Increased Hexose Transport Research Articles

Increased hexose transport in the roots of tomato plants submitted to prolonged hypoxia

We investigated the effects of prolonged hypoxia on the sugar uptake in tomato (Solanum lycopersicum L. var. MP-1) roots. Hydroponic cultures of whole tomato plants were submitted to hypoxic treatment for 1 week, and the roots were analyzed for sugar concentrations, hexose uptake and hexose transporter expression level. Contrary to what has been observed after anoxic shock or short-term hypoxic treatment, we show that sugar concentrations increase and hexose uptake is up-regulated in the roots after 1 week of hypoxic treatment. Increased hexose transport is concomitant with the induction of the hexose transporter gene LeHT2. These responses may be due either to a direct effect of low O(2) supply, or to a secondary effect associated with the increase in sugar concentrations, which, typically, develops in most hypoxic plants.

Nicotinamide is not a substrate of the facilitative hexose transporter GLUT1.

It has been proposed that GLUT1, a membrane protein that transports hexoses and the oxidized form of vitamin C, dehydroascorbic acid, is also a transporter of nicotinamide (Sofue, M., Yoshimura, Y., Nishida, M., and Kawada, J. (1992) Biochem. J. 288, 669-674). To ascertain this, we studied the transport of 2-deoxy-D-glucose, 3-O-methyl-D-glucose, and nicotinamide in human erythrocytes and right-side-out and inside-out erythrocyte membrane vesicles. The transport of nicotinamide was saturable, with a K(M) for influx and efflux of 6.1 and 6.2 mM, respectively. We found that transport of the hexoses was not competed by nicotinamide in both the erythrocytes and the erythrocyte vesicles. Likewise, the transport of nicotinamide was not affected by hexoses or by inhibitors of glucose transport such as cytochalasin B, genistein, and myricetin. On the other hand, nicotinamide blocked the binding of cytochalasin B to human erythrocyte membranes but did so in a noncompetitive manner. Using GLUT1-transfected CHO cells, we demonstrated that increased expression of GLUT1 was paralleled by a corresponding increase in hexose transport but that there were no changes in nicotinamide transport. Moreover, nicotinamide failed to affect the transport of hexoses in both control and GLUT1-transfected CHO cells. Therefore, our results indicates that GLUT1 does not transport nicotinamide, and we propose instead the existence of other systems for the translocation of nicotinamide across cell membranes.

Multiple effects of mercuric chloride on hexose transport in Xenopus oocytes

HgCl 2 had both stimulatory and inhibitory effects on [ 3H]2-deoxyglucose (DG) uptake in Xenopus laevis oocytes. The Hg dose response was complex, with 0.1–10 μM Hg increasing total DG uptake, 30–50 μM Hg inhibiting, and concentrations >100 μM increasing uptake. Analyses of the effects of Hg on DG transport kinetics and cell membrane permeability indicated that low concentrations of Hg stimulated mediated uptake, intermediate concentrations inhibited mediated uptake, but high Hg concentrations increased non-mediated uptake. 10 μM Hg increased the apparent V max for DG uptake, but caused little or no change in apparent K m. Phenylarsine oxide prevented the increase in DG uptake by 10 μM Hg, suggesting that the increase was due to transporter recruitment. Microinjecting low doses of HgCl 2 into the cell increased mediated DG uptake. Higher intracellular doses of Hg increased both mediated and non-mediated DG uptake. Both insulin and Hg cause cell swelling in isotonic media and, for insulin, this swelling has been linked to the mechanism of hormone action. Osmotically swelling Xenopus oocytes stimulated DG transport 2–5-fold and this increase was due to an increased apparent V max. Exposing cells to 10 μM Hg or 140 nM insulin both increased cellular water content by 18% and increased hexose transport 2–4-fold. These data indicate that low concentrations of Hg and insulin affect hexose transport in a similar manner and that for both an increase cellular water content could be an early event in signaling the increase in hexose transport.

Hereditary cardiomyopathy in W/SSM rats: Biochemical mechanisms of development

One mechanism shown to be responsible for the occurrence of hypertrophic cardiomyopathy in rats of the W/SSM strain, in which this disease is genetically determined, is impairment of cellular membrane integrity resulting from increased hexose transport to cells, generation of hydroxyl radicals, and intensified lipid peroxidation.

Regulation of the functional expression of hexose transporter GLUT-1 by glucose in murine fibroblasts: role of lysosomal degradation.

The nature of the membrane compartments involved in the regulation by glucose of hexose transport is not well defined. The effect of inhibitors of lysosomal protein degradation on hexose transport (i.e., uptake of [3H]-2-deoxy-D-glucose) and hexose transporter protein GLUT-1 (i.e., immunoblotting with antipeptide serum) in glucose-fed and -deprived cultured murine fibroblasts (3T3-C2 cells) was studied. The acidotropic amines chloroquine (20 microM) and ammonium chloride (10 mM) cause accumulation (both approximately 4-fold) of GLUT-1 protein and a small increase (both approximately 25%) in hexose transport in glucose-fed fibroblasts (24 h). The endopeptidase inhibitor, leupeptin (100 microM) causes accumulation (approximately 4-fold) of GLUT-1 protein in glucose-fed fibroblasts (24 h) without changing hexose transport (less than or equal to 5%). These agents do not greatly alter the electrophoretic mobility of GLUT-1. Neither chloroquine nor leupeptin augment the glucose deprivation (24 h) induced increases in hexose transport (approximately 4-fold) and GLUT-1 content (approximately 7-fold). In contrast, chloroquine or leupeptin diminish the reversal by glucose refeeding of the glucose deprivation induced accumulation of GLUT-1 protein but fail to alter the return of hexose transport to control levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibitors of protein synthesis cause increased hexose transport in cultured human fibroblasts by a mechanism other than transporter translocation.

We have investigated the effect of various inhibitors of protein synthesis on hexose transport in human skin fibroblasts using 2-deoxy-D-glucose (2-DG) and 3-0-methyl-D-glucose (3-OMG) to measure hexose transport. Exposure of glucose-fed, serum-free cultures to cycloheximide (CHX) (50 micrograms/ml) for 6 h resulted in increased 2-DG transport (3.81 +/- .53 vs. 6.62 +/- .88 nmoles/mg protein/2 min; n = 9) and 3-OMG transport (1.36 +/- .66 vs. 3.18 +/- .83 nmoles/mg protein/30 sec; n = 4) in the CHX exposed group. Under these conditions inhibition of protein synthesis was greater than 90%. This CHX induced transport increase was time dependent (approaching maximum within 1 h of exposure to CHX) and related to an increase in the Vmax of hexose transport in the CHX exposed group (18.4 +/- 2.4 vs. 4.8 +/- 1.1 nmoles 2-DG/mg protein/min) with no difference in the transport Km (1.55 +/- .63 vs. 2.92 +/- .59 mM). Further, the CHX induced increase in hexose transport was reversible. Exposure of human fibroblasts to inhibitors of protein synthesis with different mechanisms of action (e.g., puromycin, pactamycin, or CHX) all generated hexose transport increases in a concentration-dependent fashion correlating with their increasing inhibitory effects on protein synthesis. Nucleotidase enriched (i.e., plasma membrane) fractions of control and CHX-exposed cells showed no differences in D-glucose inhibitable cytochalasin B binding activity. Further, quantitative Western analysis of nucleotidase enriched fractions indicated CHX exposure resulted in no significant increase in glucose transporter mass compared with control plasma membrane fractions. Glucose deprived cells, however, which exhibited increased sugar transport comparable to the CHX-exposed group, did show increased glucose transporter mass in the plasma membrane fraction. The data indicate that inhibitors of protein synthesis can cause a significant elevation in hexose transport and that the hexose transporter mass in the isolated plasma membrane fractions did not reflect the whole cell transport change. It is suggested that a mechanism other than glucose transporter translocation to the plasma membrane may be involved in causing this sugar transport increase.

Permanent hexose transport upregulation in a respiration-deficient human fibroblast cell strain

The regulation of hexose transport was studied in a human diploid fibroblast respiration-deficient cell strain (WG750). Transport of 2-deoxy-D-glucose (2-DG) was greater than sixfold higher compared with an in vivo age-matched normal cell strain (MCH55). In addition, 3-O-methylglucose transport and 14CO2 production were elevated in the mutant cell strain. Kinetic analysis revealed that the increased sugar transport in mutant cells was due to an average 5.7-fold increase in the 2-DG maximal transport rate, with no observed differences in the transport Michaelis constant for both normal and mutant cells. Also, the inhibitor constants for D-glucose inhibition of 2-DG transport were nearly identical for both cell types. Glucose deprivation led to a similar time-dependent increase in hexose transport in both cell strains. Serum refeeding of glucose-fed serum-deprived cultures led to a progressive increase in 2-DG transport in normal cells, whereas mutant cells displayed a time-delayed increase in 2-DG transport. Exposure to 67 and 670 nM insulin stimulated 2-DG transport on average 1.99 +/- 0.25- and 2.33 +/- 0.26-fold, respectively, over basal transport in the normal cells, whereas the mutant cells were significantly less sensitive to the stimulatory effects of the hormone. Insulin binding and amino acid transport (i.e., alpha-aminoisobutyric acid uptake) in the normal and mutant cells were not different. Data obtained using Western blot analysis showed that WG750 (mutant) cells expressed an increase (approximately 4-fold) in total cellular HepG2 (erythroid-brain) transporter protein compared with normal cells, thus reflecting the changes seen in hexose transport.(ABSTRACT TRUNCATED AT 250 WORDS)

Glucose Deprivation Induces the Selective Accumulation of Hexose Transporter Protein GLUT-1 in the Plasma Membrane of Normal Rat Kidney Cells

Antibody to the carboxyl-terminal of hexose transporter protein GLUT-1 was used to localize this carrier in normal rat kidney (NRK) cells during D-glucose (Glc) deprivation. Glc-deprivation of NRK cells induces increased hexose transport, inhibits the glycosylation of GLUT-1, and increases the content of both native, 55,000 apparent mol wt (Mr) and aglyco, 38,000 Mr GLUT-1 polypeptides. The distribution of GLUT-1 protein in subcellular fractions isolated from Glc-fed NRK cells shows that the 55,000 Mr polypeptide is most abundant in intracellular membrane fractions. Glc-fed cells that have been tunicamycin treated contain principally the 38,000 Mr GLUT-1 polypeptide, which is found predominantly in intracellular membrane fractions. In Glc-deprived cells the 55,000 Mr GLUT-1 polypeptide localizes predominantly in the Golgi and plasma membrane fractions, whereas the more abundant 38,000 Mr GLUT-1 polypeptide is distributed throughout all membrane fractions. In Glc-deprived but fructose-fed cells only the 55,000 Mr GLUT-1 polypeptide is detected, and it is found predominantly in the plasma membrane and Golgi fractions. The localization of GLUT-1 protein was directly and specifically visualized in NRK cells by immunofluorescence microscopy. Glc-fed cells show little labeling of cell borders and a small punctate juxtanuclear pattern suggestive of localization to the Golgi and, perhaps, endoplasmic reticulum. Glc-fed cells that have been tunicamycin treated show large punctate intracellular accumulations suggestive of localization to distended Golgi and perhaps endoplasmic reticulum. Glc-deprived cells exhibited intense labeling of cell borders as well as intracellular accumulations. Glc-deprived but fructose-fed cells show fewer intracellular accumulations, and the labeling is, in general, limited to the cell borders. Our results suggest that Glc deprivation induces the selective accumulation of GLUT-1 in the plasma membrane of NRK cells.

Regulation of Glucose Transport by Tumor Necrosis Factor-α in Cultured Murine 3T3-L1 Fibroblasts

We have examined the ability of tumor necrosis factor-alpha (TNF) to stimulate hexose transport in 3T3-L1 fibroblasts. Activation of transport occurred in a dose- and time-dependent manner, with maximal stimulation (fourfold) occurring approximately 15 hours after exposure to 2.5 nM TNF. The protein synthesis inhibitor, cycloheximide, blunted TNF-induced 2-deoxyglucose uptake, suggesting that new synthesis was in part responsible for the observed increase. Increased hexose transport occurred concomitant with a 50% increase in general protein synthesis which preceded a fivefold stimulation of thymidine incorporation into DNA. Elevated hexose transport preceded an accumulation of glucose transporter (GLUT-1) mRNA. We have further demonstrated that the early accumulation of GLUT-1 mRNA is accompanied by accumulation of mRNA for both the prootooncogene c-fos as well as that for beta-actin. We suggest that these events represent the pleiotrophic growth factor-like properties of TNF-alpha.

Diabetes mellitus and sugar transport across the brush‐border and basolateral membranes of rat jejunal enterocytes.

1. The effects of streptozotocin-induced diabetes mellitus on active jejunal glucose uptake in vivo, and on galactose movement across the brush-border (phlorhizin-sensitive) and basolateral (phlorhizin-insensitive) membranes of isolated upper and mid-villus enterocytes has been studied. 2. Chronic diabetes increased unidirectional phlorhizin-sensitive galactose uptake by mid-villus but not upper villus cells. In contrast, phlorhizin-insensitive uptake by both cell populations was enhanced by diabetes. 3. Diabetes increased glucose absorption in vivo by mechanisms which were unrelated to hyperphagia. Mucosal hyperplasia acting together with an epithelium containing a higher proportion of mature enterocytes is the most likely explanation for the response. 4. We conclude that, during diabetes, the mid-villus region is an important site of adaptation with functional changes occurring at both the brush-border and basolateral membranes. The increased hexose transport ability of the basolateral membrane is retained during cell transit along the villus.

Reconstitution of an Insulin Signaling Pathway in Xenopus laevis Oocytes: Coexpression of a Mammalian Insulin Receptor and Three Different Mammalian Hexose Transporters

We report the functional expression of the mammalian muscle-adipocyte insulin-sensitive hexose transporter in Xenopus laevis oocytes. Oocytes microinjected with RNA synthesized in vitro showed enhanced hexose transport activity compared with uninjected controls. However, like the endogenous oocyte hexose transporter, activity was stimulated only twofold by 1 microM insulin. X. laevis oocytes injected with in vitro-synthesized RNA encoding the human insulin proreceptor expressed a functionally active insulin receptor that enhanced the insulin sensitivity of injected oocytes. This increase was not observed in oocytes expressing a mutant insulin receptor that lacked protein tyrosine kinase activity. In the presence of the coexpressed human insulin receptor, insulin induced a two- to threefold increase in hexose transport. The muscle-, brain-, and liver-type hexose carriers normally expressed in tissues with different responses to insulin exhibited the same insulin sensitivity when expressed in oocytes. This was observed whether or not the insulin signal was transduced through a coexpressed human insulin receptor or the endogenous oocyte insulin-like growth factor I receptor. We conclude that the expressed human insulin receptor is able to couple efficiently with preexisting postreceptor regulatory pathways in oocytes and that the regulation of hexose transport in these cells can be mediated through the combined actions of the expressed human insulin receptor and the endogenous oocyte insulin-like growth factor I receptor.

Reconstitution of an insulin signaling pathway in Xenopus laevis oocytes: coexpression of a mammalian insulin receptor and three different mammalian hexose transporters.

We report the functional expression of the mammalian muscle-adipocyte insulin-sensitive hexose transporter in Xenopus laevis oocytes. Oocytes microinjected with RNA synthesized in vitro showed enhanced hexose transport activity compared with uninjected controls. However, like the endogenous oocyte hexose transporter, activity was stimulated only twofold by 1 microM insulin. X. laevis oocytes injected with in vitro-synthesized RNA encoding the human insulin proreceptor expressed a functionally active insulin receptor that enhanced the insulin sensitivity of injected oocytes. This increase was not observed in oocytes expressing a mutant insulin receptor that lacked protein tyrosine kinase activity. In the presence of the coexpressed human insulin receptor, insulin induced a two- to threefold increase in hexose transport. The muscle-, brain-, and liver-type hexose carriers normally expressed in tissues with different responses to insulin exhibited the same insulin sensitivity when expressed in oocytes. This was observed whether or not the insulin signal was transduced through a coexpressed human insulin receptor or the endogenous oocyte insulin-like growth factor I receptor. We conclude that the expressed human insulin receptor is able to couple efficiently with preexisting postreceptor regulatory pathways in oocytes and that the regulation of hexose transport in these cells can be mediated through the combined actions of the expressed human insulin receptor and the endogenous oocyte insulin-like growth factor I receptor.

The Acquisition of Increased Insulin-responsive Hexose Transport in 3T3-L1 Adipocytes Correlates with Expression of a Novel Transporter Gene

The expression of two genes encoding facilitated glucose transporter proteins was studied during the differentiation of the 3T3-L1 fibroblastic cell line into adipocytes. The mRNA encoding the widely expressed HepG2/brain glucose transporter (GTI) is detectable in fibroblasts and its abundance remains unchanged during differentiation. On the other hand, the mRNA encoding a glucose transporter protein (GTIII) localized exclusively to muscle and adipose tissue is undetectable in fibroblasts but present in adipocytes. GTIII mRNA is first expressed three days after differentiation of 3T3-L1 cells has begun. Similarly, it is not until 3 days following the initiation of differentiation that GTIII protein can be detected, as assayed either by Western immunoblot or indirect immunofluorescence. The latter technique localizes GTIII predominantly to the perinuclear region of the adipocyte. The appearance of GTIII in developing fat cells correlates temporally with the acquisition of an increased stimulation of hexose uptake by maximal concentrations of insulin. These data support the concept that the marked increase in hexose transport in adipocytes in response to insulin is dependent on the expression in these cells of a specific, hormone-regulatable transport protein.

Common pathway for the induction of hexose transport by insulin and stress.

The effect of stress (heat shock, arsenite, or Semliki Forest virus [SFV] infection) on the induction of increased hexose transport has been compared with that of insulin. All four treatments increase the Vmax for transport by BHK cells three- to five-fold, with little effect (less than 40% decrease) on Km. Hydrogen peroxide and phenylarsine oxide (PAO) prevent the increase in hexose transport induced by stress treatments as effectively as they do that induced by insulin. Pinocytosis is not affected by any of the four treatments. On the other hand, the induction by insulin is sensitive to amiloride, whereas that by arsenite is not. Rat embryo fibroblasts, which respond poorly to insulin, respond well to arsenite, heat shock, or SFV infection. It is concluded that the stress response is mediated by certain compounds that may be common to those required for the action of insulin, but that those compounds act at a stage subsequent to the function of the insulin receptor.

Stress-induced increase of hexose transport as a novel index of cytopathic effects in virus-infected cells: Role of the L protein in the action of vesicular stomatitis virus

The VSV-specific increase in hexose transport by BHK cells has been measured by assay of the [ 3H]dGlc/[ 14C]AIB uptake ratio. The effect was abolished by uv-irradiation of the virus, indicating that viral gene expression is required. Cells infected with the T1026 R1 mutant of VSV, which causes only slight cytopathic changes, exhibited only a slight increase in hexose uptake. Cells infected with temperature-sensitive (ts) mutants of VSV that are defective in the function of the viral N, NS, G, or M proteins at the restrictive temperature (39.5°) exhibited increased [ 3H]dGLC/[ 14C]AIB uptake ratios typical of wild-type virus at either restrictive (39.5°) or permissive temperature (34°). Cells infected with a mutant defective in the function of the viral L protein exhibited an increased [ 3H]dGlc/[ 14C]AIB uptake ratio at permissive temperature (34°) only; at restrictive temperature (39.5°) the uptake ratio was essentially the same as that of mock-infected cells. Temperature-shift experiments indicated that the effect on hexose transport persisted for at least 6 hr in cells which no longer expressed function L protein, and that when expression of L was restricted to the first 2 hr of infection, an almost complete stimulation of hexose transport was observed 4 hr later. These results indicate that expression of the L gene is a necessary factor for inducing an increased hexose uptake in VSV-infected BHK cells. They also suggest that the action of the L protein on hexose transport is indirect, and is presumably mediated by other cellular constituents. The studies support the concept that an increased dGlc uptake may be a useful index of the cytopathic consequences of virus infection.

Effect of endotoxin-induced monokines on glucose metabolism in the muscle cell line L6.

Exposure of fully differentiated L6 myotubes to a crude monokine preparation from endotoxin-stimulated RAW 264.7 cells resulted in a rapid and substantial (70%) increase in fructose 2,6-bisphosphate concentration coincident with a depletion of cellular glycogen and an increased lactate production. During the time required for glycogen depletion (3 hr), stimulation of 3-O-methyl-D-glucose and 2-deoxy-D-glucose uptake was initiated and observed to reach a maximum enhancement of 200% 12-15 hr later. The monokine had no effect on the Km value for 2-deoxy-D-glucose uptake (1.1 mM), while Vmax was increased from 912 to 2400 pmol/min per mg of protein. The increase was cytochalasin B inhibitable and was dependent on protein synthesis. Photoaffinity labeling and equilibrium binding studies with [3H]cytochalasin B support the hypothesis that this increase in hexose transport was due to an increase in hexose transporters present in the plasma membrane. Purified recombinant interleukin-1 alpha had no effect on hexose transport, whereas purified recombinant cachetin/tumor necrosis factor did stimulate hexose uptake, with half-maximal stimulation occurring at 36 nM. Although cachetin accounts for most of the biological activity associated with the crude monokine preparations, it is not the only monokine capable of inducing glucose transport in L6 cells. Specific immunoabsorption of cachectin/tumor necrosis factor from the crude monokine preparation revealed a monokine that had a similar bioactivity at extremely low concentrations on L6 cells.

Possible involvement of a Na+/H+ antiporter in the stimulation of hexose transport in Swiss 3T3 cells by a phorbol ester and growth factors.

Phorbol-12,13-dibutyrate, epidermal growth factor, and insulin raised the intracellular pH ([pH]i), presumably through the activation of a Na+/H+ antiporter. Addition of amiloride or replacement of extra-cellular Na+ by choline which abolishes the cytoplasmic alkalinization prevented the stimulation of hexose transport by these agents. Furthermore, monensin, a Na+/H+ ionophore which increases the [pH]i, stimulated hexose transport. This stimulation was also prevented by the replacement of extra-cellular Na+ by choline. These observations suggest that stimulation of the Na+/H+ antiporter may have stimulated the increase in hexose transport.

Activation of hexose transport by antibody.

In our attempts to study alterations in cell behaviour by membrane perturbations, the effects of specific antisera on membrane functions were examined. The present communication reports the nature and kinetics of the changes in hexose transport as a result of treatment of L6 rat myoblast cells with specific antibodies. Exposure to complement-inactivated rabbit anti-myoblast antiserum resulted in an almost immediate two- to three-fold increase in hexose transport; morphological changes were observed only after prolonged incubation with the antiserum. Similar stimulation of hexose transport was observed upon exposure of cells to immunoglobulin G (IgG) from both sheep anti-myoblast and rabbit anti-myoblast plasma membrane sera. It seems that the interaction of specific IgG with cell surface components is sufficient to elicit this response. We have established that over 90% of 2-deoxyglucose is phosphorylated upon entrance into rat myoblasts and that exposure of cells to specific antibody does not cause changes in cell volume or general leakiness of the cell membrane. The antibody-stimulated hexose transport system resembles its normal counterpart in its substrate affinity and specificity; it differs from that of the control cells only in the transport capacity. This increase in hexose capacity is brought about by processes independent of protein synthesis and is not likely the result of extensive reorganization of membrane components.

Phosphotyrosine-containing proteins and expression of transformation parameters in cells infected with partial transformation mutants of Rous sarcoma virus.

We have examined the phosphorylation state of five proteins known to become phosphorylated on tyrosine during transformation by Rous sarcoma virus by using cells infected with a panel of partially transforming mutant viruses. Situations of viral mutant and growth temperature were found in which phosphorylation of some proteins occurred more extensively than that of others, indicating that mutations in the src gene had affected the specificity of pp60src for some of its substrates as well as affecting the activity of the enzyme. To obtain insight into the biological functions of these phosphorylations, comparisons were made between the degree of phosphorylation of these proteins and the expression of various indicators of the transformed phenotype. The data suggest that phosphorylation of proteins l, p, and q (Mr of 46,000, 39,000 and 28,000, respectively) is not sufficient to induce changes in adhesiveness, hexose transport or morphology. The phosphorylation of protein p or l or total phosphotyrosine content correlated well with the production of plasminogen activator, and the phosphorylation of proteins l and q correlated well with increased hexose transport. However, even when good correlations were observed, significant exceptions were sometimes noted. It thus remains possible that some phosphorylations on tyrosine observed in Rous sarcoma virus-transformed cells are not causally related to the expression of the measured parameters of transformation.

Increased hexose transport in Chinese hamster ovary cells resistant to 3-O-methyl-D-glucose.

3-O-Methyl-D-glucose-resistant mutants were selected from Chinese hamster ovary cells after mutagenesis with ethyl methanesulfonate. A mutant, MegR24, was isolated which was significantly more resistant than the parent to 3-O-methylglucose. Uptake of 50 microM D-glucose by metabolizing MegR24 cells was 2- to 3-fold higher than the parental cells in the absence or presence of 100 mM 3-O-methylglucose. A study of transport of D-[3H]glucose in ATP-depleted cells indicated an apparent Km for D-glucose transport that was 3-fold lower for the mutant (2.7 +/- 0.3 mM) than for the parent (8.9 +/- 1.0 mM). The apparent Km for transport of 3-O-methylglucose by the mutant (10.9 +/- 2.4 mM) was almost 2-fold lower than that of the parent (21.4 +/- 4.7 mM). The Vmax values for transport of D-glucose or 3-O-methylglucose by the mutant and parental cell lines were not significantly different. The MegR24 mutant also exhibited enhanced countertransport of D-[3H]glucose following preloading of ATP-depleted cells with 100 mM 3-O-methylglucose. Simple diffusion of hexoses as measured by L-glucose uptake was not altered in the mutant. These results suggest that the MegR24 hexose carrier has an increased affinity for transport of hexoses and that the resistance to the cytotoxic effects of 3-O-methylglucose exhibited by MegR24 is due to its ability to transport D-glucose 2- to 3-fold more efficiently than the parental strain.