940:Transcription Factor 4; D014157:Transcription Factors; D051176:beta Catenin; D003907:Dexamethasone