Temporal system-level organization of the switch from glycolytic to gluconeogenic operation in yeast

Abstract

• The diauxic shift involves three main events: a reduction in the glycolytic flux and the production of storage compounds before glucose depletion; the reversion of carbon flow through glycolysis and onset of the glyoxylate cycle operation upon glucose exhaustion; and the shutting down of the pentose phosphate (PP) pathway with a change in the source of NADPH regeneration.

• The redistribution of fluxes toward the production of storage compounds prior glucose depletion drives glycolytic reactions closer to equilibrium, which is essential for the reversion of fluxes upon glucose exhaustion.

• The onset of the glyoxylate cycle is quantitatively more important than the activation of the tricarboxylic acid cycle for growth on ethanol.

• Flux through the PP pathway is halted in the later stages of the adaptation and NADPH regeneration is taken over by NADP-dependent enzymes in the glyoxylate cycle and ethanol metabolism.

Abstract

The diauxic shift in Saccharomyces cerevisiae is an ideal model to study how eukaryotic cells readjust their metabolism from glycolytic to gluconeogenic operation. In this work, we generated time-resolved physiological data, quantitative metabolome (69 intracellular metabolites) and proteome (72 enzymes) profiles. We found that the diauxic shift is accomplished by three key events that are temporally organized: (i) a reduction in the glycolytic flux and the production of storage compounds before glucose depletion, mediated by downregulation of phosphofructokinase and pyruvate kinase reactions; (ii) upon glucose exhaustion, the reversion of carbon flow through glycolysis and onset of the glyoxylate cycle operation triggered by an increased expression of the enzymes that catalyze the malate synthase and cytosolic citrate synthase reactions; and (iii) in the later stages of the adaptation, the shutting down of the pentose phosphate pathway with a change in NADPH regeneration. Moreover, we identified the transcription factors associated with the observed changes in protein abundances. Taken together, our results represent an important contribution toward a systems-level understanding of how this adaptation is realized.