title: Interaction between the glutamate transporter GLT1b and the synaptic PDZ domain protein PICK1.
abstract: Synaptic plasticity is implemented by the interaction of glutamate receptors with PDZ domain proteins. Glutamate transporters provide the only known mechanism of clearance of glutamate from excitatory synapses, and GLT1 is the major glutamate transporter. We show here that GLT1 interacts with the PDZ domain protein PICK1, which plays a critical role in regulating the expression of glutamate receptors at excitatory synapses. A yeast two-hybrid screen of a neuronal library using the carboxyl tail of GLT1b yielded clones expressing PICK1. The GLT1b C-terminal peptide bound to PICK1 with high affinity (K(i) = 6.5 +/- 0.4 microM) in an in vitro fluorescence polarization assay. We also tested peptides based on other variants of GLT1 and other glutamate transporters. GLT1b co-immunoprecipitated with PICK1 from rat brain lysates and COS7 cell lysates derived from cells transfected with plasmids expressing PICK1 and GLT1b. In addition, expression of GLT1b in COS7 cells changed the distribution of PICK1, bringing it to the surface. GLT1b and PICK1 co-localized with each other and with synaptic markers in hippocampal neurons in culture. Phorbol ester, an activator of protein kinase C (PKC), a known PICK1 interactor, had no effect on glutamate transport in rat forebrain neurons in culture. However, we found that exposure of neurons to a myristolated decoy peptide with sequence identical to the C-terminal sequence of GLT1b designed to block the PICK1-GLT1b interaction rendered glutamate transport into neurons responsive to phorbol ester. These results suggest that the PICK1-GLT1b interaction regulates the modulation of GLT1 function by PKC.
authors.affiliation: Department of Neurology, Children's Hospital, Boston, MA 02115, USA.
authors.forename: Merav
authors.initials: M
authors.lastname: Bassan
authors.forename: Hongguang
authors.initials: H
authors.lastname: Liu
authors.forename: Kenneth L
authors.initials: KL
authors.lastname: Madsen
authors.forename: Wencke
authors.initials: W
authors.lastname: Armsen
authors.forename: Jiayi
authors.initials: J
authors.lastname: Zhou
authors.forename: Tara
authors.initials: T
authors.lastname: Desilva
authors.forename: Weizhi
authors.initials: W
authors.lastname: Chen
authors.forename: Allison
authors.initials: A
authors.lastname: Paradise
authors.forename: Michael A
authors.initials: MA
authors.lastname: Brasch
authors.forename: Jeff
authors.initials: J
authors.lastname: Staudinger
authors.forename: Ulrik
authors.initials: U
authors.lastname: Gether
authors.forename: Nina
authors.initials: N
authors.lastname: Irwin
authors.forename: Paul A
authors.initials: PA
authors.lastname: Rosenberg
mesh_terms: D000409:Alanine / Q000378:metabolism; D000818:Animals; D019921:Biotinylation / Q000379:methods; D001921:Brain / Q000166:cytology; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003598:Cytoskeletal Proteins; D004622:Embryo, Mammalian; D027342:Excitatory Amino Acid Transporter 2 / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D047468:Immunoprecipitation / Q000379:methods; D009154:Mutation / Q000502:physiology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D054731:PDZ Domains / Q000502:physiology*; D017434:Protein Structure, Tertiary; D051381:Rats; D020318:Rats, Long-Evans; D017207:Rats, Sprague-Dawley; D013755:Tetradecanoylphorbol Acetate / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D027342:Excitatory Amino Acid Transporter 2; D009687:Nuclear Proteins; C498298:PICK1 protein, rat; D018698:Glutamic Acid; C020922:4-O-methyl-12-O-tetradecanoylphorbol 13-acetate; D013755:Tetradecanoylphorbol Acetate; D000409:Alanine