PKM2 knockdown induces activation of Akt. (A) Si-C and Si-PKM cell lysates were analyzed by immunoblotting for phosphorylation of Akt at Thr308 and Ser473. GAPDH was used to verify equal gel loading. (B) The p-Akt levels were normalized to the loading control and presented as relative conversion to values in Si-C cells. Data are shown as means ± SEM. n = 3. Statistical analyses were carried out using Student’s t-test. Significance: *p < 0.05; **p < 0.001. (C) Si-C and Si-PKM cell lysates were analyzed by immunoblotting with antibodies against phospho-TSC2 and phospho-GSK3β. GAPDH was used as an equal loading control. (D) H1299 cells were transfected with 20 nM of scramble siRNA and PKM2-specific siRNA, respectively. 48 hours after transfection, cells were harvested and analyzed by immunoblotting with the following antibodies: p-Akt, total Akt and PKM2. GAPDH was used as an equal loading control. Similar results were obtained in three independent experiments. Representative data are shown.