Fig. 3
BIR domain-mediated inhibition of miR-200a was responsible for XIAP promotion of EGFR protein translational regulation by targeting EGFR mRNA 3′ UTR. a The indicated microRNAs, which could potential bind to EGFR mRNA 3′ UTR, were evaluated by real-time PCR. Results were presented as the mean ± SD from triplicates. The asterisk “*” indicates a significant increase as compared with nonsense transfectant (p < 0.05). b Expression levels of the miR-200a/200b/429 cluster in the indicated three stable cells were evaluated by real-time PCR. Results were presented as the mean ± SD of triplicates. The asterisk “*” indicates a significant change as compared with nonsense transfectant (p < 0.05), while symbol “※” indicates a significant decrease in comparison to vector transfectants (p < 0.05). c miR-200a expression plasmid was stably transfected to T24T cells and the expression efficiency was determined by real-time PCR. Results were presented as the mean ± SD of triplicates. The asterisk “*” indicates a significant increase as compared to that in T24T(Vector) (p < 0.05). d T24T(Vector) and T24T(miR-200a) cells were subjected to Western blotting to determine the EGFR expression. GAPDH was used as the protein loading control. e miR-200a inhibitor lentivirus was used to infect T24T(shXIAP) cells and the knockdown efficiency was determined by real-time PCR. Results were presented as the mean ± SD of triplicates. The asterisk “*” indicates a significant decrease as compared to control (p < 0.05). f The indicated cells were subjected to Western blotting to determine the EGFR expression. GAPDH was used as the protein loading control. g EGFR 3′-UTR Luciferase reporter and its miR-200a binding site point mutation were diagramed as indicated. h Wild-type (WT) and mutated (Mut) of EGFR 3′-UTR Luciferase reporters were stably co-transfected with miR-200a or its scramble vector, and the stable transfectants were used to evaluate for their reporter activity. Results were presented as the mean ± SD of triplicates. The asterisk “*” indicates a significant inhibition as compared with vector transfectant (p < 0.05). Error bars represent S.D.