Figure 3. AMPK suppresses mTOR activity under hypoxia.
(A–C) Serum-starved HEK293 cells were pretreated with 2 or 10 μM AMPK inhibitor compound C for 30 min before exposure to 2-DG (A), or 21% or 1.5% O2 for 1 or 3 hr (B–C).
(D) Serum replete HEK293 cells were preincubated with 10 mM methylpyruvate (MP) for 2 hr prior to 2-DG or 1.5% O2 (20 hr) treatment. Phosphorylation of eEF2, 4EBP1, and rpS6 are indicated.
(E–F) Kinase-inactive AMPK α2 mutant suppresses hypoxic mTOR inhibition. HEK293 cells were transfected with empty vector, wild-type AMPK-α2 (AMPK-WT), or kinase inactive K45R mutant (AMPK-DN). Clones expressing similar levels of AMPK protein were selected.
(E) Levels of AMPK protein in selected clones.
(F) Phosphorylation of p70S6K and rpS6 in cells exposed to 1.5% O2 for 0–20 hr.