The C-terminal juxtamembrane region of the delta 2 glutamate receptor controls its export from the endoplasmic reticulum.
Functions of ionotropic glutamate receptors (iGluRs) are tightly regulated by the intracellular trafficking of receptor proteins. Unlike other iGluRs that are considerably retained in the intracellular component, the delta 2 glutamate receptor (GluR delta 2) is efficiently expressed on the Purkinje cell surface. To understand the trafficking mechanism of iGluRs, we deleted various portions of the C-terminal intracellular domain of GluR delta 2 and analysed the localization of the mutant proteins in heterologous cells and neurons. Biotinylation assays indicated that GluR delta 2 lacking the C-terminal juxtamembrane region of 13 amino acids (region A) was not present on the cell surface. This mutant GluR delta 2 was sensitive to endoglycosidase H, which digests unprocessed high-mannose oligosaccharides on proteins retained in the endoplasmic reticulum (ER) or cis-Golgi. Therefore, we concluded that region A is crucial for the transport of GluR delta 2 beyond the trans-Golgi to the cell surface. Because the immunostaining pattern of GluR delta 2 lacking region A in cultured hippocampal neurons completely overlapped the pattern of fluorescence emitted by ER-resident green fluorescent protein, region A is most likely necessary for GluR delta 2's exit from the ER. Furthermore, this region is essential for the proper intracellular trafficking of GluR delta 2 in Purkinje cells. Region A does not rely on a dihydrophobic motif or positively charged residues to participate in trafficking, but its function is dependent on the juxtamembrane position. Therefore, we propose that GluR delta 2's efficient transport to the cell surface utilizes an unknown but general ER exit mechanism, which probably works in close relation to the membrane of heterologous cells and neurons.
Matsuda
Shinji
S
Department of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105-2794, USA.
Hannen
Rosalind
R
Matsuda
Keiko
K
Yamada
Nobuaki
N
Tubbs
Thomas
T
Yuzaki
Michisuke
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000502:physiology; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D064026:Calbindins; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D004622:Embryo, Mammalian; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins; D006388:Hemagglutinins / Q000378:metabolism; D006596:Hexosaminidases / Q000494:pharmacology; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D007668:Kidney; D037102:Lectins; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D011233:Precipitin Tests / Q000379:methods; D017434:Protein Structure, Tertiary / Q000502:physiology*; D021381:Protein Transport / Q000502:physiology; D011689:Purkinje Cells / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017470:Receptors, Glutamate / Q000737:chemistry / Q000378:metabolism*; D064030:S100 Calcium Binding Protein G / Q000378:metabolism; D014162:Transfection / Q000379:methods
D064026:Calbindins; D006388:Hemagglutinins; D037102:Lectins; D008164:Luminescent Proteins; D017470:Receptors, Glutamate; D064030:S100 Calcium Binding Protein G; C107913:glutamate receptor delta 2; C112408:hemagglutinin I; D049452:Green Fluorescent Proteins; D006596:Hexosaminidases