Figure 6
MiR-142a-3p binds to the coding region of p53 mRNA. (a) The scheme of the reporter constructs containing the miR-142a-binding site and luciferase reporter. The coding region of p53 was inserted downstream of the luciferase gene. The microRNA-binding sites were predicted by RNA hybrid website. (b) Luciferase reporter assay showed that there was a suppression of the luciferase activity in wild-type but not mutated p53 in a dose-dependent manner in HEK293T cells (mean±s.d., n=3, *P<0.05, ns stands for no significance). (c) Overexpression of the miR-142-3p duplex decreased the protein level of wild-type p53. The construct of Myc-tagged wild-type p53-coding region (CDS) and western blotting result, left panel; Quantification of western blot, right panel. Myc antibody serves as a tag to evaluate the expression of p53. NC stands for negative control of the miR-142-3p duplex. (d) Overexpression of the miR-142-3p duplex had no effect on the protein level of mutated p53. The construct of Myc-tagged mutated p53 CDS and western blot result, left panel; quantification of western blot analysis, right panel.
