Regulation of the neuronal glutamate transporter excitatory amino acid carrier-1 (EAAC1) by different protein kinase C subtypes.
In previous studies, we have shown that activation of protein kinase C (PKC) rapidly (within minutes) increases the activity and cell surface expression of the glutamate transporter EAAC1 in two systems that endogenously express this transporter (C6 glioma cells and cocultures of neurons and astrocytes). However, the magnitude of the increase in activity is greater than the increase in cell surface expression. In addition, certain compounds completely block the increase in cell surface expression but only partially attenuate the increase in activity. We hypothesized that PKC increases EAAC1 activity by increasing cell surface expression and catalytic efficiency and that two different subtypes of PKC mediate these effects. To address these hypotheses, the PKC subtypes expressed by C6 glioma cells were identified. Of the PKC subtypes that are activated by phorbol esters, only PKCalpha, PKCdelta, and PKCepsilon were observed. Gö6976, a compound that blocks PKCalpha at concentrations that do not inhibit PKCdelta or PKCepsilon, partially inhibited the increase in uptake but completely abolished the increase in EAAC1 cell surface expression. The 'Gö6976-insensitive' increase in activity was not associated with a change in total transporter expression but was associated with an increase in the V(max). Na(+)-dependent glycine transport was not increased, providing indirect evidence that the Gö6976-insensitive increase in activity was not caused by a change in the Na(+) electrochemical gradient required for activity. Finally, by down-regulating different subtypes of PKC, we found evidence that PKCepsilon mediates the increase in EAAC1 activity that is independent of changes in cell surface expression and found further evidence that PKCalpha mediates the increase in cell surface expression. The potential relationship of the present work with a previously identified role for PKCalpha in certain forms of synaptic plasticity is discussed.
González
Marco I
MI
Department of Pediatrics and Pharmacology, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
Kazanietz
Marcelo G
MG
Robinson
Michael B
MB
D027322:Amino Acid Transport System X-AG / Q000378:metabolism*; D000818:Animals; D054713:Bryostatins; D002227:Carbazoles / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D050580:Excitatory Amino Acid Transporter 3; D005260:Female; D050491:Glutamate Plasma Membrane Transport Proteins; D007211:Indoles / Q000494:pharmacology; D007783:Lactones / Q000494:pharmacology; D018942:Macrolides; D009474:Neurons / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000145:classification / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011956:Receptors, Cell Surface / Q000096:biosynthesis; D027981:Symporters; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D027322:Amino Acid Transport System X-AG; D054713:Bryostatins; D002227:Carbazoles; D002273:Carcinogens; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D050580:Excitatory Amino Acid Transporter 3; D050491:Glutamate Plasma Membrane Transport Proteins; D007211:Indoles; D007783:Lactones; D018942:Macrolides; D011956:Receptors, Cell Surface; C493398:Slc1a1 protein, rat; D027981:Symporters; C081021:Go 6976; C046785:bryostatin 1; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate