p53 is required for GSK-3β regulation on let-7 activity.

<p>(<b>A</b>) Forty-eight hours after transfecting BG-1 cells with Ctrl or GSK-3β siRNA as explained in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0066330#pone-0066330-g002" target="_blank">Figure 2B</a>, lysates were prepared to assess the levels of p53 and loading control GAPDH by western blot analysis. (<b>B</b>) Forty-eight hours after transfecting BG-1 cells with 50 nM Ctrl siRNA, GSK-3β siRNA, and/or p53 siRNA, lysates were prepared to measure protein levels (<i>top</i>) and cells in the culture media were collected and counted (<i>graph</i>). Signals were quantified by densitometry and normalized to the levels of β-actin (Fold). (<b>C</b>) Twenty-four hours after transfection as described in panel (B), BG-1 cells were transfected with pmirGLO-let7 and luciferase activity was measured 24 h later. (<b>D</b>) Twenty-four hours after transfection with Ctrl or GSK-3β siRNAs, HCT116 p53+/+ and p53−/− cells were transfected with pmirGLO-let7 and luciferase activity was measured 24 h later. Data in (B, C and D) are the means and S.D. from three independent experiments: * <i>P</i><0.05; NS, not significant.