Splice variants of the glutamate transporter GLT1 form hetero-oligomers that interact with PSD-95 and NMDA receptors.
The glutamate transporter GLT1 is expressed in at least two isoforms, GLT1a and GLT1b, which differ in their C termini. As GLT1 is an oligomeric protein, we have investigated whether GLT1a and GLT1b might associate as hetero-oligomers. Differential tagging (HA-GLT1a and YFP-GLT1b) revealed that these isoforms form complexes that could be immunoprecipitated when co-expressed in heterologous systems. The association of GLT1a and GLT1b was also observed in mixed primary cultures of rat brain and in the adult rat brain, where specific antibodies for GLT1a immunoprecipitated GLT1b and vice versa. Dual immunofluorescence in mixed cultures demonstrated the partial co-localization of both isoforms in neurons and in glial cells. Because GLT1b interacts with an organizer of post-synaptic densities, PSD-95, we examined the capacity of GLT1a to associate with this protein. GLT1a was immunoprecipitated from the rat brain in protein complexes that contained not only GLT1b but also PSD-95 and NMDAR. The interaction between GLT1a with PSD-95 and NMDAR was reproduced in transfected COS7 cells and it appears to be indirect as it requires the presence of GLT1b. These results indicate that the major isoform of the glutamate transporter, GLT1a, can acquire the capacity to interact with PDZ proteins through its inclusion in hetero-oligomers containing GLT1b.
González-González
Inmaculada M
IM
Facultad de Ciencias, Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid, Consejo Superior de Investigaciones Científicas, Centro de Investigación en Red de Enfermedades Raras, Madrid, Spain.
García-Tardón
Noemí
N
Giménez
Cecilio
C
Zafra
Francisco
F
D017398:Alternative Splicing / Q000235:genetics; D000818:Animals; D001921:Brain / Q000378:metabolism*; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D018920:Coculture Techniques; D000076122:Disks Large Homolog 4 Protein; D004285:Dogs; D027342:Excitatory Amino Acid Transporter 2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D009457:Neuroglia / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D011108:Polymers / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism*; D013569:Synapses / Q000378:metabolism*; D013570:Synaptic Membranes / Q000235:genetics / Q000378:metabolism; D009435:Synaptic Transmission / Q000235:genetics
D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D027342:Excitatory Amino Acid Transporter 2; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D011108:Polymers; D020033:Protein Isoforms; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D018698:Glutamic Acid