Figure 3. Regulation of ZEB2 by miR-200 family members.
(A) Phase-contrast micrographs of 2008 and ES-2 cells, showing epithelial and mesenchymal morphologies, respectively. 2008 cells express high levels of miR-200 (left), whereas ES-2 cells do not express miR-200 (right). The asterisks denote cases in which expression values were below the limit of accurate linear quantitation. (B) 2008 (left) or ES-2 (right) cells were either mock transfected, or transfected with luciferase gene constructs containing either no ZEB2 3’ UTR (Control), wild-type ZEB2 3’UTR (wt) or ZEB2 3’ UTR with mutations at all conserved, predicted miR-200 binding sites (mut). Results from luciferase assays performed 24h after transfection are shown. (C) ES-2 cells were co-transfected with one of the three luciferase gene constructs described in (A) plus different quantities of pSM30-miR-200a or pSM30-miR-200b. 1/5x, 1x and 5x indicate the molar quantity of the pSM30-miR expression plasmid relative to the luciferase gene plasmid. Results show the mean and standard error of experiments carried out in triplicate. Firefly luciferase (FL) activity normalized to Renilla luciferase (RL) activity is shown on the y-axis.