With its high-energy phosphate bonds, adenosine triphosphate (ATP) is the main intracellular energy carrier. It also functions in most signaling pathways, as a phosphate donor or a precursor for cyclic adenosine monophosphate. We show here that inositol pyrophosphates participate in the control of intracellular ATP concentration. Yeasts devoid of inositol pyrophosphates have dysfunctional mitochondria but, paradoxically, contain four times as much ATP because of increased glycolysis. We demonstrate that inositol pyrophosphates control the activity of the major glycolytic transcription factor GCR1. Thus, inositol pyrophosphates regulate ATP concentration by altering the glycolytic/mitochondrial metabolic ratio. Metabolic reprogramming through inositol pyrophosphates is an evolutionary conserved mechanism that is also preserved in mammalian systems.

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We have previously analysed the bioenergetic consequences of activating J774.A1 macrophages (MU) with interferon-c (IFN-c) and lipopolysaccharide (LPS) and found that there is a nitric oxide (NO)-dependent mitochondrial impairment and stabilization of hypoxia-inducible factor (HIF)-1a, which synergize to activate glycolysis and generate large quantities of ATP. We now show, using tetramethylrhodamine methyl ester (TMRM) fluorescence and time-lapse confocal microscopy, that these cells maintain a high mitochondrial membrane potential (DW m ) despite the complete inhibition of respiration. The maintenance of high DW m is due to the use of a significant proportion of glycolytically generated ATP as a defence mechanism against cell death. This is achieved by the reverse functioning of F o F 1 -ATP synthase and adenine nucleotide translocase (ANT). Treatment of activated MU with inhibitors of either of these enzymes, but not with inhibitors of the respiratory chain complexes I to IV, led to a collapse in DW m and to an immediate increase in intracellular [ATP], due to the prevention of ATP hydrolysis by the F o F 1 -ATP synthase. This collapse in DW m was followed by translocation of Bax from cytosol to the mitochondria, release of cytochrome c into the cytosol, activation of caspases 3 and 9 and subsequent apoptotic cell death. Our results indicate that during inflammatory activation 'glycolytically competent cells' such as MU use significant amounts of the glycolytically generated ATP to maintain DW m and thereby prevent apoptosis. Activation of murine macrophages (MF) with interferon-g (IFN-g) and lipopolysaccharide (LPS) leads to a mitochondrial defect that is dependent on the release of large quantities of nitric oxide (NO) produced by the inducible NO synthase (iNOS). This NO-dependent mitochondrial defect results in the complete arrest of mitochondrial ATP synthesis by oxidative phosphorylation (OXPHOS). In this situation, macrophages upregulate glycolysis by several fold to generate more ATP. In spite of this increase, the demand for energy of activated MF is such that the cellular [ATP] is reduced by approximately 40% after 12 h. However, at this time there is no difference in the viability of control and activated MF. 1 We have previously shown in Jurkat cells that after inhibition of respiration by exogenous NO, or by endogenous NO generated after treatment with anti-Fas antibody, there is upregulation of glycolysis and hyperpolarization of the mitochondrial membrane potential (DC m ). 2,3 In subsequent experiments we showed that such hyperpolarization occurs only in cells capable of upregulating glycolysis, such as astrocytes, but not in neurons that are unable to do so. 4 In the latter cells there is a rapid and progressive decline of DC m and increased apoptotic cell death. It is known that the onset of apoptosis is associated with a collapse in DC m and an alteration in mitochondrial matrix configuration, [5][6][7] leading to the release of pro-apoptotic proteins into the cytosol. [8][9][10][11][12]

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