Protein Kinase C Activation Decreases Cell Surface Expression of the GLT-1 Subtype of Glutamate Transporter

Abstract

Na+-dependent glutamate transporters are required for the clearance of extracellular glutamate and influence both physiological and pathological effects of this excitatory amino acid. In the present study, the effects of a protein kinase C (PKC) activator on the cell surface expression and activity of the GLT-1 subtype of glutamate transporter were examined in two model systems, primary co-cultures of neurons and astrocytes that endogenously express GLT-1 and C6 glioma cells transfected with GLT-1. In both systems, activation of PKC with phorbol ester caused a decrease in GLT-1 cell surface expression. This effect is opposite to the one observed for the EAAC1 subtype of glutamate transporter (Davis, K. E., Straff, D. J., Weinstein, E. A., Bannerman, P. G., Correale, D. M., Rothstein, J. D., and Robinson, M. B. (1998) J. Neurosci. 18, 2475–2485). Several recombinant chimeric proteins between GLT-1 and EAAC1 transporter subtypes were generated to identify domains required for the subtype-specific redistribution of GLT-1. We identified a carboxyl-terminal domain consisting of 43 amino acids (amino acids 475–517) that is required for PKC-induced GLT-1 redistribution. Mutation of a non-conserved serine residue at position 486 partially attenuated but did not completely abolish the PKC-dependent redistribution of GLT-1. Although we observed a phorbol ester-dependent incorporation of 32P into immunoprecipitable GLT-1, mutation of serine 486 did not reduce this signal. We also found that chimeras containing the first 446 amino acids of GLT-1 were not functional unless amino acids 475–517 of GLT-1 were also present. These non-functional transporters were not as efficiently expressed on the cell surface and migrated to a smaller molecular weight, suggesting that a subtype-specific interaction is required for the formation of functional transporters. These studies demonstrate a novel effect of PKC on GLT-1 activity and define a unique carboxyl-terminal domain as an important determinant in cellular localization and regulation of GLT-1. Na+-dependent glutamate transporters are required for the clearance of extracellular glutamate and influence both physiological and pathological effects of this excitatory amino acid. In the present study, the effects of a protein kinase C (PKC) activator on the cell surface expression and activity of the GLT-1 subtype of glutamate transporter were examined in two model systems, primary co-cultures of neurons and astrocytes that endogenously express GLT-1 and C6 glioma cells transfected with GLT-1. In both systems, activation of PKC with phorbol ester caused a decrease in GLT-1 cell surface expression. This effect is opposite to the one observed for the EAAC1 subtype of glutamate transporter (Davis, K. E., Straff, D. J., Weinstein, E. A., Bannerman, P. G., Correale, D. M., Rothstein, J. D., and Robinson, M. B. (1998) J. Neurosci. 18, 2475–2485). Several recombinant chimeric proteins between GLT-1 and EAAC1 transporter subtypes were generated to identify domains required for the subtype-specific redistribution of GLT-1. We identified a carboxyl-terminal domain consisting of 43 amino acids (amino acids 475–517) that is required for PKC-induced GLT-1 redistribution. Mutation of a non-conserved serine residue at position 486 partially attenuated but did not completely abolish the PKC-dependent redistribution of GLT-1. Although we observed a phorbol ester-dependent incorporation of 32P into immunoprecipitable GLT-1, mutation of serine 486 did not reduce this signal. We also found that chimeras containing the first 446 amino acids of GLT-1 were not functional unless amino acids 475–517 of GLT-1 were also present. These non-functional transporters were not as efficiently expressed on the cell surface and migrated to a smaller molecular weight, suggesting that a subtype-specific interaction is required for the formation of functional transporters. These studies demonstrate a novel effect of PKC on GLT-1 activity and define a unique carboxyl-terminal domain as an important determinant in cellular localization and regulation of GLT-1. Glutamate is the predominant excitatory neurotransmitter in the central nervous system (CNS) 1The abbreviations used for: CNS, central nervous system; PKC, protein kinase C; DMEM, Dulbecco's modified Eagle's medium; GABA, γ-aminobutyric acid; HA, hemagglutinin; PMA, phorbol 12-myristate 13-acetate; Bis II, bisindolylmaleimide II; DHK, dihydrokainate. (1Fagg G.E. Foster A.C. Neuroscience (Oxford). 1983; 9: 701-719Crossref PubMed Scopus (563) Google Scholar, 2Fonnum F. J. Neurochem. 1984; 42: 1-11Crossref PubMed Scopus (1686) Google Scholar) and is removed from the synaptic cleft by sodium-dependent glutamate transport activity. This activity is mediated by a family of five subtypes of transporters that share up to 60% sequence identity (for reviews, see Refs. 3Amara S.G. Sonders M.S. Zahniser N.R. Povlock S.L. Daniels G.M. Adv. Pharmacol. 1998; 42: 164-168Crossref PubMed Scopus (39) Google Scholar, 4Sims K.D. Robinson M.B. Crit. Rev. Neurobiol. 1999; 13: 169-197Crossref PubMed Scopus (146) Google Scholar, 5Danbolt N.C. Prog. Neurobiol. 2001; 65: 1-105Crossref PubMed Scopus (3775) Google Scholar). Expression of these transporters is generally restricted to particular cell types and brain regions. Two subtypes, GLT-1 and GLAST, are astroglial and two others, EAAC1 and EAAT4, are neuronal. Expression of the fifth transporter, EAAT5, is restricted to the retina (for reviews, see Refs. 4Sims K.D. Robinson M.B. Crit. Rev. Neurobiol. 1999; 13: 169-197Crossref PubMed Scopus (146) Google Scholar and 5Danbolt N.C. Prog. Neurobiol. 2001; 65: 1-105Crossref PubMed Scopus (3775) Google Scholar). Both the neuronal and glial glutamate transporters control the amplitude and/or duration of synaptic responses (6Mennerick S. Zorumski C.F. Nature. 1994; 368: 59-62Crossref PubMed Scopus (291) Google Scholar, 7Otis T.S. Kavanaugh M.P. Jahr C.E. Science. 1997; 277: 1515-1518Crossref PubMed Scopus (156) Google Scholar) and prevent an extracellular accumulation of this potential excitotoxin (8Robinson M.B. Djali S. Buchhalter J.R. J. Neurochem. 1993; 61: 2099-2103Crossref PubMed Scopus (100) Google Scholar, 9Rothstein J.D. Dykes-Hoberg M. Pardo C.A. Bristol L.A. Jin L. Kuncl R.W. Kanai Y. Hediger M. Wang Y. Schielke J.P. Welty D.F. Neuron. 1996; 16: 675-686Abstract Full Text Full Text PubMed Scopus Google Scholar, K. K. K. M. K. S. S. M. K. Science. 1997; PubMed Scopus Google Scholar). is that the glial transporter, GLT-1, the predominant for the clearance of extracellular glutamate in (for see M.B. Neurochem. 1999; Scopus Google Scholar). the that GLT-1 the potential to of both the and of glutamate in the Several the activity of GLT-1, and PKC (for reviews, see Refs. 4Sims K.D. Robinson M.B. Crit. Rev. Neurobiol. 1999; 13: 169-197Crossref PubMed Scopus (146) Google Scholar and 5Danbolt N.C. Prog. Neurobiol. 2001; 65: 1-105Crossref PubMed Scopus (3775) Google Scholar). the effects of PKC on activity are activation of PKC an in activity GLT-1 is expressed in cells M. F. N.C. J. 1993; Full Text PubMed Google Scholar) but effect on the activity of GLT-1 transfected into cells two cell J. J.D. Robinson M.B. J. Pharmacol. 1999; Google Scholar). In a cell that endogenously the of GLT-1 activation of PKC a decrease in activity by the for transport C.E. J. Neurochem. 1998; PubMed Scopus Google Scholar). In a study, activation of PKC caused a decrease in GLT-1 activity in transfected cells J. Neurosci. 1999; Scholar). These studies that the effects of PKC activation on the of transporter expression and/or the of cellular proteins that are not expressed in types of proteins a between the cell surface and in cell surface expression the activity of proteins by the of protein is of the subtype of transporter, is to the in to (for reviews, see Refs. M.P. S. J. 1999; Full Text Full Text PubMed Scopus Google Scholar and S. J. 1999; Full Text Full Text PubMed Scopus Google Scholar). of is of (for see Pharmacol. Rev. 2001; Google Scholar). Several that activation of PKC a in the cell surface expression of neurotransmitter the subtype of transporter J. Neurosci. 1999; Google the transporter G.M. S.G. J. 1999; Full Text Full Text PubMed Scopus Google Scholar, J. Neurosci. 1999; PubMed Google and the transporter Y. S. S. J. Neurosci. 1997; PubMed Google Scholar). In of these not that of glutamate transporters is also In studies that the cell surface expression of the neuronal glutamate transporter, is by PKC by the K.D. Robinson M.B. J. Full Text Full Text Scopus Google Scholar, J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar). In the present study, the effects of a PKC activator on GLT-1 cell surface expression were examined in co-cultures of neurons and In this model phorbol ester caused a decrease in the cell surface expression of GLT-1. GLT-1 into C6 glioma a cell that endogenously the neuronal transporter, a PKC activator caused a decrease in the cell surface expression of GLT-1 and a decrease in transport activity. In a PKC activator caused an in EAAC1 cell surface expression in these that the effects of PKC are to in primary of these two chimeras of domains of GLT-1 and we demonstrate that a carboxyl-terminal domain is both and for redistribution of GLT-1. mutation of serine and in this we that a non-conserved serine residue is partially for the decrease in GLT-1 cell surface expression. Although we found that a PKC activator incorporation of 32P into GLT-1, mutation of this serine residue did not reduce this signal. these we also that the domain of GLT-1 required for functional expression of transporter activity in these studies for PKC-dependent regulation of GLT-1 and identify a domain required for this DMEM, and were from from and were from molecular and were from from from from from and from of these were by J.D. L. Dykes-Hoberg M. Jin L. D. Kuncl R.W. Neuron. 1994; 13: Full Text PubMed Scopus Google Scholar). from from phorbol 12-myristate and were from were from of as (8Robinson M.B. Djali S. Buchhalter J.R. J. Neurochem. 1993; 61: 2099-2103Crossref PubMed Scopus (100) Google Scholar) with were and in a at were with a a neurons in these on of a of astrocytes and of the GLT-1 is endogenously expressed in these and of the with J.R. M. J.D. Robinson M.B. Pharmacol. 1998; PubMed Scopus Google Scholar). C6 glioma cells were from the and as J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar). cells were in with and at in were these in in effects were were with for and used for as glutamate transporters were GLT-1 and EAAC1 transporter subtypes, by and N.C. M. Y. L. J. E. Nature. PubMed Scopus Google Scholar, Y. M. Hediger PubMed Scopus Google Scholar). were generated unique at one of the transporters. the not in both by the present in the GLT-1 at the in In the were that the protein the amino from one of the two transporters. chimeric transporters were in this chimeras that did not the carboxyl-terminal by the (amino acids of J.D. L. Dykes-Hoberg M. Jin L. D. Kuncl R.W. Neuron. 1994; 13: Full Text PubMed Scopus Google Scholar) were with an that used to the as as chimeric transporters were into the expression of chimeras were by at the molecular at the of were and a at a unique a at a unique to of GLT-1 between were into the expression and to the C6 glioma cells were to between and 60% in to of a in were transfected with of with of of were for surface of cells in a to in of DMEM, an of modified containing and to the for an used to the and transfected were with Na+-dependent transport activity as J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar). were in a on and with of to with in for with cells were in of this were for protein and Na+-dependent as the between the observed in and in as with K.D. Robinson M.B. J. Full Text Full Text Scopus Google Scholar, J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar). cells were to between and were with containing and and were in this with for at cells were with containing and in this for at to the were with with of the cellular by proteins were of cell proteins surface and proteins were transfected C6 glioma cells were in for and were with for cells were with phorbol ester for with and into and for at cell removed by at for of the of cell first by for at with the at with at a of and by with protein for at protein were in and in of were by the to on proteins were to in containing were with and for used a of the primary were and with and proteins were by proteins with the were in for and the proteins were to a that the cell surface of neurotransmitter transporters by activation of PKC (for reviews, see Refs. Scopus Google Scholar, Neurobiol. PubMed Scopus Google Scholar, M.B. J. Neurochem. 1-11Crossref PubMed Scopus Google Scholar). effects of PKC on the activity of GLT-1 are and on cellular M. F. N.C. J. 1993; Full Text PubMed Google Scholar, J. J.D. Robinson M.B. J. Pharmacol. 1999; Google Scholar, C.E. J. Neurochem. 1998; PubMed Scopus Google Scholar, J. Neurosci. 1999; Scholar). we examined the effect of activation of PKC on the cell surface expression of GLT-1 primary co-cultures of neurons and astrocytes from effects of the PKC PMA, on the cell surface of GLT-1 were examined by of cell surface Although effect on the of GLT-1 the of cell surface GLT-1 to of control and In these caused an in the of EAAC1 Y. and M. B. Robinson, also an in EAAC1 cell surface expression in Robinson M.B. Pharmacol. PubMed Scopus Google Scholar). GLT-1 and EAAC1 expressed in cell and the effect of PKC are on the of proteins that are expressed in astrocytes and/or neurons these two transporters domains that the to PKC between these transporters the effects of PKC GLT-1 transfected into C6 a cell that endogenously the EAAC1 subtype of transporter J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar, B. 1996; PubMed Scopus Google Scholar). In C6 glioma transfected with GLT-1, a with caused a decrease in the cell surface expression of GLT-1, an in the of GLT-1, and in GLT-1 C and phorbol are to with PKC P. M. Wang Pharmacol. PubMed Scopus Google the effect of a Bis II, on GLT-1 cell surface expression In these Bis effect in the of but completely the decrease in GLT-1 cell surface expression in expression in of C6 glioma not the studies not the that the effects of a PKC activator on EAAC1 and GLT-1 in of this GLT-1 also transfected into C6 glioma and were a of cells that express both transporters. that activity that observed in cells for studies of protein with of protein in In this cell for the cell surface expression of GLT-1 of and the cell surface expression of EAAC1 in the of and These studies that the effects of on EAAC1 and GLT-1 cell surface expression are to In that the excitatory amino is as an of activity as an of transport activity to J. J.D. Robinson M.B. J. Pharmacol. 1999; Google Kavanaugh M.P. S.G. J. Neurosci. 1994; PubMed Google Scholar, and L.A. J.D. Robinson M.B. Pharmacol. 1996; Google Scholar, for see M.B. L.A. Adv. Pharmacol. 1997; PubMed Scopus Google Scholar). of activity. In the effects of of on activity were examined in transfected and C6 In these of transport activity by with a of with an of In the for of transport activity were to two with of and of the were of This of is with the observed in these cells with This used to the of effects of on transporter activity and activity were in and transfected cell In transport and effect on activity in both and cells In effect on transporter but transport activity in transport to of is in with the in GLT-1 cell surface expression observed in this cell These that activity and demonstrate that this decrease in activity is with a decrease in cell surface expression. EAAC1 and GLT-1 to in the cellular in primary for the effects of identify required for the PKC-induced redistribution of GLT-1, a family of chimeras by of domains between GLT-1 and EAAC1 were generated transporters were into the used to the amino in of a were from EAAC1 GLT-1, first the 446 amino the amino the 43 amino and the the amino In chimeras that the carboxyl-terminal domain of GLT-1, and not by a into the amino to an that used to the of these transporters. into the amino of GLT-1 to that not with of these transfected into C6 glioma cells to transport activity. and transfected C6 glioma cells were used as of of these chimeras in in transporter but were not functional of chimeric transporters and effect of on cell surface transporter of on transporter of of of in of the from cells transfected with GLT-1 chimeras that did not the EAAC1 not to to not glioma were transfected with Na+-dependent transport activity examined and expressed as a of that observed in that with effect on transport activity. functional transport activity to transfected of surface transporter examined in and of transporter that is expressed on the cell surface is expressed as a of transporter effect of on cell surface expression of functional also examined and expressed as a of that observed in are as the and the of are in EAAC1 in of the from cells transfected with GLT-1 chimeras that did not the EAAC1 not to to not in a C6 glioma were transfected with Na+-dependent transport activity examined and expressed as a of that observed in that with effect on transport activity. functional transport activity to transfected of surface transporter examined in and of transporter that is expressed on the cell surface is expressed as a of transporter effect of on cell surface expression of functional also examined and expressed as a of that observed in are as the and the of are in of of the chimeras that the first amino acids of EAAC1 in an in Na+-dependent transport activity In of the chimeras that the first 446 amino acids of GLT-1 were and of this of chimeras is the of the first and of GLT-1. and not the of GLT-1, were not This that subtype-specific between of GLT-1 are required for transport activity. interaction required for the of the transporter, for of functional and/or to the cell this we the cell surface expression of chimeras in In these the of the transporter on the cell surface that observed with of the functional and In to a smaller of transporter expressed on the cell the molecular of the that is with a smaller functional transporters in of the also observed in the and from cells transfected with functional chimeras the were in This smaller observed for non-functional by M. B. Neuron. 1998; Full Text Full Text PubMed Scopus Google Scholar, M. J. Full Text Full Text PubMed Scopus Google Scholar, D. M. P. N.C. Hediger J. 2001; Full Text Full Text PubMed Scopus (156) Google and that these proteins of the transporters. transporter, the of to the and the effect of on the cell surface expression examined a we that the not with transporter observed with GLT-1, the cell surface expression of the GLT-1 effects of on EAAC1 were also the cell surface expression of and This effect of PMA, cell surface expression to of smaller that observed in studies J.D. Robinson M.B. J. Neurosci. 1998; PubMed Google Scholar) and that observed in studies C6 glioma transfected with GLT-1 We also examined the effects of on EAAC1 cell surface expression in of the in chimeras were transfected into C6 In these we found that EAAC1 cell surface expression to of control These studies that partially the effects of on EAAC1 cell surface and in studies we found that with the effect of on EAAC1 cell surface expression not with an activator of PKC the cell surface expression of chimeric transporters that the of GLT-1, and and In chimeras that did not this were by PKC activation on the cell surface in to PKC these studies that the amino acids of GLT-1 is for to decrease GLT-1 cell surface expression. the cell surface expression of and that this domain of GLT-1 and that are not required for the PKC-dependent redistribution of GLT-1. the is in to PKC that the required for EAAC1 redistribution to the in the is that is not to this transporter domain that is for PKC-dependent of GLT-1 serine and and from EAAC1 in amino acids this effect is we on mutation of serine and as potential that important for GLT-1 containing of these to were expressed in C6 glioma cells by and for cell surface expression in to PKC mutation of both unique to effect on the redistribution of GLT-1 mutation of five serine in this domain in of the effects of serine were to to one of these serine are required for the of the to GLT-1, the effects of on the fifth were partially and to observed with the that serine is that regulation of the cell surface expression of the subtype of transporter is of J. 2001; Full Text Full Text PubMed Scopus Google Scholar). the residue that is present in this domain also to This also to GLT-1 suggesting that this residue is not required for redistribution of GLT-1. of of these in activity in C6 glioma domains that are important in of proteins J. S. J. 16: Scholar, J. J. J. 1998; Full Text Full Text PubMed Scopus Google Scholar). Although serine 486 not at a PKC the effects of on incorporation into immunoprecipitable GLT-1 were in 32P incorporation into an immunoprecipitable of observed in C6 glioma transfected with GLT-1 but not in transfected cells This that PKC GLT-1 is expressed in C6 Although mutation of five serine in the domain in incorporation of 32P into GLT-1, mutation of the serine residue that partially for redistribution effect on the of 32P a of the used to 32P incorporation were and the proteins were to and for GLT-1 of were observed in C6 glioma transfected with the GLT-1 that the in to in protein expression these studies that the in of and the serine 486 is not by this that of this residue is not required for of GLT-1. these studies that incorporation of 32P into one of the serine in this but at present is this functional effects on GLT-1. the five glutamate transporters that identified GLT-1, GLAST, and are found in in the (for reviews, see Refs. 4Sims K.D. Robinson M.B. Crit. Rev. Neurobiol. 1999; 13: 169-197Crossref PubMed Scopus (146) Google Scholar and 5Danbolt N.C. Prog. Neurobiol. 2001; 65: 1-105Crossref PubMed Scopus (3775) Google Scholar). Several of that GLT-1 the of glutamate transport in this of GLT-1 from of the activity K. M. N.C. J. 1996; Full Text Full Text PubMed Scopus Google Scholar). of GLT-1 observed in (for see M.B. L.A. Adv. Pharmacol. 1997; PubMed Scopus Google and of GLT-1 a of GLT-1 to transport activity J.D. Dykes-Hoberg M. Pardo C.A. Bristol L.A. Jin L. Kuncl R.W. Kanai Y. Hediger M. Wang Y. Schielke J.P. Welty D.F. Neuron. 1996; 16: 675-686Abstract Full Text Full Text PubMed Scopus Google Scholar). studies that activity is important for activation of glutamate and also to synaptic Jahr C.E. Neuron. 1997; Full Text Full Text PubMed Scopus Google Scholar, Science. 2001; PubMed Scopus Google Scholar, K. J. Neurosci. 2001; PubMed Google Scholar). of GLT-1 in transport activity to of control K. K. K. M. K. S. S. M. K. Science. 1997; PubMed Scopus Google Scholar). These excitatory and by of these studies that GLT-1 is for the of glutamate and that regulation of GLT-1 important for both and brain In the present study, we that with a PKC activator a decrease in GLT-1 cell surface expression in of neurons and this PKC activator also caused a decrease in the cell surface expression of GLT-1 in transfected C6 In transfected the PKC activator cell surface expression of that the effects of PKC activation are to in the primary in cellular chimeric transporters consisting of domains between these we identified a of GLT-1 that is both and for a PKC-induced decrease in cell surface expression. This is to the domain and of the carboxyl-terminal M. B. Neuron. 1998; Full Text Full Text PubMed Scopus Google Scholar, M. J. Full Text Full Text PubMed Scopus Google Scholar, S.G. Neuron. Full Text Full Text PubMed Scopus Google Scholar). cell surface expression of chimeras this domain in to with a PKC This decrease in GLT-1 cell surface expression with incorporation of into immunoprecipitable GLT-1. Mutation of two unique in this effect on the PKC-dependent redistribution of GLT-1. mutation of serine in this of GLT-1 partially attenuated the decrease in cell surface expression and the PKC-dependent These that GLT-1 is at one of these serine is not this the of the transporter, but not to required for the PKC-induced redistribution of GLT-1. Although mutation of a serine residue in this to attenuated the PKC-dependent decrease in cell surface expression to the as mutation of serine this mutation effect on of the cell surface expression of proteins is to a to activity by protein of the of this of regulation is that of activation of of these is by is caused by to by activation of a kinase caused by activation of is generally that both of are the of of the by an B. J. L. L. J. 2001; Full Text Full Text PubMed Scopus Google Scholar and Wang L. Wang J. 1998; Full Text Full Text PubMed Scopus Google for see Pharmacol. Rev. 2001; Google but is that is not required a B. J.D. J. 1996; Full Text Full Text PubMed Scopus Google Scholar). the are and for to the (for see Pharmacol. Rev. 2001; Google Scholar). model of for the of the cell (for see J. J. J. 1998; Full Text Full Text PubMed Scopus Google Scholar). studies that the activity and cell surface expression of of the neurotransmitter transporters is by of PKC the cell surface expression of the transporter Y. S. S. J. Neurosci. 1997; PubMed Google the transporter G.M. S.G. J. 1999; Full Text Full Text PubMed Scopus Google Scholar, J. Neurosci. 1999; PubMed Google and in the subtype of transporter J. Neurosci. 1999; Google Scholar). is that PKC-dependent are for these but is not transporter is is a between the for and of the transporter S. Science. 1999; PubMed Scopus Google Scholar). of serine PKC effect on PKC-dependent of transport but of the transporter were is these studies are K. M. S. J. Neurochem. 1997; PubMed Scopus Google Scholar). transporter is also in to activation of Mutation of serine at PKC PKC-dependent of this transporter but effect on PKC-dependent J. Neurochem. 2001; PubMed Scopus Google Scholar). studies that the of J. 2001; Full Text Full Text PubMed Scopus Google Scholar). In the present study, mutation of a serine residue partially the PKC-dependent redistribution of GLT-1 but did not decrease of GLT-1. This that transporter is not required for redistribution that the of is not to a mutation of this we the that serine 486 is and that mutation PKC the transporter at residue that is not in the is not required for transporter this that an protein in of GLT-1 a of proteins are with a the of and to the from (for see Rev. 1999; PubMed Scopus Google Scholar). the of the transporter, to in to PKC activation G.M. S.G. J. 1999; Full Text Full Text PubMed Scopus Google Scholar, J. Neurosci. 1999; PubMed Google is PKC of these transporters of the transporter to the cell Several amino identified as important for (for see Rev. 1999; PubMed Scopus Google Scholar). These are found on the carboxyl-terminal of the proteins as are domain identified as both and for GLT-1 is in to amino acids of the domain and amino acids of the carboxyl-terminal M. J. Full Text Full Text PubMed Scopus Google Scholar, S.G. Neuron. Full Text Full Text PubMed Scopus Google Scholar). of this sequence to the in EAAC1 amino and of these are for for and for serine residue that mutation to the effects of PKC is amino acids of an is that a serine residue by a amino acids as a sequence for of proteins (for see S. J. S. J. Full Text Full Text PubMed Scopus Google Scholar). In the present study, we not examined the effects of mutation of the and to this Two that identified for the are not found in the of GLT-1 that is required for PKC-dependent redistribution. In important to these effects of PKC are to the and to define the of GLT-1 that are required for this In we that PKC activation in a redistribution of GLT-1 from the cell surface in both primary from brain and in C6 glioma This effect of PKC is opposite to that observed with a of chimeras between EAAC1 and GLT-1, we identified a domain in GLT-1 that is required for this we identified a serine residue this domain that to partially for this redistribution. the that these transporters important for both the physiological and pathological of transporter We J. for Correale, and for with these and Hediger for of the GLT-1 and EAAC1 and the of the Robinson for and of this