Figure 1.

ATG7 knockdown affects extracellular metabolite levels. (A) A schematic diagram of energy metabolism in mammalian cells: During glycolysis, glucose (that contains 6 carbons) is converted into 2 molecules of the 3-carbon metabolite pyruvate via a series of intermediate metabolites. Pyruvate can then either be used to produce lactate (3-carbon molecule which is often secreted from cells) or transferred to the mitochondria for further breakdown in the TCA cycle. The TCA cycle is a series of chemical reactions, which leads to generation of energy through the oxidation of acetate (in the form of acetyl coenzyme A; acetyl Co-A) into carbon dioxide (CO₂) and energy in the form of ATP, which is synthesized following OXPHOS in the mitochondrial ETC. U-¹³C₆ can be used to examine flow of labeled carbons (indicated by red circles) through these pathways. During the conversion of 3-carbon pyruvate to acetyl Co-A, one carbon is converted to CO₂ such that acetyl Co-A maintains 2 pyruvate-derived carbons (labeled) when it enters the TCA cycle. This can lead to build up of labeled carbons in TCA cycle intermediates when the pathway is activated. (B to H) shATG7-expressing K562 cells were cultured in normal medium (B to E) or in the presence of U-¹³C₆ (F to H). Extracellular glucose (B and F), lactate (C and G), glutamate (D and H) and glutamine (E) were measured in the medium by LC-MS following 24 h culture. Note, the significance levels for metabolites with variable labeled carbons, (i.e. Glu+1 => glutamate + 1 × ¹³C) are indicated in figure legends to the right of the graph: +1, etc. corresponds to glutamate containing 1X, etc. ¹³C. Two independent experiments were performed in triplicate. P values: *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001.