Biochemical Society Transactions
Tissue-specific expression of facilitative glucose transporters: a rationale Helen M. Thomas, Alison M. Brant, Caroline A. Colville, Michael J. Seatter and Gwyn W. Gould* Department of Biochemistry, University of Glasgow, Glasgow G I 2 8QQ, Scotland, U.K. 538
Glucose is a major source of metabolic energy and virtually all animal cells possess a transport system for glucose of the facilitative diffusion type. l’hese transporters function to allow the movement of glucose down its chemical gradient either into or out of the cell. The application of modern molecular biological approaches to the study of glucose transport has led to the realization that rather than being mediated by a single transporter expressed in all cells glucose transport is, in fact, mediated by a family of transport proteins. These transporters are the products of distinct genes and exhibit considerable homology in their primary sequences but display ;i marked tissue-specific pattern of expression 1 1-4 1. The development and maintenance of this genetic diversity through evolution implies some underlying teleological requirement for a family of transporters, each with subtly different properties tailored t o the needs of the different tissues. As glucose transport in certain tissues is known t o be under both acute and chronic horrnonal regulation, it would seem reasonable t o propose that this tissue-specificity reflects the differing transport needs of these tissues. Summarized below are the salient features of each member of the transporter family, together u ith ;I rationale for the tissuespecific expression of this family of homologous proteins.
The glucose transporter family Several recent reviews have summarized the tissuespecific patterns of expression of the human glucose tr;insporter family I 1-4 I. ‘I’he data in Table 1 summarizes the current state of the literature. lintil very recently such results were based predominantly upon Northern blot analysis, and with the exception of (;I,IJTs 1 and 4. little was known about the level of the transporter proteins in these tissues. Recently, we have raised and characterized a panel of anti-peptide antibodies specific for each of the five human transporters and the rat GIAJT 2 and mouse GLIJT 3 isoforms. These antibodies have proven useful tools for the analysis of the transporter compliment of metabolically important tissues. Hecause a great deal has already been published Abbreviations used: GI,( ‘T?glucose transporter; 3 - 0 MG. 3- 0-methyl-i )-glucose. *Towhom correspondence should be addressed.
Volume 20
regarding (;l,IJl’s 1 and 4. here I will focus o n
GI.U’I’s 2, 3 and 5. Antibodies raised against the human isoform of GIAJT 2 specifically recognize this isotorm expressed in Xenopus oocytes, and d o not cross react with any of the other transporter tinily members. In immunoblot analysis the antibodies recognize proteins at 56. 50 and 33 kl)a. the 56 k1)a species resolving into a dinier 15 I. ‘I’his distinct pattern of cross-reacting proteins is also seen in human small intestine basolateral membranes 16 I. Moreover, similar patterns are observed in rat liver with the anti-rat GLUT 2 antibodies (56, -12 and 31 kDa). In rat islets the 56 kI)a form predominates (A. M. I h n t et al., unpublished work). ‘1’0 date ive have detected (;],I TI’ 2 in hepatocytes. pancreatic /3-cells and small intestine. Thorens ;ind colleagues have successfully used anti-peptide antibodies to localize GLLJT 2 t o the basolateral siirface of adsorptive epithelia in the small intestine and kidney [ 7, XI. From Korthern blot analysis o f human tissues (;IAJT 3 appears t o be ubiquitously expressed. ‘ h e predomiiwnt site o f expression is the brain, but it is also found at lower levels in fit, kidney, liver and muscle tissue. In i i i i effort t o further evaluate the role of (;I,( 1‘1’ 3 in these tissues, we raised anti-peptide antibodies specific for either the mouse or human isoforms of (;l,(U’ 3 15, 01. Using the anti-mouse (;l,l1‘I’ 3 ;intibody we have demonstrated that the expression of (;l,(Irl’ 3 is restricted t o brain and neural cell lines and is not immunologically detectable in highly purified mouse muscle. liver or fat membranes 10 1. ‘I’his raised the possibility that the (;l,IU’ 3 m l i N A detected in human muscle, fat ;ind liver may have been the result of neural contmiination of the tissue segments used in the KNA preparation. Indeed, we were unable to detect (;I,lJT 3 in purified ineiiibranes from human muscle or fat (1’. K. Shepherd, E. M. Gibbs. (;. W. Gould & 1% 13. Kahn. unpublished work) or in highly purified monkey fat. muscle or liver (S. McCoid & 11. M. Cibbs, personal communication). Thus it appears likely that (;I ,I 1‘1’ 3 expression is restricted mainly t o the brain and neural cells and is not ;is ubiquitously expressed as was first suggested. One exception to this conies \vith our recent observation that GLUT 3 protein is present in human small intestine adsorptive epithelia (C. A.
Membrane Transport
Table I
Major sites of expression of the different glucose transporters For reviews see [ 1-41,
539 lsoform
Tissue
Role
GLUT I
Placenta, brain, blood-tissue barrier. adipose and muscle tissue (low levels, tissue culture cells, transformed cells).
Basal glucose uptake in many cells. Kinetically asymmetric. Elevated levels in transformed cells and upon mitogen stimulation of cells in culture.
GLUT 2
Liver, pancreatic B-cell, kidney proximal tubule and small intestine (basolateral membranes).
High capacity, low affinity transporter. Important for glucose sensing in B-cells. Transepithelial glucose and fructose transport.
GLUT 3
Brain, nerve cells, small intestine
Neural tissue transporter. Role in small intestine unclear.
GLUT 4
Muscle, heart and adipose tissue.
Expressed only in tissues that exhibit acute insulin stimulated glucose transport. Translocates t o plasma membrane IYI response t o insulin.
GLUT 5
Small intestine (apical membranes), brain. muscle and adipose tissue at low levels.
Physiological role in glucose adsorption not clear. May have unusual substrate specificity?
Colville, (;. W. (;odd & S. Shirazi-Reechey, unpublished work). ‘Hie precise location o f this protein and its role in trmsepithelial glucose transport remains to be determined. Antibodies t o the human (;I,{JT 5 protein have been used t o localize the site o f expression of (;I ,LT7‘ 5 in human duodenum. Immunohistochinistry of mature rnterocytes demonstrateti that (;1,11T 5 is localized t o the luminel surfice of the cells. ‘I’his result has been confirmed using immunoblot analysis of human duodenal brushborder nienibranes I101 (G. W. (;odd & S. Schir;izi-lkechey, unpublished work). ‘I’he presence o f ;i fkditative glucose transporter on the adsorptive surface of the epithelia is some\vhat surprising, given that the tinrelated Na’-dependent glucose tramporter is h n v n to be the most important contributor to sugar uptake from the gut. Thus the role of(;I,1rl’ 5 in these cells remains t o be deterrniried. Interestingly, horthern blot analysis has shown that (;l,lJT 5 mKNA is present in adipose ;ind muscle tissue 1.3 I. We have confirmed this result using antibodies specific for (;I,ITT 5 with the protein shown t o be present in adipose tissue plasma membranes arid in highly purified muscle membr;ines (1’. K. Shepherd, E. (iibbs, C. Wesslau, G. W. (;odd b 13. I
Tissue-specific expression of facilitative glucose transporters: a rationale Helen M. Thomas, Alison M. Brant, Caroline A. Colville, Michael J. Seatter and Gwyn W. Gould* Department of Biochemistry, University of Glasgow, Glasgow G I 2 8QQ, Scotland, U.K. 538
Glucose is a major source of metabolic energy and virtually all animal cells possess a transport system for glucose of the facilitative diffusion type. l’hese transporters function to allow the movement of glucose down its chemical gradient either into or out of the cell. The application of modern molecular biological approaches to the study of glucose transport has led to the realization that rather than being mediated by a single transporter expressed in all cells glucose transport is, in fact, mediated by a family of transport proteins. These transporters are the products of distinct genes and exhibit considerable homology in their primary sequences but display ;i marked tissue-specific pattern of expression 1 1-4 1. The development and maintenance of this genetic diversity through evolution implies some underlying teleological requirement for a family of transporters, each with subtly different properties tailored t o the needs of the different tissues. As glucose transport in certain tissues is known t o be under both acute and chronic horrnonal regulation, it would seem reasonable t o propose that this tissue-specificity reflects the differing transport needs of these tissues. Summarized below are the salient features of each member of the transporter family, together u ith ;I rationale for the tissuespecific expression of this family of homologous proteins.
The glucose transporter family Several recent reviews have summarized the tissuespecific patterns of expression of the human glucose tr;insporter family I 1-4 I. ‘I’he data in Table 1 summarizes the current state of the literature. lintil very recently such results were based predominantly upon Northern blot analysis, and with the exception of (;I,IJTs 1 and 4. little was known about the level of the transporter proteins in these tissues. Recently, we have raised and characterized a panel of anti-peptide antibodies specific for each of the five human transporters and the rat GIAJT 2 and mouse GLIJT 3 isoforms. These antibodies have proven useful tools for the analysis of the transporter compliment of metabolically important tissues. Hecause a great deal has already been published Abbreviations used: GI,( ‘T?glucose transporter; 3 - 0 MG. 3- 0-methyl-i )-glucose. *Towhom correspondence should be addressed.
Volume 20
regarding (;l,IJl’s 1 and 4. here I will focus o n
GI.U’I’s 2, 3 and 5. Antibodies raised against the human isoform of GIAJT 2 specifically recognize this isotorm expressed in Xenopus oocytes, and d o not cross react with any of the other transporter tinily members. In immunoblot analysis the antibodies recognize proteins at 56. 50 and 33 kl)a. the 56 k1)a species resolving into a dinier 15 I. ‘I’his distinct pattern of cross-reacting proteins is also seen in human small intestine basolateral membranes 16 I. Moreover, similar patterns are observed in rat liver with the anti-rat GLUT 2 antibodies (56, -12 and 31 kDa). In rat islets the 56 kI)a form predominates (A. M. I h n t et al., unpublished work). ‘1’0 date ive have detected (;],I TI’ 2 in hepatocytes. pancreatic /3-cells and small intestine. Thorens ;ind colleagues have successfully used anti-peptide antibodies to localize GLLJT 2 t o the basolateral siirface of adsorptive epithelia in the small intestine and kidney [ 7, XI. From Korthern blot analysis o f human tissues (;IAJT 3 appears t o be ubiquitously expressed. ‘ h e predomiiwnt site o f expression is the brain, but it is also found at lower levels in fit, kidney, liver and muscle tissue. In i i i i effort t o further evaluate the role of (;I,( 1‘1’ 3 in these tissues, we raised anti-peptide antibodies specific for either the mouse or human isoforms of (;l,(U’ 3 15, 01. Using the anti-mouse (;l,l1‘I’ 3 ;intibody we have demonstrated that the expression of (;l,(Irl’ 3 is restricted t o brain and neural cell lines and is not immunologically detectable in highly purified mouse muscle. liver or fat membranes 10 1. ‘I’his raised the possibility that the (;l,IU’ 3 m l i N A detected in human muscle, fat ;ind liver may have been the result of neural contmiination of the tissue segments used in the KNA preparation. Indeed, we were unable to detect (;I,lJT 3 in purified ineiiibranes from human muscle or fat (1’. K. Shepherd, E. M. Gibbs. (;. W. Gould & 1% 13. Kahn. unpublished work) or in highly purified monkey fat. muscle or liver (S. McCoid & 11. M. Cibbs, personal communication). Thus it appears likely that (;I ,I 1‘1’ 3 expression is restricted mainly t o the brain and neural cells and is not ;is ubiquitously expressed as was first suggested. One exception to this conies \vith our recent observation that GLUT 3 protein is present in human small intestine adsorptive epithelia (C. A.
Membrane Transport
Table I
Major sites of expression of the different glucose transporters For reviews see [ 1-41,
539 lsoform
Tissue
Role
GLUT I
Placenta, brain, blood-tissue barrier. adipose and muscle tissue (low levels, tissue culture cells, transformed cells).
Basal glucose uptake in many cells. Kinetically asymmetric. Elevated levels in transformed cells and upon mitogen stimulation of cells in culture.
GLUT 2
Liver, pancreatic B-cell, kidney proximal tubule and small intestine (basolateral membranes).
High capacity, low affinity transporter. Important for glucose sensing in B-cells. Transepithelial glucose and fructose transport.
GLUT 3
Brain, nerve cells, small intestine
Neural tissue transporter. Role in small intestine unclear.
GLUT 4
Muscle, heart and adipose tissue.
Expressed only in tissues that exhibit acute insulin stimulated glucose transport. Translocates t o plasma membrane IYI response t o insulin.
GLUT 5
Small intestine (apical membranes), brain. muscle and adipose tissue at low levels.
Physiological role in glucose adsorption not clear. May have unusual substrate specificity?
Colville, (;. W. (;odd & S. Shirazi-Reechey, unpublished work). ‘Hie precise location o f this protein and its role in trmsepithelial glucose transport remains to be determined. Antibodies t o the human (;I,{JT 5 protein have been used t o localize the site o f expression of (;I ,LT7‘ 5 in human duodenum. Immunohistochinistry of mature rnterocytes demonstrateti that (;1,11T 5 is localized t o the luminel surfice of the cells. ‘I’his result has been confirmed using immunoblot analysis of human duodenal brushborder nienibranes I101 (G. W. (;odd & S. Schir;izi-lkechey, unpublished work). ‘I’he presence o f ;i fkditative glucose transporter on the adsorptive surface of the epithelia is some\vhat surprising, given that the tinrelated Na’-dependent glucose tramporter is h n v n to be the most important contributor to sugar uptake from the gut. Thus the role of(;I,1rl’ 5 in these cells remains t o be deterrniried. Interestingly, horthern blot analysis has shown that (;l,lJT 5 mKNA is present in adipose ;ind muscle tissue 1.3 I. We have confirmed this result using antibodies specific for (;I,ITT 5 with the protein shown t o be present in adipose tissue plasma membranes arid in highly purified muscle membr;ines (1’. K. Shepherd, E. (iibbs, C. Wesslau, G. W. (;odd b 13. I
