FIGURE 3.

Myotubularin silencing inhibits Akt signaling through downstream mTOR and FoxO effector pathways. HeLa cells were treated with control or myotubularin siRNAs, then serum-starved and stimulated with EGF. A, soluble protein extracts were analyzed by immunoblotting for the indicated proteins, and tubulin immunoblotting was used as a loading control. B, the bands on developed film corresponding to phosphorylated and total 4E-BP1, FoxO3a, FoxO1, FoxO4, and p70S6 kinase were quantified, and the values for phospho-protein/total protein in myotubularin-deficient samples are represented graphically as a percentage of EGF-stimulated control values (mean ± S.D., n = 3; *, p < 0.05; **, p < 0.01).