Increase in human placental glucose transporter-1 during pregnancy

Sakata M, Kurachi H, Imai T, Tadokoro C, Yamaguchi M, Yoshimoto Y, Oka Y, Miyaka A. Increase in human placental glucose transporter-1 during pregnancy. Eur J Endocrinol 1995;132:206–12. ISSN 0804–4643 Glucose transporter-1 (GLUT1) has been found in high abundance in human placentas. The purpose of this study was to analyze the changes in the level of GLUT1 during pregnancy. We have analzyed the gestational changes in human placental GLUT1 by [3H]cytochalasin B binding assay, immunoblot analysis and Northern blot analysis. Levels of [3H]cytochalasin B binding to placental membrane in pregnancy at 7–10, 18–20 and 38–40 weeks were 4.2 ± 0.47, 5.2 ± 0.46 and 7.2 ± 0.40 (mean±sem, N = 4) μmol/kg protein, respectively. Amounts of 49-kD and 60-kD proteins detected in immunoblot analysis increased during pregnancy in parallel with [3H]cytochalasin B binding to placental membrane. The GLUT1 mRNA levels also increased during pregnancy. Immunohistochemical examination showed that GLUT1 was localized in the plasma membranes of trophoblast cells. These findings showed that the GLUT1 level increased during pregnancy in human placentas and suggest that GLUT1 may play an important role in fetal development. Masahiro Sakata, Department of Obstetrics and Gynecology, Osaka University Medical School, 2-2 Yamadaoka, Suita, Osaka 565, Japan

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Studies of renal injury. I. Gentamicin toxicity and expression of basolateral transporters

AJP Renal Physiology ◽

10.1152/ajprenal.1996.270.2.f245 ◽

1996 ◽

Vol 270 (2) ◽

pp. F245-F253 ◽

Cited By ~ 2

Author(s):

J. H. Dominguez ◽

C. C. Hale ◽

M. Qulali

Keyword(s):

Stress Response ◽

Glucose Transporter ◽

Mrna Level ◽

Specific Protein ◽

Renal Tubules ◽

Mrna Levels ◽

Glucose Transporter 1 ◽

Protein Levels ◽

Expression Of Genes ◽

Glut1 Mrna

Gentamicin nephrotoxicity may arise in part from alterations in the expression of genes critical for renal proximal tubule metabolism. We tested the hypothesis that gentamicin suppressed the gene expression of the Na+/Ca2+ exchanger (NaCaX), glucose transporter 1 (GLUT1) and alpha 1-subunit of Na(+)-K(+)-ATPase (alpha 1-NKA) in renal tubules. The products of these genes mediate Na(+)-dependent Ca2+ efflux, glucose efflux and influx, and ATP-dependent Na+ efflux across tubular basolateral membranes, respectively. After 10 days of gentamicin intoxication (40 mg/kg ip, twice daily), levels of mRNAs encoding NaCaX and the cognate protein declined. GLUT1 mRNA levels increased, although GLUT1 protein levels were also reduced. Moreover, whereas alpha 1-NKA mRNA levels remained unchanged, alpha 1-NKA protein levels were also reduced. We suggest that the higher GLUT1 mRNA level is part of the stress response to tubular injury. However, regardless of the mRNA level, the most consistent effect of gentamicin was reduction of specific protein levels. We propose that failure to translate high levels of mRNA into proportionally high levels of protein, as in the case of GLUT1, may attenuate the expression of stress response gene products, and thus diminish the possibility of recovery in gentamicin intoxication.

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High concentration of glucose decreases glucose transporter-1 expression in mouse placenta in vitro and in vivo

Journal of Endocrinology ◽

10.1677/joe.0.1600443 ◽

1999 ◽

Vol 160 (3) ◽

pp. 443-452 ◽

Cited By ~ 27

Author(s):

K Ogura ◽

M Sakata ◽

M Yamaguchi ◽

H Kurachi ◽

Y Murata

Keyword(s):

Glucose Concentration ◽

Glucose Transporter ◽

Cytochalasin B ◽

High Concentration ◽

Glucose Transporter 1 ◽

Protein Levels ◽

Placental Cells ◽

Glut1 Mrna

Facilitative glucose transporter-1 (GLUT1) is expressed abundantly and has an important role in glucose transfer in placentas. However, little is known about the regulation of GLUT1 expression in placental cells. We studied the changes in placental GLUT1 levels in relation to changes in glucose concentration in vitro and in vivo. In in vitro experiments, dispersed mouse placental cells were incubated under control (5.5 mM) and moderately high (22 mM) glucose concentrations, and 2-deoxyglucose uptake into cells was studied on days 1-5 of culture. After 4 days of incubation under both conditions, GLUT1 mRNA and proten levels were examined by Northern and immunoblot analyses. Treatment of cells with 22 mM glucose resulted in a significant decrease in 2-deoxyglucose uptake compared with control, from day 2 to day 5 of culture. Moreover, GLUT1 mRNA and protein levels on day 4 of culture were significantly reduced in cells incubated with 22 mM glucose compared with control. Next, we rendered mice diabetic by administering 200 micrograms/g body weight streptozotocin (STZ) on day 8 of pregnancy. Animals were killed on day 12 of pregnancy and placental tissues were obtained. [3H]Cytochalasin B binding study was carried out to assess total GLUTs, and GLUT1 mRNA and protein were measured as above. [3H]Cytochalasin B binding sites in placentas from STZ-treated mice were significantly less than those in control mice. Northern and immunoblot analyses revealed a significant decrease in GLUT1 mRNA and protein levels in diabetic mice compared with the controls. These findings suggest that the glucose concentration may regulate the expression of placental GLUT1.

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Glycaemia regulates the glucose transporter number in the plasma membrane of rat skeletal muscle

Biochemical Journal ◽

10.1042/bj2840341 ◽

1992 ◽

Vol 284 (2) ◽

pp. 341-348 ◽

Cited By ~ 45

Author(s):

D Dimitrakoudis ◽

T Ramlal ◽

S Rastogi ◽

M Vranic ◽

A Klip

Keyword(s):

Skeletal Muscle ◽

Plasma Membrane ◽

Glucose Transport ◽

Transport System ◽

Plasma Membranes ◽

Glucose Transporter ◽

Cytochalasin B ◽

Glucose Transporters ◽

Mrna Levels ◽

Rat Skeletal Muscle

The number of glucose transporters was measured in isolated membranes from diabetic-rat skeletal muscle to determine the role of circulating blood glucose levels in the control of glucose uptake into skeletal muscle. Three experimental groups of animals were investigated in the post-absorptive state: normoglycaemic/normoinsulinaemic, hyperglycaemic/normoinsulinaemic and hyperglycaemic/normoinsulinaemic made normoglycaemic/normoinsulinaemic by phlorizin treatment. Hyperglycaemia caused a reversible decrease in total transporter number, as measured by cytochalasin B binding, in both plasma membranes and internal membranes of skeletal muscle. Changes in GLUT4 glucose transporter protein mirrored changes in cytochalasin B binding in plasma membranes. However, there was no recovery of GLUT4 levels in intracellular membranes with correction of glycaemia. GLUT4 mRNA levels decreased with hyperglycaemia and recovered only partially with correction of glycaemia. Conversely, GLUT1 glucose transporters were only detectable in the plasma membranes; the levels of this protein varied directly with glycaemia, i.e. in the opposite direction to GLUT4 glucose transporters. This study demonstrates that hyperglycaemia, in the absence of hypoinsulinaemia, is capable of down-regulating the glucose transport system in skeletal muscle, the major site of peripheral resistance to insulin-stimulated glucose transport in diabetes. Furthermore, correction of hyperglycaemia causes a complete restoration of the transport system in the basal state (determined by the transporter number in the plasma membrane), but possibly only an incomplete recovery of the transport system's ability to respond to insulin (since there is no recovery of GLUT4 levels in the intracellular membrane insulin-responsive transporter pool). Finally, the effect of hyperglycaemia is specific for glucose transporter isoforms, with GLUT1 and GLUT4 proteins varying respectively in parallel and opposite directions to levels of glycaemia.

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Caffeine inhibits glucose transport by binding at the GLUT1 nucleotide-binding site

AJP Cell Physiology ◽

10.1152/ajpcell.00001.2015 ◽

2015 ◽

Vol 308 (10) ◽

pp. C827-C834 ◽

Cited By ~ 14

Author(s):

Jay M. Sage ◽

Anthony J. Cura ◽

Kenneth P. Lloyd ◽

Anthony Carruthers

Keyword(s):

Binding Site ◽

Glucose Transport ◽

Glucose Transporter ◽

Cytochalasin B ◽

Sugar Transport ◽

Nucleotide Binding ◽

Nucleotide Binding Site ◽

Sugar Uptake ◽

Glucose Transporter 1 ◽

Intracellular Atp

Glucose transporter 1 (GLUT1) is the primary glucose transport protein of the cardiovascular system and astroglia. A recent study proposes that caffeine uncompetitive inhibition of GLUT1 results from interactions at an exofacial GLUT1 site. Intracellular ATP is also an uncompetitive GLUT1 inhibitor and shares structural similarities with caffeine, suggesting that caffeine acts at the previously characterized endofacial GLUT1 nucleotide-binding site. We tested this by confirming that caffeine uncompetitively inhibits GLUT1-mediated 3- O-methylglucose uptake in human erythrocytes [ Vmax and Km for transport are reduced fourfold; Ki(app) = 3.5 mM caffeine]. ATP and AMP antagonize caffeine inhibition of 3- O-methylglucose uptake in erythrocyte ghosts by increasing Ki(app) for caffeine inhibition of transport from 0.9 ± 0.3 mM in the absence of intracellular nucleotides to 2.6 ± 0.6 and 2.4 ± 0.5 mM in the presence of 5 mM intracellular ATP or AMP, respectively. Extracellular ATP has no effect on sugar uptake or its inhibition by caffeine. Caffeine and ATP displace the fluorescent ATP derivative, trinitrophenyl-ATP, from the GLUT1 nucleotide-binding site, but d-glucose and the transport inhibitor cytochalasin B do not. Caffeine, but not ATP, inhibits cytochalasin B binding to GLUT1. Like ATP, caffeine renders the GLUT1 carboxy-terminus less accessible to peptide-directed antibodies, but cytochalasin B and d-glucose do not. These results suggest that the caffeine-binding site bridges two nonoverlapping GLUT1 endofacial sites—the regulatory, nucleotide-binding site and the cytochalasin B-binding site. Caffeine binding to GLUT1 mimics the action of ATP but not cytochalasin B on sugar transport. Molecular docking studies support this hypothesis.

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Regulation of lactate production and glucose transport as well as of glucose transporter 1 and lactate dehydrogenase A mRNA levels by basic fibroblast growth factor in rat Sertoli cells

Journal of Endocrinology ◽

10.1677/joe.0.1730335 ◽

2002 ◽

Vol 173 (2) ◽

pp. 335-343 ◽

Cited By ~ 31

Author(s):

MF Riera ◽

SB Meroni ◽

HF Schteingart ◽

EH Pellizzari ◽

SB Cigorraga

Keyword(s):

Growth Factor ◽

Glucose Transport ◽

Glucose Transporter ◽

Lactate Production ◽

Mrna Levels ◽

Dependent Manner ◽

Short Term ◽

Glucose Transporter 1 ◽

Ldh Activity

By using cultured rat Sertoli cells as a model, both the action of basic fibroblast growth factor (bFGF) on lactate production and the site of this action were studied. bFGF stimulated Sertoli cell lactate production in a dose-dependent manner (basal: 7.3+/-0.5; 0.1 ng/ml bFGF: 7.5+/-0.5; 1 ng/ml bFGF: 7.5+/-0.6; 10 ng/ml bFGF: 10.3+/-1.0; 30 ng/ml bFGF: 15.2+/-1.5; 50 ng/ml bFGF: 15.4+/-1.6 microg/microg DNA). Two major sites for the action of this growth factor were identified. First, bFGF was shown to exert short- and long-term stimulatory effects on glucose transport (basal: 1170+/-102; 30 ng/ml bFGF for 120 min: 1718+/-152 and basal: 718+/-64; 30 ng/ml bFGF for 48 h: 1069+/-69 d.p.m./microg DNA respectively). Short-term bFGF stimulation of glucose transport was not inhibited by the protein synthesis inhibitor cycloheximide. These results indicate that short-term bFGF stimulation of glucose uptake does not involve an increase in the number of glucose transporters. On the other hand, stimulation with bFGF for periods of time longer than 12 h increased glucose transporter 1 (GLUT1) mRNA levels. These increased mRNA levels were probably ultimately responsible for the increments in glucose uptake that are observed in long-term treated cultures. Secondly, bFGF increased lactate dehydrogenase (LDH) activity (basal: 31.0+/-1.4; 30 ng/ml bFGF: 45.7+/- 2.4 mIU/microg DNA). The principal subunit component of those LDH isozymes that favors the transformation of pyruvate to lactate is subunit A. bFGF increased LDH A mRNA levels in a dose- and time-dependent manner. In summary, the results presented herein show that glucose transport, LDH activity and GLUT1 and LDH A mRNA levels are regulated by bFGF to achieve an increase in lactate production. These observed regulatory actions provide unequivocal evidence of the participation of bFGF in Sertoli cell lactate production which may be related to normal germ cell development.

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Cycloheximide decreases glucose transporters in rat adipocyte plasma membranes without affecting insulin-stimulated glucose transport

Biochemical Journal ◽

10.1042/bj2510491 ◽

1988 ◽

Vol 251 (2) ◽

pp. 491-497 ◽

Cited By ~ 12

Author(s):

S Matthaei ◽

J M Olefsky ◽

E Karnieli

Keyword(s):

Protein Synthesis ◽

Glucose Transport ◽

Plasma Membranes ◽

Glucose Transporter ◽

Cytochalasin B ◽

Glucose Transporters ◽

Electrophoretic Analysis ◽

Adipocyte Plasma Membranes ◽

Inhibitor Of Protein Synthesis ◽

Adipose Cells

This study examines the relationship between insulin-stimulated glucose transport and insulin-induced translocation of glucose transporters in isolated rat adipocytes. Adipose cells were incubated with or without cycloheximide, a potent inhibitor of protein synthesis, for 60 min and then for an additional 30 min with or without insulin. After the incubation we measured 3-O-methylglucose transport in the adipose cells, and subcellular membrane fractions were prepared. The numbers of glucose transporters in the various membrane fractions were determined by the cytochalasin B binding assay. Basal and insulin-stimulated 3-O-methylglucose uptakes were not affected by cycloheximide. Furthermore, cycloheximide affected neither Vmax. nor Km of insulin-stimulated 3-O-methylglucose transport. In contrast, the number of glucose transporters in plasma membranes derived from cells preincubated with cycloheximide and insulin was markedly decreased compared with those from cells incubated with insulin alone (10.5 +/- 0.8 and 22.2 +/- 1.8 pmol/mg of protein respectively; P less than 0.005). The number of glucose transporters in cells incubated with cycloheximide alone was not significantly different compared with control cells. SDS/polyacrylamide-gel-electrophoretic analysis of [3H]cytochalasin-B-photolabelled plasma-membrane fractions revealed that cycloheximide decreases the amount of labelled glucose transporters in insulin-stimulated membranes. However, the apparent molecular mass of the protein was not changed by cycloheximide treatment. The effect of cycloheximide on the two-dimensional electrophoretic profile of the glucose transporter in insulin-stimulated low-density microsomal membranes revealed a decrease in the pI-6.4 glucose-transporter isoform, whereas the insulin-translocatable isoform (pI 5.6) was decreased. Thus the observed discrepancy between insulin-stimulated glucose transport and insulin-induced translocation of glucose transporters strongly suggests that a still unknown protein-synthesis-dependent mechanism is involved in insulin activation of glucose transport.

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Insulin regulation of solute carrier family 2 member 1 (glucose transporter 1) expression and glucose uptake in decidualizing human endometrial stromal cells: an in vitro study

Reproductive Biology and Endocrinology ◽

10.1186/s12958-020-00674-0 ◽

2020 ◽

Vol 18 (1) ◽

Author(s):

Ivika Jakson ◽

Dorina Ujvari ◽

Sebastian Brusell Gidlöf ◽

Angelica Lindén Hirschberg

Keyword(s):

Glucose Uptake ◽

Stromal Cells ◽

Glucose Transporter ◽

Mrna Levels ◽

Solute Carrier Family ◽

Endometrial Stromal Cells ◽

Glucose Transporter 1 ◽

Protein Levels ◽

Solute Carrier

Abstract Background Solute carrier family 2 member 1 (SLC2A1; previously known as glucose transporter 1), is the most abundant glucose transporter in human endometrium and is up-regulated during decidualization, whereas high insulin may have a negative impact on this process. The present study aimed to investigate the effect of insulin on the expression of SLC2A1 and glucose uptake in decidualizing human endometrial stromal cells. Methods We induced in vitro decidualization of endometrial stromal cells obtained from regularly menstruating healthy non-obese women. The cells were treated with increasing concentrations of insulin, and the involvement of the transcription factor forkhead box O1 (FOXO1) was evaluated using a FOXO1 inhibitor. SLC2A1 mRNA levels were measured by Real-Time PCR and protein levels were evaluated by immunocytochemistry. Glucose uptake was estimated by an assay quantifying the cellular uptake of radioactive glucose. One-way ANOVA, Dunnett’s multiple comparisons test and paired t-test were used to determine the statistical significance of the results. Results We found that insulin dose-dependently decreased SLC2A1 mRNA levels and decreased protein levels of SLC2A1 in decidualizing human endometrial stromal cells. Transcriptional inactivation of FOXO1 seems to explain at least partly the down-regulation of SLC2A1 by insulin. Glucose uptake increased upon decidualization, whereas insulin treatment resulted in a slight inhibition of the glucose uptake, although not significant for all insulin concentrations. Conclusions These results indicate an impairment of decidualization by high concentrations of insulin. Future studies will determine the clinical significance of our results for endometrial function and decidualization in women with insulin resistance and hyperinsulinemia.

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Abstract 290: Microrna-199a and Glut1 are Differentially Expressed in Primary Human Arterial and Venous Endothelium and are Deregulated in Culture

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.33.suppl_1.a290 ◽

2013 ◽

Vol 33 (suppl_1) ◽

Author(s):

Matthew R Richardson ◽

Xianyin Lai ◽

Mervin C Yoder

Keyword(s):

Cell Fate ◽

Umbilical Cord ◽

Glucose Transporter ◽

Cell Types ◽

Enzymatic Digestion ◽

Protein Quantification ◽

Human Umbilical Cord ◽

Mrna Levels ◽

Downstream Target ◽

Glut1 Mrna

Introduction Studies over the last decade have established that the identity of arterial and venous endothelial cells (ECs) is governed by both genetic and environmental factors. Although the ephrin/Eph system is known to be a key determinant of that identity, it is unlikely that system is solely responsible for all the differences between arterial and venous ECs. Furthermore, microRNAs are becoming increasingly recognized for their important role in regulating gene expression and cell fate. We investigate here novel potential markers and regulators of arterial EC identity and the effects of environmental cues on these molecules. Methods ECs were freed from the basement membrane of human umbilical arteries and veins (HUAECs, HUVECs) (n=6) by enzymatic digestion and purified by flow cytometry (CD31+, CD45-) prior to lysis and quantitative PCR or analysis using HPLC-MS/MS. Protein quantification was performed using a proprietary software package (IdentiQuantXL TM ). HUVECs and HUAECs were harvested directly from umbilical cords and placed directly into collagen coated 6-well plates and cultured in standard conditions in EGM2. Results We found that microRNA-199a levels were 23 fold higher in HUVECs than HUAECs (p=0.0001). One of microRNA-199a’s theoretical downstream targets is GLUT1, a facilitative glucose transporter thought to be responsible for basal glucose uptake. GLUT1 mRNA and protein levels were significantly higher (5.5 and 4.3 fold, respectively) in HUAEC than HUVECs (p=0.00002). Using tissue immunohistochemistry (IHC) of we confirmed that GLUT1 expression is restricted to arterial ECs in human umbilical cord and adult peripheral artery and vein sections. Finally, GLUT1 mRNA levels are 2 fold higher in HUAECs than HUVECs in culture at P2 and levels decrease until P5 when they become equal indicating that culture has a homogenizing effect on endothelial heterogeneity. Conclusions MicroRNA-199a levels are higher in HUVECs than HUAECs and GLUT1, its downstream target, is higher in HUAECs both at the mRNA and protein level. Protein expression is restricted to arteries in both the new born umbilical cord and adult peripheral vessels. Importantly, expression in both cell types is downregulated and equilibrated by cell culture.

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The Tibetan PHD2 Polymorphism Asp4Glu Is Associated with Hypersensitivity of Erythroid Progenitors to EPO and Upregulation of HIF-1 Regulated Genes Hexokinase (HK1) and Glucose Transporter 1 (SLC2A/GLUT1),

Blood ◽

10.1182/blood.v118.21.3388.3388 ◽

2011 ◽

Vol 118 (21) ◽

pp. 3388-3388 ◽

Cited By ~ 2

Author(s):

Felipe R Lorenzo ◽

Sabina Swierczek ◽

Chad Daniel Huff ◽

Josef T. Prchal

Keyword(s):

High Altitude ◽

Glucose Transporter ◽

Conflicts Of Interest ◽

Negative Regulator ◽

Molecular Defect ◽

Mrna Levels ◽

Erythroid Progenitors ◽

Glucose Transporter 1 ◽

Exon 1 ◽

Functional Consequences

Abstract Abstract 3388 The hypoxic response, mediated by hypoxia inducible transcription factors (HIFs), is central to the control and development of many essential biological functions, including erythropoiesis. As a high-altitude population, many Tibetans have developed a remarkable ability to protect against several hypoxic complications, including polycythemia and other harmful responses exhibited by non-adapted populations upon exposure to severe hypoxia. We have identified 10 genes involved in high-altitude adaptation in Tibetans, including a principal negative regulator of HIF-1a and HIF-2 a peptides, i.e. PHD2 (EGLN1), as well as HIF2A (EPAS1) (Simonson, Science 2010). At this meeting last year (Lorenzo, Abstract# 2602 ASH 2010), we reported a novel PHD2 Asp4Glu mutation that we found in 57 of 94 Tibetan, 2 of 88 Asian and 0 of 38 Caucasian chromosomes. In most Tibetan samples, this variant is associated with a previously reported, unvalidated PHD2 polymorphism, Cys127Ser (found in 70 of 94 Tibetan, 27 of 88 Asian and 4 of 38 Caucasian chromosomes). To study the functional consequences of this PHD2 Asp4Glu mutation, we recruited five Tibetan volunteers living in Utah, four of whom were homozygous and 1 heterozygous for PHD2 Asp4Glu and Cys127Ser. We unexpectedly found that homozygotes for the exon 1 PHD2 mutation had markedly hypersensitive erythroid BFU-E (Fig.1) compared to the range of normal controls we have standardized over several decades. Interestingly, erythroid progenitors from individuals with Chuvash polycythemia or a HIF-2a gain-of-function mutation also have hypersensitive BFU-E. To determine if the Tibetan erythroid hypersensitivity data may be explained by increased HIF activity, we have quantified HIF target gene expression in subject granulocytes and found a significant increase in hexokinase (HK1) and glucose transporter (GLUT1/SLC2A) mRNA levels. These data report the first molecular defect with functional consequences that is associated with the complex Tibetan adaptation to extreme hypoxia. Disclosures: No relevant conflicts of interest to declare.

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Identification and characterization of glucose transport proteins in plasma membrane- and Golgi vesicle-enriched fractions prepared from lactating rat mammary gland

Biochemical Journal ◽

10.1042/bj2720099 ◽

1990 ◽

Vol 272 (1) ◽

pp. 99-105 ◽

Cited By ~ 28

Author(s):

R J Madon ◽

S Martin ◽

A Davies ◽

H A C Fawcett ◽

D J Flint ◽

...

Keyword(s):

Plasma Membrane ◽

Mammary Gland ◽

Membrane Protein ◽

Rat Brain ◽

Glucose Transport ◽

Binding Sites ◽

Plasma Membranes ◽

Glucose Transporter ◽

Cytochalasin B ◽

Golgi Vesicle

Plasma membrane- and Golgi vesicle-enriched membrane fractions were prepared from day-10 lactating rat mammary glands. Each fraction was found to contain a single set of D-glucose-inhibitable cytochalasin B-binding sites: plasma membranes and Golgi vesicles bound 20 +/- 2 and 53 +/- 4 pmol of cytochalasin/mg of membrane protein (means +/- S.E.M.), with dissociation constants of 259 +/- 47 and 520 +/- 47 nM respectively. Anti-peptide antibodies against the C-terminal region (residues 477-492) of the rat brain/human erythrocyte glucose transporter labelled a sharp band of apparent Mr 50,000 on Western blots of both fractions. Treatment with endoglycosidase F before blotting decreased the apparent Mr of this band to 38,000, indicating that it corresponded to a glycoprotein. Confirmation that this immunologically cross-reactive band was a glucose transporter was provided by the demonstration that it could be photoaffinity-labelled, in a D-glucose-sensitive fashion, with cytochalasin B. Quantitative Western blotting studies yielded values of 28 +/- 5 and 23 +/- 3 pmol of immunologically cross-reactive glucose transporters/mg of membrane protein in the plasma membrane and Golgi vesicle fractions respectively. From comparison with the concentration of cytochalasin B-binding sites, it is concluded that a protein homologous to the rat brain glucose transporter constitutes the major glucose transport species in the plasma membranes of mammary gland epithelial cells. Glucose transporters are also found in the Golgi membranes of these cells, at least half of them being similar, if not identical, to the transporters of the plasma membrane. However, their function in this location remains unclear.

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