Fig. 8. PRAS40 knockdown increases AMPK activity towards TSC2.
Panel A, cell extracts were collected and analyzed via Western blotting using an antibody that recognizes phosphorylated TSC2. Panel B, AMPK kinase activity was examined using an in vitro AMPK activity assay where TSC2 was utilized as the substrate in the presence of AMP. Cells were lysed in 1% NP-40, and AMPK was immunoprecipitated from 150 μg of lysate. The ability of AMPK to phosphorylate TSC2 (T1462) was determined as described under “Materials and Methods.” Results were normalized with immunoprecipitated AMPK which was assessed by immunoblotting. Results are expressed as a percentage of basal scrambled control levels. Each bar graph represents mean ± SE of 3 independent experiments consisting of 3 replicate samples per experiment. (* P< 0.05).