{"gene":"PI4KA","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":1999,"finding":"Human PI4KA (PI4K230) encodes a 2044-amino acid protein with a C-terminal catalytic domain (~260 aa) that is highly conserved among PI 4-kinases. When expressed in vitro and in Sf9 cells, it exhibits type III PI 4-kinase characteristics: not inhibited by adenosine, high Km for ATP (~300 µM), half-maximally inactivated by ~200 nM wortmannin, and specific activity of 58 µmol mg⁻¹ min⁻¹.","method":"In vitro expression, enzymatic activity assays, pharmacological characterization","journal":"Biochimica et biophysica acta","confidence":"High","confidence_rationale":"Tier 1 / Moderate — direct in vitro enzymatic reconstitution with pharmacological characterization, single lab but multiple orthogonal assays","pmids":["10101268"],"is_preprint":false},{"year":2001,"finding":"The conserved Lys-1792 in the catalytic domain of PI4K230 (PI4KA) is essential for enzymatic activity and serves as a target of affinity labeling by the ATP-analog FSBA. ATP and ATP analogues competitively protect against FSBA inactivation, confirming this residue's proximity to the ATP-binding site.","method":"Site-directed mutagenesis, affinity labeling with FSBA, enzymatic activity assays","journal":"The international journal of biochemistry & cell biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — active-site mutagenesis combined with affinity labeling and competitive protection assays, single lab with multiple orthogonal methods","pmids":["11311856"],"is_preprint":false},{"year":2000,"finding":"PI4K230 (PI4KA) localizes in neurons predominantly to the outer membrane of mitochondria and membranes of the rough endoplasmic reticulum, and is also co-localized with some multivesicular bodies. No nuclear or plasma membrane localization was detected, suggesting its role in intracellular vesicular traffic rather than direct ligand-stimulated phosphoinositide turnover at the plasma membrane.","method":"Immunocytochemistry (light and electron microscopy) with isoform-specific antibodies in rat brain sections","journal":"Experimental brain research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — direct localization by electron microscopy with specific antibodies, replicated across multiple brain regions, but no functional perturbation experiment","pmids":["11045352"],"is_preprint":false},{"year":2006,"finding":"PI4K230 (PI4KA) is detected in the nucleolus of multiple mammalian cell types, forming a Triton X-100-resistant, DNase- and RNase-sensitive complex there. siRNA knockdown of PI4K230 abolishes the nucleolar signal, confirming specificity.","method":"Immunofluorescence on ethanol-fixed cells and cryosections, siRNA knockdown, DNase/RNase treatments, Triton X-100 extraction","journal":"Cytometry. Part A","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization with siRNA validation and biochemical fractionation, single lab with multiple orthogonal methods","pmids":["17131383"],"is_preprint":false},{"year":2008,"finding":"PI4KA contains a functional monopartite NLS (NLS1: residues 916–934) that directs nuclear import via importin α1 and α3 (but not α5) through the classical importin α/β mechanism. A separate bipartite NLS2 (residues 1414–1433) functions as a nucleolar targeting signal and can mediate nuclear import via importin α1/β or α3/β complexes when present in a larger fragment (AA1166–1667).","method":"Digitonin-permeabilized HeLa cell nuclear import assays, fluorescent BSA conjugates with synthetic NLS peptides, molecular modeling, importin binding experiments","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reconstituted nuclear import assays with defined importins, multiple peptide constructs, single lab with several orthogonal approaches","pmids":["18585705"],"is_preprint":false},{"year":2011,"finding":"PI4KA is required for local enrichment of phosphatidylinositol 4-phosphate (PI4P) at the HCV membranous web and for morphologically normal web formation. PI4KA physically associates with NS5A in HCV-infected cells, whereas the related kinase PI4KB does not interact with NS5A and does not affect web morphology or PI4P enrichment upon silencing.","method":"RNAi silencing, co-immunoprecipitation (PI4KA with NS5A), immunofluorescence/confocal microscopy of PI4P at membranous webs, non-replicative web formation model","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP plus functional RNAi phenotype with PI4P localization readout, single lab with multiple orthogonal methods","pmids":["22022594"],"is_preprint":false},{"year":2014,"finding":"PI4KA is the primary enzyme maintaining plasma membrane PI4P pools and is essential for replenishing PI(4,5)P2 specifically during strong stimulation of phospholipase C-coupled receptors. Pharmacological inhibition of PI4KA in adult mice causes sudden death correlating with PI(4,5)P2 depletion after agonist stimulation; conditional genetic knockout causes severe intestinal necrosis and death.","method":"Highly specific PI4KA inhibitors (pharmacological), conditional knockout mice, phosphoinositide level measurements after PLC stimulation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — orthogonal pharmacological and genetic loss-of-function approaches with defined lipid and phenotypic readouts, multiple in vitro and in vivo experiments","pmids":["24415756"],"is_preprint":false},{"year":2015,"finding":"A missense substitution p.D1854N in the catalytic domain of PI4KA, identified in patients with polymicrogyria and cerebellar hypoplasia, abolishes kinase activity, demonstrating that residue D1854 is essential for PI4KA catalytic function.","method":"Expression of wild-type and mutant PI4KA with in vitro kinase activity assay; Sanger sequencing confirmation of segregation","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 1 / Moderate — direct in vitro kinase activity assay with disease-linked catalytic domain mutant, complemented by genetic segregation analysis","pmids":["25855803"],"is_preprint":false},{"year":2021,"finding":"PI4KA forms a heterotetrameric complex with EFR3, TTC7, and FAM126 at the plasma membrane to generate PI4P. Biallelic loss-of-function PI4KA variants reduce PI4KA protein levels, decrease PI4KA catalytic activity (assessed by immunofluorescence and targeted lipidomics in patient fibroblasts and PBMCs), and cause hypomyelinating leukodystrophy, inflammatory bowel disease, intestinal atresia, and combined immunodeficiency in humans.","method":"Western blotting, immunofluorescence, targeted lipidomics in patient-derived fibroblasts and PBMCs; structural modelling of complex interfaces; exome sequencing","journal":"Brain : a journal of neurology","confidence":"High","confidence_rationale":"Tier 2 / Strong — two independent studies (PMIDs 34415310 and 34415322) using multiple orthogonal methods in patient-derived cells, replicating PI4KA's role in the complex and PI4P production","pmids":["34415310","34415322"],"is_preprint":false},{"year":2021,"finding":"Oncogenic KRAS preferentially binds EFR3A (the PI4KA plasma membrane recruitment adaptor). Disrupting EFR3A or PI4KA reduces PI4P, phosphatidylserine, and KRAS levels at the plasma membrane, diminishing oncogenic signaling and tumorigenesis. Tethering PI4KA directly to the plasma membrane rescues these phenotypes, demonstrating that EFR3A-PI4KA axis supports KRAS plasma membrane localization and signaling.","method":"Co-immunoprecipitation (KRAS–EFR3A), siRNA/genetic disruption of EFR3A and PI4KA, PI4P and phosphatidylserine lipid measurements at PM, KRAS PM localization assays, PI4KA tethering rescue experiment, xenograft tumor assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP plus multiple functional rescue and KO experiments with defined lipid and signaling readouts, single lab with multiple orthogonal methods","pmids":["34504076"],"is_preprint":false},{"year":2023,"finding":"CXCR4 binds to PI4KA adaptor protein TTC7, and this interaction drives plasma membrane PI4P production in prostate cancer cells. Inhibiting PI4KA or TTC7 reduces PM PI4P production, cellular invasion, and bone tumor growth.","method":"Co-immunoprecipitation (CXCR4–TTC7), PI4P measurement at plasma membrane, invasion assays, in vivo bone tumor models with PI4KA/TTC7 inhibition","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP of CXCR4 with TTC7 adaptor, functional inhibition with PI4P readout and in vivo validation, single lab with multiple assays","pmids":["37996444"],"is_preprint":false},{"year":2024,"finding":"The C terminus of EFR3A undergoes a disorder-to-order transition upon binding to the PI4KA–TTC7B–FAM126A heterotrimer, directly contacting both TTC7B and FAM126A. Mutations disrupting this interface reduce PI4KA recruitment to the plasma membrane. Multiple disease-linked mutations and post-translational modifications map to this binding site.","method":"Cryo-EM structure determination, hydrogen-deuterium exchange mass spectrometry (HDX-MS), mutational analysis of complex interface","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with HDX-MS validation and mutagenesis, peer-reviewed; also independently reported as a preprint (PMID 38746453)","pmids":["39705356","38746453"],"is_preprint":false},{"year":2024,"finding":"Calcineurin (CNAβ1 isoform) directly binds to PI4KA via an evolutionarily conserved IKISVT sequence in PI4KA's horn domain, and also binds FAM126A via conserved LTLT and PSISIT sequences. These dual calcineurin-binding sites are in close proximity to phosphorylation sites in the PI4KA complex, suggesting calcineurin regulates PI4KA through dephosphorylation of the complex.","method":"Cryo-EM structure of truncated PI4KA complex with calcineurin, HDX-MS, computational analysis of binding interfaces","journal":"Structure","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with HDX-MS validation, single lab with multiple orthogonal structural methods","pmids":["39216471"],"is_preprint":false},{"year":2018,"finding":"Targeted inactivation of the Pi4ka catalytic domain or reduction in Pi4ka mRNA expression inhibits myeloid and erythroid cell differentiation in vitro and promotes anemia in vivo. The mechanism involves deregulation of AKT, MAPK, SRC, and JAK-STAT signaling pathways downstream of Pi4ka activity.","method":"Catalytic domain inactivation, mRNA knockdown, in vitro differentiation assays, in vivo anemia model, signaling pathway analysis (AKT, MAPK, SRC, JAK-STAT)","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic inactivation of catalytic domain with defined hematopoietic phenotype and pathway readouts, single lab with in vitro and in vivo experiments","pmids":["29386109"],"is_preprint":false},{"year":2022,"finding":"PI4KA forms a stable complex with TTC7 and FAM126; a novel PI4KA inhibitor (cepharanthine) undermines the stability of this PI4KA/TTC7/FAM126 complex. PI4KA depletion sensitizes drug-resistant leukemia cells to chemotherapy by regulating the ERK/AMPK/OXPHOS axis.","method":"LC-MS, ELISA kinase assay, MM/GBSA binding analysis, siRNA knockdown, in vitro and in vivo leukemia models","journal":"Theranostics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical identification of PI4KA inhibitor targeting the complex, with functional in vitro and in vivo knockdown data and signaling pathway readouts, single lab","pmids":["36276647"],"is_preprint":false},{"year":2025,"finding":"A TTC7B-selective nanobody was developed that sterically blocks EFR3 binding to TTC7B in PI4KA complexes. EFR3B phosphorylation markedly decreases its binding affinity to TTC7-FAM126. EFR3A-TTC7B-FAM126A binds with ~10-fold higher affinity than most other EFR3-TTC7-FAM126 combinations. Nanobody binding causes decreased PI4KA membrane recruitment on lipid bilayers and in cells, with decreased PM PI4P production.","method":"Cryo-EM, HDX-MS, yeast display nanobody selection, binding affinity measurements, lipid bilayer recruitment assay, cell-based PM PI4P measurement","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with HDX-MS and functional cell/bilayer assays, multiple orthogonal methods in single study; also reported as preprint (PMID 41473329)","pmids":["41197736","41473329"],"is_preprint":false},{"year":2026,"finding":"In response to Ca2+ signaling, the ER membrane tethering protein E-Syt1 recruits ER-localized PI4KA to ER-PM junctions, facilitating PI4KA's plasma membrane recruitment and assembly of the PI4KA enzyme complex, thereby enhancing PM PI4P synthesis. In hippocampal neurons, neuronal activity-induced PM localization of PI4KA and PM PI4P synthesis depend on E-Syt1 function.","method":"Live cell imaging of PI4KA localization, Ca2+ stimulation experiments, E-Syt1 knockdown/knockout, PI4P biosensor measurements at PM, hippocampal neuron synaptic potentiation assays","journal":"Science China. Life sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct subcellular localization linked to functional PI4P synthesis outcome, with genetic perturbation of E-Syt1, single lab","pmids":["42258130"],"is_preprint":false},{"year":2025,"finding":"PI4KA inhibition reduces PM PI4P levels, which triggers a compensatory increase in phospholipase D (PLD) activity and phosphatidic acid (PA) levels at the PM. This is mediated by a concomitant decrease in phosphatidylserine (PS) levels upon PI4P depletion, activating a reciprocal relationship between PS synthesis and PLD-mediated PA generation. Additionally, loss of PM PI4P upregulates the small GTPase RhoB transcriptionally and translationally, which further enhances PLD-mediated PA synthesis and actin cytoskeletal remodeling.","method":"Pharmacological PI4KA inhibition, RNA-seq, proximity labeling proteomics, lipid measurements (PI4P, PS, PA), RhoB protein/mRNA quantification, PLD activity assays","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multi-omic approach (RNA-seq, proximity proteomics, lipidomics) with pharmacological perturbation, single lab; preprint not yet peer-reviewed","pmids":["bio_10.1101_2025.09.30.679611"],"is_preprint":true},{"year":2025,"finding":"The PI4KA variant c.2819C>T (p.Ala940Val) significantly reduces PI4KA enzyme activity when assessed using purified mutant protein fragments, and a deletion variant c.2802_2863-40del produces three distinct aberrant mRNA isoforms via splicing disruption, together demonstrating catalytic domain and splice-site requirements for PI4KA function.","method":"Minigene splicing assay, purification of recombinant wild-type and mutant PI4KA fragments, in vitro kinase activity measurement","journal":"BMC medical genomics","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — direct in vitro kinase assay on purified mutant protein and minigene splicing assay, single case study, single lab","pmids":["39885450"],"is_preprint":false}],"current_model":"PI4KA (PI4KIIIα) is a type III lipid kinase that phosphorylates phosphatidylinositol at the D4 position to generate the predominant plasma membrane pool of PI4P, which is essential for PI(4,5)P2 replenishment during PLC stimulation; it exists primarily as a heterotrimer with TTC7 and FAM126, is recruited to the plasma membrane by lipidated EFR3A/B proteins (whose C-termini undergo a disorder-to-order transition upon binding both TTC7 and FAM126), can be further recruited to ER-PM junctions by Ca²⁺-activated E-Syt1, and is regulated by calcineurin binding to conserved sites on both PI4KA and FAM126A, with its catalytic activity requiring conserved residues including Lys-1792 and Asp-1854 in the kinase domain; loss of PI4KA activity disrupts plasma membrane lipid homeostasis, triggers compensatory RhoB/PLD/phosphatidic acid signaling, impairs KRAS and CXCR4 signaling at the plasma membrane, and in humans causes a spectrum of neurological, intestinal, and immunological diseases."},"narrative":{"mechanistic_narrative":"PI4KA (PI4KIIIα) is a type III phosphatidylinositol 4-kinase that generates the predominant plasma membrane pool of PI4P and is the primary enzyme replenishing PI(4,5)P2 during strong stimulation of phospholipase C-coupled receptors, making it essential for plasma membrane lipid homeostasis [PMID:10101268, PMID:24415756]. Its catalytic activity resides in a conserved C-terminal kinase domain requiring Lys-1792, which lies adjacent to the ATP-binding site, and Asp-1854 [PMID:11311856, PMID:25855803]. At the plasma membrane PI4KA functions within a heterotetrameric assembly with TTC7, FAM126, and the lipidated recruitment adaptor EFR3, whose disordered C-terminus undergoes a disorder-to-order transition to contact both TTC7B and FAM126A, an interface that controls PI4KA membrane recruitment and PI4P output [PMID:34415310, PMID:34415322, PMID:39705356, PMID:38746453, PMID:41197736, PMID:41473329]. Recruitment to ER–PM junctions is further driven by Ca²⁺-activated E-Syt1, linking neuronal activity to local PI4P synthesis [PMID:42258130], while calcineurin docks on conserved sites in the PI4KA horn domain and in FAM126A, positioning the phosphatase to regulate the complex by dephosphorylation [PMID:39216471]. Through this PI4P-generating activity PI4KA supports plasma membrane localization and signaling of oncogenic KRAS via the EFR3A adaptor and of CXCR4 via TTC7, and its inhibition triggers compensatory RhoB/PLD/phosphatidic acid signaling upon PI4P and phosphatidylserine depletion [PMID:34504076, PMID:37996444, PMID:bio_10.1101_2025.09.30.679611]. Biallelic and missense loss-of-function PI4KA variants that reduce kinase activity or protein levels cause a human spectrum of hypomyelinating leukodystrophy, polymicrogyria with cerebellar hypoplasia, inflammatory bowel disease, intestinal atresia, and combined immunodeficiency [PMID:25855803, PMID:34415310, PMID:34415322, PMID:39885450].","teleology":[{"year":1999,"claim":"Established that human PI4KA encodes a large protein whose conserved C-terminal domain confers type III PI 4-kinase enzymatic activity, defining its biochemical identity.","evidence":"In vitro expression in Sf9 cells with enzymatic and pharmacological characterization","pmids":["10101268"],"confidence":"High","gaps":["Did not define cellular substrate pool or in vivo localization","No structural model of the catalytic domain"]},{"year":2001,"claim":"Identified Lys-1792 as a catalytically essential residue near the ATP-binding site, providing the first active-site landmark for the kinase.","evidence":"Site-directed mutagenesis and FSBA affinity labeling with competitive ATP protection","pmids":["11311856"],"confidence":"High","gaps":["Did not resolve the full catalytic mechanism","No co-structure with ATP"]},{"year":2000,"claim":"Addressed where PI4KA acts in cells, initially localizing it to mitochondrial outer membrane and rough ER in neurons rather than the plasma membrane.","evidence":"Immuno-EM with isoform-specific antibodies in rat brain","pmids":["11045352"],"confidence":"Medium","gaps":["No functional perturbation linking localization to activity","Apparent conflict with later plasma membrane / nucleolar localization unresolved"]},{"year":2006,"claim":"Detected a nuclease-sensitive nucleolar pool of PI4KA, raising the possibility of functions beyond membrane lipid signaling.","evidence":"Immunofluorescence with siRNA validation, DNase/RNase treatment and detergent extraction","pmids":["17131383"],"confidence":"Medium","gaps":["No nucleolar function or substrate defined","Relationship to cytoplasmic PI4P pools unknown"]},{"year":2008,"claim":"Mapped the import determinants for the nuclear/nucleolar pool, identifying a classical NLS and a separate nucleolar targeting signal.","evidence":"Digitonin-permeabilized nuclear import assays with synthetic NLS peptides and defined importins","pmids":["18585705"],"confidence":"Medium","gaps":["No demonstrated function of nuclear-imported PI4KA","Physiological regulation of import unknown"]},{"year":2011,"claim":"Connected PI4KA to a pathogen-driven membrane remodeling event, showing it associates with HCV NS5A to enrich PI4P at the viral membranous web.","evidence":"RNAi silencing, co-IP of PI4KA with NS5A, PI4P imaging at membranous webs","pmids":["22022594"],"confidence":"Medium","gaps":["Did not establish whether NS5A directly activates the kinase","No structural basis for the interaction"]},{"year":2014,"claim":"Defined PI4KA's core physiological role as the maintainer of plasma membrane PI4P and replenisher of PI(4,5)P2 during PLC signaling, with loss being lethal.","evidence":"Specific pharmacological inhibitors plus conditional knockout mice with phosphoinositide measurements","pmids":["24415756"],"confidence":"High","gaps":["Did not resolve how the kinase is recruited to the plasma membrane","Tissue-specific requirements not fully dissected"]},{"year":2015,"claim":"Provided the first catalytic disease residue, showing the polymicrogyria-associated D1854N abolishes kinase activity.","evidence":"In vitro kinase assay of disease mutant plus segregation analysis","pmids":["25855803"],"confidence":"High","gaps":["Did not model how loss of activity causes the specific neurodevelopmental phenotype"]},{"year":2018,"claim":"Extended PI4KA function to hematopoiesis, linking its catalytic activity to myeloid/erythroid differentiation via multiple signaling pathways.","evidence":"Catalytic-domain inactivation and knockdown with in vitro differentiation and in vivo anemia assays","pmids":["29386109"],"confidence":"Medium","gaps":["Causal chain from PI4P to AKT/MAPK/SRC/JAK-STAT not directly resolved","Direct effectors unidentified"]},{"year":2021,"claim":"Established the heterotetrameric PI4KA–EFR3–TTC7–FAM126 complex as the functional plasma membrane PI4P-generating unit and tied biallelic loss-of-function variants to a multisystem human disease spectrum.","evidence":"Two independent patient-cell studies with western blot, lipidomics, structural modeling and exome sequencing","pmids":["34415310","34415322"],"confidence":"High","gaps":["Did not provide high-resolution structure of the assembled complex","Genotype-phenotype determinants across organ systems unclear"]},{"year":2021,"claim":"Demonstrated that the EFR3A–PI4KA axis supports oncogenic KRAS plasma membrane localization and signaling, implicating PI4KA in tumorigenesis.","evidence":"KRAS–EFR3A co-IP, EFR3A/PI4KA disruption, lipid and KRAS PM measurements, tethering rescue and xenograft assays","pmids":["34504076"],"confidence":"High","gaps":["Whether KRAS directly modulates kinase activity not shown","Generalizability across KRAS-driven tumors untested"]},{"year":2023,"claim":"Showed PI4KA, via its TTC7 adaptor, supports CXCR4-driven plasma membrane PI4P and prostate cancer invasion, broadening its receptor-coupled signaling roles.","evidence":"CXCR4–TTC7 co-IP, PM PI4P measurement, invasion and in vivo bone tumor models","pmids":["37996444"],"confidence":"Medium","gaps":["Single co-IP without structural mapping of the CXCR4–TTC7 interface","Direct vs indirect coupling unresolved"]},{"year":2024,"claim":"Resolved the molecular basis of plasma membrane recruitment, showing the EFR3A C-terminus folds upon binding TTC7B and FAM126A and that this interface harbors disease mutations.","evidence":"Cryo-EM, HDX-MS and interface mutagenesis of the PI4KA–TTC7B–FAM126A–EFR3A assembly","pmids":["39705356","38746453"],"confidence":"High","gaps":["Dynamics of lipidation-driven membrane targeting not fully captured","How modifications toggle the interface in vivo untested"]},{"year":2024,"claim":"Identified calcineurin as a direct regulator that docks on conserved sites in PI4KA and FAM126A near phosphorylation sites, implying phosphatase control of the complex.","evidence":"Cryo-EM of a truncated PI4KA–calcineurin complex with HDX-MS","pmids":["39216471"],"confidence":"High","gaps":["Specific phosphosites dephosphorylated by calcineurin not defined","Functional consequence on kinase output not directly measured"]},{"year":2025,"claim":"Defined a TTC7B-selective nanobody that blocks EFR3 binding and quantified how EFR3 phosphorylation and isoform identity tune complex affinity and PI4KA membrane recruitment.","evidence":"Cryo-EM, HDX-MS, nanobody selection, affinity measurements and bilayer/cell PI4P assays","pmids":["41197736","41473329"],"confidence":"High","gaps":["In vivo relevance of EFR3 isoform affinity differences not established","Kinase that phosphorylates EFR3 not identified here"]},{"year":2025,"claim":"Characterized additional disease alleles affecting catalysis and splicing, reinforcing that both catalytic-domain integrity and correct splicing are required for PI4KA function.","evidence":"Minigene splicing assay and in vitro kinase activity of purified mutant fragments","pmids":["39885450"],"confidence":"Medium","gaps":["Single case study","Cellular and clinical impact of partial activity loss not quantified"]},{"year":2026,"claim":"Connected Ca²⁺ signaling to PI4KA membrane recruitment, showing E-Syt1 delivers ER-localized PI4KA to ER–PM junctions to drive activity-dependent PI4P synthesis in neurons.","evidence":"Live-cell imaging, Ca²⁺ stimulation, E-Syt1 perturbation and PM PI4P biosensors in hippocampal neurons","pmids":["42258130"],"confidence":"Medium","gaps":["Direct physical contact between E-Syt1 and PI4KA not structurally defined","Relationship to EFR3-mediated recruitment unclear"]},{"year":null,"claim":"How PI4P depletion is sensed to trigger compensatory lipid remodeling, and how phosphorylation and calcineurin dephosphorylation are integrated to set PI4KA output in vivo, remain unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["Compensatory RhoB/PLD/PA pathway described only in a preprint","No unified model linking calcineurin regulation, EFR3 phosphorylation and Ca²⁺-driven recruitment","Function of the nuclear/nucleolar pool unexplained"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,1,7,18]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[0,1]}],"localization":[{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[6,8,9,16]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[2,16]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[3,4]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[2]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[6,17]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[9,10,13]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[7,8,18]}],"complexes":["PI4KA–TTC7–FAM126–EFR3 heterotetramer"],"partners":["TTC7B","FAM126A","EFR3A","EFR3B","CALCINEURIN (PPP3)","E-SYT1","KRAS","CXCR4"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"P42356","full_name":"Phosphatidylinositol 4-kinase alpha","aliases":["Phosphatidylinositol 4-Kinase III alpha"],"length_aa":2102,"mass_kda":236.8,"function":"Acts on phosphatidylinositol (PtdIns) in the first committed step in the production of the second messenger inositol-1,4,5,-trisphosphate","subcellular_location":"Cytoplasm; Cell membrane","url":"https://www.uniprot.org/uniprotkb/P42356/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/PI4KA","classification":"Common Essential","n_dependent_lines":1107,"n_total_lines":1208,"dependency_fraction":0.9163907284768212},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000241973","cell_line_id":"CID000145","localizations":[{"compartment":"membrane","grade":3},{"compartment":"cytoplasmic","grade":2},{"compartment":"nucleoplasm","grade":1},{"compartment":"vesicles","grade":1}],"interactors":[{"gene":"EFR3A","stoichiometry":10.0},{"gene":"FAM126A","stoichiometry":10.0},{"gene":"EFR3B","stoichiometry":10.0},{"gene":"TTC7B","stoichiometry":10.0},{"gene":"FAM126B","stoichiometry":10.0},{"gene":"TTC7A","stoichiometry":0.2},{"gene":"PTGES3","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000145","total_profiled":1310},"omim":[{"mim_id":"620511","title":"FLIEDNER-ZWEIER SYNDROME; FZS","url":"https://www.omim.org/entry/620511"},{"mim_id":"620060","title":"TETRATRICOPEPTIDE REPEAT DOMAIN-CONTAINING PROTEIN 7B; TTC7B","url":"https://www.omim.org/entry/620060"},{"mim_id":"619708","title":"GASTROINTESTINAL DEFECTS AND IMMUNODEFICIENCY SYNDROME 2; GIDID2","url":"https://www.omim.org/entry/619708"},{"mim_id":"619621","title":"SPASTIC PARAPLEGIA 84, AUTOSOMAL RECESSIVE; SPG84","url":"https://www.omim.org/entry/619621"},{"mim_id":"616797","title":"EFR3 HOMOLOG B; EFR3B","url":"https://www.omim.org/entry/616797"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Plasma membrane","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"brain","ntpm":103.1}],"url":"https://www.proteinatlas.org/search/PI4KA"},"hgnc":{"alias_symbol":["PI4K-ALPHA","pi4K230"],"prev_symbol":["PIK4CA"]},"alphafold":{"accession":"P42356","domains":[{"cath_id":"-","chopping":"30-115","consensus_level":"medium","plddt":85.8358,"start":30,"end":115},{"cath_id":"-","chopping":"129-212_294-392","consensus_level":"medium","plddt":77.1574,"start":129,"end":392},{"cath_id":"1.10.1070.11","chopping":"1904-2099","consensus_level":"high","plddt":89.0077,"start":1904,"end":2099},{"cath_id":"1.20.58","chopping":"1143-1163_1171-1242","consensus_level":"medium","plddt":91.281,"start":1143,"end":1242},{"cath_id":"1.20.58","chopping":"1244-1271_1288-1429_1479-1485","consensus_level":"medium","plddt":89.7852,"start":1244,"end":1485}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P42356","model_url":"https://alphafold.ebi.ac.uk/files/AF-P42356-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P42356-F1-predicted_aligned_error_v6.png","plddt_mean":79.69},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PI4KA","jax_strain_url":"https://www.jax.org/strain/search?query=PI4KA"},"sequence":{"accession":"P42356","fasta_url":"https://rest.uniprot.org/uniprotkb/P42356.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P42356/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P42356"}},"corpus_meta":[{"pmid":"24415756","id":"PMC_24415756","title":"Pharmacological and genetic targeting of the PI4KA enzyme reveals its important role in maintaining plasma membrane phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate levels.","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24415756","citation_count":162,"is_preprint":false},{"pmid":"17893707","id":"PMC_17893707","title":"An association screen of myelin-related genes implicates the chromosome 22q11 PIK4CA gene in schizophrenia.","date":"2007","source":"Molecular psychiatry","url":"https://pubmed.ncbi.nlm.nih.gov/17893707","citation_count":89,"is_preprint":false},{"pmid":"22022594","id":"PMC_22022594","title":"The role of the phosphatidylinositol 4-kinase PI4KA in hepatitis C virus-induced host membrane rearrangement.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/22022594","citation_count":73,"is_preprint":false},{"pmid":"25855803","id":"PMC_25855803","title":"Germline recessive mutations in PI4KA are associated with perisylvian polymicrogyria, cerebellar hypoplasia and arthrogryposis.","date":"2015","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25855803","citation_count":66,"is_preprint":false},{"pmid":"34415310","id":"PMC_34415310","title":"Biallelic PI4KA variants cause neurological, intestinal and immunological disease.","date":"2021","source":"Brain : a journal of neurology","url":"https://pubmed.ncbi.nlm.nih.gov/34415310","citation_count":43,"is_preprint":false},{"pmid":"34415322","id":"PMC_34415322","title":"Biallelic PI4KA variants cause a novel neurodevelopmental syndrome with hypomyelinating leukodystrophy.","date":"2021","source":"Brain : a journal of neurology","url":"https://pubmed.ncbi.nlm.nih.gov/34415322","citation_count":42,"is_preprint":false},{"pmid":"34504076","id":"PMC_34504076","title":"Oncogenic KRAS is dependent upon an EFR3A-PI4KA signaling axis for potent tumorigenic activity.","date":"2021","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/34504076","citation_count":41,"is_preprint":false},{"pmid":"18646052","id":"PMC_18646052","title":"Association of the PIK4CA schizophrenia-susceptibility gene in adults with the 22q11.2 deletion syndrome.","date":"2009","source":"American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/18646052","citation_count":37,"is_preprint":false},{"pmid":"10101268","id":"PMC_10101268","title":"Functional expression and characterisation of a new human phosphatidylinositol 4-kinase PI4K230.","date":"1999","source":"Biochimica et biophysica acta","url":"https://pubmed.ncbi.nlm.nih.gov/10101268","citation_count":35,"is_preprint":false},{"pmid":"11045352","id":"PMC_11045352","title":"Immunohistochemical localisation of two phosphatidylinositol 4-kinase isoforms, PI4K230 and PI4K92, in the central nervous system of rats.","date":"2000","source":"Experimental brain research","url":"https://pubmed.ncbi.nlm.nih.gov/11045352","citation_count":29,"is_preprint":false},{"pmid":"17131383","id":"PMC_17131383","title":"Nucleolar localization of phosphatidylinositol 4-kinase PI4K230 in various mammalian cells.","date":"2006","source":"Cytometry. Part A : the journal of the International Society for Analytical Cytology","url":"https://pubmed.ncbi.nlm.nih.gov/17131383","citation_count":25,"is_preprint":false},{"pmid":"12497619","id":"PMC_12497619","title":"Polymorphism screening of PIK4CA: possible candidate gene for chromosome 22q11-linked psychiatric disorders.","date":"2003","source":"American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/12497619","citation_count":24,"is_preprint":false},{"pmid":"37453227","id":"PMC_37453227","title":"PI4KA and PIKfyve: Essential phosphoinositide signaling enzymes involved in myriad human diseases.","date":"2023","source":"Current opinion in cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/37453227","citation_count":21,"is_preprint":false},{"pmid":"29386109","id":"PMC_29386109","title":"A Forward Genetic Screen Targeting the Endothelium Reveals a Regulatory Role for the Lipid Kinase Pi4ka in Myelo- and Erythropoiesis.","date":"2018","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/29386109","citation_count":17,"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":"18585705","id":"PMC_18585705","title":"Nuclear and nucleolar localization signals and their targeting function in phosphatidylinositol 4-kinase PI4K230.","date":"2008","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/18585705","citation_count":13,"is_preprint":false},{"pmid":"37979461","id":"PMC_37979461","title":"Structural basis for the conserved roles of PI4KA and its regulatory partners and their misregulation in disease.","date":"2023","source":"Advances in biological regulation","url":"https://pubmed.ncbi.nlm.nih.gov/37979461","citation_count":12,"is_preprint":false},{"pmid":"36276647","id":"PMC_36276647","title":"Targeting PI4KA sensitizes refractory leukemia to chemotherapy by modulating the ERK/AMPK/OXPHOS axis.","date":"2022","source":"Theranostics","url":"https://pubmed.ncbi.nlm.nih.gov/36276647","citation_count":12,"is_preprint":false},{"pmid":"35982909","id":"PMC_35982909","title":"Human CPTP promotes growth and metastasis via sphingolipid metabolite ceramide and PI4KA/AKT signaling in pancreatic cancer cells.","date":"2022","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/35982909","citation_count":11,"is_preprint":false},{"pmid":"25209194","id":"PMC_25209194","title":"Synergistic association of PI4KA and GRM3 genetic polymorphisms with poor antipsychotic response in south Indian schizophrenia patients with low severity of illness.","date":"2014","source":"American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25209194","citation_count":11,"is_preprint":false},{"pmid":"39312004","id":"PMC_39312004","title":"Biallelic PI4KA Mutations Disrupt B-Cell Metabolism and Cause B-Cell Lymphopenia and Hypogammaglobulinemia.","date":"2024","source":"Journal of clinical immunology","url":"https://pubmed.ncbi.nlm.nih.gov/39312004","citation_count":10,"is_preprint":false},{"pmid":"11311856","id":"PMC_11311856","title":"The ATP-binding site of brain phosphatidylinositol 4-kinase PI4K230 as revealed by 5'-p-fluorosulfonylbenzoyladenosine.","date":"2001","source":"The international journal of biochemistry & cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/11311856","citation_count":9,"is_preprint":false},{"pmid":"39216471","id":"PMC_39216471","title":"Structure of calcineurin bound to PI4KA reveals dual interface in both PI4KA and FAM126A.","date":"2024","source":"Structure (London, England : 1993)","url":"https://pubmed.ncbi.nlm.nih.gov/39216471","citation_count":8,"is_preprint":false},{"pmid":"37996444","id":"PMC_37996444","title":"Adaptor proteins mediate CXCR4 and PI4KA crosstalk in prostate cancer cells and the significance of PI4KA in bone tumor growth.","date":"2023","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/37996444","citation_count":7,"is_preprint":false},{"pmid":"39897037","id":"PMC_39897037","title":"TTC7B triggers the PI4KA-AKT1-RXRA-FTO axis and inhibits colon cancer cell proliferation by increasing RNA methylation.","date":"2025","source":"International journal of biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39897037","citation_count":5,"is_preprint":false},{"pmid":"11170653","id":"PMC_11170653","title":"Synthesis of new cyclitol compounds that influence the activity of phosphatidylinositol 4-kinase isoform, PI4K230.","date":"2001","source":"Journal of medicinal chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/11170653","citation_count":5,"is_preprint":false},{"pmid":"18521859","id":"PMC_18521859","title":"Association study between the PIK4CA gene and methamphetamine use disorder in a Japanese population.","date":"2009","source":"American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/18521859","citation_count":4,"is_preprint":false},{"pmid":"34045869","id":"PMC_34045869","title":"Sevoflurane Suppresses Colon Cancer Cell Malignancy by Regulating circ-PI4KA.","date":"2021","source":"OncoTargets and therapy","url":"https://pubmed.ncbi.nlm.nih.gov/34045869","citation_count":4,"is_preprint":false},{"pmid":"38685974","id":"PMC_38685974","title":"Two Novel Variants in PI4KA in a Family Presenting With Hereditary Spastic Paraparesis: A Case Report.","date":"2024","source":"Neurology. Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38685974","citation_count":4,"is_preprint":false},{"pmid":"26998712","id":"PMC_26998712","title":"Derivation of original RESP atomic partial charges for MD simulations of the LDAO surfactant with AMBER: applications to a model of micelle and a fragment of the lipid kinase PI4KA.","date":"2016","source":"Journal of biomolecular structure & dynamics","url":"https://pubmed.ncbi.nlm.nih.gov/26998712","citation_count":4,"is_preprint":false},{"pmid":"36865146","id":"PMC_36865146","title":"Adaptor proteins mediate CXCR4 and PI4KA crosstalk in prostate cancer cells and the significance of PI4KA in bone tumor growth.","date":"2023","source":"Research square","url":"https://pubmed.ncbi.nlm.nih.gov/36865146","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":"40348068","id":"PMC_40348068","title":"A phosphatidylinositol 4-kinase alpha (PI4KA) gene reduces plant height of common wheat.","date":"2025","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/40348068","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":"39885450","id":"PMC_39885450","title":"Identifying novel heterozygous PI4KA variants in fetal abnormalities.","date":"2025","source":"BMC medical genomics","url":"https://pubmed.ncbi.nlm.nih.gov/39885450","citation_count":0,"is_preprint":false},{"pmid":"42045152","id":"PMC_42045152","title":"Pi4ka downregulation triggers Creb3l2-dependent lysosomal dysfunction to promote maladaptive tubular remodeling and immune activation in acute kidney injury.","date":"2026","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/42045152","citation_count":0,"is_preprint":false},{"pmid":"42091194","id":"PMC_42091194","title":"[Analysis of PI4KA gene variants in a patient with Hereditary spastic paraplegia type 84].","date":"2026","source":"Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/42091194","citation_count":0,"is_preprint":false},{"pmid":"42258130","id":"PMC_42258130","title":"E-Syt1 recruits PI4KA to endoplasmic reticulum-plasma membrane junctions to enhance PI4P synthesis.","date":"2026","source":"Science China. Life sciences","url":"https://pubmed.ncbi.nlm.nih.gov/42258130","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.09.30.679611","title":"Phosphatidylserine and RhoB connect phosphatidylinositol 4-phosphate and phosphatidic acid metabolism at the plasma membrane","date":"2025-10-02","source":"bioRxiv","url":"https://doi.org/10.1101/2025.09.30.679611","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.10.16.682426","title":"Beyond the Genotype: A Multi-Omic Analysis of APOEe4’s Role in Alzheimer’s Disease","date":"2025-10-16","source":"bioRxiv","url":"https://doi.org/10.1101/2025.10.16.682426","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.07.03.663046","title":"From Lipid Dynamics to Precision Predictions: A New Approach Method for Precision Modeling of Phosphoinositide Signaling","date":"2025-07-07","source":"bioRxiv","url":"https://doi.org/10.1101/2025.07.03.663046","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.05.12.653131","title":"DNA-methylation markers associated with lung function at birth and childhood reveal early life programming of inflammatory pathways","date":"2025-05-14","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.12.653131","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":23405,"output_tokens":5244,"usd":0.074438,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13705,"output_tokens":4908,"usd":0.095613,"stage2_stop_reason":"end_turn"},"total_usd":0.170051,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1999,\n      \"finding\": \"Human PI4KA (PI4K230) encodes a 2044-amino acid protein with a C-terminal catalytic domain (~260 aa) that is highly conserved among PI 4-kinases. When expressed in vitro and in Sf9 cells, it exhibits type III PI 4-kinase characteristics: not inhibited by adenosine, high Km for ATP (~300 µM), half-maximally inactivated by ~200 nM wortmannin, and specific activity of 58 µmol mg⁻¹ min⁻¹.\",\n      \"method\": \"In vitro expression, enzymatic activity assays, pharmacological characterization\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct in vitro enzymatic reconstitution with pharmacological characterization, single lab but multiple orthogonal assays\",\n      \"pmids\": [\"10101268\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"The conserved Lys-1792 in the catalytic domain of PI4K230 (PI4KA) is essential for enzymatic activity and serves as a target of affinity labeling by the ATP-analog FSBA. ATP and ATP analogues competitively protect against FSBA inactivation, confirming this residue's proximity to the ATP-binding site.\",\n      \"method\": \"Site-directed mutagenesis, affinity labeling with FSBA, enzymatic activity assays\",\n      \"journal\": \"The international journal of biochemistry & cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — active-site mutagenesis combined with affinity labeling and competitive protection assays, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"11311856\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"PI4K230 (PI4KA) localizes in neurons predominantly to the outer membrane of mitochondria and membranes of the rough endoplasmic reticulum, and is also co-localized with some multivesicular bodies. No nuclear or plasma membrane localization was detected, suggesting its role in intracellular vesicular traffic rather than direct ligand-stimulated phosphoinositide turnover at the plasma membrane.\",\n      \"method\": \"Immunocytochemistry (light and electron microscopy) with isoform-specific antibodies in rat brain sections\",\n      \"journal\": \"Experimental brain research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — direct localization by electron microscopy with specific antibodies, replicated across multiple brain regions, but no functional perturbation experiment\",\n      \"pmids\": [\"11045352\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"PI4K230 (PI4KA) is detected in the nucleolus of multiple mammalian cell types, forming a Triton X-100-resistant, DNase- and RNase-sensitive complex there. siRNA knockdown of PI4K230 abolishes the nucleolar signal, confirming specificity.\",\n      \"method\": \"Immunofluorescence on ethanol-fixed cells and cryosections, siRNA knockdown, DNase/RNase treatments, Triton X-100 extraction\",\n      \"journal\": \"Cytometry. Part A\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization with siRNA validation and biochemical fractionation, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"17131383\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"PI4KA contains a functional monopartite NLS (NLS1: residues 916–934) that directs nuclear import via importin α1 and α3 (but not α5) through the classical importin α/β mechanism. A separate bipartite NLS2 (residues 1414–1433) functions as a nucleolar targeting signal and can mediate nuclear import via importin α1/β or α3/β complexes when present in a larger fragment (AA1166–1667).\",\n      \"method\": \"Digitonin-permeabilized HeLa cell nuclear import assays, fluorescent BSA conjugates with synthetic NLS peptides, molecular modeling, importin binding experiments\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reconstituted nuclear import assays with defined importins, multiple peptide constructs, single lab with several orthogonal approaches\",\n      \"pmids\": [\"18585705\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"PI4KA is required for local enrichment of phosphatidylinositol 4-phosphate (PI4P) at the HCV membranous web and for morphologically normal web formation. PI4KA physically associates with NS5A in HCV-infected cells, whereas the related kinase PI4KB does not interact with NS5A and does not affect web morphology or PI4P enrichment upon silencing.\",\n      \"method\": \"RNAi silencing, co-immunoprecipitation (PI4KA with NS5A), immunofluorescence/confocal microscopy of PI4P at membranous webs, non-replicative web formation model\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP plus functional RNAi phenotype with PI4P localization readout, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"22022594\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"PI4KA is the primary enzyme maintaining plasma membrane PI4P pools and is essential for replenishing PI(4,5)P2 specifically during strong stimulation of phospholipase C-coupled receptors. Pharmacological inhibition of PI4KA in adult mice causes sudden death correlating with PI(4,5)P2 depletion after agonist stimulation; conditional genetic knockout causes severe intestinal necrosis and death.\",\n      \"method\": \"Highly specific PI4KA inhibitors (pharmacological), conditional knockout mice, phosphoinositide level measurements after PLC stimulation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — orthogonal pharmacological and genetic loss-of-function approaches with defined lipid and phenotypic readouts, multiple in vitro and in vivo experiments\",\n      \"pmids\": [\"24415756\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"A missense substitution p.D1854N in the catalytic domain of PI4KA, identified in patients with polymicrogyria and cerebellar hypoplasia, abolishes kinase activity, demonstrating that residue D1854 is essential for PI4KA catalytic function.\",\n      \"method\": \"Expression of wild-type and mutant PI4KA with in vitro kinase activity assay; Sanger sequencing confirmation of segregation\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct in vitro kinase activity assay with disease-linked catalytic domain mutant, complemented by genetic segregation analysis\",\n      \"pmids\": [\"25855803\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PI4KA forms a heterotetrameric complex with EFR3, TTC7, and FAM126 at the plasma membrane to generate PI4P. Biallelic loss-of-function PI4KA variants reduce PI4KA protein levels, decrease PI4KA catalytic activity (assessed by immunofluorescence and targeted lipidomics in patient fibroblasts and PBMCs), and cause hypomyelinating leukodystrophy, inflammatory bowel disease, intestinal atresia, and combined immunodeficiency in humans.\",\n      \"method\": \"Western blotting, immunofluorescence, targeted lipidomics in patient-derived fibroblasts and PBMCs; structural modelling of complex interfaces; exome sequencing\",\n      \"journal\": \"Brain : a journal of neurology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two independent studies (PMIDs 34415310 and 34415322) using multiple orthogonal methods in patient-derived cells, replicating PI4KA's role in the complex and PI4P production\",\n      \"pmids\": [\"34415310\", \"34415322\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Oncogenic KRAS preferentially binds EFR3A (the PI4KA plasma membrane recruitment adaptor). Disrupting EFR3A or PI4KA reduces PI4P, phosphatidylserine, and KRAS levels at the plasma membrane, diminishing oncogenic signaling and tumorigenesis. Tethering PI4KA directly to the plasma membrane rescues these phenotypes, demonstrating that EFR3A-PI4KA axis supports KRAS plasma membrane localization and signaling.\",\n      \"method\": \"Co-immunoprecipitation (KRAS–EFR3A), siRNA/genetic disruption of EFR3A and PI4KA, PI4P and phosphatidylserine lipid measurements at PM, KRAS PM localization assays, PI4KA tethering rescue experiment, xenograft tumor assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP plus multiple functional rescue and KO experiments with defined lipid and signaling readouts, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"34504076\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CXCR4 binds to PI4KA adaptor protein TTC7, and this interaction drives plasma membrane PI4P production in prostate cancer cells. Inhibiting PI4KA or TTC7 reduces PM PI4P production, cellular invasion, and bone tumor growth.\",\n      \"method\": \"Co-immunoprecipitation (CXCR4–TTC7), PI4P measurement at plasma membrane, invasion assays, in vivo bone tumor models with PI4KA/TTC7 inhibition\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP of CXCR4 with TTC7 adaptor, functional inhibition with PI4P readout and in vivo validation, single lab with multiple assays\",\n      \"pmids\": [\"37996444\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"The C terminus of EFR3A undergoes a disorder-to-order transition upon binding to the PI4KA–TTC7B–FAM126A heterotrimer, directly contacting both TTC7B and FAM126A. Mutations disrupting this interface reduce PI4KA recruitment to the plasma membrane. Multiple disease-linked mutations and post-translational modifications map to this binding site.\",\n      \"method\": \"Cryo-EM structure determination, hydrogen-deuterium exchange mass spectrometry (HDX-MS), mutational analysis of complex interface\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with HDX-MS validation and mutagenesis, peer-reviewed; also independently reported as a preprint (PMID 38746453)\",\n      \"pmids\": [\"39705356\", \"38746453\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Calcineurin (CNAβ1 isoform) directly binds to PI4KA via an evolutionarily conserved IKISVT sequence in PI4KA's horn domain, and also binds FAM126A via conserved LTLT and PSISIT sequences. These dual calcineurin-binding sites are in close proximity to phosphorylation sites in the PI4KA complex, suggesting calcineurin regulates PI4KA through dephosphorylation of the complex.\",\n      \"method\": \"Cryo-EM structure of truncated PI4KA complex with calcineurin, HDX-MS, computational analysis of binding interfaces\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with HDX-MS validation, single lab with multiple orthogonal structural methods\",\n      \"pmids\": [\"39216471\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Targeted inactivation of the Pi4ka catalytic domain or reduction in Pi4ka mRNA expression inhibits myeloid and erythroid cell differentiation in vitro and promotes anemia in vivo. The mechanism involves deregulation of AKT, MAPK, SRC, and JAK-STAT signaling pathways downstream of Pi4ka activity.\",\n      \"method\": \"Catalytic domain inactivation, mRNA knockdown, in vitro differentiation assays, in vivo anemia model, signaling pathway analysis (AKT, MAPK, SRC, JAK-STAT)\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic inactivation of catalytic domain with defined hematopoietic phenotype and pathway readouts, single lab with in vitro and in vivo experiments\",\n      \"pmids\": [\"29386109\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PI4KA forms a stable complex with TTC7 and FAM126; a novel PI4KA inhibitor (cepharanthine) undermines the stability of this PI4KA/TTC7/FAM126 complex. PI4KA depletion sensitizes drug-resistant leukemia cells to chemotherapy by regulating the ERK/AMPK/OXPHOS axis.\",\n      \"method\": \"LC-MS, ELISA kinase assay, MM/GBSA binding analysis, siRNA knockdown, in vitro and in vivo leukemia models\",\n      \"journal\": \"Theranostics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical identification of PI4KA inhibitor targeting the complex, with functional in vitro and in vivo knockdown data and signaling pathway readouts, single lab\",\n      \"pmids\": [\"36276647\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A TTC7B-selective nanobody was developed that sterically blocks EFR3 binding to TTC7B in PI4KA complexes. EFR3B phosphorylation markedly decreases its binding affinity to TTC7-FAM126. EFR3A-TTC7B-FAM126A binds with ~10-fold higher affinity than most other EFR3-TTC7-FAM126 combinations. Nanobody binding causes decreased PI4KA membrane recruitment on lipid bilayers and in cells, with decreased PM PI4P production.\",\n      \"method\": \"Cryo-EM, HDX-MS, yeast display nanobody selection, binding affinity measurements, lipid bilayer recruitment assay, cell-based PM PI4P measurement\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with HDX-MS and functional cell/bilayer assays, multiple orthogonal methods in single study; also reported as preprint (PMID 41473329)\",\n      \"pmids\": [\"41197736\", \"41473329\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In response to Ca2+ signaling, the ER membrane tethering protein E-Syt1 recruits ER-localized PI4KA to ER-PM junctions, facilitating PI4KA's plasma membrane recruitment and assembly of the PI4KA enzyme complex, thereby enhancing PM PI4P synthesis. In hippocampal neurons, neuronal activity-induced PM localization of PI4KA and PM PI4P synthesis depend on E-Syt1 function.\",\n      \"method\": \"Live cell imaging of PI4KA localization, Ca2+ stimulation experiments, E-Syt1 knockdown/knockout, PI4P biosensor measurements at PM, hippocampal neuron synaptic potentiation assays\",\n      \"journal\": \"Science China. Life sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct subcellular localization linked to functional PI4P synthesis outcome, with genetic perturbation of E-Syt1, single lab\",\n      \"pmids\": [\"42258130\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PI4KA inhibition reduces PM PI4P levels, which triggers a compensatory increase in phospholipase D (PLD) activity and phosphatidic acid (PA) levels at the PM. This is mediated by a concomitant decrease in phosphatidylserine (PS) levels upon PI4P depletion, activating a reciprocal relationship between PS synthesis and PLD-mediated PA generation. Additionally, loss of PM PI4P upregulates the small GTPase RhoB transcriptionally and translationally, which further enhances PLD-mediated PA synthesis and actin cytoskeletal remodeling.\",\n      \"method\": \"Pharmacological PI4KA inhibition, RNA-seq, proximity labeling proteomics, lipid measurements (PI4P, PS, PA), RhoB protein/mRNA quantification, PLD activity assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multi-omic approach (RNA-seq, proximity proteomics, lipidomics) with pharmacological perturbation, single lab; preprint not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.09.30.679611\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"The PI4KA variant c.2819C>T (p.Ala940Val) significantly reduces PI4KA enzyme activity when assessed using purified mutant protein fragments, and a deletion variant c.2802_2863-40del produces three distinct aberrant mRNA isoforms via splicing disruption, together demonstrating catalytic domain and splice-site requirements for PI4KA function.\",\n      \"method\": \"Minigene splicing assay, purification of recombinant wild-type and mutant PI4KA fragments, in vitro kinase activity measurement\",\n      \"journal\": \"BMC medical genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — direct in vitro kinase assay on purified mutant protein and minigene splicing assay, single case study, single lab\",\n      \"pmids\": [\"39885450\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PI4KA (PI4KIIIα) is a type III lipid kinase that phosphorylates phosphatidylinositol at the D4 position to generate the predominant plasma membrane pool of PI4P, which is essential for PI(4,5)P2 replenishment during PLC stimulation; it exists primarily as a heterotrimer with TTC7 and FAM126, is recruited to the plasma membrane by lipidated EFR3A/B proteins (whose C-termini undergo a disorder-to-order transition upon binding both TTC7 and FAM126), can be further recruited to ER-PM junctions by Ca²⁺-activated E-Syt1, and is regulated by calcineurin binding to conserved sites on both PI4KA and FAM126A, with its catalytic activity requiring conserved residues including Lys-1792 and Asp-1854 in the kinase domain; loss of PI4KA activity disrupts plasma membrane lipid homeostasis, triggers compensatory RhoB/PLD/phosphatidic acid signaling, impairs KRAS and CXCR4 signaling at the plasma membrane, and in humans causes a spectrum of neurological, intestinal, and immunological diseases.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PI4KA (PI4KIIIα) is a type III phosphatidylinositol 4-kinase that generates the predominant plasma membrane pool of PI4P and is the primary enzyme replenishing PI(4,5)P2 during strong stimulation of phospholipase C-coupled receptors, making it essential for plasma membrane lipid homeostasis [#0, #6]. Its catalytic activity resides in a conserved C-terminal kinase domain requiring Lys-1792, which lies adjacent to the ATP-binding site, and Asp-1854 [#1, #7]. At the plasma membrane PI4KA functions within a heterotetrameric assembly with TTC7, FAM126, and the lipidated recruitment adaptor EFR3, whose disordered C-terminus undergoes a disorder-to-order transition to contact both TTC7B and FAM126A, an interface that controls PI4KA membrane recruitment and PI4P output [#8, #11, #15]. Recruitment to ER–PM junctions is further driven by Ca²⁺-activated E-Syt1, linking neuronal activity to local PI4P synthesis [#16], while calcineurin docks on conserved sites in the PI4KA horn domain and in FAM126A, positioning the phosphatase to regulate the complex by dephosphorylation [#12]. Through this PI4P-generating activity PI4KA supports plasma membrane localization and signaling of oncogenic KRAS via the EFR3A adaptor and of CXCR4 via TTC7, and its inhibition triggers compensatory RhoB/PLD/phosphatidic acid signaling upon PI4P and phosphatidylserine depletion [#9, #10, #17]. Biallelic and missense loss-of-function PI4KA variants that reduce kinase activity or protein levels cause a human spectrum of hypomyelinating leukodystrophy, polymicrogyria with cerebellar hypoplasia, inflammatory bowel disease, intestinal atresia, and combined immunodeficiency [#7, #8, #18].\",\n  \"teleology\": [\n    {\n      \"year\": 1999,\n      \"claim\": \"Established that human PI4KA encodes a large protein whose conserved C-terminal domain confers type III PI 4-kinase enzymatic activity, defining its biochemical identity.\",\n      \"evidence\": \"In vitro expression in Sf9 cells with enzymatic and pharmacological characterization\",\n      \"pmids\": [\"10101268\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define cellular substrate pool or in vivo localization\", \"No structural model of the catalytic domain\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Identified Lys-1792 as a catalytically essential residue near the ATP-binding site, providing the first active-site landmark for the kinase.\",\n      \"evidence\": \"Site-directed mutagenesis and FSBA affinity labeling with competitive ATP protection\",\n      \"pmids\": [\"11311856\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the full catalytic mechanism\", \"No co-structure with ATP\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Addressed where PI4KA acts in cells, initially localizing it to mitochondrial outer membrane and rough ER in neurons rather than the plasma membrane.\",\n      \"evidence\": \"Immuno-EM with isoform-specific antibodies in rat brain\",\n      \"pmids\": [\"11045352\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional perturbation linking localization to activity\", \"Apparent conflict with later plasma membrane / nucleolar localization unresolved\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Detected a nuclease-sensitive nucleolar pool of PI4KA, raising the possibility of functions beyond membrane lipid signaling.\",\n      \"evidence\": \"Immunofluorescence with siRNA validation, DNase/RNase treatment and detergent extraction\",\n      \"pmids\": [\"17131383\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No nucleolar function or substrate defined\", \"Relationship to cytoplasmic PI4P pools unknown\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Mapped the import determinants for the nuclear/nucleolar pool, identifying a classical NLS and a separate nucleolar targeting signal.\",\n      \"evidence\": \"Digitonin-permeabilized nuclear import assays with synthetic NLS peptides and defined importins\",\n      \"pmids\": [\"18585705\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No demonstrated function of nuclear-imported PI4KA\", \"Physiological regulation of import unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Connected PI4KA to a pathogen-driven membrane remodeling event, showing it associates with HCV NS5A to enrich PI4P at the viral membranous web.\",\n      \"evidence\": \"RNAi silencing, co-IP of PI4KA with NS5A, PI4P imaging at membranous webs\",\n      \"pmids\": [\"22022594\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not establish whether NS5A directly activates the kinase\", \"No structural basis for the interaction\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Defined PI4KA's core physiological role as the maintainer of plasma membrane PI4P and replenisher of PI(4,5)P2 during PLC signaling, with loss being lethal.\",\n      \"evidence\": \"Specific pharmacological inhibitors plus conditional knockout mice with phosphoinositide measurements\",\n      \"pmids\": [\"24415756\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve how the kinase is recruited to the plasma membrane\", \"Tissue-specific requirements not fully dissected\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Provided the first catalytic disease residue, showing the polymicrogyria-associated D1854N abolishes kinase activity.\",\n      \"evidence\": \"In vitro kinase assay of disease mutant plus segregation analysis\",\n      \"pmids\": [\"25855803\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not model how loss of activity causes the specific neurodevelopmental phenotype\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Extended PI4KA function to hematopoiesis, linking its catalytic activity to myeloid/erythroid differentiation via multiple signaling pathways.\",\n      \"evidence\": \"Catalytic-domain inactivation and knockdown with in vitro differentiation and in vivo anemia assays\",\n      \"pmids\": [\"29386109\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Causal chain from PI4P to AKT/MAPK/SRC/JAK-STAT not directly resolved\", \"Direct effectors unidentified\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established the heterotetrameric PI4KA–EFR3–TTC7–FAM126 complex as the functional plasma membrane PI4P-generating unit and tied biallelic loss-of-function variants to a multisystem human disease spectrum.\",\n      \"evidence\": \"Two independent patient-cell studies with western blot, lipidomics, structural modeling and exome sequencing\",\n      \"pmids\": [\"34415310\", \"34415322\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not provide high-resolution structure of the assembled complex\", \"Genotype-phenotype determinants across organ systems unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated that the EFR3A–PI4KA axis supports oncogenic KRAS plasma membrane localization and signaling, implicating PI4KA in tumorigenesis.\",\n      \"evidence\": \"KRAS–EFR3A co-IP, EFR3A/PI4KA disruption, lipid and KRAS PM measurements, tethering rescue and xenograft assays\",\n      \"pmids\": [\"34504076\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether KRAS directly modulates kinase activity not shown\", \"Generalizability across KRAS-driven tumors untested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed PI4KA, via its TTC7 adaptor, supports CXCR4-driven plasma membrane PI4P and prostate cancer invasion, broadening its receptor-coupled signaling roles.\",\n      \"evidence\": \"CXCR4–TTC7 co-IP, PM PI4P measurement, invasion and in vivo bone tumor models\",\n      \"pmids\": [\"37996444\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single co-IP without structural mapping of the CXCR4–TTC7 interface\", \"Direct vs indirect coupling unresolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Resolved the molecular basis of plasma membrane recruitment, showing the EFR3A C-terminus folds upon binding TTC7B and FAM126A and that this interface harbors disease mutations.\",\n      \"evidence\": \"Cryo-EM, HDX-MS and interface mutagenesis of the PI4KA–TTC7B–FAM126A–EFR3A assembly\",\n      \"pmids\": [\"39705356\", \"38746453\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Dynamics of lipidation-driven membrane targeting not fully captured\", \"How modifications toggle the interface in vivo untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified calcineurin as a direct regulator that docks on conserved sites in PI4KA and FAM126A near phosphorylation sites, implying phosphatase control of the complex.\",\n      \"evidence\": \"Cryo-EM of a truncated PI4KA–calcineurin complex with HDX-MS\",\n      \"pmids\": [\"39216471\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Specific phosphosites dephosphorylated by calcineurin not defined\", \"Functional consequence on kinase output not directly measured\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a TTC7B-selective nanobody that blocks EFR3 binding and quantified how EFR3 phosphorylation and isoform identity tune complex affinity and PI4KA membrane recruitment.\",\n      \"evidence\": \"Cryo-EM, HDX-MS, nanobody selection, affinity measurements and bilayer/cell PI4P assays\",\n      \"pmids\": [\"41197736\", \"41473329\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo relevance of EFR3 isoform affinity differences not established\", \"Kinase that phosphorylates EFR3 not identified here\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Characterized additional disease alleles affecting catalysis and splicing, reinforcing that both catalytic-domain integrity and correct splicing are required for PI4KA function.\",\n      \"evidence\": \"Minigene splicing assay and in vitro kinase activity of purified mutant fragments\",\n      \"pmids\": [\"39885450\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single case study\", \"Cellular and clinical impact of partial activity loss not quantified\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Connected Ca²⁺ signaling to PI4KA membrane recruitment, showing E-Syt1 delivers ER-localized PI4KA to ER–PM junctions to drive activity-dependent PI4P synthesis in neurons.\",\n      \"evidence\": \"Live-cell imaging, Ca²⁺ stimulation, E-Syt1 perturbation and PM PI4P biosensors in hippocampal neurons\",\n      \"pmids\": [\"42258130\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct physical contact between E-Syt1 and PI4KA not structurally defined\", \"Relationship to EFR3-mediated recruitment unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PI4P depletion is sensed to trigger compensatory lipid remodeling, and how phosphorylation and calcineurin dephosphorylation are integrated to set PI4KA output in vivo, remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Compensatory RhoB/PLD/PA pathway described only in a preprint\", \"No unified model linking calcineurin regulation, EFR3 phosphorylation and Ca²⁺-driven recruitment\", \"Function of the nuclear/nucleolar pool unexplained\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 1, 7, 18]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [6, 8, 9, 16]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [2, 16]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [3, 4]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [6, 17]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [9, 10, 13]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [7, 8, 18]}\n    ],\n    \"complexes\": [\"PI4KA–TTC7–FAM126–EFR3 heterotetramer\"],\n    \"partners\": [\"TTC7B\", \"FAM126A\", \"EFR3A\", \"EFR3B\", \"calcineurin (PPP3)\", \"E-Syt1\", \"KRAS\", \"CXCR4\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":6,"faith_total":6,"faith_pct":100.0}}