Evaluation of glucose transport and its regulation by insulin in human monocytes using flow cytometry.

BACKGROUND: We investigated the effects of insulin on glucose transport in human monocytes using flow cytometry, a method with several advantages over previously used techniques. We hypothesized that monocytes could be used as tools to study insulin action at the cellular level and facilitate the investigation of mechanisms that lead to insulin resistance. METHODS: Blood was withdrawn from 38 healthy subjects. The expression of glucose transporter (GLUT) isoforms in plasma membrane and the rates of glucose transport were determined with and without insulin (10 to 1,000 mU/L). Anti-CD14 phycoerythrin monoclonal antibody was used for monocyte gating. GLUT isoforms were determined after staining cells with specific antisera to GLUT1, GLUT3, and GLUT4. Glucose transport was monitored with 6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-6-deoxyglucose (NBDG). RESULTS: Insulin increased the uptake of NBDG (median effective dose 20 mU/L) and the expression of GLUT3 and GLUT4 isoforms in the plasma membrane (median effective doses 20 and 35 mU/L, respectively) but had no effect on GLUT1. Maximal effects were always reached at 100 mU/L of insulin. CONCLUSIONS: Monocytes may be a valid model system to study the effects of insulin on glucose transport. Further, flow cytometry is suitable for this investigation and can be used as an alternative to radiotracer methods.     

Current understanding of glucose transporter 4 expression and functional mechanisms

Glucose is used aerobically and anaerobically to generate energy for cells. Glucose transporters (GLUTs) are transmembrane proteins that transport glucose across the cell membrane. Insulin promotes glucose utilization in part through promoting glucose entry into the skeletal and adipose tissues. This has been thought to be achieved through insulin-induced GLUT4 translocation from intracellular compartments to the cell membrane, which increases the overall rate of glucose flux into a cell. The insulin-induced GLUT4 translocation has been investigated extensively. Recently, significant progress has been made in our understanding of GLUT4 expression and translocation. Here, we summarized the methods and reagents used to determine the expression levels of Slc2a4 mRNA and GLUT4 protein, and GLUT4 translocation in the skeletal muscle, adipose tissues, heart and brain. Overall, a variety of methods such real-time polymerase chain reaction, immunohistochemistry, fluorescence microscopy, fusion proteins, stable cell line and transgenic animals have been used to answer particular questions related to GLUT4 system and insulin action. It seems that insulin-induced GLUT4 translocation can be observed in the heart and brain in addition to the skeletal muscle and adipocytes. Hormones other than insulin can induce GLUT4 translocation. Clearly, more studies of GLUT4 are warranted in the future to advance of our understanding of glucose homeostasis.     

Subcellular localization and trafficking of the GLUT4 glucose transporter isoform in insulin-responsive cells

The rate-limiting step in the uptake and metabolism of Dglucose by insulin target cells is thought to be glucose transport mediated by glucose transporters (primarily the GLUT4 isoform) localized to the plasma membrane. However, subcellular fractionation, photolabelling and immunocytochemical studies have shown that the pool of GLUT4 present in the plasma membrane is only one of many subcellular pools of this protein. GLUT4 has been found in occluded vesicles at the plasma membrane, clathrin-coated pits and vesicles, early endosomes, and tubulo-vesicular structures; the latter are analogous to known specialized secretory compartments. Tracking the movement of GLUT4 through these compartments, and defining the mechanism and site of action of insulin in stimulating this subcellular trafficking, are major topics of current investigation. Recent evidence focuses attention on the exocytosis of GLUT4 as the major site of insulin action. Increased exocytosis may be due to decreased retention of glucose transporters in an intracellular pool, or possibly to increased assembly of a vesicle docking and fusion complex. Although details are unknown, the presence in GLUT4 vesicles of a synaptobrevin homologue leads us to propose that a process analogous to that occurring in synaptic vesicle trafficking is involved in the assembly of GLUT4 vesicles into a form suitable for fusion with the plasma membrane. Evidence that the pathways of signalling from the insulin receptor and of GLUT4 vesicle exocytosis may converge at the level of the key signalling enzyme, phosphatidylinositol 3-kinase, is discussed.     

GLUT4 translocation: the last 200 nanometers

Insulin regulates circulating glucose levels by suppressing hepatic glucose production and increasing glucose transport into muscle and adipose tissues. Defects in these processes are associated with elevated vascular glucose levels and can lead to increased risk for the development of Type 2 diabetes mellitus and its associated disease complications. At the cellular level, insulin stimulates glucose uptake by inducing the translocation of the glucose transporter 4 (GLUT4) from intracellular storage sites to the plasma membrane, where the transporter facilitates the diffusion of glucose into striated muscle and adipocytes. Although the immediate downstream molecules that function proximal to the activated insulin receptor have been relatively well-characterized, it remains unknown how the distal insulin-signaling cascade interfaces with and recruits GLUT4 to the cell surface. New biochemical assays and imaging techniques, however, have focused attention on the plasma membrane as a potential target of insulin action leading to GLUT4 translocation. Indeed, it now appears that insulin specifically regulates the docking and/or fusion of GLUT4-vesicles with the plasma membrane. Future work will focus on identifying the key insulin targets that regulate the GLUT4 docking/fusion processes.     

Cyclic AMP acutely stimulates translocation of the major insulin-regulatable glucose transporter GLUT4.

Facilitated glucose transport across plasma membranes is mediated by a family of transporters (GLUT1-GLUT5) that have different tissue distributions and Km values for transport. It has been shown that insulin stimulates glucose transport in fat and muscle tissues by causing the redistribution of one of these proteins (GLUT4) from inside the cell to the plasma membrane. Previous studies have shown that agents that change cAMP levels are able to modulate glucose transport in fat cells. The aim of this study was to investigate the mechanisms responsible for modulation of glucose transport by cAMP. 2-Deoxyglucose transport and insulin-regulatable glucose transporter (GLUT4) immunoreactivity in plasma and low density microsomal membranes were measured in adipocytes incubated for 30 min with insulin or dibutyryl-cAMP (Bt2cAMP). Low concentrations of Bt2cAMP (10 microM) increased 2-deoxyglucose uptake by translocating GLUT4 from low density microsomal membranes to the plasma membranes. Bt2cAMP at 1000 microM inhibited glucose transport below basal but further increased translocation of GLUT4. The effect of Bt2cAMP on translocation was additive to that of 7 nM insulin. We conclude that in rat adipocytes, Bt2cAMP acutely translocates GLUT4 but inhibits its activity to transport glucose.     

Gallic acid induces GLUT4 translocation and glucose uptake activity in 3T3-L1 cells

GLUT4, a 12 transmembrane protein, plays a major role in insulin mediated glucose transport in muscle and adipocytes. For glucose transport, the GLUT4 protein needs to be translocated to the plasma membrane from the intracellular pool and it is possible that certain compounds may be able to enhance this process. In the present work, we have shown that gallic acid can increase GLUT4 translocation and glucose uptake activity in an Akt-independent but wortmannin-sensitive manner. Further analysis suggested the role of atypical protein kinase Cζ/λ in gallic acid mediated GLUT4 translocation and glucose uptake.     

Role of protein kinase C in the regulation of glucose transport in the rat adipose cell. Translocation of glucose transporters without stimulation of glucose transport activity

The possible role of protein kinase C in the regulation of glucose transport in the rat adipose cell has been examined. Both insulin and phorbol 12-myristate 13-acetate (PMA) stimulate 3-O-methylglucose transport in the intact cell ein association with the subcellular redistribution of glucose transporters from the low density microsomes to the plasma membranes, as assessed by cytochalasin B binding. In addition, the actions of insulin and PMA on glucose transport activity and glucose transporter redistribution are additive. Furthermore, PMA accelerates insulin's stimulation of glucose transport activity, reducing the t1/2 from 3.2 +/- 0.4 to 2.1 +/- 0.2 min (mean +/- S.E.). However, the effect of PMA on glucose transport activity is approximately 10% of that for insulin whereas its effect on glucose transporter redistribution is approximately 50% of the insulin response. Immunoblots of the GLUT1 and GLUT4 glucose transporter isoforms in subcellular membrane fractions also demonstrate that the translocations of GLUT1 in response to PMA and insulin are of similar magnitude whereas the translocation of GLUT4 in response to insulin is markedly greater than that in response to PMA. Thus, glucose transport activity in the intact cell with PMA and insulin correlates more closely with the appearance of GLUT4 in the plasma membrane than cytochalasin B-assayable glucose transporters. Although these data do not clarify the potential role of protein kinase C in the mechanism of insulin action, they do suggest that the mechanisms through which insulin and PMA stimulate glucose transport are distinct but interactive.     

Regulated transport of the glucose transporter GLUT4

In muscle and fat cells, insulin stimulates the delivery of the glucose transporter GLUT4 from an intracellular location to the cell surface, where it facilitates the reduction of plasma glucose levels. Understanding the molecular mechanisms that mediate this translocation event involves integrating our knowledge of two fundamental processes--the signal transduction pathways that are triggered when insulin binds to its receptor and the membrane transport events that need to be modified to divert GLUT4 from intracellular storage to an active plasma membrane shuttle service.     

Insulin-stimulated glucose uptake involves the transition of glucose transporters to a caveolae-rich fraction within the plasma membrane: implications for type II diabetes.

BACKGROUND: Adipose and muscle tissues express an insulin-sensitive glucose transporter (GLUT4). This transporter has been shown to translocate from intracellular stores to the plasma membrane following insulin stimulation. The molecular mechanisms signalling this event and the details of the translocation pathway remain unknown. In type II diabetes, the cellular transport of glucose in response to insulin is impaired, partly explaining why blood-glucose levels in patients are not lowered by insulin as in normal individuals. MATERIALS AND METHODS: Isolated rat epididymal adipocytes were stimulated with insulin and subjected to subcellular fractionation and to measurement of glucose uptake. A caveolae-rich fraction was isolated from the plasma membranes after detergent solubilization and ultracentrifugal floatation in a sucrose gradient. Presence of GLUT4 and caveolin was determined by immunoblotting after SDS-PAGE. RESULTS: In freshly isolated adipocytes, insulin induced a rapid translocation of GLUT4 to the plasma membrane fraction, which was followed by a slower transition of the transporter into a detergent resistant caveolae-rich region of the plasma membrane. The insulin-stimulated appearance of transporters in the caveolae-rich fraction occurred in parallel with enhanced glucose uptake by cells. Treatment with isoproterenol plus adenosine deaminase rapidly inhibited insulin-stimulated glucose transport by 40%, and at the same time GLUT4 disappeared from the caveolae-rich fraction and from plasma membranes as a whole. CONCLUSIONS: Insulin stimulates glucose uptake in adipocytes by rapidly translocating GLUT4 from intracellular stores to the plasma membrane. This is followed by a slower transition of GLUT4 to the caveolae-rich regions of the plasma membrane, where glucose transport appears to take place. These results have implications for an understanding of the defect in glucose transport involved in type II diabetes.     

Regulation of glucose transporters by insulin and extracellular glucose in C2C12 myotubes

It is well established that insulin stimulation of glucose uptake in skeletal muscle cells is mediated through translocation of GLUT4 from intracellular storage sites to the cell surface. However, the established skeletal muscle cell lines, with the exception of L6 myocytes, reportedly show minimal insulin-dependent glucose uptake and GLUT4 translocation. Using C(2)C(12) myocytes expressing exofacial-Myc-GLUT4-enhanced cyan fluorescent protein, we herein show that differentiated C(2)C(12) myotubes are equipped with basic GLUT4 translocation machinery that can be activated by insulin stimulation ( approximately 3-fold increase as assessed by anti-Myc antibody uptake and immunostaining assay). However, this insulin stimulation of GLUT4 translocation was difficult to demonstrate with a conventional 2-deoxyglucose uptake assay because of markedly elevated basal glucose uptake via other glucose transporter(s). Intriguingly, the basal glucose transport activity in C(2)C(12) myotubes appeared to be acutely suppressed within 5 min by preincubation with a pathophysiologically high level of extracellular glucose (25 mM). In contrast, this activity was augmented by acute glucose deprivation via an unidentified mechanism that is independent of GLUT4 translocation but is dependent on phosphatidylinositol 3-kinase activity. Taken together, these findings indicate that regulation of the facilitative glucose transport system in differentiated C(2)C(12) myotubes can be achieved through surprisingly acute glucose-dependent modulation of the activity of glucose transporter(s), which apparently contributes to obscuring the insulin augmentation of glucose uptake elicited by GLUT4 translocation. We herein also describe several methods of monitoring insulin-dependent glucose uptake in C(2)C(12) myotubes and propose this cell line to be a useful model for analyzing GLUT4 translocation in skeletal muscle.     

The glucose transporter 4-regulating protein TUG is essential for highly insulin-responsive glucose uptake in 3T3-L1 adipocytes

Insulin stimulates glucose uptake in fat and muscle by redistributing GLUT4 glucose transporters from intracellular membranes to the cell surface. We previously proposed that, in 3T3-L1 adipocytes, TUG retains GLUT4 within unstimulated cells and insulin mobilizes this retained GLUT4 by stimulating its dissociation from TUG. Yet the relative importance of this action in the overall control of glucose uptake remains uncertain. Here we report that transient, small interfering RNA-mediated depletion of TUG causes GLUT4 translocation and enhances glucose uptake in unstimulated 3T3-L1 adipocytes, similar to insulin. Stable TUG depletion or expression of a dominant negative fragment likewise stimulates GLUT4 redistribution and glucose uptake, and insulin causes a 2-fold further increase. Microscopy shows that TUG governs the accumulation of GLUT4 in perinuclear membranes distinct from endosomes and indicates that it is this pool of GLUT4 that is mobilized by TUG disruption. Interestingly, in addition to translocating GLUT4 and enhancing glucose uptake, TUG disruption appears to accelerate the degradation of GLUT4 in lysosomes. Finally, we find that TUG binds directly and specifically to a large intracellular loop in GLUT4. Together, these findings demonstrate that TUG is required to retain GLUT4 intracellularly in 3T3-L1 adipocytes in the absence of insulin and further implicate the insulin-stimulated dissociation of TUG and GLUT4 as an important action by which insulin stimulates glucose uptake.     

Identification of a Distal GLUT4 Trafficking Event Controlled by Actin Polymerization

The insulin-stimulated trafficking of GLUT4 to the plasma membrane in muscle and fat tissue constitutes a central process in blood glucose homeostasis. The tethering, docking, and fusion of GLUT4 vesicles with the plasma membrane (PM) represent the most distal steps in this pathway and have been recently shown to be key targets of insulin action. However, it remains unclear how insulin influences these processes to promote the insertion of the glucose transporter into the PM. In this study we have identified a previously uncharacterized role for cortical actin in the distal trafficking of GLUT4. Using high-frequency total internal reflection fluorescence microscopy (TIRFM) imaging, we show that insulin increases actin polymerization near the PM and that disruption of this process inhibited GLUT4 exocytosis. Using TIRFM in combination with probes that could distinguish between vesicle transport and fusion, we found that defective actin remodeling was accompanied by normal insulin-regulated accumulation of GLUT4 vesicles close to the PM, but the final exocytotic fusion step was impaired. These data clearly resolve multiple steps of the final stages of GLUT4 trafficking, demonstrating a crucial role for actin in the final stage of this process.     

Effect of insulin and contraction up on glucose transport in skeletal muscle

The major glucose transporter protein expressed in skeletal muscle is GLUT4. Both muscle contraction and insulin induce translocation of GLUT4 from the intracellular pool to the plasma membrane. The intracellular pathways that lead to contraction- and insulin-stimulated GLUT4 translocation seem to be different, allowing the attainment of a maximal effect when acting together. Insulin utilizes a phosphatidylinositol 3-kinase-dependent mechanism, whereas the exercise signal may be initiated by calcium release from the sarcoplasmic reticulum or from autocrine- or paracrine-mediated activation of glucose transport. During exercise skeletal muscle utilizes more glucose than when at rest. However, endurance training leads to decreased glucose utilization during sub-maximal exercise, in spite of a large increase in the total GLUT4 content associated with training. The mechanisms involved in this reduction have not been totally elucidated, but appear to cause the decrease of the amount of GLUT4 translocated to the plasma membrane by altering the exercise-induced enhancement of glucose transport capacity. On the other hand, the effect of resistance training is controversial. Recent studies, however, demonstrated the improvement in insulin sensitivity correlated with increasing muscle mass. New studies should be designed to define the molecular basis for these important adaptations to skeletal muscle. Since during exercise the muscle may utilize insulin-independent mechanisms to increase glucose uptake, the mechanisms involved should provide important knowledge to the understanding and managing peripheral insulin resistance.     

Elucidation of the glucose transport pathway in glucose transporter 4 via steered molecular dynamics simulations

Background

GLUT4 is a predominant insulin regulated glucose transporter expressed in major glucose disposal tissues such as adipocytes and muscles. Under the unstimulated state, GLUT4 resides within intracellular vesicles. Various stimuli such as insulin translocate this protein to the plasma membrane for glucose transport. In the absence of a crystal structure for GLUT4, very little is known about the mechanism of glucose transport by this protein. Earlier we proposed a homology model for GLUT4 and performed a conventional molecular dynamics study revealing the conformational rearrangements during glucose and ATP binding. However, this study could not explain the transport of glucose through the permeation tunnel.

Methodology/Principal Findings

To elucidate the molecular mechanism of glucose transport and its energetic, a steered molecular dynamics study (SMD) was used. Glucose was pulled from the extracellular end of GLUT4 to the cytoplasm along the pathway using constant velocity pulling method. We identified several key residues within the tunnel that interact directly with either the backbone ring or the hydroxyl groups of glucose. A rotation of glucose molecule was seen near the sugar binding site facilitating the sugar recognition process at the QLS binding site.

Conclusions/Significance

This study proposes a possible glucose transport pathway and aids the identification of several residues that make direct interactions with glucose during glucose transport. Mutational studies are required to further validate the observation made in this study.     

Chromium activates glucose transporter 4 trafficking and enhances insulin-stimulated glucose transport in 3T3-L1 adipocytes via a cholesterol-dependent mechanism

Evidence suggests that chromium supplementation may alleviate symptoms associated with diabetes, such as high blood glucose and lipid abnormalities, yet a molecular mechanism remains unclear. Here, we report that trivalent chromium in the chloride (CrCl3) or picolinate (CrPic) salt forms mobilize the glucose transporter, GLUT4, to the plasma membrane in 3T3-L1 adipocytes. Concomitant with an increase in GLUT4 at the plasma membrane, insulin-stimulated glucose transport was enhanced by chromium treatment. In contrast, the chromium-mobilized pool of transporters was not active in the absence of insulin. Microscopic analysis of an exofacially Myc-tagged enhanced green fluorescent protein-GLUT4 construct revealed that the chromium-induced accumulation of GLUT4-containing vesicles occurred adjacent to the inner cell surface membrane. With insulin these transporters physically incorporated into the plasma membrane. Regulation of GLUT4 translocation by chromium did not involve known insulin signaling proteins such as the insulin receptor, insulin receptor substrate-1, phosphatidylinositol 3-kinase, and Akt. Consistent with a reported effect of chromium on increasing membrane fluidity, we found that chromium treatment decreased plasma membrane cholesterol. Interestingly, cholesterol add-back to the plasma membrane prevented the beneficial effect of chromium on both GLUT4 mobilization and insulin-stimulated glucose transport. Furthermore, chromium action was absent in methyl-beta-cyclodextrin-pretreated cells already displaying reduced plasma membrane cholesterol and increased GLUT4 translocation. Together, these data reveal a novel mechanism by which chromium may enhance GLUT4 trafficking and insulin-stimulated glucose transport. Moreover, these findings at the level of the cell are consistent with in vivo observations of improved glucose tolerance and decreased circulating cholesterol levels after chromium supplementation.     

Cell surface labeling of glucose transporter isoform GLUT4 by bis-mannose photolabel. Correlation with stimulation of glucose transport in rat adipose cells by insulin and phorbol ester

A new impermeant photoaffinity label has been used for identifying cell surface glucose transporters in isolated rat adipose cells. This compound is 2-N-4(1-azi-2,2,2-trifluoroethyl)benzoyl-1,3-bis(D-mannos-4- yloxy)-2- propylamine. We have used this reagent in combination with immunoprecipitation by specific antibodies against the GLUT4 and GLUT1 glucose transporter isoforms to estimate the relative abundance of these two transporters on the surface of the intact adipose cell following stimulation by insulin and phorbol 12-myristate 13-acetate (PMA). In the basal state, GLUT4 and GLUT1 are both present at the cell surface but GLUT4 is more abundant than GLUT1. In response to insulin, GLUT4 increases 15-20-fold and GLUT1 increases approximately 5-fold while 3-O-methyl-D-glucose transport is stimulated 20-30-fold. By contrast, PMA only induces a approximately 4-fold increase in GLUT4 while GLUT1 increases approximately 5-fold to the same level as seen with insulin. In addition, PMA stimulates 3-O-methyl-D-glucose transport approximately 3-fold to only 13% of the insulin-stimulated state. Thus GLUT4 is the major glucose transporter isoform under all conditions, and it is selectively and markedly enriched in response to insulin but not PMA which increases GLUT1 and GLUT4 equally. Furthermore, stimulation of glucose transport activity correlates closely with the appearance of GLUT4 on the cell surface in response to both insulin and PMA but does not correlate with the sum of GLUT1 and GLUT4 appearance. These results suggest that GLUT4 may be inherently more active than GLUT1 due to a higher TK (turnover/Km).     

Identification of a Role for CLASP2 in Insulin Action

Insulin stimulates the mobilization of glucose transporter 4 (GLUT4) storage vesicles to the plasma membrane, resulting in an influx of glucose into target tissues such as muscle and fat. We present evidence that CLIP-associating protein 2 (CLASP2), a protein previously unassociated with insulin action, is responsive to insulin stimulation. Using mass spectrometry-based protein identification combined with phosphoantibody immunoprecipitation in L6 myotubes, we detected a 4.8-fold increase of CLASP2 in the anti-phosphoserine immunoprecipitates upon insulin stimulation. Western blotting of CLASP2 immunoprecipitates with the phosphoantibody confirmed the finding that CLASP2 undergoes insulin-stimulated phosphorylation, and a number of novel phosphorylation sites were identified. Confocal imaging of L6 myotubes revealed that CLASP2 colocalizes with GLUT4 at the plasma membrane within areas of insulin-mediated cortical actin remodeling. CLASP2 is responsible for directing the distal end of microtubules to the cell cortex, and it has been shown that GLUT4 travels along microtubule tracks. In support of the concept that CLASP2 plays a role in the trafficking of GLUT4 at the cell periphery, CLASP2 knockdown by siRNA in L6 myotubes interfered with insulin-stimulated GLUT4 localization to the plasma membrane. Furthermore, siRNA mediated knockdown of CLASP2 in 3T3-L1 adipocytes inhibited insulin-stimulated glucose transport. We therefore propose a new model for CLASP2 in insulin action, where CLASP2 directs the delivery of GLUT4 to cell cortex landing zones important for insulin action.     

Topology Mapping of Insulin-Regulated Glucose Transporter GLUT4 Using Computational Biology

The type 2 diabetes is increasing rapidly around the globe. The primary cause for this is insulin resistance due to the disruption of the insulin signal transduction mechanism. Insulin signal transduction stimulates glucose transport through the glucose transporter GLUT4, by promoting the exocytosis process. Understanding the structural topology of GLUT4 mechanism will increase our understanding of the dynamic activities about glucose transport and its regulation in the membrane environment. However, little is known about the topology of GLUT4. In this article, we have determined the amino acid composition, disulfide topology, structure conformation pattern of GLUT4. The amino acid composition portrays that leucine composition is the highest contributing to 15.5 % among all other amino acids. Three cysteine residues such as Cys223, Cys361, and Cys363 were observed and the last two were associated with one disulfide bond formation. We have generated surface cavities to know the clefts/pockets on the surface of this protein that showed few irregular cavities placed mostly in the transmembrane-helical part. Besides, topology mapping of 12 transmembrane-helixes was done to predict N- and O-glycosylation sites and to show the highly glycosylated GLUT4 that includes both N- and O-glycosylation sites. Furthermore, hydrophobic segment and molecular charge distribution were analyzed. This article shows that bioinformatics tools can provide a rapid methodology to predict the topology of GLUT4. It also provides insights into the structural details and structural functioning relationships in the human GLUT4. The results can be of great help to advance future drug development research using GLUT4 as a target protein.     

GLUT4 trafficking in insulin-sensitive cells

In recent years, there have been major advances in the under-standing of both the cell biology of vesicle trafficking between intracellular compartments and the molecular targeting signals intrinsic to the trafficking proteins themselves. One system to which these advances have been profitably applied is the regulation of the trafficking of a glucose transporter, GLUT4, from intracellular compartment(s) to the cell surface in response to insulin. The unique nature of the trafficking of GLUT4 and its expression in highly differentiated cells makes this a question of considerable interest to cell biologists. Unraveling the tangled web of molecular events coordinating GLUT4 trafficking will eventually lead to a greater understanding of mammalian glucose metabolism, as well as fundamental cell biological principles related to organelle biogenesis and protein trafficking.     

Glucose transporters: structure, function, and regulation

Glucose is transported into the cell by facilitated diffusion via a family of structurally related proteins, whose expression is tissue-specific. One of these transporters, GLUT4, is expressed specifically in insulin-sensitive tissues. A possible change in the synthesis and/or in the amount of GLUT4 has therefore been studied in situations associated with an increase or a decrease in the effect of insulin on glucose transport. Chronic hyperinsulinemia in rats produces a hyper-response of white adipose tissue to insulin and resistance in skeletal muscle. The hyper-response of white adipose tissue is associated with an increase in GLUT4 mRNA and protein. In contrast, in skeletal muscle, a decrease in GLUT4 mRNA and a decrease (tibialis) or no change (diaphragm) in GLUT4 protein are measured, suggesting a divergent regulation by insulin of glucose transport and transporters in the 2 tissues. In rodents, brown adipose tissue is very sensitive to insulin. The response of this tissue to insulin is decreased in obese insulin-resistant fa/fa rats. Treatment with a beta-adrenergic agonist increases insulin-stimulated glucose transport, GLUT4 protein and mRNA. The data suggest that transporter synthesis can be modulated in vivo by insulin (muscle, white adipose tissue) or by catecholamines (brown adipose tissue).