The Trehalose Pathway Regulates Mitochondrial Respiratory Chain Content through Hexokinase 2 and cAMP in Saccharomyces cerevisiae

In yeast, trehalose is synthesized by a multimeric enzymatic complex: TPS1 encodes trehalose 6-phosphate synthase, which belongs to a complex that is composed of at least three other subunits, including trehalose 6-phosphate phosphatase Tps2 and the redundant regulatory subunits Tps3 and Tsl1. The product of the TPS1 gene plays an essential role in the control of the glycolytic pathway by restricting the influx of glucose into glycolysis. In this paper, we investigated whether the trehalose synthesis pathway could be involved in the control of the other energy-generating pathway: oxidative phosphorylation. We show that the different mutants of the trehalose synthesis pathway (tps1Δ, tps2Δ, and tps1,2Δ) exhibit modulation in the amount of respiratory chains, in terms of cytochrome content and maximal respiratory activity. Furthermore, these variations in mitochondrial enzymatic content are positively linked to the intracellular concentration in cAMP that is modulated by Tps1p through hexokinase2. This is the first time that a pathway involved in sugar storage, i.e. trehalose, is shown to regulate the mitochondrial enzymatic content.The control of glycolysis in the yeast Saccharomyces cerevisiae has been extensively studied. First, allosteric regulation of the irreversible steps catalyzed by phosphofructokinase (1), pyruvate kinase (review in Ref. 2), and fructose-1,6-bisphosphatase (1) has been proposed, even though the overexpression of these key enzymes does not increase the glycolytic flux (3). Other mechanisms of control have been proposed such as futile cycle activity (4) and an inhibitory effect of ATP (5). Indeed, it seems likely that the regulation of glycolysis is a complex process involving different hierarchical events leading from gene expression to the metabolic fluxes via protein levels, enzyme activities, and metabolite effects (6, 7). Among these actors, the product of the TPS1 gene has been shown to play an essential role in the control of the glycolytic pathway by restricting the influx of glucose into glycolysis (8). TPS1 encodes trehalose 6-phosphate (Tre6P)³ synthase (9–12). This enzyme is part of a multimeric protein complex composed of at least three other subunits, i.e. Tre6P phosphatase encoded by TPS2 (13) and the redundant regulatory subunits Tps3 and Tsl1 (14).A particularly intriguing finding is that tps1Δ mutants are defective not only for Tre6P synthesis but also for growth on glucose or related rapidly fermented sugars (8, 11, 15). This may be explained by an uncontrolled influx of glucose into the glycolytic pathway. This phenomenon is characterized by hyperaccumulation of glucose 6-phosphate, fructose 6-phosphate, and fructose 1,6-bisphosphate (Fru1,6bP) (8, 16–18) and depletion of ATP, P_i, and all intermediates of glycolysis downstream of glyceraldehyde-3-phosphate dehydrogenase (19). Several mutations have been described that suppress the growth defect of tps1Δ mutants apparently by reducing sugar influx into glycolysis (16, 20) or by diverting the excess sugar phosphate into glycerol synthesis through overexpression of the GPD1-encoded NAD-dependent glycerol-3-phosphate dehydrogenase (17, 21). Reconstitution of ethanolic fermentation in permeabilized yeast spheroplasts indicated that in addition to Tre6P, the Tps1 protein itself also seems to play a role in restricting glucose influx into glycolysis (22).Whatever the mechanism by which the multimeric complex involved in trehalose synthesis controls glycolytic flux in yeast, such a regulation is associated with modification of the cellular content of sugar phosphates. Moreover, in a recent paper, we have shown that in yeast, low physiological concentrations of glucose 6-phosphate and fructose 6-phosphate slightly (20%) stimulate the respiratory flux and that this effect was strongly antagonized by Fru1,6bP (18). On the other hand, Fru1,6bP by itself is able to inhibit mitochondrial respiration only in mitochondria isolated from a Crabtree-positive strain. Taken together, these results indicate that besides the thermodynamic link between glycolysis and mitochondrial respiration (i.e. the cytosolic ATP/ADP and NADH/NAD+ ratio), a kinetic control of oxidative phosphorylation activity is exerted by the level of glycolytic sugar phosphates (18, 23). This raises the question of a possible direct or indirect regulation of oxidative phosphorylation by the trehalose synthesis pathway.