{"gene":"EFR3B","run_date":"2026-06-09T23:54:42","timeline":{"discoveries":[{"year":2014,"finding":"EFR3B (and EFR3A) are palmitoylated plasma membrane proteins; siRNA-mediated depletion of both EFR3A and EFR3B impaired the sustained phase of cytosolic Ca2+ response to angiotensin II (AngII) in HEK293 cells, and caused higher basal receptor phosphorylation and faster Gq desensitization after AngII stimulation, suggesting Efr3s contribute to controlling the phosphorylation state and desensitization of AT1a receptors.","method":"siRNA knockdown, cytosolic Ca2+ measurements, GPCR phosphorylation assays, Gq activation assays in HEK293 cells","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional knockdown with defined cellular phenotype, multiple readouts (Ca2+ response, receptor phosphorylation, Gq desensitization), single lab","pmids":["25380825"],"is_preprint":false},{"year":2021,"finding":"EFR3B is a component of the plasma membrane PI4KA (PI4KIIIα) complex (containing PI4KA, TTC7B, and FAM126A); palmitoylation of CNAβ1 recruits it to this complex, and calcineurin dephosphorylates FAM126A to promote PI4KA complex activity.","method":"Palmitoylation-dependent co-immunoprecipitation, hydrogen-deuterium exchange mass spectrometry, calcineurin substrate assay, PI4P production measurement during GPCR signaling","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods (Co-IP, HDX-MS, substrate phosphorylation assay, PI4P production assay) in a single rigorous study","pmids":["34663815"],"is_preprint":false},{"year":2021,"finding":"A tri-cysteine motif (Cys5, Cys7, Cys8) in EFR3B is differentially palmitoylated, creating distinct 'lipoforms'; spacing of palmitoyl groups controls (1) interaction between EFR3B and TMEM150A (a transmembrane protein governing PI4KIIIα complex formation for PI(4,5)P2 resynthesis after PLC signaling), and (2) EFR3B partitioning between liquid-ordered and liquid-disordered plasma membrane regions.","method":"Mutagenesis of palmitoylation sites, co-immunoprecipitation with TMEM150A, membrane domain partitioning assays, phosphoinositide homeostasis measurements","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mutagenesis of specific palmitoylation sites combined with protein-protein interaction and membrane partitioning assays, multiple orthogonal methods in one study","pmids":["34569608"],"is_preprint":false},{"year":2024,"finding":"The C-terminus of EFR3A (and by extension EFR3B as a homolog) undergoes a disorder-to-order transition upon binding to the PI4KA-TTC7B-FAM126A complex, making direct contacts with both TTC7B and FAM126A; complex-disrupting mutations in TTC7B, FAM126A, and EFR3 decrease PI4KA recruitment to the plasma membrane.","method":"Cryo-EM structure determination, hydrogen-deuterium exchange mass spectrometry, mutational analysis of binding interface, plasma membrane recruitment assays","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure plus HDX-MS plus mutagenesis in one study, extensive validation of the EFR3–PI4KA complex interface","pmids":["39705356"],"is_preprint":false},{"year":2025,"finding":"EFR3B phosphorylation markedly decreases its binding affinity to TTC7-FAM126, thereby reducing PI4KA recruitment to the plasma membrane; EFR3A-TTC7B-FAM126A binds with ~10-fold higher affinity than most other EFR3-TTC7-FAM126 combinations.","method":"Binding affinity measurements across EFR3 isoform and TTC7/FAM126 combinations, phosphorylation assays, cryo-EM, HDX-MS, nanobody competition assay, lipid bilayer and cell-based PI4P production assays","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods (binding assays, cryo-EM, HDX-MS, functional PI4P production), rigorous comparison of isoform combinations","pmids":["41197736"],"is_preprint":false},{"year":2024,"finding":"Conditional knockout of Efr3b in mouse brain (Nestin-cre) leads to hypoexcitability of CA2 pyramidal neurons and deficits in social novelty recognition; knockdown specifically in CA2 PNs recapitulates this phenotype, and re-expression of Efr3b in CA2 PNs restores excitability and social behavior, demonstrating a cell-autonomous role for Efr3b in regulating CA2 PN excitability.","method":"Conditional knockout (Nestin-cre x Efr3bf/f), region-specific knockdown, Efr3b re-expression rescue, chemogenetic (DREADD) activation, behavioral testing (social novelty recognition), electrophysiology of CA2 pyramidal neurons","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO, knockdown, rescue, and chemogenetic experiments with defined electrophysiological and behavioral readouts in multiple complementary experiments","pmids":["38190534"],"is_preprint":false},{"year":2024,"finding":"EFR3B (via its C-terminus, studied as EFR3A ortholog) recruits PI4KA to the plasma membrane by binding the PI4KA-TTC7B-FAM126A complex; this is the preprint version of the same cryo-EM/HDX-MS/mutagenesis study later published in Science Advances.","method":"Cryo-EM, HDX-MS, mutational analysis","journal":"bioRxiv","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure plus HDX-MS plus mutagenesis; superseded by peer-reviewed publication PMID:39705356","pmids":["38746453"],"is_preprint":true}],"current_model":"EFR3B is a palmitoylated peripheral plasma membrane protein that recruits the PI4KA (PI4KIIIα)–TTC7–FAM126 complex to the plasma membrane via a disorder-to-order transition of its C-terminus that contacts both TTC7B and FAM126A directly; differential palmitoylation of a tri-cysteine motif in EFR3B controls its partitioning between plasma membrane domains and its interaction with TMEM150A to regulate PI(4,5)P2 resynthesis, while phosphorylation of EFR3B markedly reduces its affinity for TTC7-FAM126, providing a regulatory switch for PI4KA membrane recruitment; EFR3B also controls AT1a receptor desensitization and, in the brain, maintains the excitability of CA2 hippocampal pyramidal neurons required for social novelty recognition."},"narrative":{"mechanistic_narrative":"EFR3B is a palmitoylated peripheral plasma membrane protein that serves as the membrane anchor recruiting the lipid kinase PI4KA to the plasma membrane, where it governs phosphoinositide homeostasis [PMID:34663815, PMID:39705356]. It does so as a component of the plasma membrane PI4KA complex containing PI4KA, TTC7B, and FAM126A, with its C-terminus undergoing a disorder-to-order transition upon binding to make direct contacts with both TTC7B and FAM126A; complex-disrupting mutations in EFR3, TTC7B, or FAM126A reduce PI4KA recruitment to the plasma membrane [PMID:39705356]. This recruitment is regulated by two layers of post-translational control: differential palmitoylation of a tri-cysteine motif (Cys5/Cys7/Cys8) creates distinct lipoforms that control both EFR3B partitioning between liquid-ordered and liquid-disordered membrane domains and its interaction with TMEM150A to tune PI(4,5)P2 resynthesis after PLC signaling [PMID:34569608], and phosphorylation of EFR3B markedly decreases its affinity for TTC7–FAM126, providing a switch that downregulates PI4KA membrane recruitment [PMID:41197736]. Through control of plasma membrane phosphoinositide signaling, EFR3B (with EFR3A) contributes to the desensitization and phosphorylation state of AT1a angiotensin receptors and the sustained Ca2+ response to angiotensin II [PMID:25380825]. In the brain, EFR3B acts cell-autonomously to maintain the excitability of CA2 hippocampal pyramidal neurons required for social novelty recognition [PMID:38190534].","teleology":[{"year":2014,"claim":"Established a cellular function for EFR3 proteins in GPCR signaling, showing they constrain receptor desensitization rather than acting as inert membrane proteins.","evidence":"siRNA double-knockdown of EFR3A/EFR3B with Ca2+, receptor phosphorylation, and Gq desensitization readouts in HEK293 cells","pmids":["25380825"],"confidence":"Medium","gaps":["Did not resolve the molecular intermediary linking EFR3 to receptor phosphorylation state","Redundancy between EFR3A and EFR3B not dissected","Direct binding partners not identified"]},{"year":2021,"claim":"Placed EFR3B within the plasma membrane PI4KA complex and linked its assembly to palmitoylation and to calcineurin-mediated dephosphorylation of FAM126A, defining a regulated lipid-kinase recruitment module.","evidence":"Palmitoylation-dependent Co-IP, HDX-MS, calcineurin substrate assay, and PI4P production measurement during GPCR signaling","pmids":["34663815"],"confidence":"High","gaps":["Structural basis of EFR3B contacts within the complex not yet resolved","Did not establish EFR3B's own phosphoregulation"]},{"year":2021,"claim":"Demonstrated that differential palmitoylation of an EFR3B tri-cysteine motif encodes distinct lipoforms controlling membrane-domain partitioning and TMEM150A interaction, connecting lipid modification to PI(4,5)P2 resynthesis.","evidence":"Mutagenesis of palmitoylation sites, Co-IP with TMEM150A, membrane partitioning and phosphoinositide homeostasis assays","pmids":["34569608"],"confidence":"High","gaps":["Enzymes setting the differential palmitoylation pattern not identified","Quantitative coupling between lipoform and downstream PI4P output not fully defined"]},{"year":2024,"claim":"Resolved the structural mechanism of recruitment, showing the EFR3 C-terminus folds upon binding to contact both TTC7B and FAM126A and that interface mutations reduce PI4KA membrane targeting.","evidence":"Cryo-EM, HDX-MS, mutational interface analysis, and plasma membrane recruitment assays (peer-reviewed PMID:39705356; preprint PMID:38746453)","pmids":["39705356","38746453"],"confidence":"High","gaps":["Structural work conducted largely on EFR3A with extension to EFR3B by homology","Does not address how palmitoylation modulates the ordered C-terminal contact"]},{"year":2024,"claim":"Defined a cell-autonomous in vivo role for EFR3B in maintaining CA2 pyramidal neuron excitability and social novelty recognition, linking the molecular module to a circuit-level behavior.","evidence":"Conditional knockout, region-specific knockdown, re-expression rescue, chemogenetic activation, electrophysiology and behavioral testing in mouse","pmids":["38190534"],"confidence":"High","gaps":["Did not directly tie the behavioral phenotype to PI4KA/phosphoinositide signaling in CA2 neurons","Effector channels or pathways downstream of EFR3B in CA2 neurons unidentified"]},{"year":2025,"claim":"Identified phosphorylation of EFR3B as a regulatory switch that lowers its affinity for TTC7–FAM126, and quantified isoform-specific binding hierarchies governing PI4KA recruitment strength.","evidence":"Binding affinity measurements across EFR3/TTC7/FAM126 combinations, phosphorylation assays, cryo-EM, HDX-MS, nanobody competition, and bilayer/cell-based PI4P assays","pmids":["41197736"],"confidence":"High","gaps":["Kinase(s) phosphorylating EFR3B in vivo not identified","Physiological context where this switch operates not defined"]},{"year":null,"claim":"How EFR3B's palmitoylation-, phosphorylation-, and TMEM150A-dependent regulation of PI4KA is integrated to produce the neuronal excitability and behavioral phenotypes remains unresolved.","evidence":"No single study connects the biochemical regulatory mechanisms to the CA2 neuron and AT1a receptor phenotypes","pmids":[],"confidence":"High","gaps":["No demonstration that phosphoinositide signaling underlies the CA2 excitability defect","Tissue-specific upstream regulators of EFR3B modification unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[1,3,4]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[0,2]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[0,1,2,3]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[1,2]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0]},{"term_id":"R-HSA-112316","term_label":"Neuronal System","supporting_discovery_ids":[5]}],"complexes":["PI4KA-TTC7B-FAM126A complex"],"partners":["PI4KA","TTC7B","FAM126A","TMEM150A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y2G0","full_name":"Protein EFR3 homolog B","aliases":[],"length_aa":817,"mass_kda":92.5,"function":"Component of a complex required to localize phosphatidylinositol 4-kinase (PI4K) to the plasma membrane (PubMed:23229899, PubMed:25608530, PubMed:26571211). The complex acts as a regulator of phosphatidylinositol 4-phosphate (PtdIns(4)P) synthesis (Probable). In the complex, EFR3B probably acts as the membrane-anchoring component (PubMed:23229899). Also involved in responsiveness to G-protein-coupled receptors; it is however unclear whether this role is direct or indirect (PubMed:25380825)","subcellular_location":"Cell membrane; Cytoplasm, cytosol","url":"https://www.uniprot.org/uniprotkb/Q9Y2G0/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/EFR3B","classification":"Not Classified","n_dependent_lines":23,"n_total_lines":1208,"dependency_fraction":0.01903973509933775},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"PI4KA","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/search/EFR3B","total_profiled":1310},"omim":[{"mim_id":"620060","title":"TETRATRICOPEPTIDE REPEAT DOMAIN-CONTAINING PROTEIN 7B; TTC7B","url":"https://www.omim.org/entry/620060"},{"mim_id":"616797","title":"EFR3 HOMOLOG B; EFR3B","url":"https://www.omim.org/entry/616797"},{"mim_id":"616757","title":"TRANSMEMBRANE PROTEIN 150A; TMEM150A","url":"https://www.omim.org/entry/616757"},{"mim_id":"611798","title":"EFR3 HOMOLOG A; EFR3A","url":"https://www.omim.org/entry/611798"},{"mim_id":"610531","title":"HYCCIN, PI4KA LIPID KINASE COMPLEX, SUBUNIT 1; HYCC1","url":"https://www.omim.org/entry/610531"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Plasma membrane","reliability":"Supported"},{"location":"Actin filaments","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"brain","ntpm":32.5},{"tissue":"skeletal muscle","ntpm":18.6}],"url":"https://www.proteinatlas.org/search/EFR3B"},"hgnc":{"alias_symbol":["FLJ37871"],"prev_symbol":["KIAA0953"]},"alphafold":{"accession":"Q9Y2G0","domains":[{"cath_id":"1.20.930","chopping":"1-130","consensus_level":"medium","plddt":85.8313,"start":1,"end":130},{"cath_id":"1.10.4140","chopping":"483-686","consensus_level":"high","plddt":90.5532,"start":483,"end":686}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y2G0","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y2G0-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y2G0-F1-predicted_aligned_error_v6.png","plddt_mean":81.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=EFR3B","jax_strain_url":"https://www.jax.org/strain/search?query=EFR3B"},"sequence":{"accession":"Q9Y2G0","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y2G0.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y2G0/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y2G0"}},"corpus_meta":[{"pmid":"21980299","id":"PMC_21980299","title":"A genome-wide meta-analysis of six type 1 diabetes cohorts identifies multiple associated loci.","date":"2011","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/21980299","citation_count":261,"is_preprint":false},{"pmid":"25380825","id":"PMC_25380825","title":"EFR3s are palmitoylated plasma membrane proteins that control responsiveness to G-protein-coupled receptors.","date":"2014","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/25380825","citation_count":44,"is_preprint":false},{"pmid":"34663815","id":"PMC_34663815","title":"Palmitoylation targets the calcineurin phosphatase to the phosphatidylinositol 4-kinase complex at the plasma membrane.","date":"2021","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/34663815","citation_count":42,"is_preprint":false},{"pmid":"34569608","id":"PMC_34569608","title":"A palmitoylation code controls PI4KIIIα complex formation and PI(4,5)P2 homeostasis at the plasma membrane.","date":"2021","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/34569608","citation_count":21,"is_preprint":false},{"pmid":"39705356","id":"PMC_39705356","title":"Molecular basis for plasma membrane recruitment of PI4KA by EFR3.","date":"2024","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/39705356","citation_count":13,"is_preprint":false},{"pmid":"38190534","id":"PMC_38190534","title":"Efr3b is essential for social recognition by modulating the excitability of CA2 pyramidal neurons.","date":"2024","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/38190534","citation_count":10,"is_preprint":false},{"pmid":"31798789","id":"PMC_31798789","title":"Type 1 diabetes loci display a variety of native American and African ancestries in diseased individuals from Northwest Colombia.","date":"2019","source":"World journal of diabetes","url":"https://pubmed.ncbi.nlm.nih.gov/31798789","citation_count":5,"is_preprint":false},{"pmid":"39439909","id":"PMC_39439909","title":"IMPC-based screening revealed that ROBO1 can regulate osteoporosis by inhibiting osteogenic differentiation.","date":"2024","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/39439909","citation_count":3,"is_preprint":false},{"pmid":"39596826","id":"PMC_39596826","title":"Cross-Tissue Regulatory Network Analyses Reveal Novel Susceptibility Genes and Potential Mechanisms for Endometriosis.","date":"2024","source":"Biology","url":"https://pubmed.ncbi.nlm.nih.gov/39596826","citation_count":2,"is_preprint":false},{"pmid":"41610418","id":"PMC_41610418","title":"Common variation at 1q23.3, 2p23.3, 2q33.3, and 2p21 influences the risk of acute myeloid leukemia.","date":"2026","source":"Blood","url":"https://pubmed.ncbi.nlm.nih.gov/41610418","citation_count":2,"is_preprint":false},{"pmid":"38746453","id":"PMC_38746453","title":"Molecular basis for plasma membrane recruitment of PI4KA by EFR3.","date":"2024","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/38746453","citation_count":1,"is_preprint":false},{"pmid":"41197736","id":"PMC_41197736","title":"Development of an inhibitory TTC7B selective nanobody that blocks EFR3 recruitment of PI4KA.","date":"2025","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/41197736","citation_count":0,"is_preprint":false},{"pmid":"40305161","id":"PMC_40305161","title":"A Potential Role of EFR3A in Human Disease States.","date":"2025","source":"Biomolecules","url":"https://pubmed.ncbi.nlm.nih.gov/40305161","citation_count":0,"is_preprint":false},{"pmid":"41473329","id":"PMC_41473329","title":"Development of an inhibitory TTC7B selective nanobody that blocks EFR3 recruitment of PI4KA.","date":"2025","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/41473329","citation_count":0,"is_preprint":false},{"pmid":"40629317","id":"PMC_40629317","title":"The related SNPs and genes to body size using GWAS- latent variable modeling in dromedaries.","date":"2025","source":"BMC genomics","url":"https://pubmed.ncbi.nlm.nih.gov/40629317","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.10.29.24316311","title":"Transcriptome-wide association study of Alzheimer disease reveals many differentially expressed genes and multiple biological pathways in brain tissue from African American donors","date":"2024-11-04","source":"bioRxiv","url":"https://doi.org/10.1101/2024.10.29.24316311","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":10537,"output_tokens":2111,"usd":0.031638,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9064,"output_tokens":2723,"usd":0.056697,"stage2_stop_reason":"end_turn"},"total_usd":0.088335,"stage1_batch_id":"msgbatch_01AsR7g8dGTEiEw9U83bJeWn","stage2_batch_id":"msgbatch_01YV5ZyxtFNUZEjxRcpd9jzb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"EFR3B (and EFR3A) are palmitoylated plasma membrane proteins; siRNA-mediated depletion of both EFR3A and EFR3B impaired the sustained phase of cytosolic Ca2+ response to angiotensin II (AngII) in HEK293 cells, and caused higher basal receptor phosphorylation and faster Gq desensitization after AngII stimulation, suggesting Efr3s contribute to controlling the phosphorylation state and desensitization of AT1a receptors.\",\n      \"method\": \"siRNA knockdown, cytosolic Ca2+ measurements, GPCR phosphorylation assays, Gq activation assays in HEK293 cells\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional knockdown with defined cellular phenotype, multiple readouts (Ca2+ response, receptor phosphorylation, Gq desensitization), single lab\",\n      \"pmids\": [\"25380825\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"EFR3B is a component of the plasma membrane PI4KA (PI4KIIIα) complex (containing PI4KA, TTC7B, and FAM126A); palmitoylation of CNAβ1 recruits it to this complex, and calcineurin dephosphorylates FAM126A to promote PI4KA complex activity.\",\n      \"method\": \"Palmitoylation-dependent co-immunoprecipitation, hydrogen-deuterium exchange mass spectrometry, calcineurin substrate assay, PI4P production measurement during GPCR signaling\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods (Co-IP, HDX-MS, substrate phosphorylation assay, PI4P production assay) in a single rigorous study\",\n      \"pmids\": [\"34663815\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"A tri-cysteine motif (Cys5, Cys7, Cys8) in EFR3B is differentially palmitoylated, creating distinct 'lipoforms'; spacing of palmitoyl groups controls (1) interaction between EFR3B and TMEM150A (a transmembrane protein governing PI4KIIIα complex formation for PI(4,5)P2 resynthesis after PLC signaling), and (2) EFR3B partitioning between liquid-ordered and liquid-disordered plasma membrane regions.\",\n      \"method\": \"Mutagenesis of palmitoylation sites, co-immunoprecipitation with TMEM150A, membrane domain partitioning assays, phosphoinositide homeostasis measurements\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis of specific palmitoylation sites combined with protein-protein interaction and membrane partitioning assays, multiple orthogonal methods in one study\",\n      \"pmids\": [\"34569608\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The C-terminus of EFR3A (and by extension EFR3B as a homolog) undergoes a disorder-to-order transition upon binding to the PI4KA-TTC7B-FAM126A complex, making direct contacts with both TTC7B and FAM126A; complex-disrupting mutations in TTC7B, FAM126A, and EFR3 decrease PI4KA recruitment to the plasma membrane.\",\n      \"method\": \"Cryo-EM structure determination, hydrogen-deuterium exchange mass spectrometry, mutational analysis of binding interface, plasma membrane recruitment assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure plus HDX-MS plus mutagenesis in one study, extensive validation of the EFR3–PI4KA complex interface\",\n      \"pmids\": [\"39705356\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"EFR3B phosphorylation markedly decreases its binding affinity to TTC7-FAM126, thereby reducing PI4KA recruitment to the plasma membrane; EFR3A-TTC7B-FAM126A binds with ~10-fold higher affinity than most other EFR3-TTC7-FAM126 combinations.\",\n      \"method\": \"Binding affinity measurements across EFR3 isoform and TTC7/FAM126 combinations, phosphorylation assays, cryo-EM, HDX-MS, nanobody competition assay, lipid bilayer and cell-based PI4P production assays\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods (binding assays, cryo-EM, HDX-MS, functional PI4P production), rigorous comparison of isoform combinations\",\n      \"pmids\": [\"41197736\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Conditional knockout of Efr3b in mouse brain (Nestin-cre) leads to hypoexcitability of CA2 pyramidal neurons and deficits in social novelty recognition; knockdown specifically in CA2 PNs recapitulates this phenotype, and re-expression of Efr3b in CA2 PNs restores excitability and social behavior, demonstrating a cell-autonomous role for Efr3b in regulating CA2 PN excitability.\",\n      \"method\": \"Conditional knockout (Nestin-cre x Efr3bf/f), region-specific knockdown, Efr3b re-expression rescue, chemogenetic (DREADD) activation, behavioral testing (social novelty recognition), electrophysiology of CA2 pyramidal neurons\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO, knockdown, rescue, and chemogenetic experiments with defined electrophysiological and behavioral readouts in multiple complementary experiments\",\n      \"pmids\": [\"38190534\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"EFR3B (via its C-terminus, studied as EFR3A ortholog) recruits PI4KA to the plasma membrane by binding the PI4KA-TTC7B-FAM126A complex; this is the preprint version of the same cryo-EM/HDX-MS/mutagenesis study later published in Science Advances.\",\n      \"method\": \"Cryo-EM, HDX-MS, mutational analysis\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure plus HDX-MS plus mutagenesis; superseded by peer-reviewed publication PMID:39705356\",\n      \"pmids\": [\"38746453\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"EFR3B is a palmitoylated peripheral plasma membrane protein that recruits the PI4KA (PI4KIIIα)–TTC7–FAM126 complex to the plasma membrane via a disorder-to-order transition of its C-terminus that contacts both TTC7B and FAM126A directly; differential palmitoylation of a tri-cysteine motif in EFR3B controls its partitioning between plasma membrane domains and its interaction with TMEM150A to regulate PI(4,5)P2 resynthesis, while phosphorylation of EFR3B markedly reduces its affinity for TTC7-FAM126, providing a regulatory switch for PI4KA membrane recruitment; EFR3B also controls AT1a receptor desensitization and, in the brain, maintains the excitability of CA2 hippocampal pyramidal neurons required for social novelty recognition.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"EFR3B is a palmitoylated peripheral plasma membrane protein that serves as the membrane anchor recruiting the lipid kinase PI4KA to the plasma membrane, where it governs phosphoinositide homeostasis [#1, #3]. It does so as a component of the plasma membrane PI4KA complex containing PI4KA, TTC7B, and FAM126A, with its C-terminus undergoing a disorder-to-order transition upon binding to make direct contacts with both TTC7B and FAM126A; complex-disrupting mutations in EFR3, TTC7B, or FAM126A reduce PI4KA recruitment to the plasma membrane [#3]. This recruitment is regulated by two layers of post-translational control: differential palmitoylation of a tri-cysteine motif (Cys5/Cys7/Cys8) creates distinct lipoforms that control both EFR3B partitioning between liquid-ordered and liquid-disordered membrane domains and its interaction with TMEM150A to tune PI(4,5)P2 resynthesis after PLC signaling [#2], and phosphorylation of EFR3B markedly decreases its affinity for TTC7\\u2013FAM126, providing a switch that downregulates PI4KA membrane recruitment [#4]. Through control of plasma membrane phosphoinositide signaling, EFR3B (with EFR3A) contributes to the desensitization and phosphorylation state of AT1a angiotensin receptors and the sustained Ca2+ response to angiotensin II [#0]. In the brain, EFR3B acts cell-autonomously to maintain the excitability of CA2 hippocampal pyramidal neurons required for social novelty recognition [#5].\",\n  \"teleology\": [\n    {\n      \"year\": 2014,\n      \"claim\": \"Established a cellular function for EFR3 proteins in GPCR signaling, showing they constrain receptor desensitization rather than acting as inert membrane proteins.\",\n      \"evidence\": \"siRNA double-knockdown of EFR3A/EFR3B with Ca2+, receptor phosphorylation, and Gq desensitization readouts in HEK293 cells\",\n      \"pmids\": [\"25380825\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not resolve the molecular intermediary linking EFR3 to receptor phosphorylation state\", \"Redundancy between EFR3A and EFR3B not dissected\", \"Direct binding partners not identified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Placed EFR3B within the plasma membrane PI4KA complex and linked its assembly to palmitoylation and to calcineurin-mediated dephosphorylation of FAM126A, defining a regulated lipid-kinase recruitment module.\",\n      \"evidence\": \"Palmitoylation-dependent Co-IP, HDX-MS, calcineurin substrate assay, and PI4P production measurement during GPCR signaling\",\n      \"pmids\": [\"34663815\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of EFR3B contacts within the complex not yet resolved\", \"Did not establish EFR3B's own phosphoregulation\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated that differential palmitoylation of an EFR3B tri-cysteine motif encodes distinct lipoforms controlling membrane-domain partitioning and TMEM150A interaction, connecting lipid modification to PI(4,5)P2 resynthesis.\",\n      \"evidence\": \"Mutagenesis of palmitoylation sites, Co-IP with TMEM150A, membrane partitioning and phosphoinositide homeostasis assays\",\n      \"pmids\": [\"34569608\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Enzymes setting the differential palmitoylation pattern not identified\", \"Quantitative coupling between lipoform and downstream PI4P output not fully defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Resolved the structural mechanism of recruitment, showing the EFR3 C-terminus folds upon binding to contact both TTC7B and FAM126A and that interface mutations reduce PI4KA membrane targeting.\",\n      \"evidence\": \"Cryo-EM, HDX-MS, mutational interface analysis, and plasma membrane recruitment assays (peer-reviewed PMID:39705356; preprint PMID:38746453)\",\n      \"pmids\": [\"39705356\", \"38746453\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural work conducted largely on EFR3A with extension to EFR3B by homology\", \"Does not address how palmitoylation modulates the ordered C-terminal contact\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined a cell-autonomous in vivo role for EFR3B in maintaining CA2 pyramidal neuron excitability and social novelty recognition, linking the molecular module to a circuit-level behavior.\",\n      \"evidence\": \"Conditional knockout, region-specific knockdown, re-expression rescue, chemogenetic activation, electrophysiology and behavioral testing in mouse\",\n      \"pmids\": [\"38190534\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not directly tie the behavioral phenotype to PI4KA/phosphoinositide signaling in CA2 neurons\", \"Effector channels or pathways downstream of EFR3B in CA2 neurons unidentified\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified phosphorylation of EFR3B as a regulatory switch that lowers its affinity for TTC7\\u2013FAM126, and quantified isoform-specific binding hierarchies governing PI4KA recruitment strength.\",\n      \"evidence\": \"Binding affinity measurements across EFR3/TTC7/FAM126 combinations, phosphorylation assays, cryo-EM, HDX-MS, nanobody competition, and bilayer/cell-based PI4P assays\",\n      \"pmids\": [\"41197736\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase(s) phosphorylating EFR3B in vivo not identified\", \"Physiological context where this switch operates not defined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How EFR3B's palmitoylation-, phosphorylation-, and TMEM150A-dependent regulation of PI4KA is integrated to produce the neuronal excitability and behavioral phenotypes remains unresolved.\",\n      \"evidence\": \"No single study connects the biochemical regulatory mechanisms to the CA2 neuron and AT1a receptor phenotypes\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No demonstration that phosphoinositide signaling underlies the CA2 excitability defect\", \"Tissue-specific upstream regulators of EFR3B modification unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [1, 3, 4]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [0, 2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [0, 1, 2, 3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [1, 2]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"R-HSA-112316\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"complexes\": [\"PI4KA-TTC7B-FAM126A complex\"],\n    \"partners\": [\"PI4KA\", \"TTC7B\", \"FAM126A\", \"TMEM150A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}