TARP γ-8 glycosylation regulates the surface expression of AMPA receptors.

TARP [transmembrane AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor regulatory protein] γ-8 is an auxiliary subunit of AMPA receptors that is widely distributed in the hippocampus. It has been shown that TARP γ-8 promotes surface expression of AMPA receptors; however, how TARP γ-8 regulates the expression of AMPA receptors remains unclear. In the present study, we examined the effect of TARP glycosylation on AMPA receptor trafficking.

Cornichon proteins determine the subunit composition of synaptic AMPA receptors.

Cornichon-2 and cornichon-3 (CNIH-2/-3) are AMPA receptor (AMPAR) binding proteins that promote receptor trafficking and markedly slow AMPAR deactivation in heterologous cells, but their role in neurons is unclear. Using CNIH-2 and CNIH-3 conditional knockout mice, we find a profound reduction of AMPAR synaptic transmission in the hippocampus. This deficit is due to the selective loss of surface GluA1-containing AMPARs (GluA1A2 heteromers), leaving a small residual pool of synaptic GluA2A3 heteromers.

Fluorescence recovery after photobleaching (FRAP) of fluorescence tagged proteins in dendritic spines of cultured hippocampal neurons.

FRAP has been used to quantify the mobility of GFP-tagged proteins. Using a strong excitation laser, the fluorescence of a GFP-tagged protein is bleached in the region of interest. The fluorescence of the region recovers when the unbleached GFP-tagged protein from outside of the region diffuses into the region of interest.

Differential localization of SAP102 and PSD-95 is revealed in hippocampal spines using super-resolution light microscopy.

Synapse-associated protein 102 (SAP102) and postsynaptic density 95 (PSD-95) are two major cytoskeleton proteins in the postsynaptic density (PSD). Both of them belong to the membrane-associated guanylate kinase (MAGUK) family, which clusters and anchors glutamate receptors and other proteins at synapses. In our previous study, we found that SAP102 and PSD-95 have different distributions, using combined light/electron microscopy (LM/EM) methods.

MAGUKs, synaptic development, and synaptic plasticity.

MAGUKs are proteins that act as key scaffolds in surface complexes containing receptors, adhesion proteins, and various signaling molecules. These complexes evolved prior to the appearance of multicellular animals and play key roles in cell-cell intercommunication. A major example of this is the neuronal synapse, which contains several presynaptic and postsynaptic MAGUKs including PSD-95, SAP102, SAP97, PSD-93, CASK, and MAGIs.

SAP102 is a highly mobile MAGUK in spines.

Membrane-associated guanylate kinases (MAGUKs), which are essential proteins in the postsynaptic density (PSD), cluster and anchor glutamate receptors and other proteins at synapses. The MAGUK family includes PSD-95, PSD-93, SAP102, and SAP97. Individual family members can compensate for one another in their ability to recruit and retain receptors at the postsynaptic membrane as shown through deletion and knock-down studies.

Phorbol-induced surface expression of NR2A subunit homologues in HEK293 cells.

Aim: N-methyl-D-aspartate receptors (NMDAR) are heteromeric complexes primarily assembled from NR1 and NR2 subunits. In normal conditions, NR2 subunits assemble into homodimers in the endoplasmic reticulum (ER). These homodimers remain in the ER until they coassemble with NR1 dimers and are trafficked to the cell surface.

[Construction of expression vectors for C-terminally deleted NR2B subunit mutants and their application in the study of assembling of NMDA receptors].

Objective: To investigate the role of NR2B subunit C-terminus in assembling and surface expression of NMDA receptor subtype composed of NR1-1a/NR2B subunits.

Methods: Eight vectors NR2BDelta1-Delta8) expressing GFP-tagged NR2B subunit mutants with various deletion in the carboxyl-terminal region were generated by conventional molecular cloning techniques. Each of these vectors was transfected alone, or co-transfected with NR1-1a into HEK293 cells.

[Surface expression of NMDA receptors composed of NR1 subunit and NR2A subunit mutants with partially deleted C-terminus in HEK293 cells].

Objective: To examine the potential function of NMDA receptor NR2A subunit C-terminus in assembling and surface expression of the receptor in HEK293 cells.

Methods: Five vectors GFP- NR2ADeltaC1- DeltaC5 were constructed for expressing N-terminally GFP-tagged NR2A with C-terminal deletion at different regions by using conventional techniques of molecular cloning. The deleted region for NR2ADeltaC1-Delta C5 was 897L-1017S, 1024D-1142P, 1149D-1347G, 1354S-1464V, and 897L-1464V.