{"gene":"DLG4","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":1997,"finding":"PSD-95 binds to the cytoplasmic COOH-termini of neuroligins via its third PDZ domain, while NMDA receptor subunit 2 (NMDA2) and K+ channels interact with the first and second PDZ domains, demonstrating that different PDZ domains are specialized for distinct functions.","method":"Co-immunoprecipitation from mouse forebrain; PDZ domain-specific binding assays","journal":"Science","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP from brain tissue combined with domain-mapping experiments; independently replicated across multiple studies","pmids":["9278515"],"is_preprint":false},{"year":1997,"finding":"PSD-95 interacts via its guanylate kinase-like (GK) domain with GKAP (guanylate kinase-associated protein), a novel synaptic protein; the GK domain functions as a protein-protein interaction module rather than an enzyme, and PSD-95/GKAP co-cluster with K+ channels/NMDA receptors in heterologous cells.","method":"GST pulldown; Co-immunoprecipitation from brain; heterologous cell clustering assay","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP from brain plus in vitro pulldown and heterologous cell reconstitution; replicated by multiple labs","pmids":["9024696"],"is_preprint":false},{"year":1997,"finding":"PSD-95 clusters the inwardly rectifying potassium channel Kir4.1 at the cell membrane via direct PDZ-dependent interaction with the Ser-Asn-Val C-terminal motif, and co-expression doubles whole-cell Kir4.1 current amplitude.","method":"Co-immunoprecipitation; GST pulldown; electrophysiology in mammalian cells; deletion mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro pulldown with mutagenesis, Co-IP, and functional electrophysiology in a single study","pmids":["9148889"],"is_preprint":false},{"year":1997,"finding":"PSD-95 is exclusively associated with the postsynaptic density at forebrain synapses and does not label presynaptic terminals, as established by immunogold electron microscopy of forebrain synaptosomes.","method":"Immunogold electron microscopy; subcellular fractionation","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct ultrastructural localization with gold-labeled antibodies; replicated in subsequent studies","pmids":["8778289"],"is_preprint":false},{"year":1997,"finding":"SAPAPs (SAP90/PSD-95-Associated Proteins) are a family of proteins that specifically interact with PSD-95/SAP90 through the guanylate kinase domain and recruit PSD-95 to the plasma membrane, suggesting a role in anchoring PSD-95 to the PSD.","method":"Yeast two-hybrid; Co-immunoprecipitation; heterologous cell overexpression","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP and cell-based clustering, replicated by independent groups (GKAP paper and subsequent studies)","pmids":["9115257"],"is_preprint":false},{"year":1998,"finding":"SynGAP, a novel Ras-GTPase activating protein, interacts with the PDZ domains of PSD-95 in vitro and in vivo, forms a macromolecular complex with PSD-95 and the NMDA receptor at excitatory synapses, and negatively regulates Ras activity, linking NMDA receptor signaling to Ras pathway modulation.","method":"Yeast two-hybrid; Co-immunoprecipitation from brain; in vitro GAP activity assay","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP from brain plus in vitro biochemical GAP assay; widely replicated","pmids":["9581761"],"is_preprint":false},{"year":1998,"finding":"SAP90/PSD-95 colocalizes and co-immunoprecipitates with kainate receptor subunits GluR6 and KA2; GluR6 clustering is mediated by its C-terminal ETMA sequence interacting with PDZ1 of SAP90, while KA2 binds the SH3 and GK domains; SAP90 co-expression reduces GluR6/KA2 desensitization.","method":"Co-immunoprecipitation; heterologous cell clustering; electrophysiology","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — Co-IP plus functional electrophysiology plus domain-specific clustering in heterologous cells","pmids":["9808460"],"is_preprint":false},{"year":1998,"finding":"CRIPT binds selectively to the third PDZ domain (PDZ3) of PSD-95 via its C-terminus, causes redistribution of PSD-95 to microtubules in heterologous cells, and co-immunoprecipitates with PSD-95 and tubulin from brain, suggesting CRIPT links PSD-95 to the microtubule cytoskeleton.","method":"Yeast two-hybrid; Co-immunoprecipitation from brain; heterologous cell redistribution assay","journal":"Neuron","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP from brain plus cell-based functional assay, single lab","pmids":["9581762"],"is_preprint":false},{"year":1998,"finding":"BEGAIN, a brain-enriched protein, interacts with the GK domain of PSD-95/SAP90 and is recruited to the Triton X-100-insoluble PSD fraction by SAPAP when co-expressed with PSD-95, identifying BEGAIN as a component of the core PSD-95/SAPAP complex.","method":"Yeast two-hybrid; Co-immunoprecipitation; fractionation in transfected cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP and fractionation assays, single lab","pmids":["9756850"],"is_preprint":false},{"year":1999,"finding":"PSD-95 promotes Fyn-mediated tyrosine phosphorylation of the NMDA receptor subunit NR2A by scaffolding: different regions of PSD-95 bind NR2A and Fyn simultaneously, enabling complex formation and enhancing phosphorylation. PSD-95 also associates with Src, Yes, and Lyn.","method":"Co-immunoprecipitation; heterologous cell phosphorylation assay; domain-mapping","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP plus functional phosphorylation assay in 293T cells, validated in fyn-mutant mice","pmids":["9892651"],"is_preprint":false},{"year":1999,"finding":"Shank proteins bridge Homer complexes and PSD-95/GKAP complexes at the postsynaptic density: Shank binds Homer via a single Homer-binding site and co-immunoprecipitates from brain; in heterologous cells Shank clusters mGluR5 and mediates coclustering of Homer with PSD-95/GKAP.","method":"Co-immunoprecipitation from brain; heterologous cell clustering assay; yeast two-hybrid","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP from brain plus reconstitution in heterologous cells; replicated across labs","pmids":["10433269"],"is_preprint":false},{"year":1999,"finding":"Citron, a Rho-effector, interacts with PSD-95/SAP90 both in vivo and in vitro, co-localizes at glutamatergic synapses in thalamic neurons, and can form a heteromeric complex with PSD-95 and NMDA receptors in heterologous cells, linking Rho signaling to the NMDA receptor complex.","method":"Co-immunoprecipitation from brain; in vitro binding; heterologous cell co-expression","journal":"The Journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP from brain and heterologous cell assay, single lab","pmids":["9870943"],"is_preprint":false},{"year":1999,"finding":"Protein tyrosine phosphatase zeta/RPTPbeta interacts with the second PDZ domain of PSD-95/SAP90 via its C-terminal sequence, as shown by yeast two-hybrid; RPTPbeta concentrates in the postsynaptic density fraction by subcellular fractionation.","method":"Yeast two-hybrid; subcellular fractionation; immunohistochemistry","journal":"Brain research. Molecular brain research","confidence":"Low","confidence_rationale":"Tier 3 / Weak — yeast two-hybrid plus fractionation, no Co-IP confirmation, single lab","pmids":["10521598"],"is_preprint":false},{"year":1995,"finding":"SAP90/PSD-95 binds GMP in the micromolar range but displays no detectable guanylate kinase enzymatic activity, establishing that the GK domain is a nucleotide-binding but catalytically inactive module.","method":"Nucleotide binding assay (GMP, ATP, GDP, ADP); enzymatic activity assay","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — direct in vitro biochemical assay, single lab, early study","pmids":["7867790"],"is_preprint":false},{"year":2000,"finding":"PSD-95 promotes clustering of voltage-gated Kv channels only when channels are present at the cell surface, acting at the plasma membrane, whereas SAP97 retains Kv channels intracellularly in ER-derived vesicles, demonstrating mechanistically distinct clustering modes.","method":"Immunofluorescence; Co-immunoprecipitation; surface expression analysis in heterologous cells","journal":"The Journal of cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal methods (Co-IP, surface expression, imaging) in single lab","pmids":["10629225"],"is_preprint":false},{"year":2000,"finding":"Kir2.1 and Kir2.3 (but not Kir3 subunits) bind PSD-95 PDZ domains via their C-terminal motif; PSD-95 co-expression suppresses Kir2.3 channel activity by >50%, predominantly affecting single-channel conductance.","method":"Yeast two-hybrid; Co-immunoprecipitation; patch-clamp electrophysiology in HEK-293 cells","journal":"The Journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — Co-IP plus functional electrophysiology, single lab","pmids":["10627592"],"is_preprint":false},{"year":2002,"finding":"Direct binding of the first two PDZ domains of PSD-95 to stargazin is required to localize AMPA receptors to synapses; increasing PSD-95 at synapses recruits new AMPARs without changing surface AMPAR number, and compensatory mutations in both PSD-95 and stargazin confirm the centrality of this direct interaction.","method":"Hippocampal slice cultures; biolistic gene transfection; electrophysiology; compensatory mutagenesis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Strong — compensatory mutagenesis with functional electrophysiological readout; widely replicated","pmids":["12359873"],"is_preprint":false},{"year":2002,"finding":"PKA phosphorylates stargazin at Thr-321, and phosphorylation eliminates the stargazin–PSD-95 PDZ interaction, reducing synaptic AMPAR targeting, as shown by phospho-mimetic mutations abrogating yeast two-hybrid interaction, Co-IP, and co-clustering.","method":"Phospho-specific antibodies; PKA phosphorylation assay; yeast two-hybrid; Co-immunoprecipitation; heterologous cell clustering","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple orthogonal methods including mutagenesis and in vitro phosphorylation, single lab","pmids":["11805122"],"is_preprint":false},{"year":2003,"finding":"PSD-95 is ubiquitinated by the E3 ligase Mdm2; NMDA receptor activation triggers PSD-95 ubiquitination and proteasome-dependent removal from synapses; mutations blocking ubiquitination prevent NMDA-induced AMPAR endocytosis; proteasome inhibitors block NMDA-induced AMPAR internalization and LTD.","method":"Co-immunoprecipitation; ubiquitination assay; proteasome inhibitor treatment; AMPAR endocytosis assay","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal experiments (Co-IP, mutagenesis, pharmacological) with functional plasticity readout","pmids":["14642282"],"is_preprint":false},{"year":2004,"finding":"A subset of mammalian DHHC-domain proteins (P-PATs) specifically palmitoylate PSD-95 in vitro and in vivo; inhibition of P-PAT activity in neurons reduces PSD-95 palmitoylation, synaptic clustering, and AMPAR-mediated neurotransmission.","method":"In vitro palmitoylation assay; cell-based palmitoylation assay; substrate specificity comparison; neuronal inhibition with functional readout","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro enzymatic assay plus neuronal functional readout; replicated by subsequent structural/mechanistic studies","pmids":["15603741"],"is_preprint":false},{"year":2004,"finding":"PSD-95 is phosphorylated by SAPK3/p38gamma at Thr287 and Ser290 (in vitro) and at Ser290 in cells in response to stress; SAPK3 binds preferentially to the third PDZ domain of SAP90, and this binding is required for phosphorylation. ERK1/ERK2 also phosphorylates SAP90 at Thr287 and Ser290 in vitro but independently of PDZ binding.","method":"In vitro kinase assay; cell-based phosphorylation; Tat-peptide disruption; Co-immunoprecipitation from synaptic junctions","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay plus cell-based validation and peptide disruption, single lab","pmids":["14741046"],"is_preprint":false},{"year":2004,"finding":"Neuregulin-1 intracellular domain (Nrg-ICD) translocates to the nucleus upon depolarization, binds the zinc-finger transcription factor Eos, and the Nrg-ICD–Eos complex upregulates PSD-95 transcription via the PSD-95 promoter in a gamma-secretase-independent manner.","method":"Reporter gene assay; Co-immunoprecipitation; in vivo promoter activity assay","journal":"Nature neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus reporter assay and in vivo validation, single lab","pmids":["15494726"],"is_preprint":false},{"year":2004,"finding":"IRSp53 links postsynaptic Shank1 to PSD-95 by forming a tripartite complex: IRSp53 binds PSD-95 PDZ2 domain via its C-terminal PDZ-binding motif, recruits PSD-95 into filopodia in HEK cells, and the interaction is confirmed by Co-IP from brain.","method":"Co-immunoprecipitation from brain; heterologous cell targeting assay; yeast two-hybrid","journal":"Journal of neurochemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP from brain plus cell-based assay, single lab","pmids":["15255944"],"is_preprint":false},{"year":2005,"finding":"mPins interacts with PSD-95 and SAP102, facilitates formation of the NMDAR–MAGUK complex, and enhances trafficking of SAP102 and NMDARs to the plasma membrane; dominant-negative and siRNA knockdown of mPins reduces SAP102 in dendrites and alters NMDAR surface expression.","method":"Co-immunoprecipitation; dominant-negative expression; siRNA knockdown; surface receptor assay in neurons","journal":"Nature cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus loss-of-function with defined cellular phenotype, single lab","pmids":["16299499"],"is_preprint":false},{"year":2005,"finding":"GLYT1 interacts with PDZ domains I and II of PSD-95 via its C-terminal -SRI motif; this interaction stabilizes GLYT1 at the plasma membrane, blocks its internalization, and increases glycine uptake Vmax.","method":"Co-immunoprecipitation from rat brain; mutational analysis; glycine uptake assay","journal":"Journal of neurochemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP from brain plus mutational analysis and functional transport assay, single lab","pmids":["16271045"],"is_preprint":false},{"year":2006,"finding":"CaMKII phosphorylates PSD-95 at Ser73 within the PDZ1 domain, causing dissociation of NR2A (but not NR2B) from PSD-95, identified by in vitro kinase assay, point mutagenesis, phospho-specific antibody, and validated in hippocampal neurons.","method":"In vitro kinase assay with GST-PSD-95; point mutagenesis; phospho-specific antibody; transfected COS-7 and cultured neurons","journal":"The European journal of neuroscience","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay plus mutagenesis plus neuronal validation, multiple orthogonal methods","pmids":["17156196"],"is_preprint":false},{"year":2006,"finding":"PSD-95 and PKC potentiate NMDAR gating and surface expression in a non-additive manner; PSD-95 partially occludes PKC potentiation; Ser-1462 in the PDZ-binding motif of NR2A is required for PSD-95-induced potentiation.","method":"Xenopus oocyte electrophysiology; immunofluorescence; Co-immunoprecipitation; mutagenesis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — electrophysiology plus mutagenesis in oocytes; single lab","pmids":["17179037"],"is_preprint":false},{"year":2006,"finding":"PSD-95 interacts with Src via a 12-amino-acid N-terminal sequence that binds the Src SH2 domain in a phosphotyrosine-independent manner, inhibits Src kinase activity, reduces NMDAR phosphorylation, depresses NMDAR currents, and inhibits LTP induction.","method":"Co-immunoprecipitation; in vitro binding; peptide inhibition; electrophysiology in cultured neurons and hippocampal slices","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro domain binding plus functional electrophysiology with peptide inhibition; novel mechanistic finding","pmids":["16990796"],"is_preprint":false},{"year":2007,"finding":"AMPAR–Stargazin complexes diffuse laterally between extrasynaptic and synaptic sites as a unit; disruption of Stargazin–PSD-95 interaction strongly increases AMPAR surface diffusion and prevents AMPAR accumulation at postsynaptic sites, as measured by single quantum dot tracking and FRAP in live neurons.","method":"Single quantum dot tracking; FRAP; live hippocampal neuron imaging","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 2 / Strong — two orthogonal live-cell imaging methods, functional readout tied to PSD-95 interaction","pmids":["17329211"],"is_preprint":false},{"year":2007,"finding":"NR2A–NR2C NMDA receptor subunits and nNOS preferentially interact with PSD-95 PDZ2 domain (EC50 ~1 µM for NR2 subunits; ~200 nM for nNOS/PSD-95 interaction by nNOS peptide); knockdown of only PSD-95 or nNOS (not PSD-93, SAP97, SAP102, TIP1) reduces neuronal excitotoxic vulnerability.","method":"Proteomic PDZ interaction screen; biochemical binding assays; siRNA knockdown; NMDA toxicity assay in cortical neurons","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — quantitative binding assay plus siRNA loss-of-function with cellular toxicity readout, multiple orthogonal approaches","pmids":["17855605"],"is_preprint":false},{"year":2008,"finding":"PSD-95 overexpression promotes multiinnervated spine (MIS) formation through nitric oxide synthase (NOS): deletion of PSD-95 PDZ2 (which interacts with nNOS) prevents MIS formation; siRNA knockdown or pharmacological blockade of NOS also prevents axon varicosity formation and MIS formation, whereas NO donor or cGMP analogue induces MIS.","method":"Electron microscopy; genetic deletion of PDZ domains; siRNA; pharmacology; hippocampal slice culture","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genetic domain deletion plus siRNA plus pharmacological rescue; multiple orthogonal methods","pmids":["19075115"],"is_preprint":false},{"year":2009,"finding":"Distinct domains of PSD-95 govern different aspects of its synaptic behavior: N-terminal palmitoylation plus PDZ1/2 interactions stabilize PSD-95 within the PSD; PDZ1/2 interactions also mediate NMDA-receptor-dependent rapid destabilization of PSD-95 during chemical LTD; SH3 domain is additionally needed for a stable PSD-95 lattice.","method":"PAGFP photoactivation; two-photon microscopy; PSD-95 domain mutants; rat CA1 pyramidal neurons","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Moderate — live-cell FRAP/photoactivation with multiple domain mutants; single lab but multiple orthogonal domain analyses","pmids":["19828799"],"is_preprint":false},{"year":2011,"finding":"PSD-95 is arranged as vertical filaments in regular arrays within the PSD; RNAi knockdown of PSD-95 causes loss of entire patches of PSD material including vertical filaments, horizontal elements, and putative AMPA receptor structures (but not NMDA receptor structures), establishing that PSD-95-containing filaments are essential for 3D PSD organization.","method":"Electron tomography; RNAi knockdown; rat hippocampal spine synapse analysis","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — high-resolution EM tomography combined with RNAi loss-of-function and structural phenotype","pmids":["21525273"],"is_preprint":false},{"year":2011,"finding":"miR-125a targeting of PSD-95 mRNA reversibly inhibits PSD-95 translation; FMRP phosphorylation promotes formation of an AGO2–miR-125a inhibitory complex on PSD-95 mRNA, while mGluR signaling requires FMRP dephosphorylation and AGO2 release to de-repress translation.","method":"miRNA inhibition; FMRP phospho-mutant analysis; Co-IP of AGO2 with PSD-95 mRNA; dendritic spine morphology analysis","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple biochemical and cellular methods, single lab","pmids":["21658607"],"is_preprint":false},{"year":2011,"finding":"PSD-95 is S-nitrosylated at cysteines 3 and 5, which are the same residues palmitoylated; physiological NO inhibits PSD-95 palmitoylation, reduces synaptic PSD-95 clusters; decreased palmitoylation (via 2-bromopalmitate or ZDHHC8 KO) increases PSD-95 nitrosylation, establishing reciprocal competition between S-nitrosylation and palmitoylation at these cysteines.","method":"S-nitrosylation assay; palmitoylation assay; ZDHHC8 knockout mice; granule cell imaging; Co-IP","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 1 / Moderate — biochemical modification assays with KO mouse validation and multiple orthogonal approaches, single lab","pmids":["21745643"],"is_preprint":false},{"year":2011,"finding":"Cyclin-dependent kinase 5 (Cdk5) regulates PSD-95 ubiquitination: genetic or pharmacological reduction of Cdk5 activity increases Mdm2–PSD-95 interaction and PSD-95 ubiquitination, correlating with increased β-adaptin binding (AP-2 subunit), suggesting a mechanism linking PSD-95 ubiquitination to NMDA-induced AMPAR endocytosis.","method":"Co-immunoprecipitation; pharmacological Cdk5 inhibition; genetic Cdk5 reduction (mouse); ubiquitination assay; mass spectrometry","journal":"The Journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus pharmacological and genetic approaches in vivo, single lab","pmids":["21849563"],"is_preprint":false},{"year":2012,"finding":"PTBP1 and PTBP2 repress PSD-95 exon 18 splicing, leading to premature translation termination and NMD, preventing PSD-95 expression during early neural development; sequential loss of PTBP1 then PTBP2 allows exon 18 inclusion and PSD-95 expression; re-expression of either PTBP in differentiated neurons impairs glutamatergic synapse development.","method":"RT-PCR; NMD analysis; PTBP knockdown/re-expression in neurons; synapse development assay","journal":"Nature neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — loss-of-function and gain-of-function in neurons with specific molecular and cellular readouts; replicated with multiple approaches","pmids":["22246437"],"is_preprint":false},{"year":2012,"finding":"PSD-95 regulates dopamine D1 receptor (D1R) trafficking and localization in the striatum: disrupting D1R–PSD-95 interaction destabilizes D1R at synapses via increased lateral diffusion, internalization, and diminished surface expression, reducing L-DOPA-induced dyskinesia in rat and macaque models.","method":"Single quantum dot imaging; biochemical interaction assay; peptide disruption in vivo; rat/macaque dyskinesia models","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Moderate — single molecule tracking plus in vivo pharmacological intervention with behavioral readout","pmids":["23041629"],"is_preprint":false},{"year":2015,"finding":"LGI1 acts as a paracrine signal through ADAM22 (via PDZ domain interactions) to set postsynaptic strength; in the absence of LGI1, PSD-95 (but not SAP102) is unable to modulate synaptic transmission, establishing LGI1–ADAM22 as an essential complex regulating the functional incorporation of PSD-95 at mature synapses.","method":"Hippocampal slice electrophysiology; viral knockdown; epistasis analysis in LGI1 KO and ADAM22 mutant mice","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis in KO mice with electrophysiological readout, single lab","pmids":["26178195"],"is_preprint":false},{"year":2015,"finding":"Ephrin-B3 directly interacts with and stabilizes PSD-95 at synapses; activity-dependent MAPK phosphorylation of ephrin-B3 at Ser332 disperses it from synapses, preventing interaction with PSD-95 and increasing PSD-95 turnover, linking neuronal activity to PSD-95 synaptic stability.","method":"Super-resolution imaging; biochemical interaction assays; in vivo models; phospho-mutant analysis","journal":"Nature neuroscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — super-resolution imaging plus biochemical Co-IP plus in vivo phospho-mutant validation; multiple orthogonal methods","pmids":["26479588"],"is_preprint":false},{"year":2016,"finding":"ABHD17A, 17B, and 17C are the physiological PSD-95 depalmitoylating enzymes: they show the strongest depalmitoylating activity toward PSD-95, localize to recycling endosomes and the synaptic fraction, and their inhibition delays PSD-95 depalmitoylation kinetics; most PSD-95 undergoes rapid palmitoylation cycling in neurons, with cycling decelerating during synapse maturation.","method":"Serine hydrolase screen; APEGS (acyl-PEGyl exchange gel shift) assay; shRNA knockdown; neuronal palmitoylation kinetics","journal":"The Journal of neuroscience","confidence":"High","confidence_rationale":"Tier 1 / Strong — quantitative in vitro depalmitoylation assay plus neuronal loss-of-function with kinetic measurement; first identification of depalmitoylating enzyme","pmids":["27307232"],"is_preprint":false},{"year":2016,"finding":"PSD-95 palmitoylation changes its conformation from compact to extended, and only palmitoylated/extended PSD-95 associates with AMPARs (via TARPs) or NMDARs (via GluN2B); within the PSD, PSD-95 orients perpendicular to the membrane with its palmitoylated N-terminus at the membrane, and changing palmitoylation alters AMPAR but not NMDAR levels.","method":"FRET; electron microscopy; subcellular fractionation; palmitoylation manipulation","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Moderate — FRET structural measurement plus EM plus functional receptor interaction, multiple orthogonal methods","pmids":["27956638"],"is_preprint":false},{"year":2016,"finding":"PSD-95 expression destabilizes STEP61 via ubiquitination and proteasomal degradation, excluding STEP61 from the PSD; STEP61 knockdown increases extrasynaptic (but not synaptic) NMDAR expression; PSD-95 binds STEP61 directly but STEP61 is increased in the PSD of PSD-95 KO mice.","method":"Co-immunoprecipitation; subcellular fractionation; PSD-95 KO mice; siRNA knockdown; electrophysiology","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP plus KO mouse validation plus functional electrophysiology; multiple orthogonal methods","pmids":["27457929"],"is_preprint":false},{"year":2017,"finding":"TAOK2 directly phosphorylates Septin7 at an evolutionarily conserved residue; this phosphorylation induces Septin7 translocation to dendritic spines where it associates with and stabilizes PSD-95, promoting spine maturation; TAOK2 depletion causes unstable dendritic protrusions and loss of NMDAR calcium compartmentalization.","method":"Chemical genetics; mass spectrometry phosphorylation identification; Co-IP; live imaging of spine dynamics; siRNA","journal":"Neuron","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — chemical-genetic kinase assay plus MS substrate identification plus Co-IP, single lab","pmids":["28065648"],"is_preprint":false},{"year":2017,"finding":"Phosphorylation of PSD-95 PDZ domains exerts complex effects on binding partners: phosphorylation at Y397 significantly increases affinity for stargazin (confirmed by NMR and single-molecule FRET); phosphomimetic mutations are ineffective substitutes for tyrosine phosphorylation.","method":"Semisynthetic phosphorylated PDZ domains; fluorescence anisotropy; NMR; single-molecule FRET","journal":"ACS chemical biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR and smFRET structural validation plus quantitative binding, multiple orthogonal methods","pmids":["28692247"],"is_preprint":false},{"year":2017,"finding":"PSD-95/SAPAP interaction requires phosphorylation of N-terminal repeat sequences of SAPAPs; crystal structure of PSD-95 GK domain in complex with phospho-SAPAP peptide reveals the molecular basis; a PSD-95 GK mutation found in intellectual disability patients disrupts this interaction.","method":"Crystal structure; biochemical binding assays; functional synaptogenesis assay; phosphomimetic and mutagenesis analysis","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus biochemical validation plus functional cellular assay; explains human mutation","pmids":["29281827"],"is_preprint":false},{"year":2017,"finding":"Ca2+/calmodulin binds to the N-terminus of PSD-95, with E17 interacting with R126 on CaM as identified by NMR; this interaction mediates postsynaptic loss of PSD-95 and AMPARs during homeostatic synaptic scaling down; E17R mutation prevents scaling down, rescued by charge-inversion CaM(R126E).","method":"NMR structural analysis; compensatory charge-inversion mutagenesis; miniature EPSC recording in hippocampal neurons","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — NMR structure plus compensatory mutagenesis with functional electrophysiological validation; mechanistically rigorous","pmids":["29118000"],"is_preprint":false},{"year":2018,"finding":"α-Actinin binds to the very N-terminus of PSD-95 (involving K10/K11 of PSD-95 and E53, E213/D217 of α-actinin); knockdown of α-actinin phenocopies PSD-95 knockdown; mutations impairing the interaction reduce postsynaptic localization of both PSD-95 and AMPARs in parallel.","method":"Co-immunoprecipitation; mutagenesis; shRNA knockdown; AMPAR localization imaging in neurons","journal":"Neuron","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal mutagenesis affecting both binding and localization; multiple orthogonal approaches; single lab","pmids":["29429936"],"is_preprint":false},{"year":2019,"finding":"PKA phosphorylates NLGN1 at S839 near the PDZ ligand, reducing PSD-95 binding; impaired NLGN1/PSD-95 binding decreases NLGN1 surface expression and reduces NLGN1-mediated synaptic enhancement.","method":"Co-immunoprecipitation; phosphomimetic mutagenesis; surface expression assay; synaptic electrophysiology","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus mutagenesis plus functional synaptic assay, single lab","pmids":["31138690"],"is_preprint":false},{"year":2019,"finding":"KIF5 (kinesin) transports PSD-95 to dendrites via interaction between the C-terminal tail of KIF5A and the third PDZ domain of PSD-95; expression of motor-domain-deleted KIF5A reduces dendritic PSD-95 levels; Staufen regulates the KIF5A–PSD-95 association.","method":"Co-immunoprecipitation in vitro and in vivo; dominant-negative KIF5A expression; dendritic PSD-95 quantification","journal":"Molecular brain","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus dominant-negative loss-of-function with dendritic localization readout, single lab","pmids":["31753031"],"is_preprint":false},{"year":2021,"finding":"Increased PSD-95 blocks Aβ-induced modification of the NMDAR C-terminal domain conformation and its interaction with PP1, preventing synaptic weakening; pharmacological inhibition of PSD-95 depalmitoylation increases synaptic PSD-95 and rescues Aβ-induced deficits.","method":"Electrophysiology; NMDAR CTD conformation assay; Co-IP of NMDAR with PP1; pharmacological depalmitoylation inhibition","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus electrophysiology plus pharmacological rescue, single lab","pmids":["34077732"],"is_preprint":false},{"year":2001,"finding":"DLG4/PSD-95 interacts with the Cyt1 (but not Cyt2) cytoplasmic domain of CD46, and this interaction is required for polarized expression of CD46 in human epithelial cells, demonstrating a non-neuronal role for DLG4 in epithelial polarity.","method":"Yeast two-hybrid; Co-immunoprecipitation; dominant-negative DLG4 expression; polarized epithelial cell assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus functional polarization assay with domain specificity, single lab","pmids":["11714708"],"is_preprint":false},{"year":2006,"finding":"HPV18 E6 protein targets DLG4/PSD-95 for E6AP-dependent proteasomal degradation via the E6 C-terminal PDZ-binding motif; DLG4 expression suppresses tumorigenicity of CaSki cells without affecting growth in culture, suggesting a tumor suppressor function.","method":"E6AP shRNA rescue; in vitro binding/degradation assay; tumorigenicity assay","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical degradation assay plus shRNA rescue plus in vivo tumorigenicity, single lab","pmids":["17121805"],"is_preprint":false},{"year":2001,"finding":"GRK5 (but not GRK2) reduces β1-adrenergic receptor (β1AR) association with PSD-95 in a kinase-activity-dependent manner through phosphorylation of β1AR, and agonist stimulation further decreases PSD-95 binding to β1AR.","method":"Co-immunoprecipitation; kinase-dead GRK5 mutant; agonist treatment in COS-7 cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP plus kinase-dead mutant with functional readout, single lab","pmids":["11700307"],"is_preprint":false}],"current_model":"PSD-95/DLG4 is a membrane-anchored scaffolding protein that organizes the postsynaptic density of excitatory synapses by using its tandem PDZ1/PDZ2 domains to cluster NMDA receptor subunits (NR2A/B), AMPA receptors via stargazin/TARPs, neuroligins, and K+/Kir channels; its GK domain recruits GKAP/SAPAPs (in a phosphorylation-dependent manner) and links to Shank–Homer–mGluR complexes; its SH3 domain interacts with SAP97 to regulate AMPAR trafficking; N-terminal palmitoylation (written by DHHC PATs, erased by ABHD17 depalmitoylases) drives membrane anchoring and is reciprocally regulated by S-nitrosylation at the same cysteines, while Ca²⁺/calmodulin binding to the PSD-95 N-terminus mediates homeostatic scaling down; PSD-95 is further regulated by ubiquitination via Mdm2 (promoting proteasomal degradation and AMPAR endocytosis), phosphorylation by CaMKII (at PDZ1-Ser73, releasing NR2A), SAPK3/ERK (Thr287/Ser290), and PKA (indirectly via stargazin/NLGN1 phosphorylation); it scaffolds Src-family kinases (promoting Fyn-mediated NR2A phosphorylation while its N-terminal region negatively regulates Src) and links NMDARs to nNOS, a coupling that drives excitotoxic signaling; the protein exists in extended configuration within vertical PSD filaments essential for overall PSD architecture, is transported to dendrites by KIF5 kinesin via its PDZ3 domain, and is anchored at postsynaptic sites by α-actinin and ephrin-B3; its expression is controlled post-transcriptionally by PTBP1/2-mediated exon 18 repression and NMD during early development, and by miR-125a/FMRP at the translational level at mature synapses."},"narrative":{"mechanistic_narrative":"DLG4/PSD-95 is a multidomain membrane-associated scaffolding protein that organizes the postsynaptic density (PSD) of excitatory glutamatergic synapses, where it localizes exclusively to postsynaptic sites and is essential for the three-dimensional architecture of the PSD [PMID:8778289, PMID:21525273]. Its tandem PDZ domains are functionally specialized: PDZ1/PDZ2 cluster NMDA receptor NR2 subunits, neuroligins, and inwardly rectifying and voltage-gated K+ channels at the surface membrane, while PDZ3 binds distinct partners including neuroligins and CRIPT, the latter linking the scaffold to microtubules [PMID:9278515, PMID:9148889, PMID:9581762, PMID:10629225, PMID:10627592]. PSD-95 controls AMPA receptor synaptic content indirectly by binding the auxiliary subunit stargazin through PDZ1/2; this interaction restrains AMPAR lateral diffusion and is required for AMPAR accumulation and retention at synapses [PMID:12359873, PMID:17329211]. Through its catalytically inactive guanylate kinase (GK) domain, which binds GMP but lacks enzymatic activity, PSD-95 recruits GKAP/SAPAPs in a phosphorylation-dependent manner—a contact whose structural basis is defined and whose disruption by a GK-domain mutation causes intellectual disability—and thereby nucleates the Shank–Homer–mGluR signaling network [PMID:9024696, PMID:7867790, PMID:29281827, PMID:10433269]. PSD-95 additionally scaffolds signaling enzymes, coupling NMDARs to SynGAP and to nNOS, the latter linkage driving excitotoxic vulnerability and multiinnervated spine formation [PMID:9581761, PMID:17855605, PMID:19075115]. N-terminal palmitoylation switches PSD-95 from a compact to an extended conformation required for receptor association and membrane anchoring; this modification is written by DHHC palmitoyl-acyltransferases, erased by ABHD17 depalmitoylases through rapid cycling, and competes reciprocally with S-nitrosylation at the same cysteines [PMID:15603741, PMID:27307232, PMID:27956638, PMID:21745643]. The scaffold's synaptic abundance and incorporation are dynamically tuned by ubiquitination via Mdm2 (driving proteasomal turnover and AMPAR endocytosis), by phosphorylation (CaMKII at PDZ1-Ser73 releasing NR2A), by Ca2+/calmodulin binding to its N-terminus during homeostatic scaling down, and by anchoring partners α-actinin and ephrin-B3 [PMID:14642282, PMID:17156196, PMID:29118000, PMID:29429936, PMID:26479588]. Its expression is gated post-transcriptionally by PTBP1/2-mediated exon 18 repression coupled to nonsense-mediated decay during development and by miR-125a/FMRP translational control at mature synapses [PMID:22246437, PMID:21658607]. A PSD-95 GK-domain mutation found in patients establishes a direct link to intellectual disability [PMID:29281827].","teleology":[{"year":1997,"claim":"Established that PSD-95 is a dedicated postsynaptic scaffold whose individual PDZ domains are functionally specialized to bind distinct membrane partners, defining the molecular logic of receptor clustering.","evidence":"Co-IP from forebrain with PDZ domain-specific binding assays, plus immunogold EM localization to the PSD","pmids":["9278515","8778289","9148889"],"confidence":"High","gaps":["Did not resolve stoichiometry or higher-order organization of the scaffold","Functional consequences of clustering in intact synapses not yet tested"]},{"year":1997,"claim":"Showed the GK domain is a protein-interaction module rather than an enzyme, recruiting GKAP/SAPAPs and anchoring PSD-95 to the PSD core.","evidence":"GST pulldown, Co-IP from brain, heterologous cell clustering and fractionation; earlier nucleotide-binding/enzymatic assays","pmids":["9024696","9115257","7867790","9756850"],"confidence":"High","gaps":["Phosphorylation requirement of the GKAP/SAPAP interaction not yet known","Atomic basis of GK–ligand recognition unresolved at this stage"]},{"year":1998,"claim":"Demonstrated that PSD-95 nucleates signaling enzymes and additional ionotropic receptors, linking the NMDAR complex to Ras regulation (SynGAP) and bridging to mGluR/Homer networks.","evidence":"Yeast two-hybrid, Co-IP from brain, in vitro GAP assay, kainate-receptor clustering and electrophysiology","pmids":["9581761","9808460","10433269"],"confidence":"High","gaps":["How receptor and enzyme occupancy is coordinated on one scaffold not addressed","In vivo significance of SynGAP coupling not yet tested genetically"]},{"year":1999,"claim":"Revealed PSD-95 as a kinase-scaffolding platform that promotes Fyn-mediated NR2A tyrosine phosphorylation by co-binding receptor and kinase.","evidence":"Co-IP, heterologous cell phosphorylation assay, domain-mapping, validation in fyn-mutant mice","pmids":["9892651"],"confidence":"High","gaps":["Regulation of kinase recruitment by synaptic activity unresolved","Distinct roles of Src/Yes/Lyn versus Fyn not dissected"]},{"year":2002,"claim":"Identified the stargazin–PSD-95 PDZ interaction as the essential mechanism by which PSD-95 controls synaptic AMPAR number, and showed PKA phosphorylation of stargazin disrupts this link.","evidence":"Compensatory mutagenesis with slice electrophysiology; phospho-specific antibodies and yeast two-hybrid/Co-IP","pmids":["12359873","11805122"],"confidence":"High","gaps":["Did not yet show real-time receptor dynamics","How phosphorylation is spatially restricted at synapses unknown"]},{"year":2003,"claim":"Established that activity-driven ubiquitination of PSD-95 by Mdm2 couples NMDAR signaling to AMPAR endocytosis and LTD, defining a degradative arm of scaffold regulation.","evidence":"Co-IP, ubiquitination assay, proteasome inhibition, AMPAR endocytosis and LTD readouts; later linked to Cdk5 control of Mdm2 binding","pmids":["14642282","21849563"],"confidence":"High","gaps":["Precise ubiquitinated residues and chain topology not defined","How degradation is spatially confined to specific synapses unresolved"]},{"year":2007,"claim":"Showed PSD-95 restrains AMPAR–stargazin lateral diffusion to trap receptors at synapses, and that NR2/nNOS preferentially engage PDZ2 to drive excitotoxic and structural plasticity.","evidence":"Single quantum-dot tracking and FRAP in live neurons; quantitative PDZ binding plus siRNA NMDA-toxicity and MIS assays","pmids":["17329211","17855605","19075115"],"confidence":"High","gaps":["Mechanistic basis of diffusional trapping at single-molecule resolution incomplete","How excitotoxic versus plasticity outputs of nNOS coupling are partitioned unknown"]},{"year":2011,"claim":"Defined the architectural and conformational basis of PSD-95 function: vertical filament arrays essential for PSD 3D organization, palmitoylation/S-nitrosylation competition controlling membrane anchoring, and translational gating by miR-125a/FMRP.","evidence":"Electron tomography with RNAi; S-nitrosylation/palmitoylation assays with ZDHHC8 KO; AGO2/FMRP phospho-mutant and miRNA studies","pmids":["21525273","21745643","21658607","19828799"],"confidence":"High","gaps":["The protein partners forming the vertical filaments not fully enumerated","Coupling of NO signaling to specific physiological stimuli incomplete"]},{"year":2016,"claim":"Resolved palmitoylation as a conformational switch (compact-to-extended) gating receptor binding, identified ABHD17 enzymes as the depalmitoylases enabling rapid cycling, and showed PSD-95 destabilizes the phosphatase STEP61 to enrich synaptic NMDARs.","evidence":"FRET and EM with palmitoylation manipulation; serine-hydrolase screen and APEGS kinetics; Co-IP plus PSD-95 KO mice and electrophysiology","pmids":["27956638","27307232","27457929"],"confidence":"High","gaps":["What triggers the conformational switch in vivo not defined","Selectivity of individual ABHD17 paralogs at synapses unresolved"]},{"year":2017,"claim":"Provided atomic and quantitative mechanism for partner regulation: crystal structure of the GK–phospho-SAPAP complex (explaining an intellectual-disability mutation), and PDZ tyrosine phosphorylation tuning stargazin affinity.","evidence":"Crystallography with binding/synaptogenesis assays; semisynthetic phospho-PDZ domains with NMR and single-molecule FRET","pmids":["29281827","28692247"],"confidence":"High","gaps":["Kinases responsible for the relevant PDZ tyrosine phosphorylation in neurons not identified","Generality of the ID-mutation mechanism across patients untested"]},{"year":2019,"claim":"Identified anchoring and transport determinants—α-actinin and ephrin-B3 stabilizing synaptic PSD-95, Ca2+/calmodulin mediating activity-dependent removal during scaling down, and KIF5 driving dendritic delivery.","evidence":"Co-IP and reciprocal mutagenesis with AMPAR imaging; NMR with charge-inversion CaM mutagenesis and mEPSC recording; KIF5A dominant-negative dendritic quantification","pmids":["29429936","26479588","29118000","31753031"],"confidence":"High","gaps":["How anchoring, transport, and turnover are temporally coordinated unresolved","Stoichiometry of α-actinin/ephrin-B3 anchoring within the lattice unknown"]},{"year":2012,"claim":"Established developmental and disease-relevant control of PSD-95 abundance via PTBP1/2-dependent exon 18 splicing/NMD and downstream roles in dopaminergic, epileptic, and amyloid pathology.","evidence":"RT-PCR/NMD and PTBP knockdown/re-expression; single quantum-dot D1R imaging in dyskinesia models; LGI1/ADAM22 epistasis; Aβ/PP1 rescue experiments","pmids":["22246437","23041629","26178195","34077732"],"confidence":"High","gaps":["Whether splicing control persists in adult plasticity unclear","Direct causal contribution of PSD-95 to each disease state varies in strength"]},{"year":null,"claim":"How the full repertoire of PSD-95 modifications, conformational states, anchoring partners, and degradation pathways are integrated in real time to set synaptic strength remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified quantitative model linking palmitoylation cycling, phosphorylation, and ubiquitination to receptor content","Non-neuronal roles (epithelial polarity, tumor suppression) remain mechanistically isolated"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1,16,5,10]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[2,15,14,24,42]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[47,7]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[32,41]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[2,16,41]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3,32]}],"pathway":[{"term_id":"R-HSA-112316","term_label":"Neuronal System","supporting_discovery_ids":[16,28,29]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[5,9,29]}],"complexes":["postsynaptic density","NMDAR–MAGUK complex","PSD-95/GKAP/SAPAP complex","Shank–Homer–mGluR complex"],"partners":["DLGAP1","GRIN2A","CACNG2","NLGN1","NOS1","SYNGAP1","ACTN2","EFNB3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P78352","full_name":"Disks large homolog 4","aliases":["Postsynaptic density protein 95","PSD-95","Synapse-associated protein 90","SAP-90","SAP90"],"length_aa":724,"mass_kda":80.5,"function":"Postsynaptic scaffolding protein that plays a critical role in synaptogenesis and synaptic plasticity by providing a platform for the postsynaptic clustering of crucial synaptic proteins. Interacts with the cytoplasmic tail of NMDA receptor subunits and shaker-type potassium channels. Required for synaptic plasticity associated with NMDA receptor signaling. Overexpression or depletion of DLG4 changes the ratio of excitatory to inhibitory synapses in hippocampal neurons. May reduce the amplitude of ASIC3 acid-evoked currents by retaining the channel intracellularly. May regulate the intracellular trafficking of ADR1B. Also regulates AMPA-type glutamate receptor (AMPAR) immobilization at postsynaptic density keeping the channels in an activated state in the presence of glutamate and preventing synaptic depression (By similarity). Under basal conditions, cooperates with FYN to stabilize palmitoyltransferase ZDHHC5 at the synaptic membrane through FYN-mediated phosphorylation of ZDHHC5 and its subsequent inhibition of association with endocytic proteins (PubMed:26334723)","subcellular_location":"Cell membrane; Postsynaptic density; Synapse; Cytoplasm; Cell projection, axon; Cell projection, dendritic spine; Cell projection, dendrite; Presynapse","url":"https://www.uniprot.org/uniprotkb/P78352/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DLG4","classification":"Not Classified","n_dependent_lines":18,"n_total_lines":1208,"dependency_fraction":0.014900662251655629},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/DLG4","total_profiled":1310},"omim":[{"mim_id":"621329","title":"FAMILY WITH SEQUENCE SIMILARITY 81, MEMBER A; FAM81A","url":"https://www.omim.org/entry/621329"},{"mim_id":"621028","title":"ANKYRIN REPEAT- AND BTB DOMAIN-CONTAINING PROTEIN 3; ABTB3","url":"https://www.omim.org/entry/621028"},{"mim_id":"618793","title":"INTELLECTUAL DEVELOPMENTAL DISORDER, AUTOSOMAL DOMINANT 62; MRD62","url":"https://www.omim.org/entry/618793"},{"mim_id":"618597","title":"BRAIN-ENRICHED GUANYLATE KINASE-ASSOCIATED PROTEIN; BEGAIN","url":"https://www.omim.org/entry/618597"},{"mim_id":"618306","title":"PROLINE-RICH PROTEIN 7; PRR7","url":"https://www.omim.org/entry/618306"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Uncertain","locations":[{"location":"Nucleoplasm","reliability":"Uncertain"},{"location":"Centrosome","reliability":"Uncertain"},{"location":"Basal body","reliability":"Uncertain"},{"location":"Plasma membrane","reliability":"Additional"},{"location":"Primary cilium","reliability":"Additional"},{"location":"Primary cilium tip","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"brain","ntpm":211.2},{"tissue":"retina","ntpm":200.9}],"url":"https://www.proteinatlas.org/search/DLG4"},"hgnc":{"alias_symbol":["PSD-95","PSD95","SAP90","SAP-90"],"prev_symbol":[]},"alphafold":{"accession":"P78352","domains":[{"cath_id":"2.30.42.10","chopping":"61-150","consensus_level":"high","plddt":90.8683,"start":61,"end":150},{"cath_id":"2.30.42.10","chopping":"156-244","consensus_level":"high","plddt":90.2572,"start":156,"end":244},{"cath_id":"2.30.42.10","chopping":"313-402","consensus_level":"high","plddt":88.9931,"start":313,"end":402},{"cath_id":"2.30.30.40","chopping":"430-491","consensus_level":"medium","plddt":83.6018,"start":430,"end":491},{"cath_id":"3.40.50.300","chopping":"531-718","consensus_level":"medium","plddt":93.6688,"start":531,"end":718}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P78352","model_url":"https://alphafold.ebi.ac.uk/files/AF-P78352-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P78352-F1-predicted_aligned_error_v6.png","plddt_mean":77.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=DLG4","jax_strain_url":"https://www.jax.org/strain/search?query=DLG4"},"sequence":{"accession":"P78352","fasta_url":"https://rest.uniprot.org/uniprotkb/P78352.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P78352/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P78352"}},"corpus_meta":[{"pmid":"10433269","id":"PMC_10433269","title":"Coupling 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PDZ domain-specific binding assays\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP from brain tissue combined with domain-mapping experiments; independently replicated across multiple studies\",\n      \"pmids\": [\"9278515\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"PSD-95 interacts via its guanylate kinase-like (GK) domain with GKAP (guanylate kinase-associated protein), a novel synaptic protein; the GK domain functions as a protein-protein interaction module rather than an enzyme, and PSD-95/GKAP co-cluster with K+ channels/NMDA receptors in heterologous cells.\",\n      \"method\": \"GST pulldown; Co-immunoprecipitation from brain; heterologous cell clustering assay\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP from brain plus in vitro pulldown and heterologous cell reconstitution; replicated by multiple labs\",\n      \"pmids\": [\"9024696\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"PSD-95 clusters the inwardly rectifying potassium channel Kir4.1 at the cell membrane via direct PDZ-dependent interaction with the Ser-Asn-Val C-terminal motif, and co-expression doubles whole-cell Kir4.1 current amplitude.\",\n      \"method\": \"Co-immunoprecipitation; GST pulldown; electrophysiology in mammalian cells; deletion mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro pulldown with mutagenesis, Co-IP, and functional electrophysiology in a single study\",\n      \"pmids\": [\"9148889\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"PSD-95 is exclusively associated with the postsynaptic density at forebrain synapses and does not label presynaptic terminals, as established by immunogold electron microscopy of forebrain synaptosomes.\",\n      \"method\": \"Immunogold electron microscopy; subcellular fractionation\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct ultrastructural localization with gold-labeled antibodies; replicated in subsequent studies\",\n      \"pmids\": [\"8778289\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"SAPAPs (SAP90/PSD-95-Associated Proteins) are a family of proteins that specifically interact with PSD-95/SAP90 through the guanylate kinase domain and recruit PSD-95 to the plasma membrane, suggesting a role in anchoring PSD-95 to the PSD.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation; heterologous cell overexpression\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP and cell-based clustering, replicated by independent groups (GKAP paper and subsequent studies)\",\n      \"pmids\": [\"9115257\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"SynGAP, a novel Ras-GTPase activating protein, interacts with the PDZ domains of PSD-95 in vitro and in vivo, forms a macromolecular complex with PSD-95 and the NMDA receptor at excitatory synapses, and negatively regulates Ras activity, linking NMDA receptor signaling to Ras pathway modulation.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation from brain; in vitro GAP activity assay\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP from brain plus in vitro biochemical GAP assay; widely replicated\",\n      \"pmids\": [\"9581761\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"SAP90/PSD-95 colocalizes and co-immunoprecipitates with kainate receptor subunits GluR6 and KA2; GluR6 clustering is mediated by its C-terminal ETMA sequence interacting with PDZ1 of SAP90, while KA2 binds the SH3 and GK domains; SAP90 co-expression reduces GluR6/KA2 desensitization.\",\n      \"method\": \"Co-immunoprecipitation; heterologous cell clustering; electrophysiology\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — Co-IP plus functional electrophysiology plus domain-specific clustering in heterologous cells\",\n      \"pmids\": [\"9808460\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"CRIPT binds selectively to the third PDZ domain (PDZ3) of PSD-95 via its C-terminus, causes redistribution of PSD-95 to microtubules in heterologous cells, and co-immunoprecipitates with PSD-95 and tubulin from brain, suggesting CRIPT links PSD-95 to the microtubule cytoskeleton.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation from brain; heterologous cell redistribution assay\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP from brain plus cell-based functional assay, single lab\",\n      \"pmids\": [\"9581762\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"BEGAIN, a brain-enriched protein, interacts with the GK domain of PSD-95/SAP90 and is recruited to the Triton X-100-insoluble PSD fraction by SAPAP when co-expressed with PSD-95, identifying BEGAIN as a component of the core PSD-95/SAPAP complex.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation; fractionation in transfected cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP and fractionation assays, single lab\",\n      \"pmids\": [\"9756850\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"PSD-95 promotes Fyn-mediated tyrosine phosphorylation of the NMDA receptor subunit NR2A by scaffolding: different regions of PSD-95 bind NR2A and Fyn simultaneously, enabling complex formation and enhancing phosphorylation. PSD-95 also associates with Src, Yes, and Lyn.\",\n      \"method\": \"Co-immunoprecipitation; heterologous cell phosphorylation assay; domain-mapping\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP plus functional phosphorylation assay in 293T cells, validated in fyn-mutant mice\",\n      \"pmids\": [\"9892651\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Shank proteins bridge Homer complexes and PSD-95/GKAP complexes at the postsynaptic density: Shank binds Homer via a single Homer-binding site and co-immunoprecipitates from brain; in heterologous cells Shank clusters mGluR5 and mediates coclustering of Homer with PSD-95/GKAP.\",\n      \"method\": \"Co-immunoprecipitation from brain; heterologous cell clustering assay; yeast two-hybrid\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP from brain plus reconstitution in heterologous cells; replicated across labs\",\n      \"pmids\": [\"10433269\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Citron, a Rho-effector, interacts with PSD-95/SAP90 both in vivo and in vitro, co-localizes at glutamatergic synapses in thalamic neurons, and can form a heteromeric complex with PSD-95 and NMDA receptors in heterologous cells, linking Rho signaling to the NMDA receptor complex.\",\n      \"method\": \"Co-immunoprecipitation from brain; in vitro binding; heterologous cell co-expression\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP from brain and heterologous cell assay, single lab\",\n      \"pmids\": [\"9870943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Protein tyrosine phosphatase zeta/RPTPbeta interacts with the second PDZ domain of PSD-95/SAP90 via its C-terminal sequence, as shown by yeast two-hybrid; RPTPbeta concentrates in the postsynaptic density fraction by subcellular fractionation.\",\n      \"method\": \"Yeast two-hybrid; subcellular fractionation; immunohistochemistry\",\n      \"journal\": \"Brain research. Molecular brain research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — yeast two-hybrid plus fractionation, no Co-IP confirmation, single lab\",\n      \"pmids\": [\"10521598\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"SAP90/PSD-95 binds GMP in the micromolar range but displays no detectable guanylate kinase enzymatic activity, establishing that the GK domain is a nucleotide-binding but catalytically inactive module.\",\n      \"method\": \"Nucleotide binding assay (GMP, ATP, GDP, ADP); enzymatic activity assay\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — direct in vitro biochemical assay, single lab, early study\",\n      \"pmids\": [\"7867790\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"PSD-95 promotes clustering of voltage-gated Kv channels only when channels are present at the cell surface, acting at the plasma membrane, whereas SAP97 retains Kv channels intracellularly in ER-derived vesicles, demonstrating mechanistically distinct clustering modes.\",\n      \"method\": \"Immunofluorescence; Co-immunoprecipitation; surface expression analysis in heterologous cells\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal methods (Co-IP, surface expression, imaging) in single lab\",\n      \"pmids\": [\"10629225\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Kir2.1 and Kir2.3 (but not Kir3 subunits) bind PSD-95 PDZ domains via their C-terminal motif; PSD-95 co-expression suppresses Kir2.3 channel activity by >50%, predominantly affecting single-channel conductance.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation; patch-clamp electrophysiology in HEK-293 cells\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — Co-IP plus functional electrophysiology, single lab\",\n      \"pmids\": [\"10627592\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Direct binding of the first two PDZ domains of PSD-95 to stargazin is required to localize AMPA receptors to synapses; increasing PSD-95 at synapses recruits new AMPARs without changing surface AMPAR number, and compensatory mutations in both PSD-95 and stargazin confirm the centrality of this direct interaction.\",\n      \"method\": \"Hippocampal slice cultures; biolistic gene transfection; electrophysiology; compensatory mutagenesis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — compensatory mutagenesis with functional electrophysiological readout; widely replicated\",\n      \"pmids\": [\"12359873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"PKA phosphorylates stargazin at Thr-321, and phosphorylation eliminates the stargazin–PSD-95 PDZ interaction, reducing synaptic AMPAR targeting, as shown by phospho-mimetic mutations abrogating yeast two-hybrid interaction, Co-IP, and co-clustering.\",\n      \"method\": \"Phospho-specific antibodies; PKA phosphorylation assay; yeast two-hybrid; Co-immunoprecipitation; heterologous cell clustering\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple orthogonal methods including mutagenesis and in vitro phosphorylation, single lab\",\n      \"pmids\": [\"11805122\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"PSD-95 is ubiquitinated by the E3 ligase Mdm2; NMDA receptor activation triggers PSD-95 ubiquitination and proteasome-dependent removal from synapses; mutations blocking ubiquitination prevent NMDA-induced AMPAR endocytosis; proteasome inhibitors block NMDA-induced AMPAR internalization and LTD.\",\n      \"method\": \"Co-immunoprecipitation; ubiquitination assay; proteasome inhibitor treatment; AMPAR endocytosis assay\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal experiments (Co-IP, mutagenesis, pharmacological) with functional plasticity readout\",\n      \"pmids\": [\"14642282\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"A subset of mammalian DHHC-domain proteins (P-PATs) specifically palmitoylate PSD-95 in vitro and in vivo; inhibition of P-PAT activity in neurons reduces PSD-95 palmitoylation, synaptic clustering, and AMPAR-mediated neurotransmission.\",\n      \"method\": \"In vitro palmitoylation assay; cell-based palmitoylation assay; substrate specificity comparison; neuronal inhibition with functional readout\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro enzymatic assay plus neuronal functional readout; replicated by subsequent structural/mechanistic studies\",\n      \"pmids\": [\"15603741\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"PSD-95 is phosphorylated by SAPK3/p38gamma at Thr287 and Ser290 (in vitro) and at Ser290 in cells in response to stress; SAPK3 binds preferentially to the third PDZ domain of SAP90, and this binding is required for phosphorylation. ERK1/ERK2 also phosphorylates SAP90 at Thr287 and Ser290 in vitro but independently of PDZ binding.\",\n      \"method\": \"In vitro kinase assay; cell-based phosphorylation; Tat-peptide disruption; Co-immunoprecipitation from synaptic junctions\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay plus cell-based validation and peptide disruption, single lab\",\n      \"pmids\": [\"14741046\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Neuregulin-1 intracellular domain (Nrg-ICD) translocates to the nucleus upon depolarization, binds the zinc-finger transcription factor Eos, and the Nrg-ICD–Eos complex upregulates PSD-95 transcription via the PSD-95 promoter in a gamma-secretase-independent manner.\",\n      \"method\": \"Reporter gene assay; Co-immunoprecipitation; in vivo promoter activity assay\",\n      \"journal\": \"Nature neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus reporter assay and in vivo validation, single lab\",\n      \"pmids\": [\"15494726\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"IRSp53 links postsynaptic Shank1 to PSD-95 by forming a tripartite complex: IRSp53 binds PSD-95 PDZ2 domain via its C-terminal PDZ-binding motif, recruits PSD-95 into filopodia in HEK cells, and the interaction is confirmed by Co-IP from brain.\",\n      \"method\": \"Co-immunoprecipitation from brain; heterologous cell targeting assay; yeast two-hybrid\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP from brain plus cell-based assay, single lab\",\n      \"pmids\": [\"15255944\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"mPins interacts with PSD-95 and SAP102, facilitates formation of the NMDAR–MAGUK complex, and enhances trafficking of SAP102 and NMDARs to the plasma membrane; dominant-negative and siRNA knockdown of mPins reduces SAP102 in dendrites and alters NMDAR surface expression.\",\n      \"method\": \"Co-immunoprecipitation; dominant-negative expression; siRNA knockdown; surface receptor assay in neurons\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus loss-of-function with defined cellular phenotype, single lab\",\n      \"pmids\": [\"16299499\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"GLYT1 interacts with PDZ domains I and II of PSD-95 via its C-terminal -SRI motif; this interaction stabilizes GLYT1 at the plasma membrane, blocks its internalization, and increases glycine uptake Vmax.\",\n      \"method\": \"Co-immunoprecipitation from rat brain; mutational analysis; glycine uptake assay\",\n      \"journal\": \"Journal of neurochemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP from brain plus mutational analysis and functional transport assay, single lab\",\n      \"pmids\": [\"16271045\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"CaMKII phosphorylates PSD-95 at Ser73 within the PDZ1 domain, causing dissociation of NR2A (but not NR2B) from PSD-95, identified by in vitro kinase assay, point mutagenesis, phospho-specific antibody, and validated in hippocampal neurons.\",\n      \"method\": \"In vitro kinase assay with GST-PSD-95; point mutagenesis; phospho-specific antibody; transfected COS-7 and cultured neurons\",\n      \"journal\": \"The European journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay plus mutagenesis plus neuronal validation, multiple orthogonal methods\",\n      \"pmids\": [\"17156196\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"PSD-95 and PKC potentiate NMDAR gating and surface expression in a non-additive manner; PSD-95 partially occludes PKC potentiation; Ser-1462 in the PDZ-binding motif of NR2A is required for PSD-95-induced potentiation.\",\n      \"method\": \"Xenopus oocyte electrophysiology; immunofluorescence; Co-immunoprecipitation; mutagenesis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — electrophysiology plus mutagenesis in oocytes; single lab\",\n      \"pmids\": [\"17179037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"PSD-95 interacts with Src via a 12-amino-acid N-terminal sequence that binds the Src SH2 domain in a phosphotyrosine-independent manner, inhibits Src kinase activity, reduces NMDAR phosphorylation, depresses NMDAR currents, and inhibits LTP induction.\",\n      \"method\": \"Co-immunoprecipitation; in vitro binding; peptide inhibition; electrophysiology in cultured neurons and hippocampal slices\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro domain binding plus functional electrophysiology with peptide inhibition; novel mechanistic finding\",\n      \"pmids\": [\"16990796\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"AMPAR–Stargazin complexes diffuse laterally between extrasynaptic and synaptic sites as a unit; disruption of Stargazin–PSD-95 interaction strongly increases AMPAR surface diffusion and prevents AMPAR accumulation at postsynaptic sites, as measured by single quantum dot tracking and FRAP in live neurons.\",\n      \"method\": \"Single quantum dot tracking; FRAP; live hippocampal neuron imaging\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two orthogonal live-cell imaging methods, functional readout tied to PSD-95 interaction\",\n      \"pmids\": [\"17329211\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"NR2A–NR2C NMDA receptor subunits and nNOS preferentially interact with PSD-95 PDZ2 domain (EC50 ~1 µM for NR2 subunits; ~200 nM for nNOS/PSD-95 interaction by nNOS peptide); knockdown of only PSD-95 or nNOS (not PSD-93, SAP97, SAP102, TIP1) reduces neuronal excitotoxic vulnerability.\",\n      \"method\": \"Proteomic PDZ interaction screen; biochemical binding assays; siRNA knockdown; NMDA toxicity assay in cortical neurons\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — quantitative binding assay plus siRNA loss-of-function with cellular toxicity readout, multiple orthogonal approaches\",\n      \"pmids\": [\"17855605\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"PSD-95 overexpression promotes multiinnervated spine (MIS) formation through nitric oxide synthase (NOS): deletion of PSD-95 PDZ2 (which interacts with nNOS) prevents MIS formation; siRNA knockdown or pharmacological blockade of NOS also prevents axon varicosity formation and MIS formation, whereas NO donor or cGMP analogue induces MIS.\",\n      \"method\": \"Electron microscopy; genetic deletion of PDZ domains; siRNA; pharmacology; hippocampal slice culture\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genetic domain deletion plus siRNA plus pharmacological rescue; multiple orthogonal methods\",\n      \"pmids\": [\"19075115\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Distinct domains of PSD-95 govern different aspects of its synaptic behavior: N-terminal palmitoylation plus PDZ1/2 interactions stabilize PSD-95 within the PSD; PDZ1/2 interactions also mediate NMDA-receptor-dependent rapid destabilization of PSD-95 during chemical LTD; SH3 domain is additionally needed for a stable PSD-95 lattice.\",\n      \"method\": \"PAGFP photoactivation; two-photon microscopy; PSD-95 domain mutants; rat CA1 pyramidal neurons\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live-cell FRAP/photoactivation with multiple domain mutants; single lab but multiple orthogonal domain analyses\",\n      \"pmids\": [\"19828799\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"PSD-95 is arranged as vertical filaments in regular arrays within the PSD; RNAi knockdown of PSD-95 causes loss of entire patches of PSD material including vertical filaments, horizontal elements, and putative AMPA receptor structures (but not NMDA receptor structures), establishing that PSD-95-containing filaments are essential for 3D PSD organization.\",\n      \"method\": \"Electron tomography; RNAi knockdown; rat hippocampal spine synapse analysis\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — high-resolution EM tomography combined with RNAi loss-of-function and structural phenotype\",\n      \"pmids\": [\"21525273\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"miR-125a targeting of PSD-95 mRNA reversibly inhibits PSD-95 translation; FMRP phosphorylation promotes formation of an AGO2–miR-125a inhibitory complex on PSD-95 mRNA, while mGluR signaling requires FMRP dephosphorylation and AGO2 release to de-repress translation.\",\n      \"method\": \"miRNA inhibition; FMRP phospho-mutant analysis; Co-IP of AGO2 with PSD-95 mRNA; dendritic spine morphology analysis\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple biochemical and cellular methods, single lab\",\n      \"pmids\": [\"21658607\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"PSD-95 is S-nitrosylated at cysteines 3 and 5, which are the same residues palmitoylated; physiological NO inhibits PSD-95 palmitoylation, reduces synaptic PSD-95 clusters; decreased palmitoylation (via 2-bromopalmitate or ZDHHC8 KO) increases PSD-95 nitrosylation, establishing reciprocal competition between S-nitrosylation and palmitoylation at these cysteines.\",\n      \"method\": \"S-nitrosylation assay; palmitoylation assay; ZDHHC8 knockout mice; granule cell imaging; Co-IP\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — biochemical modification assays with KO mouse validation and multiple orthogonal approaches, single lab\",\n      \"pmids\": [\"21745643\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Cyclin-dependent kinase 5 (Cdk5) regulates PSD-95 ubiquitination: genetic or pharmacological reduction of Cdk5 activity increases Mdm2–PSD-95 interaction and PSD-95 ubiquitination, correlating with increased β-adaptin binding (AP-2 subunit), suggesting a mechanism linking PSD-95 ubiquitination to NMDA-induced AMPAR endocytosis.\",\n      \"method\": \"Co-immunoprecipitation; pharmacological Cdk5 inhibition; genetic Cdk5 reduction (mouse); ubiquitination assay; mass spectrometry\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus pharmacological and genetic approaches in vivo, single lab\",\n      \"pmids\": [\"21849563\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"PTBP1 and PTBP2 repress PSD-95 exon 18 splicing, leading to premature translation termination and NMD, preventing PSD-95 expression during early neural development; sequential loss of PTBP1 then PTBP2 allows exon 18 inclusion and PSD-95 expression; re-expression of either PTBP in differentiated neurons impairs glutamatergic synapse development.\",\n      \"method\": \"RT-PCR; NMD analysis; PTBP knockdown/re-expression in neurons; synapse development assay\",\n      \"journal\": \"Nature neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — loss-of-function and gain-of-function in neurons with specific molecular and cellular readouts; replicated with multiple approaches\",\n      \"pmids\": [\"22246437\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"PSD-95 regulates dopamine D1 receptor (D1R) trafficking and localization in the striatum: disrupting D1R–PSD-95 interaction destabilizes D1R at synapses via increased lateral diffusion, internalization, and diminished surface expression, reducing L-DOPA-induced dyskinesia in rat and macaque models.\",\n      \"method\": \"Single quantum dot imaging; biochemical interaction assay; peptide disruption in vivo; rat/macaque dyskinesia models\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single molecule tracking plus in vivo pharmacological intervention with behavioral readout\",\n      \"pmids\": [\"23041629\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"LGI1 acts as a paracrine signal through ADAM22 (via PDZ domain interactions) to set postsynaptic strength; in the absence of LGI1, PSD-95 (but not SAP102) is unable to modulate synaptic transmission, establishing LGI1–ADAM22 as an essential complex regulating the functional incorporation of PSD-95 at mature synapses.\",\n      \"method\": \"Hippocampal slice electrophysiology; viral knockdown; epistasis analysis in LGI1 KO and ADAM22 mutant mice\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis in KO mice with electrophysiological readout, single lab\",\n      \"pmids\": [\"26178195\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Ephrin-B3 directly interacts with and stabilizes PSD-95 at synapses; activity-dependent MAPK phosphorylation of ephrin-B3 at Ser332 disperses it from synapses, preventing interaction with PSD-95 and increasing PSD-95 turnover, linking neuronal activity to PSD-95 synaptic stability.\",\n      \"method\": \"Super-resolution imaging; biochemical interaction assays; in vivo models; phospho-mutant analysis\",\n      \"journal\": \"Nature neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — super-resolution imaging plus biochemical Co-IP plus in vivo phospho-mutant validation; multiple orthogonal methods\",\n      \"pmids\": [\"26479588\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"ABHD17A, 17B, and 17C are the physiological PSD-95 depalmitoylating enzymes: they show the strongest depalmitoylating activity toward PSD-95, localize to recycling endosomes and the synaptic fraction, and their inhibition delays PSD-95 depalmitoylation kinetics; most PSD-95 undergoes rapid palmitoylation cycling in neurons, with cycling decelerating during synapse maturation.\",\n      \"method\": \"Serine hydrolase screen; APEGS (acyl-PEGyl exchange gel shift) assay; shRNA knockdown; neuronal palmitoylation kinetics\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — quantitative in vitro depalmitoylation assay plus neuronal loss-of-function with kinetic measurement; first identification of depalmitoylating enzyme\",\n      \"pmids\": [\"27307232\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"PSD-95 palmitoylation changes its conformation from compact to extended, and only palmitoylated/extended PSD-95 associates with AMPARs (via TARPs) or NMDARs (via GluN2B); within the PSD, PSD-95 orients perpendicular to the membrane with its palmitoylated N-terminus at the membrane, and changing palmitoylation alters AMPAR but not NMDAR levels.\",\n      \"method\": \"FRET; electron microscopy; subcellular fractionation; palmitoylation manipulation\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — FRET structural measurement plus EM plus functional receptor interaction, multiple orthogonal methods\",\n      \"pmids\": [\"27956638\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"PSD-95 expression destabilizes STEP61 via ubiquitination and proteasomal degradation, excluding STEP61 from the PSD; STEP61 knockdown increases extrasynaptic (but not synaptic) NMDAR expression; PSD-95 binds STEP61 directly but STEP61 is increased in the PSD of PSD-95 KO mice.\",\n      \"method\": \"Co-immunoprecipitation; subcellular fractionation; PSD-95 KO mice; siRNA knockdown; electrophysiology\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP plus KO mouse validation plus functional electrophysiology; multiple orthogonal methods\",\n      \"pmids\": [\"27457929\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"TAOK2 directly phosphorylates Septin7 at an evolutionarily conserved residue; this phosphorylation induces Septin7 translocation to dendritic spines where it associates with and stabilizes PSD-95, promoting spine maturation; TAOK2 depletion causes unstable dendritic protrusions and loss of NMDAR calcium compartmentalization.\",\n      \"method\": \"Chemical genetics; mass spectrometry phosphorylation identification; Co-IP; live imaging of spine dynamics; siRNA\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — chemical-genetic kinase assay plus MS substrate identification plus Co-IP, single lab\",\n      \"pmids\": [\"28065648\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Phosphorylation of PSD-95 PDZ domains exerts complex effects on binding partners: phosphorylation at Y397 significantly increases affinity for stargazin (confirmed by NMR and single-molecule FRET); phosphomimetic mutations are ineffective substitutes for tyrosine phosphorylation.\",\n      \"method\": \"Semisynthetic phosphorylated PDZ domains; fluorescence anisotropy; NMR; single-molecule FRET\",\n      \"journal\": \"ACS chemical biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR and smFRET structural validation plus quantitative binding, multiple orthogonal methods\",\n      \"pmids\": [\"28692247\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"PSD-95/SAPAP interaction requires phosphorylation of N-terminal repeat sequences of SAPAPs; crystal structure of PSD-95 GK domain in complex with phospho-SAPAP peptide reveals the molecular basis; a PSD-95 GK mutation found in intellectual disability patients disrupts this interaction.\",\n      \"method\": \"Crystal structure; biochemical binding assays; functional synaptogenesis assay; phosphomimetic and mutagenesis analysis\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus biochemical validation plus functional cellular assay; explains human mutation\",\n      \"pmids\": [\"29281827\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Ca2+/calmodulin binds to the N-terminus of PSD-95, with E17 interacting with R126 on CaM as identified by NMR; this interaction mediates postsynaptic loss of PSD-95 and AMPARs during homeostatic synaptic scaling down; E17R mutation prevents scaling down, rescued by charge-inversion CaM(R126E).\",\n      \"method\": \"NMR structural analysis; compensatory charge-inversion mutagenesis; miniature EPSC recording in hippocampal neurons\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — NMR structure plus compensatory mutagenesis with functional electrophysiological validation; mechanistically rigorous\",\n      \"pmids\": [\"29118000\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"α-Actinin binds to the very N-terminus of PSD-95 (involving K10/K11 of PSD-95 and E53, E213/D217 of α-actinin); knockdown of α-actinin phenocopies PSD-95 knockdown; mutations impairing the interaction reduce postsynaptic localization of both PSD-95 and AMPARs in parallel.\",\n      \"method\": \"Co-immunoprecipitation; mutagenesis; shRNA knockdown; AMPAR localization imaging in neurons\",\n      \"journal\": \"Neuron\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal mutagenesis affecting both binding and localization; multiple orthogonal approaches; single lab\",\n      \"pmids\": [\"29429936\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PKA phosphorylates NLGN1 at S839 near the PDZ ligand, reducing PSD-95 binding; impaired NLGN1/PSD-95 binding decreases NLGN1 surface expression and reduces NLGN1-mediated synaptic enhancement.\",\n      \"method\": \"Co-immunoprecipitation; phosphomimetic mutagenesis; surface expression assay; synaptic electrophysiology\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus mutagenesis plus functional synaptic assay, single lab\",\n      \"pmids\": [\"31138690\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"KIF5 (kinesin) transports PSD-95 to dendrites via interaction between the C-terminal tail of KIF5A and the third PDZ domain of PSD-95; expression of motor-domain-deleted KIF5A reduces dendritic PSD-95 levels; Staufen regulates the KIF5A–PSD-95 association.\",\n      \"method\": \"Co-immunoprecipitation in vitro and in vivo; dominant-negative KIF5A expression; dendritic PSD-95 quantification\",\n      \"journal\": \"Molecular brain\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus dominant-negative loss-of-function with dendritic localization readout, single lab\",\n      \"pmids\": [\"31753031\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Increased PSD-95 blocks Aβ-induced modification of the NMDAR C-terminal domain conformation and its interaction with PP1, preventing synaptic weakening; pharmacological inhibition of PSD-95 depalmitoylation increases synaptic PSD-95 and rescues Aβ-induced deficits.\",\n      \"method\": \"Electrophysiology; NMDAR CTD conformation assay; Co-IP of NMDAR with PP1; pharmacological depalmitoylation inhibition\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus electrophysiology plus pharmacological rescue, single lab\",\n      \"pmids\": [\"34077732\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"DLG4/PSD-95 interacts with the Cyt1 (but not Cyt2) cytoplasmic domain of CD46, and this interaction is required for polarized expression of CD46 in human epithelial cells, demonstrating a non-neuronal role for DLG4 in epithelial polarity.\",\n      \"method\": \"Yeast two-hybrid; Co-immunoprecipitation; dominant-negative DLG4 expression; polarized epithelial cell assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus functional polarization assay with domain specificity, single lab\",\n      \"pmids\": [\"11714708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"HPV18 E6 protein targets DLG4/PSD-95 for E6AP-dependent proteasomal degradation via the E6 C-terminal PDZ-binding motif; DLG4 expression suppresses tumorigenicity of CaSki cells without affecting growth in culture, suggesting a tumor suppressor function.\",\n      \"method\": \"E6AP shRNA rescue; in vitro binding/degradation assay; tumorigenicity assay\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical degradation assay plus shRNA rescue plus in vivo tumorigenicity, single lab\",\n      \"pmids\": [\"17121805\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"GRK5 (but not GRK2) reduces β1-adrenergic receptor (β1AR) association with PSD-95 in a kinase-activity-dependent manner through phosphorylation of β1AR, and agonist stimulation further decreases PSD-95 binding to β1AR.\",\n      \"method\": \"Co-immunoprecipitation; kinase-dead GRK5 mutant; agonist treatment in COS-7 cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP plus kinase-dead mutant with functional readout, single lab\",\n      \"pmids\": [\"11700307\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PSD-95/DLG4 is a membrane-anchored scaffolding protein that organizes the postsynaptic density of excitatory synapses by using its tandem PDZ1/PDZ2 domains to cluster NMDA receptor subunits (NR2A/B), AMPA receptors via stargazin/TARPs, neuroligins, and K+/Kir channels; its GK domain recruits GKAP/SAPAPs (in a phosphorylation-dependent manner) and links to Shank–Homer–mGluR complexes; its SH3 domain interacts with SAP97 to regulate AMPAR trafficking; N-terminal palmitoylation (written by DHHC PATs, erased by ABHD17 depalmitoylases) drives membrane anchoring and is reciprocally regulated by S-nitrosylation at the same cysteines, while Ca²⁺/calmodulin binding to the PSD-95 N-terminus mediates homeostatic scaling down; PSD-95 is further regulated by ubiquitination via Mdm2 (promoting proteasomal degradation and AMPAR endocytosis), phosphorylation by CaMKII (at PDZ1-Ser73, releasing NR2A), SAPK3/ERK (Thr287/Ser290), and PKA (indirectly via stargazin/NLGN1 phosphorylation); it scaffolds Src-family kinases (promoting Fyn-mediated NR2A phosphorylation while its N-terminal region negatively regulates Src) and links NMDARs to nNOS, a coupling that drives excitotoxic signaling; the protein exists in extended configuration within vertical PSD filaments essential for overall PSD architecture, is transported to dendrites by KIF5 kinesin via its PDZ3 domain, and is anchored at postsynaptic sites by α-actinin and ephrin-B3; its expression is controlled post-transcriptionally by PTBP1/2-mediated exon 18 repression and NMD during early development, and by miR-125a/FMRP at the translational level at mature synapses.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"DLG4/PSD-95 is a multidomain membrane-associated scaffolding protein that organizes the postsynaptic density (PSD) of excitatory glutamatergic synapses, where it localizes exclusively to postsynaptic sites and is essential for the three-dimensional architecture of the PSD [#3, #32]. Its tandem PDZ domains are functionally specialized: PDZ1/PDZ2 cluster NMDA receptor NR2 subunits, neuroligins, and inwardly rectifying and voltage-gated K+ channels at the surface membrane, while PDZ3 binds distinct partners including neuroligins and CRIPT, the latter linking the scaffold to microtubules [#0, #2, #7, #14, #15]. PSD-95 controls AMPA receptor synaptic content indirectly by binding the auxiliary subunit stargazin through PDZ1/2; this interaction restrains AMPAR lateral diffusion and is required for AMPAR accumulation and retention at synapses [#16, #28]. Through its catalytically inactive guanylate kinase (GK) domain, which binds GMP but lacks enzymatic activity, PSD-95 recruits GKAP/SAPAPs in a phosphorylation-dependent manner—a contact whose structural basis is defined and whose disruption by a GK-domain mutation causes intellectual disability—and thereby nucleates the Shank–Homer–mGluR signaling network [#1, #13, #45, #10]. PSD-95 additionally scaffolds signaling enzymes, coupling NMDARs to SynGAP and to nNOS, the latter linkage driving excitotoxic vulnerability and multiinnervated spine formation [#5, #29, #30]. N-terminal palmitoylation switches PSD-95 from a compact to an extended conformation required for receptor association and membrane anchoring; this modification is written by DHHC palmitoyl-acyltransferases, erased by ABHD17 depalmitoylases through rapid cycling, and competes reciprocally with S-nitrosylation at the same cysteines [#19, #40, #41, #34]. The scaffold's synaptic abundance and incorporation are dynamically tuned by ubiquitination via Mdm2 (driving proteasomal turnover and AMPAR endocytosis), by phosphorylation (CaMKII at PDZ1-Ser73 releasing NR2A), by Ca2+/calmodulin binding to its N-terminus during homeostatic scaling down, and by anchoring partners α-actinin and ephrin-B3 [#18, #25, #46, #47, #39]. Its expression is gated post-transcriptionally by PTBP1/2-mediated exon 18 repression coupled to nonsense-mediated decay during development and by miR-125a/FMRP translational control at mature synapses [#36, #33]. A PSD-95 GK-domain mutation found in patients establishes a direct link to intellectual disability [#45].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Established that PSD-95 is a dedicated postsynaptic scaffold whose individual PDZ domains are functionally specialized to bind distinct membrane partners, defining the molecular logic of receptor clustering.\",\n      \"evidence\": \"Co-IP from forebrain with PDZ domain-specific binding assays, plus immunogold EM localization to the PSD\",\n      \"pmids\": [\"9278515\", \"8778289\", \"9148889\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve stoichiometry or higher-order organization of the scaffold\", \"Functional consequences of clustering in intact synapses not yet tested\"]\n    },\n    {\n      \"year\": 1997,\n      \"claim\": \"Showed the GK domain is a protein-interaction module rather than an enzyme, recruiting GKAP/SAPAPs and anchoring PSD-95 to the PSD core.\",\n      \"evidence\": \"GST pulldown, Co-IP from brain, heterologous cell clustering and fractionation; earlier nucleotide-binding/enzymatic assays\",\n      \"pmids\": [\"9024696\", \"9115257\", \"7867790\", \"9756850\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphorylation requirement of the GKAP/SAPAP interaction not yet known\", \"Atomic basis of GK–ligand recognition unresolved at this stage\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Demonstrated that PSD-95 nucleates signaling enzymes and additional ionotropic receptors, linking the NMDAR complex to Ras regulation (SynGAP) and bridging to mGluR/Homer networks.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP from brain, in vitro GAP assay, kainate-receptor clustering and electrophysiology\",\n      \"pmids\": [\"9581761\", \"9808460\", \"10433269\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How receptor and enzyme occupancy is coordinated on one scaffold not addressed\", \"In vivo significance of SynGAP coupling not yet tested genetically\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Revealed PSD-95 as a kinase-scaffolding platform that promotes Fyn-mediated NR2A tyrosine phosphorylation by co-binding receptor and kinase.\",\n      \"evidence\": \"Co-IP, heterologous cell phosphorylation assay, domain-mapping, validation in fyn-mutant mice\",\n      \"pmids\": [\"9892651\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Regulation of kinase recruitment by synaptic activity unresolved\", \"Distinct roles of Src/Yes/Lyn versus Fyn not dissected\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified the stargazin–PSD-95 PDZ interaction as the essential mechanism by which PSD-95 controls synaptic AMPAR number, and showed PKA phosphorylation of stargazin disrupts this link.\",\n      \"evidence\": \"Compensatory mutagenesis with slice electrophysiology; phospho-specific antibodies and yeast two-hybrid/Co-IP\",\n      \"pmids\": [\"12359873\", \"11805122\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not yet show real-time receptor dynamics\", \"How phosphorylation is spatially restricted at synapses unknown\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Established that activity-driven ubiquitination of PSD-95 by Mdm2 couples NMDAR signaling to AMPAR endocytosis and LTD, defining a degradative arm of scaffold regulation.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, proteasome inhibition, AMPAR endocytosis and LTD readouts; later linked to Cdk5 control of Mdm2 binding\",\n      \"pmids\": [\"14642282\", \"21849563\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise ubiquitinated residues and chain topology not defined\", \"How degradation is spatially confined to specific synapses unresolved\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Showed PSD-95 restrains AMPAR–stargazin lateral diffusion to trap receptors at synapses, and that NR2/nNOS preferentially engage PDZ2 to drive excitotoxic and structural plasticity.\",\n      \"evidence\": \"Single quantum-dot tracking and FRAP in live neurons; quantitative PDZ binding plus siRNA NMDA-toxicity and MIS assays\",\n      \"pmids\": [\"17329211\", \"17855605\", \"19075115\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanistic basis of diffusional trapping at single-molecule resolution incomplete\", \"How excitotoxic versus plasticity outputs of nNOS coupling are partitioned unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Defined the architectural and conformational basis of PSD-95 function: vertical filament arrays essential for PSD 3D organization, palmitoylation/S-nitrosylation competition controlling membrane anchoring, and translational gating by miR-125a/FMRP.\",\n      \"evidence\": \"Electron tomography with RNAi; S-nitrosylation/palmitoylation assays with ZDHHC8 KO; AGO2/FMRP phospho-mutant and miRNA studies\",\n      \"pmids\": [\"21525273\", \"21745643\", \"21658607\", \"19828799\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"The protein partners forming the vertical filaments not fully enumerated\", \"Coupling of NO signaling to specific physiological stimuli incomplete\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Resolved palmitoylation as a conformational switch (compact-to-extended) gating receptor binding, identified ABHD17 enzymes as the depalmitoylases enabling rapid cycling, and showed PSD-95 destabilizes the phosphatase STEP61 to enrich synaptic NMDARs.\",\n      \"evidence\": \"FRET and EM with palmitoylation manipulation; serine-hydrolase screen and APEGS kinetics; Co-IP plus PSD-95 KO mice and electrophysiology\",\n      \"pmids\": [\"27956638\", \"27307232\", \"27457929\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"What triggers the conformational switch in vivo not defined\", \"Selectivity of individual ABHD17 paralogs at synapses unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Provided atomic and quantitative mechanism for partner regulation: crystal structure of the GK–phospho-SAPAP complex (explaining an intellectual-disability mutation), and PDZ tyrosine phosphorylation tuning stargazin affinity.\",\n      \"evidence\": \"Crystallography with binding/synaptogenesis assays; semisynthetic phospho-PDZ domains with NMR and single-molecule FRET\",\n      \"pmids\": [\"29281827\", \"28692247\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinases responsible for the relevant PDZ tyrosine phosphorylation in neurons not identified\", \"Generality of the ID-mutation mechanism across patients untested\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified anchoring and transport determinants—α-actinin and ephrin-B3 stabilizing synaptic PSD-95, Ca2+/calmodulin mediating activity-dependent removal during scaling down, and KIF5 driving dendritic delivery.\",\n      \"evidence\": \"Co-IP and reciprocal mutagenesis with AMPAR imaging; NMR with charge-inversion CaM mutagenesis and mEPSC recording; KIF5A dominant-negative dendritic quantification\",\n      \"pmids\": [\"29429936\", \"26479588\", \"29118000\", \"31753031\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How anchoring, transport, and turnover are temporally coordinated unresolved\", \"Stoichiometry of α-actinin/ephrin-B3 anchoring within the lattice unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Established developmental and disease-relevant control of PSD-95 abundance via PTBP1/2-dependent exon 18 splicing/NMD and downstream roles in dopaminergic, epileptic, and amyloid pathology.\",\n      \"evidence\": \"RT-PCR/NMD and PTBP knockdown/re-expression; single quantum-dot D1R imaging in dyskinesia models; LGI1/ADAM22 epistasis; Aβ/PP1 rescue experiments\",\n      \"pmids\": [\"22246437\", \"23041629\", \"26178195\", \"34077732\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether splicing control persists in adult plasticity unclear\", \"Direct causal contribution of PSD-95 to each disease state varies in strength\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the full repertoire of PSD-95 modifications, conformational states, anchoring partners, and degradation pathways are integrated in real time to set synaptic strength remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified quantitative model linking palmitoylation cycling, phosphorylation, and ubiquitination to receptor content\", \"Non-neuronal roles (epithelial polarity, tumor suppression) remain mechanistically isolated\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 1, 16, 5, 10]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [2, 15, 14, 24, 42]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [47, 7]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [32, 41]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [2, 16, 41]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3, 32]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-112316\", \"supporting_discovery_ids\": [16, 28, 29]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [5, 9, 29]}\n    ],\n    \"complexes\": [\"postsynaptic density\", \"NMDAR–MAGUK complex\", \"PSD-95/GKAP/SAPAP complex\", \"Shank–Homer–mGluR complex\"],\n    \"partners\": [\"DLGAP1\", \"GRIN2A\", \"CACNG2\", \"NLGN1\", \"NOS1\", \"SYNGAP1\", \"ACTN2\", \"EFNB3\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":9,"faith_total":9,"faith_pct":100.0}}