S6 kinase (S6K) deletion in metazoans causes small cell size, insulin hypersensitivity, and metabolic adaptations; however, the underlying molecular mechanisms are unclear. Here we show that S6K-deficient skeletal muscle cells have increased AMP and inorganic phosphate levels relative to ATP and phosphocreatine, causing AMP-activated protein kinase (AMPK) upregulation. Energy stress and muscle cell atrophy are specifically triggered by the S6K1 deletion, independent of S6K2 activity. Two known AMPK-dependent functions, mitochondrial biogenesis and fatty acid β-oxidation, are upregulated in S6K-deficient muscle cells, leading to a sharp depletion of lipid content, while glycogen stores are spared. Strikingly, AMPK inhibition in S6K-deficient cells restores cell growth and sensitivity to nutrient signals. These data indicate that S6K1 controls the energy state of the cell and the AMPK-dependent metabolic program, providing a mechanism for cell mass accumulation under high-calorie diet.