Adaptations in metabolism and protein translation give rise to the Crabtree effect in yeast

Abstract

Aerobic fermentation, also referred to as the Crabtree effect in yeast, is a well-studied phenomenon that allows many eukaryal cells to attain higher growth rates at high glucose availability. Not all yeasts exhibit the Crabtree effect, and it is not known why Crabtree-negative yeasts can grow at rates comparable to Crabtree-positive yeasts. Here, we quantitatively compared two Crabtree-positive yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe, and two Crabtree-negative yeasts, Kluyveromyces marxianus and Scheffersomyces stipitis, cultivated under glucose excess conditions. Combining physiological and proteome quantification with genome-scale metabolic modeling, we found that the two groups differ in energy metabolism and translation efficiency. In Crabtree-positive yeasts, the central carbon metabolism flux and proteome allocation favor a glucose utilization strategy minimizing proteome cost as proteins translation parameters, including ribosomal content and/or efficiency, are lower. Crabtree-negative yeasts, however, use a strategy of maximizing ATP yield, accompanied by higher protein translation parameters. Our analyses provide insight into the underlying reasons for the Crabtree effect, demonstrating a coupling to adaptations in both metabolism and protein translation.

Keywords: Crabtree effect; constraint-based modeling; proteomics; systems biology; yeast.

Fig. 1.

Characterization of physiological differences between Crabtree-positive and -negative yeast. (A–D) Cell density (optical density at 600 nm [OD600]) and extracellular concentrations of glucose, ethanol, glycerol, and acetate during the course of the batch cultivation. Mean values ± SD of biological triplicates are shown. See SI Appendix, Fig. S2 A–D for additional metabolites. (E) The fractional consumption of glucose, C-mmol/C-mol glucose, normalized against the total carbon recovery. Mean values of biological triplicates are shown. The carbon recovery was between 91 and 100% for all four yeasts. (F) GUR. Mean values ± SD of biological triplicates are shown. (G) Protein content (P_tot), biomass yield (Y_sx), and growth rate (µ) plotted against GUR. Mean values ± SD and individual values of biological triplicates are shown. Colored boxed represent the organism, as indicated in panel F. (H) OUR, CER, and RQ plotted against GUR. Mean values ± SD and individual values of biological triplicates are shown for S. stipitis and mean values, and individual values for biological duplicates are shown for S. pombe, S. cerevisiae, and K. marxianus. Colored boxes represent the organism, as indicated in panel F. Spo, S. pombe; Sce, S. cerevisiae; Kma, K. marxianus; Sstip, S. stipitis.

Fig. 2.

Adaptations in metabolism and proteome allocation reflect the trade-off between glucose utilization and ATP yield. (A) Flux distribution of central carbon metabolism as predicted by FBA using condition-specific enzyme-constrained models, constrained with experimentally measured exchange fluxes and protein levels. (B) Summed proteome allocation to selected processes plotted against GUR. Protein members of each process were defined by GO annotation. Individual values and mean values of biological triplicates are shown. Colored boxes represent the organism, as specified in Fig. 1F. See SI Appendix, Fig. S3 for absolute abundances of the pathways. (C) Protein abundance of glucose transporters measured in this study. Mean values ± SD of biological triplicates are shown. (D) ATP production rate calculated based on FBA simulation results. (E) ATP yield calculated from FBA simulation results. Spo, S. pombe; Sce, S. cerevisiae; Kma, K. marxianus; Sstip, S. stipitis.

Fig. 3.

Limitations in the ETC and ATP synthase are characteristic for the Crabtree effect. (A) Capacity usage, calculated as the model-predicted enzyme levels divided by the experimentally measured enzyme levels, of the main pathways of central carbon metabolism (CCM). (B) Summed proteome allocation to processes of CCM plotted against GUR. Protein members of each process were defined by GO annotation. Mean values of biological triplicates are shown. Colored boxes represent the organism, as specified in panel C. See SI Appendix, Fig. S7A for absolute abundances of the processes. (C) Absolute protein abundances of the components constituting the ETC and ATP synthase. Mean values ± SD of biological triplicates are shown. See SI Appendix, Fig. S7B for proteome allocation of the components. PPP, pentose phosphate pathway; Spo, S. pombe; Sce, S. cerevisiae; Kma, K. marxianus; Sstip, S. stipitis.

Fig. 4.

The Crabtree effect is accompanied by differences in RP content and protein translation efficiencies. (A) Comparison of the overall ribosomal translation efficiency, calculated as milligrams total protein per nanomoles ribosomes per hour. (B) Comparison of the expression levels of MRPs, expressed as percent out of total proteome. (C) Comparison of the mitoribosomal protein translation efficiency, calculated based on the mean molar abundance of all subunits of the mitochondrially expressed protein complexes, including cytochrome c oxidase, ATP synthase, and mitoribosomes. (D) Comparison of the expression levels of cytochrome c oxidase and ATP synthase, expressed in percent of total cellular protein. C, Spo, S. pombe; Sce, S. cerevisiae; Kma, K. marxianus; Sstip, S. stipitis.