Elzbieta Petelenz-Kurdziel, Clemens Kuehn, Bodil Nordlander, Dagmara Klein, Kuk-Ki Hong, Therese Jacobson, Peter Dahl, Jörg Schaber, Jens Nielsen, Stefan Hohmann, Edda Klipp

We provide an integrated dynamic view on a eukaryotic osmolyte system, linking signaling with regulation of gene expression, metabolic control and growth. Adaptation to osmotic changes enables cells to adjust cellular activity and turgor pressure to an altered environment. The yeast Saccharomyces cerevisiae adapts to hyperosmotic stress by activating the HOG signaling cascade, which controls glycerol accumulation. The Hog1 kinase stimulates transcription of genes encoding enzymes required for glycerol production (Gpd1, Gpp2) and glycerol import (Stl1) and activates a regulatory enzyme in glycolysis (Pfk26/27). In addition, glycerol outflow is prevented by closure of the Fps1 glycerol facilitator. In order to better understand the contributions to glycerol accumulation of these different mechanisms and how redox and energy metabolism as well as biomass production are maintained under such conditions we collected an extensive dataset. Over a period of 180 min after hyperosmotic shock we monitored in wild type and different mutant cells the concentrations of key metabolites and proteins relevant for osmoadaptation. The dataset was used to parameterize an ODE model that reproduces the generated data very well. A detailed computational analysis using time-dependent response coefficients showed that Pfk26/27 contributes to rerouting glycolytic flux towards lower glycolysis. The transient growth arrest following hyperosmotic shock further adds to redirecting almost all glycolytic flux from biomass towards glycerol production. Osmoadaptation is robust to loss of individual adaptation pathways because of the existence and upregulation of alternative routes of glycerol accumulation. For instance, the Stl1 glycerol importer contributes to glycerol accumulation in a mutant with diminished glycerol production capacity. In addition, our observations suggest a role for trehalose accumulation in osmoadaptation and that Hog1 probably directly contributes to the regulation of the Fps1 glycerol facilitator. Taken together, we elucidated how different metabolic adaptation mechanisms cooperate and provide hypotheses for further experimental studies.