PSD-95 family MAGUKs are essential for anchoring AMPA and NMDA receptor complexes at the postsynaptic density

Chen, Xiaobing; Levy, Jay M.; Hou, Austin; Winters, Christine A.; Azzam, Rita; Sousa, Alioscka A.; Leapman, Richard D.; Nicoll, Roger A.; Reese, Thomas S.

doi:10.1073/pnas.1517045112

Cited by 252 publications

(220 citation statements)

References 87 publications

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“…EM tomography suggested that the NMDAR nanodomains contained NMDAR-PSD95 complexes in 1:2 stoichiometry and that other PSD regions with AMPARs, but without NMDARs, contained extended PSD95-AMPAR complexes in a 1:1 stoichiometry (2). When PSD95 was acutely knocked down, AMPAR nanodomains were lost, whereas NMDAR nanodomains in PSDs were relatively preserved, consistent with the NMDAR nanodomains being more stable (1,41). The presence of separate AMPAR and NMDAR domains in PSDs provides a simple explanation for why PSD95 palmitoylation and depalmitoylation are different at AMPARs and NMDARs.…”

Section: Discussionmentioning

confidence: 81%

Palmitoylation regulates glutamate receptor distributions in postsynaptic densities through control of PSD95 conformation and orientation

Jeyifous

Lin

Chen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Postsynaptic density protein 95 (PSD95) and synapse-associated protein 97 (SAP97) are homologous scaffold proteins with different N-terminal domains, possessing either a palmitoylation site (PSD95) or an L27 domain (SAP97). Here, we measured PSD95 and SAP97 conformation in vitro and in postsynaptic densities (PSDs) using FRET and EM, and examined how conformation regulated interactions with AMPA-type and NMDA-type glutamate receptors (AMPARs/NMDARs). Palmitoylation of PSD95 changed its conformation from a compact to an extended configuration. PSD95 associated with AMPARs (via transmembrane AMPAR regulatory protein subunits) or NMDARs [via glutamate ionotropic receptor NMDA-type subunit 2B (GluN2B) subunits] only in its palmitoylated and extended conformation. In contrast, in its extended conformation, SAP97 associates with NMDARs, but not with AMPARs. Within PSDs, PSD95 and SAP97 were largely in the extended conformation, but had different orientations. PSD95 oriented perpendicular to the PSD membrane, with its palmitoylated, N-terminal domain at the membrane. SAP97 oriented parallel to the PSD membrane, likely as a dimer through interactions of its N-terminal L27 domain. Changing PSD95 palmitoylation in PSDs altered PSD95 and AMPAR levels but did not affect NMDAR levels. These results indicate that in PSDs, PSD95 palmitoylation, conformation, and its interactions are dynamic when associated with AMPARs and more stable when associated with NMDARs. Altogether, our results are consistent with differential regulation of PSD95 palmitoylation in PSDs resulting from the clustering of palmitoylating and depalmitoylating enzymes into AMPAR nanodomains segregated away from NMDAR nanodomains.P ostsynaptic densities (PSDs) at glutamatergic synapses organize and hold NMDA receptors (NMDARs), AMPA receptors (AMPARs), and other signaling molecules in place, apposed to sites of neurotransmitter release. Just below the PSD plasma membrane lies a latticework of vertical and parallel filaments that provides a structural scaffold to stabilize synaptic signaling molecules within PSDs (1, 2). Postsynaptic density protein 95 (PSD95) and synapse-associated protein 97 (SAP97) are members of a family of membrane-associated guanylate kinases (MAGUKs) (3). PSD95 is the most abundant scaffold protein in adult synapses, with ∼300 PSD95 molecules (2.3% of the mass of the PSD) in the average PSD, and is part of the lattice forming the core of the PSD (4). SAP97 is also a component of the PSD lattice. Estimates of its PSD copy numbers range from 90 SAP97 molecules per average PSD [0.9% of the mass of the PSD (4)] to lower values (5). As MAGUKs, PSD95 and SAP97 share a series of highly homologous protein-interacting domains but diverge at their N-terminal domains, which affects their trafficking into and out of the PSD, as well as interactions with AMPARs and NMDARs (3, 6, 7). The SAP97β-isoform, like almost all SAP97 molecules, contains an N-terminal L27 domain that interacts with other L27 domaincontaining proteins, particularly with a differ...

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Section: Discussionmentioning

confidence: 81%

Palmitoylation regulates glutamate receptor distributions in postsynaptic densities through control of PSD95 conformation and orientation

Jeyifous

Lin

Chen

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…For n channels, the probability of at least one receptor opening is the same as the complement of the probability that all receptors fail to open (1− P o ) n , or P event = 1−(1− P o ) n . Although we do not know how many NMDARs are present on each MSN spine, studies from CA1 pyramidal neurons suggest between 1 and 10 functional NMDARs at the synapse (Chen et al, 2015; Nimchinsky et al, 2004; Spruston et al, 1995). Thus the likelihood of observing an event, given a steady-state concentration of 25 nM glutamate, is estimated to be between 0.004 and 0.041 (for n = 1 or n = 10, respectively).…”

Section: Discussionmentioning

confidence: 90%

Extracellular Glutamate in the Nucleus Accumbens Is Nanomolar in Both Synaptic and Non-synaptic Compartments

Chiu

Jahr

2017

Cell Reports

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Summary In the CNS, glutamate is both phasically and tonically released into the extracellular space, and must be removed by excitatory amino acid transporters (EAATs) to prevent excitotoxic accumulation. There remains uncertainty, however, regarding the functional steady-state concentration, with estimates ranging from tens of nanomolar to tens of micromolar. Efforts to reconcile these disparate values have led to a hypothesis that the extracellular space is comprised of distinct compartments in which basal glutamate concentrations are maintained independently. We used electrophysiology and two-photon Ca2+ imaging to test this hypothesis in the nucleus accumbens (NAc), where it has been proposed that micromolar extracellular glutamate is necessary for normal function. We found that the average concentration of synaptic glutamate is nanomolar, in agreement with previous electrophysiological estimates. Furthermore, this held true when glutamate uptake was inhibited, indicating that extracellular glutamate is not compartmentalized by EAATs.

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“…Extending these findings, we performed immuno-EM analysis from mouse primary neuronal cultures, as these preparations are largely devoid of non-neuronal cell types and can provide higher resolution analysis 20,21 . No immunogold label was observed in samples treated with secondary gold-conjugated antibodies alone (Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

A mammalian nervous-system-specific plasma membrane proteasome complex that modulates neuronal function

Ramachandran

Margolis

2017

Nat Struct Mol Biol

120

133

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In the nervous system, rapidly occurring processes such as neuronal transmission and calcium signaling are affected by short-term inhibition of proteasome function. It remains unclear how proteasomes can acutely regulate such processes, as this is inconsistent with their canonical role in proteostasis. Here, we made the discovery of a mammalian nervous system-specific membrane proteasome complex that directly and rapidly modulates neuronal function by degrading intracellular proteins into extracellular peptides that can stimulate neuronal signaling. This proteasome complex is tightly associated with neuronal plasma membranes, exposed to the extracellular space, and catalytically active. Selective inhibition of this membrane proteasome complex by a cell-impermeable proteasome inhibitor blocked extracellular peptide production and attenuated neuronal activity-induced calcium signaling. Moreover, membrane proteasome-derived peptides are sufficient to induce neuronal calcium signaling. Our discoveries challenge the prevailing notion that proteasomes primarily function to maintain proteostasis, and highlight a form of neuronal communication through a membrane proteasome complex.

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PSD-95 family MAGUKs are essential for anchoring AMPA and NMDA receptor complexes at the postsynaptic density

Cited by 252 publications

References 87 publications

Palmitoylation regulates glutamate receptor distributions in postsynaptic densities through control of PSD95 conformation and orientation

Palmitoylation regulates glutamate receptor distributions in postsynaptic densities through control of PSD95 conformation and orientation

Extracellular Glutamate in the Nucleus Accumbens Is Nanomolar in Both Synaptic and Non-synaptic Compartments

A mammalian nervous-system-specific plasma membrane proteasome complex that modulates neuronal function

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