Lost in transition startup of glycolysis yields subpopulations of non - growing cells In collaboration with :

Cells need to adapt to dynamic environments. Yeast that fail to cope with dynamic changes in the abundance of glucose can undergo growth arrest. We show that this failure is caused by imbalanced reactions in glycolysis, the essential pathway in energy metabolism in most organisms. The imbalance arises largely from the fundamental design of glycolysis, making this state of glycolysis a generic risk. Cells with unbalanced glycolysis co-existed with vital cells. Spontaneous, non-genetic metabolic variability among individual cells determines which state is reached and consequently which cells survive. Transient ATP hydrolysis through futile cycling reduces the probability of reaching the imbalanced state. Our results reveal dynamic behavior of glycolysis and indicate that cell fate can be determined by heterogeneity purely at the metabolic level. Main text Key properties of biological systems are adaptability and robustness the ability to maintain the physiological state in response to perturbations or dynamic conditions (Stelling et al , 2004). Cells use multilayered regulation to respond adequately to changing environments or sudden perturbations. Failures in regulation underlie cellular malfunctioning, loss of fitness, or disease. In recent years, there has been a revived interest in metabolic processes as many diseases are associated with metabolic aberrations, such as diabetes and cancer (DeBerardinis and Thompson, 2012). Glycolysis is the central pathway in energy metabolism, which converts glucose to pyruvate with a net production of two adenosine 5’-triphosphate (ATP) molecules per glucose molecule. However, this net formation of ATP in “lower glycolysis” (Fig. 7.1) is preceded by an initial ATP investment at the first steps in the pathway (“upper glycolysis”, Fig. 7.1). This sequence of enzymatic steps in glycolysis, which is adopted by many organisms (Flamholz et al , 2013), implies a serious risk. If upper glycolysis outpaces lower glycolysis, a massive accumulation of glycolytic intermediates can occur with much reduced ATP production (Teusink et al , 1998). This phenotype is observed in pancreatic beta-cells overexpressing glucokinase (coined acute glucose intolerance) (Iynedjian, 1998), the enzyme that catalyzes the first step of glycolysis. Similarly, Saccharomyces cerevisiae mutants defective in the biosynthesis of the disaccharide trehalose, that branches off from glycolysis at the level of glucose-6-phosphate (Fig. 7.1), show accumulation of the glycolytic intermediate fructose 1,6-bisphosphate (FBP) at low concentrations of ATP (Van Aelst et al , 1993). We call this detrimental state of glycolysis an imbalanced state. We find that this imbalanced state is a risk also for normal cells. Through detailed system-level analysis of yeast glycolysis, we explain how phosphate dynamics is at the origin of the imbalance, and reveal how specific regulatory mechanisms affect the probability for cells to get trapped in it. S. cerevisiae mutants with a defect in trehalose 6-phosphate synthase (Tps1), the first committed step in trehalose biosynthesis (Fig. 7.1), exhibit the imbalanced-state phenotype and are unable to grow on glucose (Thevelein and Hohmann, 1995). The molecular mechanism underlying this imbalanced phenotype has proven challenging to elucidate (we provide an extensive account of the literature in the supplementary online text). Trehalose 6-phoshate (T6P), the product of Tps1, acts in vitro as a competitive inhibitor of the hexoki82