Fig. 5

Sema5A is regulated by the miR-204/c-Myb axis. a Quantitative RT-PCR of miR-204 expression in M14 cells 72 h after transfection with mimic miR-204 or Pre-miRNA Precursor-Negative Control. Values are expressed as means of ratio ± SEM where ratio was calculated considering mimic miR-204 transfected cells versus control. b Western blot analysis of Sema5A, Bcl-2 and c-Myb expression after mir-204 transfection in M14 cells. c Quantitative RT-PCR of miR-204 expression in M14 human melanoma control (empty) and Bcl-2 overexpressing (bcl-2) clones. Samples were normalized using RNU19 as endogenous control. Values are expressed as means of ratio ± SEM where ratio was calculated considering Bcl-2-overexpressing versus control clones. *p-values < 0.05. d Western blot analysis of Sema5A, Bcl-2 and c-Myb expression after 48 h of miR-204 transfection in Bcl-2 overexpressing (bcl-2) clones from M14 and A375SM-SC1 cells. b, d, β-actin or HSP70/72 expression was evaluated to confirm equivalent transfer and loading. Representative images of three independent experiments are reported. a, c Experiments were performed at least three times (three technical replicates). *p-values< 0.05