Figure 3
MiR-203 posttranscriptionally regulates ΔNp63 expression by targeting the 3'-UTR of ΔNp63. (A, B) 1 × 106 cells (Eca109 or TE-1) were cotransfected with 50 pmol of miR-203 (or control microRNA) and 1 μg of Luc-ΔNp63 (or Luc-ΔNp63-mut) plasmid, respectively. Luciferase reporter assay were performed at 48 hr posttransfection. Results showed that cells cotransfected with miR-203 and Luc-ΔNp63 plasmid exhibited a significant decrease of reporter activity in comparison with those cotransfected with the control microRNA and Luc-ΔNp63 plasmid (a). However, the reporter activity of cells cotransfected with miR-203 and Luc-ΔNp63-mut plasmid showed no significant difference with that of cells cotransfected with control microRNA and Luc-ΔNp63-mut plasmid (b). (C, D) Cells (1 × 106) of Eca109 and TE-1 were transfected with 50 pmol of miR-203 (or control microRNA), respectively. The expression level of ΔNp63 protein was detected by Western Blot at 48 hr posttransfection and normalized to that of β-actin. Results showed that the level of ΔNp63 protein was significantly decreased in cells transfected with miR-203 as compared to the cells transfected with control microRNA. Results showed that the expression level of ΔNp63 protein was significantly decreased in cells transfected with miR-203 as compared to the cells transfected with control microRNA. (E) Cells (1 × 106) of Eca109 and TE-1 were transfected with 50 pmol of miR-203 (or control microRNA), respectively. The expression level of ΔNp63 mRNA was detected by qRT-PCR at 48 hr posttransfection and normalized to that of GAPDH. Results showed that the expression level of ΔNp63 mRNA exhibited no significantly difference between cells transfected with miR-203 and those transfected with control microRNA. Data represent mean ± SEM from 4 independent experiments; **, P < 0.01 by t test.