Research Paper Volume 12, Issue 16 pp 16304—16325

Figure 6. MiR-520c-3p was a direct regulator of MYCN in CCA cells. (A) MiR-520c-3p restrained MYCN mRNA expression certified by qRT-PCR. (B) MiR-520c-3p refrained MYCN protein expression testified via western blot. (C) The expression of MYCN mRNA in CCA tissues and paired adjacent nontumor bile duct tissues. (D) The correlation between relative MYCN mRNA expression and relative miR-520c-3p expression in CCA tissues. (E) The MYCN mRNA expression in CCLP-1, QBC939, TFK-1, RBE and normal HIBEC. (F) The MYCN protein expression in CCA cells (CCLP-1, QBC939, TFK-1, RBE) and normal HIBEC. (G) Luciferase reporter plasmids were constructed with miR-520c-3p-binding site region of MYCN sequence, including wild type and mutant type. (H) The luciferase activity of MYCN wild type was observably inhibited by miR-520c-3p mimics cotransfection. (I) AGO2 RIP assays were conducted to further demonstrate the binding of miR-520c-3p to 3’UTR of MYCN. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001.