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Louis Hue - One of the best experts on this subject based on the ideXlab platform.
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5 aminoimidazole 4 carboxamide 1 β d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
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5 aminoimidazole 4 carboxamide 1 beta d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
Bruno Guigas - One of the best experts on this subject based on the ideXlab platform.
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5 aminoimidazole 4 carboxamide 1 β d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
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5 aminoimidazole 4 carboxamide 1 beta d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
Pierre Cardol - One of the best experts on this subject based on the ideXlab platform.
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atypical subunit composition of the chlorophycean mitochondrial f1fo atp synthase and role of asa7 protein in stability and Oligomycin resistance of the enzyme
Molecular Biology and Evolution, 2010Co-Authors: Marie Lapaille, Diego Gonzalezhalphen, Adelma Escobarramirez, Herve Degand, Denis Baurain, Elizabeth Rodriguezsalinas, Nadine Coosemans, Marc Boutry, Claire Remacle, Pierre CardolAbstract:In yeast, mammals, and land plants, mitochondrial F1FO-ATP synthase (complex V) is a remarkable enzymatic machinery that comprises about 15 conserved subunits. Peculiar among eukaryotes, complex V from Chlamydomonadales algae (order of chlorophycean class) has an atypical subunit composition of its peripheral stator and dimerization module, with nine subunits of unknown evolutionary origin (Asa subunits). In vitro, this enzyme exhibits an increased stability of its dimeric form, and in vivo, Chlamydomonas reinhardtii cells are insensitive to Oligomycins, which are potent inhibitors of proton translocation through the F-O moiety. In this work, we showed that the atypical features of the Chlamydomonadales complex V enzyme are shared by the other chlorophycean orders. By biochemical and in silico analyses, we detected several atypical Asa subunits in Scenedesmus obliquus (Sphaeropleales) and Chlorococcum ellipsoideum (Chlorococcales). In contrast, complex V has a canonical subunit composition in other classes of Chlorophytes (Trebouxiophyceae, Prasinophyceae, and Ulvophyceae) as well as in Streptophytes (land plants), and in Rhodophytes (red algae). Growth, respiration, and ATP levels in Chlorophyceae were also barely affected by Oligomycin concentrations that affect representatives of the other classes of Chlorophytes. We finally studied the function of the Asa7 atypical subunit by using RNA interference in C. reinhardtii. Although the loss of Asa7 subunit has no impact on cell bioenergetics or mitochondrial structures, it destabilizes in vitro the enzyme dimeric form and renders growth, respiration, and ATP level sensitive to Oligomycins. Altogether, our results suggest that the loss of canonical components of the complex V stator happened at the root of chlorophycean lineage and was accompanied by the recruitment of novel polypeptides. Such a massive modification of complex V stator features might have conferred novel properties, including the stabilization of the enzyme dimeric form and the shielding of the proton channel. In these respects, we discuss an evolutionary scenario for F1FO-ATP synthase in the whole green lineage (i.e., Chlorophyta and Streptophyta).
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The fully-active and structurally-stable form of the mitochondrial ATP synthase of Polytomella sp. is dimeric
Journal of Bioenergetics and Biomembranes, 2009Co-Authors: Alexa Villavicencio-queijeiro, Julio A Mignaco, Debora Foguel, Miriam Vázquez-acevedo, Araceli Cano-estrada, Mariel Zarco-zavala, Marietta Tuena De Gómez, Monica M. Freire, Helena M. Scofano, Pierre CardolAbstract:Mitochondrial F_1F_O-ATP synthase of chlorophycean algae is a stable dimeric complex of 1,600 kDa. It lacks the classic subunits that constitute the peripheral stator-stalk and the orthodox polypeptides involved in the dimerization of the complex. Instead, it contains nine polypeptides of unknown evolutionary origin named ASA1 to ASA9. The isolated enzyme exhibited a very low ATPase activity (0.03 Units/mg), that increased upon heat treatment, due to the release of the F_1 sector. Oligomycin was found to stabilize the dimeric structure of the enzyme, providing partial resistance to heat dissociation. Incubation in the presence of low concentrations of several non-ionic detergents increased the Oligomycin-sensitive ATPase activity up to 7.0–9.0 Units/mg. Incubation with 3% ( w / v ) taurodeoxycholate monomerized the enzyme. The monomeric form of the enzyme exhibited diminished activity in the presence of detergents and diminished Oligomycin sensitivity. Cross-linking experiments carried out with the dimeric and monomeric forms of the ATP synthase suggested the participation of the ASA6 subunit in the dimerization of the enzyme. The dimeric enzyme was more resistant to heat treatment, high hydrostatic pressures, and protease digestion than the monomeric enzyme, which was readily disrupted by these treatments. We conclude that the fully-active algal mitochondrial ATP synthase is a stable catalytically active dimer; the monomeric form is less active and less stable. Monomer-monomer interactions could be mediated by the membrane-bound subunits ASA6 and ASA9, and may be further stabilized by other polypeptides such as ASA1 and ASA5.
Luc Bertrand - One of the best experts on this subject based on the ideXlab platform.
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5 aminoimidazole 4 carboxamide 1 beta d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
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5 aminoimidazole 4 carboxamide 1 β d ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an amp activated protein kinase independent effect on glucokinase translocation
Diabetes, 2006Co-Authors: Bruno Guigas, Luc Bertrand, Nellie Taleux, Marc Foretz, Nicolas Wiernsperger, Didier Vertommen, Fabrizio Andreelli, Benoit Viollet, Louis HueAbstract:AMP-activated protein kinase (AMPK) controls glucose uptake and glycolysis in muscle. Little is known about its role in liver glucose uptake, which is controlled by glucokinase. We report here that 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), metformin, and Oligomycin activated AMPK and inhibited glucose phosphorylation and glycolysis in rat hepatocytes. In vitro experiments demonstrated that this inhibition was not due to direct phosphorylation of glucokinase or its regulatory protein by AMPK. By contrast, AMPK phosphorylated liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase without affecting activity. Inhibitors of the endothelial nitric oxide synthase, stress kinases, and phosphatidylinositol 3-kinase pathways did not counteract the effects of AICAR, metformin, or Oligomycin, suggesting that these signaling pathways were not involved. Interestingly, the inhibitory effect on glucose phosphorylation of these well-known AMPK activators persisted in primary cultured hepatocytes from newly engineered mice lacking both liver alpha1 and alpha2 AMPK catalytic subunits, demonstrating that this effect was clearly not mediated by AMPK. Finally, AICAR, metformin, and Oligomycin were found to inhibit the glucose-induced translocation of glucokinase from the nucleus to the cytosol by a mechanism that could be related to the decrease in intracellular ATP concentrations observed in these conditions.
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phosphorylation and activation of heart pfk 2 by ampk has a role in the stimulation of glycolysis during ischaemia
Current Biology, 2000Co-Authors: Annesophie Marsin, Luc Bertrand, Mark H Rider, Johan Deprez, Christophe Beauloye, Mariefrancoise Vincent, G Van Den Berghe, David CarlingAbstract:BACKGROUND: The role of protein phosphorylation in the Pasteur effect--the phenomenon whereby anaerobic conditions stimulate glycolysis--has not been addressed. The AMP-activated protein kinase (AMPK) is activated when the oxygen supply is restricted. AMPK acts as an energy-state sensor and inhibits key biosynthetic pathways, thus conserving ATP. Here, we studied whether AMPK is involved in the Pasteur effect in the heart by phosphorylating and activating 6-phosphofructo-2-kinase (PFK-2), the enzyme responsible for the synthesis of fructose 2,6-bisphosphate, a potent stimulator of glycolysis. RESULTS: Heart PFK-2 was phosphorylated on Ser466 and activated by AMPK in vitro. In perfused rat hearts, anaerobic conditions or inhibitors of oxidative phosphorylation (Oligomycin and antimycin) induced AMPK activation, which correlated with PFK-2 activation and with an increase in fructose 2,6-bisphosphate concentration. Moreover, in cultured cells transfected with heart PFK-2, Oligomycin treatment resulted in a parallel activation of endogenous AMPK and PFK-2. In these cells, the activation of PFK-2 was due to the phosphorylation of Ser466. A dominant-negative construct of AMPK abolished the activation of endogenous and cotransfected AMPK, and prevented both the activation and phosphorylation of transfected PFK-2 by Oligomycin. CONCLUSIONS: AMPK phosphorylates and activates heart PFK-2 in vitro and in intact cells. AMPK-mediated PFK-2 activation is likely to be involved in the stimulation of heart glycolysis during ischaemia.
Michael L J Ashford - One of the best experts on this subject based on the ideXlab platform.
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dual actions of the metabolic inhibitor sodium azide on katp channel currents in the rat cri g1 insulinoma cell line
British Journal of Pharmacology, 1999Co-Authors: J Harvey, S C Hardy, Michael L J AshfordAbstract:The effects of various inhibitors of the mitochondrial electron transport chain on the activity of ATP-sensitive K+ channels were examined in the Cambridge rat insulinoma G1 (CRI-G1) cell line using a combination of whole cell and single channel recording techniques. Whole cell current clamp recordings, with 5 mM ATP in the pipette, demonstrate that the mitochondrial uncoupler sodium azide (3 mM) rapidly hyperpolarizes CRI-G1 cells with a concomitant increase in K+ conductance. This is due to activation of KATP channels as the sulphonylurea tolbutamide (100 μM) completely reversed the actions of azide. Other inhibitors of the mitochondrial electron transport chain, rotenone (10 μM) or Oligomycin (2 μM) did not hyperpolarize CRI-G1 cells or increase K+ conductance. In cell-attached recordings, bath application of 3 mM sodium azide (in the absence of glucose) resulted in a rapid increase in KATP channel activity, an action readily reversible by tolbutamide (100 μM). Application of sodium azide (3 mM), in the presence of Mg-ATP, to the intracellular surface of excised inside-out patches also increased KATP channel activity, in a reversible manner. In contrast, rotenone (10 μM) or Oligomycin (2 μM) did not increase KATP channel activity in either cell-attached, in the absence of glucose, or inside-out membrane patch recordings. Addition of sodium azide (3 mM) to the intracellular surface of inside-out membrane patches in the presence of Mg-free ATP or the non-hydrolysable analogue 5′-adenylylimidodiphosphate (AMP-PNP) inhibited, rather than increased, KATP channel activity. In conclusion, sodium azide, but not rotenone or Oligomycin, directly activates KATP channels in CRI-G1 insulin secreting cells. This action of azide is similar to that reported previously for diazoxide. British Journal of Pharmacology (1999) 126, 51–60; doi:10.1038/sj.bjp.0702267
