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Anthony Carruthers - One of the best experts on this subject based on the ideXlab platform.
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red wine and green tea flavonoids are cis allosteric activators and competitive inhibitors of glucose transporter 1 glut1 mediated sugar uptake
Journal of Biological Chemistry, 2018Co-Authors: Ogooluwa A Ojelabi, Julie K De Zutter, Kenneth P Lloyd, Anthony CarruthersAbstract:: The antioxidant- and flavonoid-rich contents of red wine and green tea are reported to offer protection against cancer, cardiovascular disease, and diabetes. Some studies, however, show that flavonoids inhibit GLUT1-mediated, facilitative glucose transport, raising the possibility that their interaction with GLUT1 and subsequent downstream effects on carbohydrate metabolism may also impact health. The present study explores the structure-function relationships of flavonoid-GLUT1 interactions. We find that low concentrations of flavonoids act as cis-allosteric activators of sugar uptake, whereas higher concentrations competitively inhibit sugar uptake and noncompetitively inhibit sugar exit. Studies with heterologously expressed human GLUT1, -3, or -4 reveal that quercetin-GLUT1 and -GLUT4 interactions are stronger than quercetin-GLUT3 interactions, that epicatechin gallate (ECG) is more selective for GLUT1, and that epigallocatechin gallate (EGCG) is less GLUT isoform-selective. Docking studies suggest that only one flavonoid can bind to GLUT1 at any instant, but sugar transport and ligand-binding studies indicate that human erythrocyte GLUT1 can bind at least two flavonoid molecules simultaneously. Quercetin and EGCG are each characterized by positive, cooperative binding, whereas ECG shows negative cooperative binding. These findings support recent studies suggesting that GLUT1 forms an oligomeric complex of interacting, allosteric, alternating access transporters. We discuss how modulation of facilitative glucose transporters could contribute to the protective actions of the flavonoids against diabetes and Alzheimer's disease.
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wzb117 2 fluoro 6 m hydroxybenzoyloxy phenyl m hydroxybenzoate inhibits glut1 mediated sugar transport by binding reversibly at the exofacial sugar binding site
Journal of Biological Chemistry, 2016Co-Authors: Ogooluwa A Ojelabi, Julie K De Zutter, Kenneth P Lloyd, Andrew H Simon, Anthony CarruthersAbstract:Abstract WZB117 (2-fluoro-6-(m-hydroxybenzoyloxy) phenyl m-hydroxybenzoate) inhibits passive sugar transport in human erythrocytes and cancer cell lines and, by limiting glycolysis, inhibits tumor growth in mice. This study explores how WZB117 inhibits the erythrocyte sugar transporter glucose transport protein 1 (GLUT1) and examines the transporter isoform specificity of inhibition. WZB117 reversibly and competitively inhibits erythrocyte 3-O-methylglucose (3MG) uptake with Ki(app) = 6 μm but is a noncompetitive inhibitor of sugar exit. Cytochalasin B (CB) is a reversible, noncompetitive inhibitor of 3MG uptake with Ki(app) = 0.3 μm but is a competitive inhibitor of sugar exit indicating that WZB117 and CB bind at exofacial and endofacial sugar binding sites, respectively. WZB117 inhibition of GLUTs expressed in HEK293 cells follows the order of potency: insulin-regulated GLUT4 ≫ GLUT1 ≈ neuronal GLUT3. This may explain WZB117-induced murine lipodystrophy. Molecular docking suggests the following. 1) The WZB117 binding envelopes of exofacial GLUT1 and GLUT4 conformers differ significantly. 2) GLUT1 and GLUT4 exofacial conformers present multiple, adjacent glucose binding sites that overlap with WZB117 binding envelopes. 3) The GLUT1 exofacial conformer lacks a CB binding site. 4) The inward GLUT1 conformer presents overlapping endofacial WZB117, d-glucose, and CB binding envelopes. Interrogating the GLUT1 mechanism using WZB117 reveals that subsaturating WZB117 and CB stimulate erythrocyte 3MG uptake. Extracellular WZB117 does not affect CB binding to GLUT1, but intracellular WZB117 inhibits CB binding. These findings are incompatible with the alternating conformer carrier for glucose transport but are consistent with either a multisubunit, allosteric transporter, or a transporter in which each subunit presents multiple, interacting ligand binding sites.
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sequence determinants of glut1 oligomerization analysis by homology scanning mutagenesis
Journal of Biological Chemistry, 2013Co-Authors: Julie K De Zutter, Kara B Levine, Di Deng, Anthony CarruthersAbstract:Abstract The human blood-brain barrier glucose transport protein (GLUT1) forms homodimers and homotetramers in detergent micelles and in cell membranes, where the GLUT1 oligomeric state determines GLUT1 transport behavior. GLUT1 and the neuronal glucose transporter GLUT3 do not form heterocomplexes in human embryonic kidney 293 (HEK293) cells as judged by co-immunoprecipitation assays. Using homology-scanning mutagenesis in which GLUT1 domains are substituted with equivalent GLUT3 domains and vice versa, we show that GLUT1 transmembrane helix 9 (TM9) is necessary for optimal association of GLUT1-GLUT3 chimeras with parental GLUT1 in HEK cells. GLUT1 TMs 2, 5, 8, and 11 also contribute to a less abundant heterocomplex. Cell surface GLUT1 and GLUT3 containing GLUT1 TM9 are 4-fold more catalytically active than GLUT3 and GLUT1 containing GLUT3 TM9. GLUT1 and GLUT3 display allosteric transport behavior. Size exclusion chromatography of detergent solubilized, purified GLUT1 resolves GLUT1/lipid/detergent micelles as 6- and 10-nm Stokes radius particles, which correspond to GLUT1 dimers and tetramers, respectively. Studies with GLUTs expressed in and solubilized from HEK cells show that HEK cell GLUT1 resolves as 6- and 10-nm Stokes radius particles, whereas GLUT3 resolves as a 6-nm particle. Substitution of GLUT3 TM9 with GLUT1 TM9 causes chimeric GLUT3 to resolve as 6- and 10-nm Stokes radius particles. Substitution of GLUT1 TM9 with GLUT3 TM9 causes chimeric GLUT1 to resolve as a mixture of 6- and 4-nm particles. We discuss these findings in the context of determinants of GLUT oligomeric structure and transport function.
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role of monosaccharide transport proteins in carbohydrate assimilation distribution metabolism and homeostasis
Comprehensive Physiology, 2012Co-Authors: Anthony J Cura, Anthony CarruthersAbstract:The facilitated diffusion of glucose, galactose, fructose, urate, myoinositol, and dehydroascorbic acid in mammals is catalyzed by a family of 14 monosaccharide transport proteins called GLUTs. These transporters may be divided into three classes according to sequence similarity and function/substrate specificity. GLUT1 appears to be highly expressed in glycolytically active cells and has been coopted in vitamin C auxotrophs to maintain the redox state of the blood through transport of dehydroascorbate. Several GLUTs are definitive glucose/galactose transporters, GLUT2 and GLUT5 are physiologically important fructose transporters, GLUT9 appears to be a urate transporter while GLUT13 is a proton/myoinositol cotransporter. The physiologic substrates of some GLUTs remain to be established. The GLUTs are expressed in a tissue specific manner where affinity, specificity, and capacity for substrate transport are paramount for tissue function. Although great strides have been made in characterizing GLUT-catalyzed monosaccharide transport and mapping GLUT membrane topography and determinants of substrate specificity, a unifying model for GLUT structure and function remains elusive. The GLUTs play a major role in carbohydrate homeostasis and the redistribution of sugar-derived carbons among the various organ systems. This is accomplished through a multiplicity of GLUT-dependent glucose sensing and effector mechanisms that regulate monosaccharide ingestion, absorption, distribution, cellular transport and metabolism, and recovery/retention. Glucose transport and metabolism have coevolved in mammals to support cerebral glucose utilization. © 2012 American Physiological Society. Compr Physiol 2:863-914, 2012.
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role of monosaccharide transport proteins in carbohydrate assimilation distribution metabolism and homeostasis
Comprehensive Physiology, 2012Co-Authors: Anthony J Cura, Anthony CarruthersAbstract:The facilitated diffusion of glucose, galactose, fructose, urate, myoinositol, and dehydroascorbicacid in mammals is catalyzed by a family of 14 monosaccharide transport proteins called GLUTs. These transporters may be divided into three classes according to sequence similarity and function/substrate specificity. GLUT1 appears to be highly expressed in glycolytically active cells and has been coopted in vitamin C auxotrophs to maintain the redox state of the blood through transport of dehydroascorbate. Several GLUTs are definitive glucose/galactose transporters, GLUT2 and GLUT5 are physiologically important fructose transporters, GLUT9 appears to be a urate transporter while GLUT13 is a proton/myoinositol cotransporter. The physiologic substrates of some GLUTs remain to be established. The GLUTs are expressed in a tissue specific manner where affinity, specificity, and capacity for substrate transport are paramount for tissue function. Although great strides have been made in characterizing GLUT-catalyzed monosaccharide transport and mapping GLUT membrane topography and determinants of substrate specificity, a unifying model for GLUT structure and function remains elusive. The GLUTs play a major role in carbohydrate homeostasis and the redistribution of sugar-derived carbons among the various organ systems. This is accomplished through a multiplicity of GLUT-dependent glucose sensing and effector mechanisms that regulate monosaccharide ingestion, absorption,distribution, cellular transport and metabolism, and recovery/retention. Glucose transport and metabolism have coevolved in mammals to support cerebral glucose utilization.
Nidaa Mikail - One of the best experts on this subject based on the ideXlab platform.
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differences in alimentary glucose absorption and intestinal disposal of blood glucose after roux en y gastric bypass vs sleeve gastrectomy
Gastroenterology, 2016Co-Authors: Jeanbaptiste Cavin, Robert Ducroc, Konstantinos Arapis, Eglantine Voitellier, Laura Gillard, Muriel Hourseau, Francoise Cluzeaud, Anne Couvelard, Rachida Lebtahi, Nidaa MikailAbstract:Background & Aims Bariatric procedures, such as Roux-en-Y gastric bypass (RYGB) or vertical sleeve gastrectomy (VSG), are the most effective approaches to resolve type 2 diabetes in obese individuals. Alimentary glucose absorption and intestinal disposal of blood glucose have not been directly compared between individuals or animals that underwent RYGB vs VSG. We evaluated in rats and humans how the gut epithelium adapts after surgery and the consequences on alimentary glucose absorption and intestinal disposal of blood glucose. Methods Obese male rats underwent RYGB, VSG, or sham (control) operations. We collected intestine segments from all rats; we performed histologic analyses and measured levels of messenger RNAs encoding the sugar transporters SGLT1, GLUT1, GLUT2, GLUT3, GLUT4, and GLUT5. Glucose transport and consumption were assayed using ex vivo jejunal loops. Histologic analyses were also performed on Roux limb sections from patients who underwent RYGB 1−5 years after surgery. Roux limb glucose consumption was assayed after surgery by positron emission and computed tomography imaging. Results In rats and humans that underwent RYGB, the Roux limb became hyperplasic, with an increased number of incretin-producing cells compared with the corresponding jejunal segment of controls. Furthermore, expression of sugar transporters and hypoxia-related genes increased and the nonintestinal glucose transporter GLUT1 appeared at the basolateral membrane of enterocytes. Ingested and circulating glucose was trapped within the intestinal epithelial cells of rats and humans that underwent RYGB. By contrast, there was no hyperplasia of the intestine after VSG, but the intestinal absorption of alimentary glucose was reduced and density of endocrine cells secreting glucagon-like peptide-1 increased. Conclusions The intestine adapts differently to RYGB vs VSG. RYGB increases intestinal glucose disposal and VSG delays glucose absorption; both contribute to observed improvements in glycemia.
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differences in alimentary glucose absorption and intestinal disposal of blood glucose after roux en y gastric bypass vs sleeve gastrectomy
Gastroenterology, 2016Co-Authors: Jeanbaptiste Cavin, Robert Ducroc, Konstantinos Arapis, Eglantine Voitellier, Laura Gillard, Muriel Hourseau, Francoise Cluzeaud, Anne Couvelard, Rachida Lebtahi, Nidaa MikailAbstract:Background & Aims: Bariatric surgeries, such as Roux-en-Y gastric bypass (RYGB) or vertical sleeve gastrectomy (VSG), are the most effective approaches to resolve type 2 diabetes in obese individuals. Alimentary glucose absorption and intestinal disposal of blood glucose have not been directly compared between individuals or animals that underwent RYGB vs VSG. We evaluated in rats and humans how the gut epithelium adapts following surgery and the consequences on alimentary glucose absorption and intestinal disposal of blood glucose. Methods: Obese male rats underwent RYGB, VSG, or sham (control) surgeries. We collected intestine segments from all rats; we performed histologic analyses and measured levels of mRNAs encoding the sugar transporters SGLT1, GLUT1, GLUT2, GLUT3, GLUT4 and GLUT5. Glucose transport and consumption were assayed using ex vivo jejunal loops. Histologic analyses were also performed on Roux limb sections from patients who underwent RYGB, 1–5 years after surgery. Roux limb glucose consumption was assayed following surgery by positron emission and computed tomography imaging. Results: In rats and humans that underwent RYGB, the Roux limb became hyperplasic, with an increased number of incretin-producing cells, compared with the corresponding jejunal segment of controls. Furthermore, expression of sugar transporters and hypoxia-related genes increased and the non-intestinal glucose transporter GLUT1 appeared at the basolateral membrane of enterocytes. Ingested and circulating glucose was trapped within the intestinal epithelial cells of rats and humans that underwent RYGB. By contrast, there was no hyperplasia of the intestine after VSG, but the intestinal absorption of alimentary glucose was reduced and density of endocrine cells secreting glucagon-like peptide-1 (GLP1) increased. Conclusions: The intestine adapts differently to RYGB vs VSG. RYGB increases intestinal glucose disposal, whereas VSG delays glucose absorption; both contribute to observed improvements in glycemia.
Anthony J Cura - One of the best experts on this subject based on the ideXlab platform.
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role of monosaccharide transport proteins in carbohydrate assimilation distribution metabolism and homeostasis
Comprehensive Physiology, 2012Co-Authors: Anthony J Cura, Anthony CarruthersAbstract:The facilitated diffusion of glucose, galactose, fructose, urate, myoinositol, and dehydroascorbic acid in mammals is catalyzed by a family of 14 monosaccharide transport proteins called GLUTs. These transporters may be divided into three classes according to sequence similarity and function/substrate specificity. GLUT1 appears to be highly expressed in glycolytically active cells and has been coopted in vitamin C auxotrophs to maintain the redox state of the blood through transport of dehydroascorbate. Several GLUTs are definitive glucose/galactose transporters, GLUT2 and GLUT5 are physiologically important fructose transporters, GLUT9 appears to be a urate transporter while GLUT13 is a proton/myoinositol cotransporter. The physiologic substrates of some GLUTs remain to be established. The GLUTs are expressed in a tissue specific manner where affinity, specificity, and capacity for substrate transport are paramount for tissue function. Although great strides have been made in characterizing GLUT-catalyzed monosaccharide transport and mapping GLUT membrane topography and determinants of substrate specificity, a unifying model for GLUT structure and function remains elusive. The GLUTs play a major role in carbohydrate homeostasis and the redistribution of sugar-derived carbons among the various organ systems. This is accomplished through a multiplicity of GLUT-dependent glucose sensing and effector mechanisms that regulate monosaccharide ingestion, absorption, distribution, cellular transport and metabolism, and recovery/retention. Glucose transport and metabolism have coevolved in mammals to support cerebral glucose utilization. © 2012 American Physiological Society. Compr Physiol 2:863-914, 2012.
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role of monosaccharide transport proteins in carbohydrate assimilation distribution metabolism and homeostasis
Comprehensive Physiology, 2012Co-Authors: Anthony J Cura, Anthony CarruthersAbstract:The facilitated diffusion of glucose, galactose, fructose, urate, myoinositol, and dehydroascorbicacid in mammals is catalyzed by a family of 14 monosaccharide transport proteins called GLUTs. These transporters may be divided into three classes according to sequence similarity and function/substrate specificity. GLUT1 appears to be highly expressed in glycolytically active cells and has been coopted in vitamin C auxotrophs to maintain the redox state of the blood through transport of dehydroascorbate. Several GLUTs are definitive glucose/galactose transporters, GLUT2 and GLUT5 are physiologically important fructose transporters, GLUT9 appears to be a urate transporter while GLUT13 is a proton/myoinositol cotransporter. The physiologic substrates of some GLUTs remain to be established. The GLUTs are expressed in a tissue specific manner where affinity, specificity, and capacity for substrate transport are paramount for tissue function. Although great strides have been made in characterizing GLUT-catalyzed monosaccharide transport and mapping GLUT membrane topography and determinants of substrate specificity, a unifying model for GLUT structure and function remains elusive. The GLUTs play a major role in carbohydrate homeostasis and the redistribution of sugar-derived carbons among the various organ systems. This is accomplished through a multiplicity of GLUT-dependent glucose sensing and effector mechanisms that regulate monosaccharide ingestion, absorption,distribution, cellular transport and metabolism, and recovery/retention. Glucose transport and metabolism have coevolved in mammals to support cerebral glucose utilization.
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Acute Modulation of Sugar Transport in Brain Capillary Endothelial Cell Cultures during Activation of the Metabolic Stress Pathway
The Journal of biological chemistry, 2010Co-Authors: Anthony J Cura, Anthony CarruthersAbstract:Abstract GLUT1-catalyzed equilibrative sugar transport across the mammalian blood-brain barrier is stimulated during acute and chronic metabolic stress; however, the mechanism of acute transport regulation is unknown. We have examined acute sugar transport regulation in the murine brain microvasculature endothelial cell line bEnd.3. Acute cellular metabolic stress was induced by glucose depletion, by potassium cyanide, or by carbonyl cyanide p-trifluoromethoxyphenylhydrazone, which reduce or deplete intracellular ATP within 15 min. This results in a 1.7–7-fold increase in Vmax for zero-trans 3-O-methylglucose uptake (sugar uptake into sugar-free cells) and a 3–10-fold increase in Vmax for equilibrium exchange transport (intracellular [sugar] = extracellular [sugar]). GLUT1, GLUT8, and GLUT9 mRNAs are detected in bEnd.3 cells where GLUT1 mRNA levels are 33-fold greater than levels of GLUT8 or GLUT9 mRNA. Neither GLUT1 mRNA nor total protein levels are affected by acute metabolic stress. Cell surface biotinylation reveals that plasma membrane GLUT1 levels are increased 2–3-fold by metabolic depletion, although cell surface Na+,K+-ATPase levels remain unaffected by ATP depletion. Treatment with the AMP-activated kinase agonist, AICAR, increases Vmax for net 3-O-methylglucose uptake by 2-fold. Glucose depletion and treatment with potassium cyanide, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and AICAR also increase AMP-dependent kinase phosphorylation in bEnd.3 cells. These results suggest that metabolic stress rapidly stimulates blood-brain barrier endothelial cell sugar transport by acute up-regulation of plasma membrane GLUT1 levels, possibly involving AMP-activated kinase activity.
Thomas Jansson - One of the best experts on this subject based on the ideXlab platform.
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insulin stimulates GLUT4 trafficking to the syncytiotrophoblast basal plasma membrane in the human placenta
The Journal of Clinical Endocrinology and Metabolism, 2019Co-Authors: Laura B Jamesallan, Theresa L Powell, Jaron Arbet, Stephanie B Teal, Thomas JanssonAbstract:Context Placental transport capacity influences fetal glucose supply. The syncytiotrophoblast is the transporting epithelium in the human placenta, expressing glucose transporters (GLUT) and insulin receptors (IR) in its maternal-facing microvillous (MVM) and fetal-facing basal plasma membrane (BM). Objective The objectives of this study were to (1) determine the expression of the insulin-sensitive GLUT4 glucose transporter and IR in the syncytiotrophoblast plasma membranes across gestation in normal pregnancy and in pregnancies complicated by maternal obesity and (2) assess the effect of insulin on GLUT4 plasma membrane trafficking in human placental explants. Design, setting, participants Placental tissue was collected across gestation from women with normal body mass index (BMI) and obese mothers with appropriate for gestational age (AGA) and macrosomic infants. MVM and BM were isolated. Main outcome measures Protein expression of GLUT4, GLUT1 and IR were determined by western blot. Results GLUT4 was exclusively expressed in the BM and IR was predominantly expressed in the MVM, with increasing expression across gestation. BM GLUT1 expression was increased and BM GLUT4 expression was decreased in obese women delivering macrosomic babies. In placental villous explants incubation with insulin stimulated Akt (S473) phosphorylation (+76%, p=0.0003, n=13) independent of maternal BMI and increased BM GLUT4 protein expression (+77%, p=0.0013, n=7) in placentas from lean but not obese women. Conclusion We propose that maternal insulin stimulates placental glucose transport by promoting GLUT4 trafficking to the BM, which may enhance glucose transfer to the fetus in response to postprandial hyperinsulinemia in women with normal BMI.
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glucose transporter isoform 4 is expressed in the syncytiotrophoblast of first trimester human placenta
Human Reproduction, 2005Co-Authors: Anette Ericsson, Bengt Hamark, Theresa L Powell, Thomas JanssonAbstract:BACKGROUND: Placental glucose transport mechanisms in early pregnancy are poorly understood. The aims of this study were to investigate the expression of glucose transporter (GLUT) isoforms 1, 3 and 4 in first trimester villous tissue, to assess the effects of insulin on glucose uptake and compare them with term. METHODS: The expression of GLUT isoforms was investigated using immunohistochemistry, Western blot and reverse transcription (RT) ‐PCR in trophoblast tissue from terminations at 6 ‐ 13 weeks gestation and term. The effects of insulin (300 ng/ml, 1 h) on glucose uptake were studied in villous fragments. RESULTS: In the first trimester, GLUT1 and GLUT3 were present in the microvillous membrane and the cytotrophoblast, and GLUT4 in perinuclear membranes in the cytosol of the syncytiotrophoblast (ST). GLUT4 protein (48 kDa) and mRNA were identified in trophoblast homogenates. Whereas GLUT1 was expressed abundantly in term placenta, the expression of GLUT3 and 4 was markedly lower at term compared with first trimester. Insulin increased glucose uptake by 182% (n 5 6, P< 0.05) in first trimester fragments, but not in term fragments. CONCLUSIONS: The insulin-regulatable GLUT4 is expressed in the cytosol of first trimester ST compatible with a role for GLUT4 in placental glucose transport in early pregnancy. The placental expression pattern of GLUT isoforms in early pregnancy is distinct from that later in pregnancy.
Mark Levine - One of the best experts on this subject based on the ideXlab platform.
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intestinal dehydroascorbic acid dha transport mediated by the facilitative sugar transporters glut2 and glut8
Journal of Biological Chemistry, 2013Co-Authors: Christopher Corpe, Jin Wang, Hadi Alhasani, Mark LevineAbstract:Intestinal vitamin C (Asc) absorption was believed to be mediated by the Na+-dependent ascorbic acid transporter SVCT1. However, Asc transport across the intestines of SVCT1 knock-out mice is normal indicating that alternative ascorbic acid transport mechanisms exist. To investigate these mechanisms, rodents were gavaged with Asc or its oxidized form dehydroascorbic acid (DHA), and plasma Asc concentrations were measured. Asc concentrations doubled following DHA but not Asc gavage. We hypothesized that the transporters responsible were facilitated glucose transporters (GLUTs). Using Xenopus oocyte expression, we investigated whether facilitative glucose transporters GLUT2 and GLUT5–12 transported DHA. Only GLUT2 and GLUT8, known to be expressed in intestines, transported DHA with apparent transport affinities (Km) of 2.33 and 3.23 mm and maximal transport rates (Vmax) of 25.9 and 10.1 pmol/min/oocyte, respectively. Maximal rates for DHA transport mediated by GLUT2 and GLUT8 in oocytes were lower than maximal rates for 2-deoxy-d-glucose (Vmax of 224 and 32 pmol/min/oocyte for GLUT2 and GLUT8, respectively) and fructose (Vmax of 406 and 116 pmol/min/oocyte for GLUT2 and GLUT8, respectively). These findings may be explained by differences in the exofacial binding of substrates, as shown by inhibition studies with ethylidine glucose. DHA transport activity in GLUT2- and GLUT8-expressing oocytes was inhibited by glucose, fructose, and by the flavonoids phloretin and quercetin. These studies indicate intestinal DHA transport may be mediated by the facilitative sugar transporters GLUT2 and GLUT8. Furthermore, dietary sugars and flavonoids in fruits and vegetables may modulate Asc bioavailability via inhibition of small intestinal GLUT2 and GLUT8.
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glucose transporter isoforms glut1 and glut3 transport dehydroascorbic acid
Journal of Biological Chemistry, 1997Co-Authors: Steven C Rumsey, Guo Wei Xu, Oran Kwon, Charles F Burant, Ian A. Simpson, Mark LevineAbstract:Abstract Dehydroascorbic acid (DHA) is rapidly taken up by cells and reduced to ascorbic acid (AA). Using the Xenopus laevis oocyte expression system we examined transport of DHA and AA via glucose transporter isoforms GLUT1–5 and SGLT1. The apparentK m of DHA transport via GLUT1 and GLUT3 was 1.1 ± 0.2 and 1.7 ± 0.3 mm, respectively. High performance liquid chromatography analysis confirmed 100% reduction of DHA to AA within oocytes. GLUT4 transport of DHA was only 2–4-fold above control and transport kinetics could not be calculated. GLUT2, GLUT5, and SGLT1 did not transport DHA and none of the isoforms transported AA. Radiolabeled sugar transport confirmed transporter function and identity of all cDNA clones was confirmed by restriction fragment mapping. GLUT1 and GLUT3 cDNA were further verified by polymerase chain reaction. DHA transport activity in both GLUT1 and GLUT3 was inhibited by 2-deoxyglucose, d-glucose, and 3-O-methylglucose among other hexoses while fructose and l-glucose showed no inhibition. Inhibition by the endofacial inhibitor, cytochalasin B, was non-competitive and inhibition by the exofacial inhibitor, 4,6-O-ethylidene-α-glucose, was competitive. Expressed mutant constructs of GLUT1 and GLUT3 did not transport DHA. DHA and 2-deoxyglucose uptake by Chinese hamster ovary cells overexpressing either GLUT1 or GLUT3 was increased 2–8-fold over control cells. These studies suggest GLUT1 and GLUT3 isoforms are the specific glucose transporter isoforms which mediate DHA transport and subsequent accumulation of AA.