Tamalin, a PDZ domain-containing protein, links a protein complex formation of group 1 metabotropic glutamate receptors and the guanine nucleotide exchange factor cytohesins.
In this investigation, we report identification and characterization of a 95 kDa postsynaptic density protein (PSD-95)/discs-large/ZO-1 (PDZ) domain-containing protein termed tamalin, also recently named GRP1-associated scaffold protein (GRASP), that interacts with group 1 metabotropic glutamate receptors (mGluRs). The yeast two-hybrid system and in vitro pull-down assays indicated that the PDZ domain-containing, amino-terminal half of tamalin directly binds to the class I PDZ-binding motif of group 1 mGluRs. The C-terminal half of tamalin also bound to cytohesins, the members of guanine nucleotide exchange factors (GEFs) specific for the ADP-ribosylation factor (ARF) family of small GTP-binding proteins. Tamalin mRNA is expressed predominantly in the telencephalic region and highly overlaps with the expression of group 1 mGluR mRNAs. Both tamalin and cytohesin-2 were enriched and codistributed with mGluR1a in postsynaptic membrane fractions. Importantly, recombinant and native mGluR1a/tamalin/cytohesin-2 complexes were coimmunoprecipitated from transfected COS-7 cells and rat brain tissue, respectively. Transfection of tamalin and mutant tamalin lacking a cytohesin-binding domain caused an increase and decrease in cell-surface expression of mGluR1a in COS-7 cells, respectively. Furthermore, adenovirus-mediated expression of tamalin and dominant-negative tamalin facilitated and reduced the neuritic distribution of endogenous mGluR5 in cultured hippocampal neurons, respectively. The results indicate that tamalin plays a key role in the association of group 1 mGluRs with the ARF-specific GEF proteins and contributes to intracellular trafficking and the macromolecular organization of group 1 mGluRs at synapses.
Kitano
Jun
J
Department of Biological Sciences, Faculty of Medicine, Graduate School of Biostudies, Kyoto University, Kyoto, 606-8501, Japan.
Kimura
Kouji
K
Yamazaki
Yoshimitsu
Y
Soda
Takeshi
T
Shigemoto
Ryuichi
R
Nakajima
Yoshiaki
Y
Nakanishi
Shigetada
S
D020727:ADP-Ribosylation Factors / Q000378:metabolism; D000818:Animals; D001923:Brain Chemistry; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism*; D006801:Humans; D017403:In Situ Hybridization; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D009928:Organ Specificity; D011233:Precipitin Tests; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; D015815:Cell Adhesion Molecules; C574553:GRASP protein, rat; C578935:GRM5 protein, human; C578936:Grm5 protein, mouse; C578937:Grm5 protein, rat; D020662:Guanine Nucleotide Exchange Factors; D046911:Macromolecular Substances; D008565:Membrane Proteins; D012333:RNA, Messenger; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate; C104077:metabotropic glutamate receptor type 1; C410230:tamalin protein, human; D020727:ADP-Ribosylation Factors