title: Protein kinase C-mediated phosphorylation of a single serine residue on the rat glial glutamine transporter SN1 governs its membrane trafficking.
abstract: Molecular mechanisms involved in the replenishment of the fast neurotransmitters glutamate and GABA are poorly understood. Glutamine sustains their generation. However, glutamine formation from the recycled transmitters is confined to glial processes and requires facilitators for its translocation across the glial and neuronal membranes. Indeed, glial processes are enriched with the system N transporter SN1 (Slc38a3), which, by bidirectional transport, maintains steady extracellular glutamine levels and thereby furnishes neurons with the primary precursor for fast neurotransmitters. We now demonstrate that SN1 is phosphorylated by protein kinase Cα (PKCα) and PKCγ. Electrophysiological characterization shows that phosphorylation reduces V(max) dramatically, whereas no significant effects are seen on the K(m). Phosphorylation occurs specifically at a single serine residue (S52) in the N-terminal rat (Rattus norvegicus) SN1 and results in sequestration of the protein into intracellular reservoirs. Prolonged activation of PKC results in partial degradation of SN1. These results provide the first demonstration of phosphorylation of SN1 and regulation of its activity at the plasma membrane. Interestingly, membrane trafficking of SN1 resembles that of the glutamate transporter GLT and the glutamate-aspartate transporter GLAST: it involves the same PKC isoforms and occurs in the same glial processes. This suggests that the glutamate/GABA-glutamine cycle may be modified at two key points by similar signaling events and unmasks a prominent role for PKC-dependent phosphorylation. Our data suggest that extracellular glutamine levels may be fine-tuned by dynamic regulation of glial SN1 activity, which may impact on transmitter generation, contribute to defining quantal size, and have profound effects on synaptic plasticity.
authors.affiliation: The Biotechnology Centre and Centre for Molecular Biology and Neuroscience, University of Oslo, N-0317 Oslo, Norway.
authors.forename: Lise Sofie H
authors.initials: LS
authors.lastname: Nissen-Meyer
authors.forename: Mark C
authors.initials: MC
authors.lastname: Popescu
authors.forename: El Hassan
authors.initials: el H
authors.lastname: Hamdani
authors.forename: Farrukh A
authors.initials: FA
authors.lastname: Chaudhry
mesh_terms: D026921:Amino Acid Transport Systems, Neutral / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001216:Asparagine / Q000494:pharmacology; D019921:Biotinylation; D002522:Chlorocebus aethiops; D019295:Computational Biology; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005973:Glutamine / Q000494:pharmacology; D006657:Histones / Q000378:metabolism; D047468:Immunoprecipitation / Q000379:methods; D007211:Indoles / Q000494:pharmacology; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics; D008958:Models, Molecular; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D017354:Point Mutation / Q000187:drug effects / Q000502:physiology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000737:chemistry / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D050765:Qa-SNARE Proteins / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012694:Serine / Q000378:metabolism*; D014162:Transfection / Q000379:methods; D014982:Xenopus laevis
chemical_list: D026921:Amino Acid Transport Systems, Neutral; D004791:Enzyme Inhibitors; D006657:Histones; D007211:Indoles; D008301:Maleimides; D010703:Phorbol Esters; D020033:Protein Isoforms; D050765:Qa-SNARE Proteins; C403222:system N protein 1; D005973:Glutamine; D012694:Serine; D001216:Asparagine; C032974:12-O-retinoylphorbol-13-acetate; D011493:Protein Kinase C; C070515:bisindolylmaleimide I