In numerous cell types, tumoral cells, proliferating cells, bacteria, and yeast, respiration is inhibited when high concentrations of glucose are added to the culture medium. This phenomenon has been named the "Crabtree effect." We used yeast to investigate (i) the short term event(s) associated with the Crabtree effect and (ii) a putative role of hexose phosphates in the inhibition of respiration. Indeed, yeast divide into "Crabtree-positive," where the Crabtree effect occurs, and "Crabtree-negative," where it does not. In mitochondria isolated from these two categories of yeast, we found that low, physiological concentrations of glucose 6-phosphate and fructose 6-phosphate slightly (20%) stimulated the respiratory flux and that this effect was strongly antagonized by fructose 1,6-bisphosphate (F16bP). On the other hand, F16bP by itself was able to inhibit mitochondrial respiration only in mitochondria isolated from a Crabtree-positive strain. Using permeabilized spheroplasts from Crabtree-positive yeast, we have shown that the sole effect observed at physiological concentrations of hexose phosphates is an inhibition of oxidative phosphorylation by F16bP. This F16bP-mediated inhibition was also observed in isolated rat liver mitochondria, extending this process to mammalian cells. From these results and taking into account that F16bP is able to accumulate in the cell cytoplasm, we propose that F16bP regulates oxidative phosphorylation and thus participates in the establishment of the Crabtree effect.

No abstract

In the Saccharomyces cerevisiae glycolytic pathway, 11 enzymes catalyze the stepwise conversion of glucose to two molecules of ethanol plus two CO 2 molecules. In the highly crowded cytoplasm, this pathway would be very inefficient if it were dependent on substrate/enzyme diffusion. Therefore, the existence of a multi-enzymatic glycolytic complex has been suggested. This complex probably uses the cytoskeleton to stabilize the interaction of the various enzymes. Here, the role of filamentous actin (F-actin) in stabilization of a putative glycolytic metabolon is reported. Experiments were performed in isolated enzyme/actin mixtures, cytoplasmic extracts and permeabilized yeast cells. Polymerization of actin was promoted using phalloidin or inhibited using cytochalasin D or latrunculin. The polymeric filamentous F-actin, but not the monomeric globular G-actin, stabilized both the interaction of isolated glycolytic pathway enzyme mixtures and the whole fermentation pathway, leading to higher fermentation activity. The associated complexes were resistant against inhibition as a result of viscosity (promoted by the disaccharide trehalose) or inactivation (using specific enzyme antibodies). In S. cerevisiae, a glycolytic metabolon appear to assemble in association with F-actin. In this complex, fermentation activity is enhanced and enzymes are partially protected against inhibition by trehalose or by antibodies. Structured digital abstract• ALD physically interacts with PGK and GAPDH by anti bait coimmunoprecipitation (View interaction)• ALD physically interacts with GAPDH and PGK by affinity chromatography technology (View interaction) IntroductionThe cytoplasm is a highly concentrated suspension of proteins, polysaccharides, nucleic acids and small solutes [1,2]. It has been proposed that saturation promotes specific protein-protein interactions [1,3], and, once associated, enzymes in a given pathway team up to catalyze several consecutive reactions; these enzyme complexes are called metabolons [4,5]. In metabolons, intermediaries are channeled, i.e. enzymes that catalyze consecutive reactions transfer intermediaries directly to each other [2,6,7]. Substrate channeling confers a number of benefits, including altered reaction kinetics, preservation of cellular solvation capacity [8] or sequestration of toxic intermediaries [9]. The highly dynamic nature of enzyme-enzyme interactions Abbreviations ADH, alcohol dehydrogenase; ALD, aldolase; ENO, enolase; F-actin, filamentous actin; G-actin, globular (monomeric) actin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GPI, glucose-6-phosphate isomerase; HXK, hexokinase; PFK, phosphofructokinase; PGAM, phosphoglyceromutase; PGK, phosphoglycerate kinase; PK, pyruvate kinase; TPI, triosephosphate isomerase. 3887[10] probably regulates their reaction rate, and channels substrates through specific pathway(s) [2,6,11]. Various groups have described metabolons in the cysteine synthase complex, the Calvin cycle, cyanogenic glucoside synthesis and the phenylpropanoid pathway of ...

No abstract

Non-alcoholic fatty liver disease (NAFLD) encompasses a broad spectrum of histopathological changes ranging from non-inflammatory intracellular fat deposition to non-alcoholic steatohepatitis (NASH), which may progress into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. Recent data suggest that impaired hepatic cholesterol homeostasis and its accumulation are relevant to the pathogenesis of NAFLD/NASH. Despite a vital physiological function of cholesterol, mitochondrial dysfunction is an important consequence of dietary-induced hypercholesterolemia and was, subsequently, linked to many pathophysiological conditions. The aim in the current study was to evaluate the morphological and molecular changes of cholesterol overload in mouse liver and particularly, in mitochondria, induced by a high-cholesterol (HC) diet for one month. Histopathological studies revealed microvesicular hepatic steatosis and significantly elevated levels of liver cholesterol and triglycerides leading to impaired liver synthesis. Further, high levels of oxidative stress could be determined in liver tissue as well as primary hepatocyte culture.Transcriptomic changes induced by the HC diet involved disruption in key pathways related to cell death and oxidative stress as well as upregulation of genes related to glutathione homeostasis. Impaired liver function could be associated with a decrease in mitochondrial membrane potential and ATP content and significant alterations in mitochondrial dynamics. We demonstrate that cholesterol overload in the liver leads to mitochondrial changes which may render damaged hepatocytes proliferative and resistant to cell death whereby perpetuating liver damage. K E Y W O R D S apoptosis, cholesterol, mitochondrial dynamics, oxidative stress J Cell Physiol. 2019;234:7213-7223. wileyonlinelibrary.com/journal/jcp Additional supporting information may be found online in the Supporting Information section at the end of the article. How to cite this article: Domínguez-Pérez M, Simoni-Nieves A, Rosales P, et al. Cholesterol burden in the liver induces mitochondrial dynamic changes and resistance to apoptosis.