SAP97 Is Associated with the α-Amino-3-hydroxy-5-methylisoxazole-4-propionic Acid Receptor GluR1 Subunit

Abstract

Rapid glutamatergic synaptic transmission is mediated by ionotropic glutamate receptors and depends on their precise localization at postsynaptic membranes opposing the presynaptic neurotransmitter release sites. Postsynaptic localization of N-methyl-D-aspartatetype glutamate receptors may be mediated by the synapse-associated proteins (SAPs) SAP90, SAP102, and chapsyn-110. SAPs contain three PDZ domains that can interact with the C termini of proteins such as N-methyl-D-aspartate receptor subunits that carry a serine or threonine at the -2 position and a valine, isoleucine, or leucine at the very C terminus (position 0). We now show that SAP97, a SAP whose function at the synapse has been unclear, is associated with ␣-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptors. AMPA receptors are probably tetramers and are formed by two or more of the four AMPA receptor subunits GluR1-4. GluR1 possesses a C-terminal consensus sequence for interactions with PDZ domains of SAPs. SAP97 was present in AMPA receptor complexes immunoprecipitated from detergent extracts of rat brain. After treatment of rat brain membrane fractions with the cross-linker dithiobis(succinimidylpropionate) and solubilization with sodium dodecylsulfate, SAP97 was associated with GluR1 but not GluR2 or GluR3. In vitro experiments with recombinant proteins indicate that SAP97 specifically associates with the C terminus of GluR1 but not other AMPA receptor subunits. Our findings suggest that SAP97 may be involved in localizing AMPA receptors at postsynaptic sites through its interaction with the GluR1 subunit.The prevailing excitatory neurotransmitter in the mammalian brain is glutamate (1, 2). Upon its release from presynaptic sites, this neurotransmitter binds to ionotropic glutamate receptors that mediate rapid excitatory synaptic transmission in the mammalian brain (1, 2). Several immuno-electron microscopic studies have demonstrated that ionotropic glutamate receptors are clustered at postsynaptic sites of excitatory synapses (3-5). Two major glutamate receptor families exist, namely N-methyl-D-aspartate (NMDA) 1 receptors, which mediate Ca 2ϩ influx, and non-NMDA receptors, which are usually not Ca 2ϩ -permeable (1, 2, 6). Non-NMDA receptors are further divided into ␣-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors and kainate receptors. At low frequency, synaptic transmission normally depends nearly exclusively upon AMPA receptors. On the other hand, kainate and NMDA receptors require higher frequencies for activation. NMDA receptor-mediated Ca 2ϩ influx is necessary for different forms of synaptic plasticity, such as long term potentiation (1,7,8). At different synapses in the hippocampus and in other brain areas, a few bursts of high frequency electric stimulation that activate NMDA receptors induce a long lasting increase in synaptic transmission, the hallmark of long term potentiation.Glutamate receptors are thought to be heterotetramers consisting of homologous subunits (39). AMPA recepto...

Selective reduction of a PDZ protein, SAP‐97, in the prefrontal cortex of patients with chronic schizophrenia

“…For example, Pick 1 and GRIP1, which both interact with the C-terminal terminal of the AMPA receptor subunits, GluR2/3, are involved in their internalization and synaptic stabilization (Dong et al 1999;Dev et al 1999;Chung et al 2000;Daw et al 2000;El-Husseini et al 2000;Osten et al 2000). SAP97 interacts with the AMPA receptor subunit GluR1 and stabilizes receptor protein expression at postsynaptic sites (Leonard et al 1998;Jourdi et al 2001;Sans et al 2001). Among the various PDZ proteins, levels of the PDZ protein SAP97 were specifically decreased in the prefrontal cortex of schizophrenics (59%, p < 0.05) compared to those in controls in the present experiment.…”

“…Similarly, a group of PDZ molecules interacting with a-amino-3-hydroxy-5-methyl-4-isoxzolepropionic acid (AMPA)-type glutamate receptors has been identified. SAP97 associates with the C-terminal motif of glutamate receptor 1 (GluR1) (Leonard et al 1998;Sans et al 2001) and GRIP, AMPA receptorbinding protein (ABP), stargazin and Pick1 bind to the C-terminal motif of GluR2/3 (Dong et al 1997;Srivastava et al 1998;Dong et al 1999;Xia et al 1999;Chen et al 2000). The PDZ domain of PSD-95 and other proteins also interacts with a variety of other signaling molecules at synapses (O'Brien et al 1998;Nagano et al 1998;Fanning and Anderson 1999).…”

Distinct molecular mechanisms and divergent endocytotic pathways of AMPA receptor internalization

“…To date, studies of AMPA receptor trafficking have focused on the presence or absence of AMPA receptors at synaptic sites, or on internal versus surface expression [3][4][5][6][7][8][9][10][15][16][17][18] . However, the multiple subunits of AMPA receptors interact differentially with diverse cytoplasmic proteins including GRIP/ABP, PICK1, NSF and SAP97, all of which have been implicated in synaptic targeting of AMPA receptors 3,5,17,[19][20][21][22][23][24][25][26][27] . The diversity of AMPA receptor protein interactions, and their regulation by phosphorylation 28 , suggest that AMPA receptor trafficking is likely to be regulated by varied mechanisms.…”

Elevated GRIA1 mRNA expression in Layer II/III and V pyramidal cells of the DLPFC in schizophrenia

“…Due to limitations in resource material, we were unable to assess alterations in the mRNAs encoding glutamate receptor trafficking proteins in these cells. Given the interaction between GRIA1 and synapse associated protein 97 (SAP97) (Leonard et al, 1998) and the presumed role of SAP97 in trafficking of GRIA1 to the membrane (Rumbaugh et al, 2003;Sans et al, 2001), future studies investigating the expression of SAP97 and other binding proteins may provide additional insight into biochemical correlates of increased GRIA1 subunit expression in Layer III and V pyramidal in the DLPFC.…”