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SORTING OF GABA TRANSPORTER PROTEINS IN POLARIZED CELLS
Pergamon
0197-0186(95)O0159-X
Neurochem. Int, Vol. 29, No. 4, pp. 357-360, 1996 Copyright 01996 Elsevier Science Ltd Printed in Great Britain. All rights reserved 01974186/96 $15.00+0.00
CRITIQUE SORTING OF GABA TRANSPORTER PROTEINS IN POLARIZED CELLS MICHAEL J. CAPLAN Department of Cellular and Molecular Physiology,Yale UniversitySchoolof Medicine, 333Cedar Street, New Haven, CT 06510,U.S.A.
The past several years have witnessed an unprecedented explosion in our knowledge of the cellular machinery which controls communication across the neuronal synapse. Many of the proteins which initiate and regulate the fusion of synaptic vesicles with the pre-synaptic plasma membrane have been cloned and characterized (Ferro-Novick and Jahn, 1994). The complex assemblies in which these polypeptides participate have been dissected and reconstituted, producing a remarkably rich picture of the chain of events which couples the arrival of an action potential to synaptic vesicle discharge (Sudhof, 1995). In order to develop a complete understanding of the critically important process of synaptic transmission, however, it is not sufficient to elucidate the molecular mechanisms which contribute to the delivery of neurotransmitters to the synaptic space. It is equally important to chart the cellular pathways which remove neurotransmitters from the synaptic space, resulting in the termination of synaptic communication. All of the elegant specializations which permit extremely rapid and exquisitely modulated synaptic release would be useless if there were not similarly sophisticated mechanisms available to prevent neurotransmitters from accumulating in the vicinity of their receptors. In those pharmacologically or pathologically induced settings in which neurotransmitters are allowed to persist within the synaptic space, the temporal and spatial accuracy upon which effective synaptic transmission depends cannot be maintained (Iversenj 1971; Ritz and Kuhar, 1993). The cellular components which terminate synaptic transmission have, perhaps, received somewhat less attention than
they deserve in light of the current euphoria over the unraveling of the exocytotic machinery. This state of affairs is especially unfortunate, since research into the machinery responsible for neurotransmitter removal from the synapse has recently undergone its own renaissance. Most neurons in the central nervous system terminate synaptic transmission by importing their respective neurotransmitter from the synaptic space back into their axon terminus. This retrieval process makes use of secondary active transport systems, which exploit the energy stored in trans-plasmalemmal ion gradients to drive the rapid uptake of neurotransmitters from the synaptic space into the pre-synaptic cytoplasm (Kanner, 1994). The recovered neurotransmitter can then be re-packaged into synaptic vesicles and employed in subsequent rounds of neuronal communication. A great deal of research has defined many of the physiological properties of these transporters and established their importance as the primary pharmacological targets for a number of therapeutic and abused substances (Schloss et al., 1992). Biochemical studies have revealed that each neurotransmitter is retrieved by a distinct class of transport proteins, which manifest unique ionic requirements and inhibitor sensitivities. In the past 4 years the genes encoding the transport systems specific for a number of different neurotransmitters have been cloned and their molecular structures revealed (Amara and Arriza, 1993). These genes all belong to a large family, within which a number of subfamilies can be defined. The preceding review by Laurence Borden provides 357
358
Critique
an excellent and remarkably thorough summary of our current understanding of one of these subfamilies. At least four genes encode proteins which are capable of transporting the neurotransmitter GABA (Guastella et al., 1990; Borden et al., 1992; Yamauchi et al., 1992). To some extent, this multiplicity illustrates the embarrassment of riches that can accompany the successful application of molecular techniques to problems which were previously susceptible only to pharmacological or biochemical analysis. While pharmacological studies had subdivided GABA transporters into two sub-classes (“neuronal” and “glial”) based on their inhibitor sensitivities, cloning revealed the existence of much greater diversity. Furthermore, correlating the “glial” transport activity with the behaviors of any of the cloned transporters expressed by transection in heterologous systems has proven difficult (see Dr Borden’s review for a detailed discussion of this issue). Thus, in a sense, the wealth of information provided by molecular biological analysis has created as many problems as it has resolved. How does the number and variety of newly-identified GABA transporter proteins correspond to the GABA transport activities measured in actual tissues? Furthermore, why has evolution gone to the trouble of creating this molecular diversity? Dr Borden has explored, in great depth, the physiological properties of the newly cloned transporters and correlated these behaviors with the functional characteristics and expression patterns of GABA transporters in situ. It is clear from his discussion that the relationship between the cloned transporters and the identified activities is not simple. It also appears likely that the number and diversity of GABA transport proteins may have arisen in order to satisfy biological requirements which are not reflected in substrate affinities or inhibitor sensitivities. Dr Borden suggests the possibility that different members of the GABA transporter family maybe targeted to distinct subcellular localization. He further hypothesizes that the sequence differences among the GABA transporter isoforms may account for their differential distributions. Results from recent experiments in which GABA transporter isoforms were exogenously expressed in epithelial cells and neurons suggest that these propositions may, in fact, be correct. The cell surface membranes of polarized epithelial cells are divided into apical and basolateral domains which are separated from one another by tight junctions. These two regions of the plasmalemma confront distinct body compartments, subserve different physiological functions and are endowed with distinct sets of integral membrane proteins (Caplan and
Matlin, 1989; Rodriguez-Boulan and Nelson, 1989). We have expressed the cDNAs encoding the GAT-1, GAT-2, GAT-3 and BGT-1 proteins by transection in the polarized MDCK epithelial cell line, which derives from and resembles the canine renal tubule. We find that the GAT-1 and GAT-3 polypeptides are restricted to the apical surface of the transected cells (Pietrini et al., 1994; Ahn et al., in press). This distribution is confirmed by immunofluorescence analysis as well as by transport assays and biochemical surface labelling techniques. In contrast, GAT-2 and BGT-1 are restricted to the basolateral plasma membrane domains when expressed in MDCK cells (Pietrini et al., 1994; Ahn et al., in press). Once again, immunofluorescence, transport assays and surface labelling methods support this conclusion. As noted in Dr Borden’s review, the sorting of BGT1 to the basolateral surfaces of transected MDCK cells is not surprising, since physiological studies had previously demonstrated the presence of the BGT-1 transport activity in the MDCK basolateral plasmaIemma (Yamauchi et al., 1991). BGT-I appears to be involved in importing betaine into cells from the extracellular fluid compartment in order to assist renal epithelial cells in adapting to osmotic stress (Nakanishi et al., 1990). In order to serve in this function, BGT-1 must, therefore, be exposed to the extracellular fluid compartment and, consequently, must be sorted to the basolateral membrane. The targeting of the three other GAT isoforms was not as easy to predict from basic principles. Immunocytochemical studies have demonstrated that GAT-1 is restricted to the pre-synaptic membranes of axon termini in the GABA-ergic neurons which express it (Radian et al., 1990). Recent studies on the mechanisms of neuronal membrane protein sorting suggest that the signals and machinery involved in sorting proteins to axons may be similar or analogous to ‘the signals and machinery which participates in apical targeting in epithelial cells (Dotti and Simons, 1990; Dotti et al., 1991). Similarly, sorting to the somatodendritic surfaces of neurons appears to be mechanistically related to the process of basolateral targeting in epithelial cells. Our observation that GAT-1 is apically sorted in epithelial cells lends support to this hypothesis. Further support for the model relating epithelial and neuronal sorting mechanisms can be found in the results of experiments in which we expressed the cDNAs encoding GAT-3 and BGT-1 in polarized hippocampal neurons in culture (Ahn et al., in press). We developed a protocol in which thecDNAs of interest were microinjected directly into the nuclei of
——
..
.—..———.. .
Critique cultured post-mitotic neurons derived from the embryonic rat hippocampus. Immunfluorescence analysis revealed that while the expressed BGT-1 protein was restricted to somatodendritic regions, GAT-3 could gain access to axons. It would appear, therefore, that at least with respect to the members of the GAT transporter family, there is a close correlation between neuronal and epithelial sorting behaviors. It is important to note, however, that studies of the sorting behaviors of other neurotransmitter re-uptake systems suggest that this model may not be generally applicable. Recent experiments performed by the laboratory of Dr Gary Rudnick at Yale University, in collaboration with our own, suggest that the strict correlation between axonal and apical sorting may not apply as simply to the dopamine, serotonin and norepinepherine transporters. Only a single isoform has been identified for both the norepinepherine and serotonin transporters. Furthermore, the functional roles of both transporters require that they be present in the axonal plasmalemma. When expressed in MDCK cells, however, both of these proteins are sorted exclusively to the basolateral surface. It would appear, therefore, that the relationship between epithelial and neuronal sorting is less straightforward than had originally been assumed. Clearly, a great deal of research remains to be done in order to understand more completely the cell-type specific specializations which ensure that membrane protein targeting pathways meet the physiological requirements of each individual polarized tissue. Dr Borden’s review points out another interesting correlation relevant to the sorting behavior of the GAT transporters. The GAT-1 and GAT-3 proteins, which are apically sorted in transected epithelia, appear to be expressed exclusively in neurons in situ. In contrast, BGT-1 and GAT-2, which are normally expressed in epithelia as well as in cells of the nervous system, behave as basolateral proteins in transected MDCK cells. As noted above, BGT-1 requires access to the basolateral surface in order to carry out its physiological function in renal epithelial cells. It is quite possible that GAT-2 serves in a similar or related capacity in the epithelial cells in which it is expressed. If GAT-2 imports GABA or a related substance from the extracellular fluid compartment into epithelial cells, it is logical to predict that it, too, would need to enjoy a basolateral localization. It will be extremely interesting to monitor the results of efforts to immunolocalize GAT-3 and GAT-2 in situ in order to determine if these predictions are borne out. Finally, it should be noted that the distinct sorting behaviors exhibited by the GAT isoforms must reflect
359
the presence of distinct sorting information embedded within the structures of these proteins (Caplan and Matlin, 1989). Presumably, GAT-1 and GAT-3 encode apical sorting signals, whereas GAT-2 and BGT-1 possess sorting signals which specify their basolateral distribution. The high degree of homology which relates the GAT family members to one another provides us with a valuable system with which to explore the nature and composition of these sorting signals. We have begun to generate molecular constructs in which portions of GAT transporters are removed or replaced by corresponding portions of oppositely-sorted isoforms. While still quite preliminary, this analysis suggests that both the cytosolic amino and carboxyl termini of these proteins may contribute to the presentation of these proteins’ sorting signals. Hopefully, further chimera and deletion studies will allow us to identify narrowly defined sequence domains which account for the GAT transporters’ differential sorting behaviors in both epithelial cells and in neurons. Elucidating these signals may further our understanding of the information transfer processes involved in membrane protein sorting and help us to identify the cellular apparatus which is responsible. Insight into the nature of these signals may also shed light on the evolutionary need for the multiplicity of GABA transporters and suggest their individual biological roles.
REFERENCES
Ahn J., Mundigl O., Muth R. R., Rudnick G. and Caplan M. J. (1996)Polarized expressionof GABA transporters in MDCK cells and cultured hippocampal neurons. .J. Bid. Chem.In press. Amara S. G. and Arriza J. L. (1993)Neurotransmitter transporters: three distinct gene families. Curr. Opin. Neurol. 337-344. Borden L. A., Smith K. E., Hartig P. R., Branchek T. A. and Weinshank R. L. (1992)Molecular heterogeneityof they-aminobutyricacid (GABA)transport system.J. BioL Chem.267,21,098–21,104. Caplan M. J. and Matlin K. S. (1989)Sorting of membrane and secretoryproteinsin polarizedepithelialcells.In FunctionalEpithelialCellsin Culture(Matlin K. S. and Valentich J. D., eds.), pp. 71–127.Alan R. Liss, New York. Dotti C. G., Parton R. G. and SimonsK. (1991)Polarized sorting of glypiated proteins in hippocampal neurons. Nature 349, 158–161. Dotti C. G. and SimonsK. (1990)Polarized sorting of viral glycoproteinsto the axon and dendrites of hippocampal neurons in culture. Cell62, 63–72. Ferro-Novick S. and Jahn R. (1994) Vesicle fusion from yeast to man. Nature370, 191-193. Guastella J., Nelson N., Nelson H., CzyzykL., Kenyan S.,
360
Critique
Miedel M., Davidson N., Lester H. and Kanner B. I. (1990)Cloningand expressionofa rat brain GABAtransporter. Science249, 1303-1306. IversenL. L. (1971)Role of transmitter uptake mechanisms in synaptic neurotransmission. Br. J. Pharmac.41, 571– 591, Kanner B. L (1994)Sodium-coupledneurotransmitter transport: structure, functionand regulation.J. Exp. Biol. 196, 237-249. Nakanishi T., Turner R. J. and Burg M. B. (1990)Osmoregulation of betaine transporter in mammalian renal medullarycells.Am.1 Physiol.258,F1061–F1O67. Pietrini G., SuhY. J., EdelmannL., RudnickG. and Caplan M. J. (1994)The axonal y-aminobutyricacid transporter GAT-I is sorted to the apical membranes of polarized epithelialcells.J. Biol. Chem.269,4668&4674. Radian R., Ottersen O. P., Storm-Mathisen J., Castel M. and Kanner B.I. (1990)Immunocytochemicallocalization of the GABA transporter in rat brain. J. Neurosci. 10, 1319-1330.
Ritz M. C. and Kuhar M. J. (1993)Psychostimulantdrugs and a dopamine hypothesis regarding addiction: update on recent research. Biochem.Soc. Symp. 59, 51L64. Rodriguez-BoulanE. and Nelson W. J. (1989) Morphogenesisof the polarized epithelial cell phenotype. Science 245,718-725. SchlossP., Mayser W. and BetzH. (1992)Neurotransmitter transporters: a novelfamilyof integral plasma membrane proteins. REBS Lett. 307,7G80. SudhofT. C. (1995)The synapticvesiclecycle: a cascade of protein-proteininteractions. Nature 375,645+53. Yamauchi A., Kwon H. M., Uchida S., Preston A. and HandlerJ. S. (1991)Myo-inositoland betainetransporters regulated by tonicity are basolateral in MDCK cells.Am. J. Physiol.261,F197-F202. YamauchiA., Uchida S., KwonH. M., Preston A. S., Robey R. B., Garcia-Perez A., Burg M. B. and Handler J. S. (1992)Cloning of a Na and Cl-dependentbetaine transporter that is regulated by hypertonicity. J. BioL Chem. 267,649%652.
0197-0186(95)O0159-X
Neurochem. Int, Vol. 29, No. 4, pp. 357-360, 1996 Copyright 01996 Elsevier Science Ltd Printed in Great Britain. All rights reserved 01974186/96 $15.00+0.00
CRITIQUE SORTING OF GABA TRANSPORTER PROTEINS IN POLARIZED CELLS MICHAEL J. CAPLAN Department of Cellular and Molecular Physiology,Yale UniversitySchoolof Medicine, 333Cedar Street, New Haven, CT 06510,U.S.A.
The past several years have witnessed an unprecedented explosion in our knowledge of the cellular machinery which controls communication across the neuronal synapse. Many of the proteins which initiate and regulate the fusion of synaptic vesicles with the pre-synaptic plasma membrane have been cloned and characterized (Ferro-Novick and Jahn, 1994). The complex assemblies in which these polypeptides participate have been dissected and reconstituted, producing a remarkably rich picture of the chain of events which couples the arrival of an action potential to synaptic vesicle discharge (Sudhof, 1995). In order to develop a complete understanding of the critically important process of synaptic transmission, however, it is not sufficient to elucidate the molecular mechanisms which contribute to the delivery of neurotransmitters to the synaptic space. It is equally important to chart the cellular pathways which remove neurotransmitters from the synaptic space, resulting in the termination of synaptic communication. All of the elegant specializations which permit extremely rapid and exquisitely modulated synaptic release would be useless if there were not similarly sophisticated mechanisms available to prevent neurotransmitters from accumulating in the vicinity of their receptors. In those pharmacologically or pathologically induced settings in which neurotransmitters are allowed to persist within the synaptic space, the temporal and spatial accuracy upon which effective synaptic transmission depends cannot be maintained (Iversenj 1971; Ritz and Kuhar, 1993). The cellular components which terminate synaptic transmission have, perhaps, received somewhat less attention than
they deserve in light of the current euphoria over the unraveling of the exocytotic machinery. This state of affairs is especially unfortunate, since research into the machinery responsible for neurotransmitter removal from the synapse has recently undergone its own renaissance. Most neurons in the central nervous system terminate synaptic transmission by importing their respective neurotransmitter from the synaptic space back into their axon terminus. This retrieval process makes use of secondary active transport systems, which exploit the energy stored in trans-plasmalemmal ion gradients to drive the rapid uptake of neurotransmitters from the synaptic space into the pre-synaptic cytoplasm (Kanner, 1994). The recovered neurotransmitter can then be re-packaged into synaptic vesicles and employed in subsequent rounds of neuronal communication. A great deal of research has defined many of the physiological properties of these transporters and established their importance as the primary pharmacological targets for a number of therapeutic and abused substances (Schloss et al., 1992). Biochemical studies have revealed that each neurotransmitter is retrieved by a distinct class of transport proteins, which manifest unique ionic requirements and inhibitor sensitivities. In the past 4 years the genes encoding the transport systems specific for a number of different neurotransmitters have been cloned and their molecular structures revealed (Amara and Arriza, 1993). These genes all belong to a large family, within which a number of subfamilies can be defined. The preceding review by Laurence Borden provides 357
358
Critique
an excellent and remarkably thorough summary of our current understanding of one of these subfamilies. At least four genes encode proteins which are capable of transporting the neurotransmitter GABA (Guastella et al., 1990; Borden et al., 1992; Yamauchi et al., 1992). To some extent, this multiplicity illustrates the embarrassment of riches that can accompany the successful application of molecular techniques to problems which were previously susceptible only to pharmacological or biochemical analysis. While pharmacological studies had subdivided GABA transporters into two sub-classes (“neuronal” and “glial”) based on their inhibitor sensitivities, cloning revealed the existence of much greater diversity. Furthermore, correlating the “glial” transport activity with the behaviors of any of the cloned transporters expressed by transection in heterologous systems has proven difficult (see Dr Borden’s review for a detailed discussion of this issue). Thus, in a sense, the wealth of information provided by molecular biological analysis has created as many problems as it has resolved. How does the number and variety of newly-identified GABA transporter proteins correspond to the GABA transport activities measured in actual tissues? Furthermore, why has evolution gone to the trouble of creating this molecular diversity? Dr Borden has explored, in great depth, the physiological properties of the newly cloned transporters and correlated these behaviors with the functional characteristics and expression patterns of GABA transporters in situ. It is clear from his discussion that the relationship between the cloned transporters and the identified activities is not simple. It also appears likely that the number and diversity of GABA transport proteins may have arisen in order to satisfy biological requirements which are not reflected in substrate affinities or inhibitor sensitivities. Dr Borden suggests the possibility that different members of the GABA transporter family maybe targeted to distinct subcellular localization. He further hypothesizes that the sequence differences among the GABA transporter isoforms may account for their differential distributions. Results from recent experiments in which GABA transporter isoforms were exogenously expressed in epithelial cells and neurons suggest that these propositions may, in fact, be correct. The cell surface membranes of polarized epithelial cells are divided into apical and basolateral domains which are separated from one another by tight junctions. These two regions of the plasmalemma confront distinct body compartments, subserve different physiological functions and are endowed with distinct sets of integral membrane proteins (Caplan and
Matlin, 1989; Rodriguez-Boulan and Nelson, 1989). We have expressed the cDNAs encoding the GAT-1, GAT-2, GAT-3 and BGT-1 proteins by transection in the polarized MDCK epithelial cell line, which derives from and resembles the canine renal tubule. We find that the GAT-1 and GAT-3 polypeptides are restricted to the apical surface of the transected cells (Pietrini et al., 1994; Ahn et al., in press). This distribution is confirmed by immunofluorescence analysis as well as by transport assays and biochemical surface labelling techniques. In contrast, GAT-2 and BGT-1 are restricted to the basolateral plasma membrane domains when expressed in MDCK cells (Pietrini et al., 1994; Ahn et al., in press). Once again, immunofluorescence, transport assays and surface labelling methods support this conclusion. As noted in Dr Borden’s review, the sorting of BGT1 to the basolateral surfaces of transected MDCK cells is not surprising, since physiological studies had previously demonstrated the presence of the BGT-1 transport activity in the MDCK basolateral plasmaIemma (Yamauchi et al., 1991). BGT-I appears to be involved in importing betaine into cells from the extracellular fluid compartment in order to assist renal epithelial cells in adapting to osmotic stress (Nakanishi et al., 1990). In order to serve in this function, BGT-1 must, therefore, be exposed to the extracellular fluid compartment and, consequently, must be sorted to the basolateral membrane. The targeting of the three other GAT isoforms was not as easy to predict from basic principles. Immunocytochemical studies have demonstrated that GAT-1 is restricted to the pre-synaptic membranes of axon termini in the GABA-ergic neurons which express it (Radian et al., 1990). Recent studies on the mechanisms of neuronal membrane protein sorting suggest that the signals and machinery involved in sorting proteins to axons may be similar or analogous to ‘the signals and machinery which participates in apical targeting in epithelial cells (Dotti and Simons, 1990; Dotti et al., 1991). Similarly, sorting to the somatodendritic surfaces of neurons appears to be mechanistically related to the process of basolateral targeting in epithelial cells. Our observation that GAT-1 is apically sorted in epithelial cells lends support to this hypothesis. Further support for the model relating epithelial and neuronal sorting mechanisms can be found in the results of experiments in which we expressed the cDNAs encoding GAT-3 and BGT-1 in polarized hippocampal neurons in culture (Ahn et al., in press). We developed a protocol in which thecDNAs of interest were microinjected directly into the nuclei of
——
..
.—..———.. .
Critique cultured post-mitotic neurons derived from the embryonic rat hippocampus. Immunfluorescence analysis revealed that while the expressed BGT-1 protein was restricted to somatodendritic regions, GAT-3 could gain access to axons. It would appear, therefore, that at least with respect to the members of the GAT transporter family, there is a close correlation between neuronal and epithelial sorting behaviors. It is important to note, however, that studies of the sorting behaviors of other neurotransmitter re-uptake systems suggest that this model may not be generally applicable. Recent experiments performed by the laboratory of Dr Gary Rudnick at Yale University, in collaboration with our own, suggest that the strict correlation between axonal and apical sorting may not apply as simply to the dopamine, serotonin and norepinepherine transporters. Only a single isoform has been identified for both the norepinepherine and serotonin transporters. Furthermore, the functional roles of both transporters require that they be present in the axonal plasmalemma. When expressed in MDCK cells, however, both of these proteins are sorted exclusively to the basolateral surface. It would appear, therefore, that the relationship between epithelial and neuronal sorting is less straightforward than had originally been assumed. Clearly, a great deal of research remains to be done in order to understand more completely the cell-type specific specializations which ensure that membrane protein targeting pathways meet the physiological requirements of each individual polarized tissue. Dr Borden’s review points out another interesting correlation relevant to the sorting behavior of the GAT transporters. The GAT-1 and GAT-3 proteins, which are apically sorted in transected epithelia, appear to be expressed exclusively in neurons in situ. In contrast, BGT-1 and GAT-2, which are normally expressed in epithelia as well as in cells of the nervous system, behave as basolateral proteins in transected MDCK cells. As noted above, BGT-1 requires access to the basolateral surface in order to carry out its physiological function in renal epithelial cells. It is quite possible that GAT-2 serves in a similar or related capacity in the epithelial cells in which it is expressed. If GAT-2 imports GABA or a related substance from the extracellular fluid compartment into epithelial cells, it is logical to predict that it, too, would need to enjoy a basolateral localization. It will be extremely interesting to monitor the results of efforts to immunolocalize GAT-3 and GAT-2 in situ in order to determine if these predictions are borne out. Finally, it should be noted that the distinct sorting behaviors exhibited by the GAT isoforms must reflect
359
the presence of distinct sorting information embedded within the structures of these proteins (Caplan and Matlin, 1989). Presumably, GAT-1 and GAT-3 encode apical sorting signals, whereas GAT-2 and BGT-1 possess sorting signals which specify their basolateral distribution. The high degree of homology which relates the GAT family members to one another provides us with a valuable system with which to explore the nature and composition of these sorting signals. We have begun to generate molecular constructs in which portions of GAT transporters are removed or replaced by corresponding portions of oppositely-sorted isoforms. While still quite preliminary, this analysis suggests that both the cytosolic amino and carboxyl termini of these proteins may contribute to the presentation of these proteins’ sorting signals. Hopefully, further chimera and deletion studies will allow us to identify narrowly defined sequence domains which account for the GAT transporters’ differential sorting behaviors in both epithelial cells and in neurons. Elucidating these signals may further our understanding of the information transfer processes involved in membrane protein sorting and help us to identify the cellular apparatus which is responsible. Insight into the nature of these signals may also shed light on the evolutionary need for the multiplicity of GABA transporters and suggest their individual biological roles.
REFERENCES
Ahn J., Mundigl O., Muth R. R., Rudnick G. and Caplan M. J. (1996)Polarized expressionof GABA transporters in MDCK cells and cultured hippocampal neurons. .J. Bid. Chem.In press. Amara S. G. and Arriza J. L. (1993)Neurotransmitter transporters: three distinct gene families. Curr. Opin. Neurol. 337-344. Borden L. A., Smith K. E., Hartig P. R., Branchek T. A. and Weinshank R. L. (1992)Molecular heterogeneityof they-aminobutyricacid (GABA)transport system.J. BioL Chem.267,21,098–21,104. Caplan M. J. and Matlin K. S. (1989)Sorting of membrane and secretoryproteinsin polarizedepithelialcells.In FunctionalEpithelialCellsin Culture(Matlin K. S. and Valentich J. D., eds.), pp. 71–127.Alan R. Liss, New York. Dotti C. G., Parton R. G. and SimonsK. (1991)Polarized sorting of glypiated proteins in hippocampal neurons. Nature 349, 158–161. Dotti C. G. and SimonsK. (1990)Polarized sorting of viral glycoproteinsto the axon and dendrites of hippocampal neurons in culture. Cell62, 63–72. Ferro-Novick S. and Jahn R. (1994) Vesicle fusion from yeast to man. Nature370, 191-193. Guastella J., Nelson N., Nelson H., CzyzykL., Kenyan S.,
360
Critique
Miedel M., Davidson N., Lester H. and Kanner B. I. (1990)Cloningand expressionofa rat brain GABAtransporter. Science249, 1303-1306. IversenL. L. (1971)Role of transmitter uptake mechanisms in synaptic neurotransmission. Br. J. Pharmac.41, 571– 591, Kanner B. L (1994)Sodium-coupledneurotransmitter transport: structure, functionand regulation.J. Exp. Biol. 196, 237-249. Nakanishi T., Turner R. J. and Burg M. B. (1990)Osmoregulation of betaine transporter in mammalian renal medullarycells.Am.1 Physiol.258,F1061–F1O67. Pietrini G., SuhY. J., EdelmannL., RudnickG. and Caplan M. J. (1994)The axonal y-aminobutyricacid transporter GAT-I is sorted to the apical membranes of polarized epithelialcells.J. Biol. Chem.269,4668&4674. Radian R., Ottersen O. P., Storm-Mathisen J., Castel M. and Kanner B.I. (1990)Immunocytochemicallocalization of the GABA transporter in rat brain. J. Neurosci. 10, 1319-1330.
Ritz M. C. and Kuhar M. J. (1993)Psychostimulantdrugs and a dopamine hypothesis regarding addiction: update on recent research. Biochem.Soc. Symp. 59, 51L64. Rodriguez-BoulanE. and Nelson W. J. (1989) Morphogenesisof the polarized epithelial cell phenotype. Science 245,718-725. SchlossP., Mayser W. and BetzH. (1992)Neurotransmitter transporters: a novelfamilyof integral plasma membrane proteins. REBS Lett. 307,7G80. SudhofT. C. (1995)The synapticvesiclecycle: a cascade of protein-proteininteractions. Nature 375,645+53. Yamauchi A., Kwon H. M., Uchida S., Preston A. and HandlerJ. S. (1991)Myo-inositoland betainetransporters regulated by tonicity are basolateral in MDCK cells.Am. J. Physiol.261,F197-F202. YamauchiA., Uchida S., KwonH. M., Preston A. S., Robey R. B., Garcia-Perez A., Burg M. B. and Handler J. S. (1992)Cloning of a Na and Cl-dependentbetaine transporter that is regulated by hypertonicity. J. BioL Chem. 267,649%652.