{"gene":"ATP5F1B","run_date":"2026-04-28T17:12:37","timeline":{"discoveries":[{"year":1994,"finding":"Crystal structure of bovine mitochondrial F1-ATPase at 2.8 Å resolution revealed that the three catalytic β-subunits (ATP5F1B orthologs) differ in conformation and bound nucleotide, supporting a rotary catalytic mechanism in which the three catalytic subunits are in different states of the catalytic cycle at any instant, with interconversion achieved by rotation of the α3β3 subassembly relative to the γ-subunit.","method":"X-ray crystallography at 2.8 Å resolution","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 — landmark crystal structure, foundational paper with >2500 citations providing direct structural evidence for rotary catalysis","pmids":["8065448"],"is_preprint":false},{"year":1999,"finding":"The β-subunit of ATP synthase (ATP5F1B) is present on the surface of human endothelial cells and functions as a binding protein for angiostatin; angiostatin binding to cell-surface ATP synthase α/β-subunits mediates its antiproliferative and antiangiogenic effects on endothelial cells.","method":"Ligand blot analysis, amino-terminal sequencing, peptide mass fingerprinting, immunologic analysis, flow cytometry, immunofluorescence, anti-subunit antibody inhibition of angiostatin antiproliferative effect","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (ligand blot, sequencing, flow cytometry, functional antibody inhibition) in a highly cited study","pmids":["10077593"],"is_preprint":false},{"year":2003,"finding":"The β-chain of ATP synthase (ATP5F1B) is ectopically expressed on the surface of hepatocytes and functions as a high-affinity receptor for apolipoprotein A-I (apoA-I), mediating HDL endocytosis; receptor stimulation by apoA-I triggers holo-HDL endocytosis via a mechanism strictly dependent on ADP generation by cell-surface ATP hydrolase activity.","method":"Biochemical receptor isolation, immunologic confirmation of ectopic localization, cell-surface ATP hydrolase activity assay, endocytosis assay with ATP synthase inhibitor in perfused rat liver ex vivo","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1-2 — multiple orthogonal biochemical and functional approaches including ex vivo perfused liver experiments, >389 citations","pmids":["12511957"],"is_preprint":false},{"year":2011,"finding":"miR-101 directly binds to the 3'UTR of ATP5F1B (ATP5B) mRNA and negatively regulates ATP5B protein expression; knockdown of ATP5B significantly inhibits HSV-1 replication, demonstrating that ATP5B functions as a pro-viral host factor for HSV-1.","method":"Luciferase 3'UTR reporter assay, siRNA knockdown, plaque assay, real-time PCR","journal":"Antiviral research","confidence":"Medium","confidence_rationale":"Tier 2 — multiple methods (reporter, RNAi, functional viral assay) in single study","pmids":["21291913"],"is_preprint":false},{"year":2011,"finding":"ATP5F1B (ATP5B) interacts with the calcium channel α2/δ1 subunit at the plasma membrane of developing myotubes, forming a functional signaling complex that accelerates the rate of decline of calcium transients, particularly during trains of stimulation pulses.","method":"FRET, coimmunoprecipitation, fluorescence imaging, calcium transient measurements in myotubes","journal":"American journal of physiology. Cell physiology","confidence":"Medium","confidence_rationale":"Tier 2 — reciprocal Co-IP and FRET confirming interaction, with functional readout (calcium transient kinetics)","pmids":["21490313"],"is_preprint":false},{"year":2012,"finding":"In a mouse model of multiple sclerosis (ND4 mice), the mRNA export factor REF undergoes loss of deimination (arginine-to-citrulline conversion), which impairs its binding strength to ATP5F1B mRNA and reduces ATP5B mRNA transport to mitochondria; pharmacological inhibition of deimination in PC12 cells reduced mitochondrial ATP synthase activity.","method":"mRNA binding assay, mRNA transport assay, ATP synthase activity assay after deimination inhibition","journal":"EMBO reports","confidence":"Medium","confidence_rationale":"Tier 2 — multiple orthogonal assays linking REF deimination to ATP5B mRNA transport and ATP synthase activity","pmids":["22261716"],"is_preprint":false},{"year":2016,"finding":"Knockdown of Atp5b in mouse spermatocytes (GC2 cells) decreased mitochondrial membrane potential and induced apoptosis, demonstrating that Atp5b is required for maintaining mitochondrial integrity in spermatocytes.","method":"siRNA knockdown, flow cytometry (mitochondrial membrane potential, apoptosis assay)","journal":"Environmental toxicology and pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 — direct knockdown with defined mechanistic phenotype (MMP loss, apoptosis)","pmids":["27525561"],"is_preprint":false},{"year":2019,"finding":"ATP5F1B (ATP5B) binds to the 3'UTR consensus sequences of rotavirus (RV) RNA with high affinity; during RV infection, ATP5B co-localizes with viral RNA and viroplasm; siRNA-mediated depletion of ATP5B (or other ATP synthase subunits) reduces production of infectious viral progeny without altering intracellular viral RNA levels or translation, placing ATP5B as a positive regulator of late-stage RV maturation/particle formation.","method":"RaPID proteomics-based RNA-protein interaction screen, siRNA knockdown, viral plaque assay, chemical inhibition in human intestinal enteroids","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 — RNA-protein interaction screen, confirmed by siRNA and chemical inhibition in both cell culture and primary enteroids with mechanistic specificity (late-stage maturation, not RNA replication)","pmids":["30770472"],"is_preprint":false},{"year":2021,"finding":"ATP5F1B (ATP5B) overexpression in gastric cancer cells elevates intracellular ATP content, increases extracellular ATP secretion, activates the P2X7 purinergic receptor, and thereby activates the FAK/AKT/MMP2 signaling pathway to promote cell migration, invasion, and proliferation; inhibitors of P2X7, FAK, AKT, and MMP2 suppress these effects.","method":"Overexpression and knockdown in gastric cancer cell lines, ATP content assay, migration/invasion assays, Western blotting for pathway components, pharmacological inhibitor experiments","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2-3 — gain- and loss-of-function with multiple downstream pathway readouts in a single study","pmids":["33715234"],"is_preprint":false},{"year":2021,"finding":"Inhibition of ATP5F1B (ATP5B) by siRNA lentivirus impairs osteoclast differentiation, suppresses osteoclast-related gene and protein expression, significantly impairs F-actin ring formation, decreases adhesion-associated proteins, causes mitochondrial dysfunction, and impairs vacuolar proton secretion and MMP9 secretion, thereby protecting arthritic mouse joints from bone erosion.","method":"Lentiviral siRNA delivery in vitro and intra-articular in vivo, gene/protein expression analysis, F-actin staining, bone resorption pit assay, mitochondrial function assay","journal":"Pharmacological research","confidence":"Medium","confidence_rationale":"Tier 2 — in vitro and in vivo loss-of-function with multiple mechanistic readouts","pmids":["33515708"],"is_preprint":false},{"year":2021,"finding":"Ectopic ATP5F1B (ATP5B) at the plasma membrane co-localizes and physically interacts with caveolin-1 (Cav-1) in MDA-MB-231 breast cancer cells; Cav-1 knockdown reduces migration and invasion abilities and is required for the pro-migratory/invasive function of ectopic ATP5B.","method":"Coimmunoprecipitation, double immunofluorescence staining, siRNA knockdown, migration/invasion assays","journal":"Medical oncology","confidence":"Medium","confidence_rationale":"Tier 3 — Co-IP confirmed interaction with functional siRNA data but single-lab study","pmids":["34009483"],"is_preprint":false},{"year":2022,"finding":"Cell-surface ATP5F1B (ATP5B) on hepatocellular carcinoma cells binds the myristoylated (but not non-myristoylated) preS1 2-47 peptide of hepatitis B virus; knockdown of ATP5B in NTCP-expressing HepG2 cells reduces HBV infectivity with less cccDNA formation, establishing ATP5B as an essential factor for HBV cell entry.","method":"Flow cytometry (cell surface expression), binding assay with myristoylated preS1 peptide, siRNA knockdown, HBV infection assay (cccDNA quantification)","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 — binding specificity established plus functional RNAi with mechanistic endpoint (cccDNA)","pmids":["36076968"],"is_preprint":false},{"year":2023,"finding":"Two heterozygous missense variants in ATP5F1B (p.Thr334Pro and p.Val482Ala) segregate with autosomal dominant early-onset isolated dystonia with incomplete penetrance; functional studies in patient fibroblasts showed preserved ATP5F1B protein levels but severe reduction of complex V (ATP synthase) activity and impaired mitochondrial membrane potential, consistent with a dominant-negative mechanism.","method":"Genetic segregation analysis, protein quantification, complex V enzymatic activity assay, mitochondrial membrane potential measurement in patient fibroblasts","journal":"Brain","confidence":"High","confidence_rationale":"Tier 2 — disease variants with functional validation using multiple mitochondrial assays in patient-derived cells, establishing dominant-negative mechanism","pmids":["36860166"],"is_preprint":false},{"year":2023,"finding":"ATP5F1B (ATP5B) binds HIF-1α mRNA; small molecules (HI-derivatives containing an adamantaniline moiety) promote this binding and thereby inhibit HIF-1α translation without affecting mRNA levels; target identification was achieved via affinity-based protein profiling of the probe HI-102.","method":"Affinity-based protein profiling (chemoproteomic target ID), mRNA-binding assay, HIF-1α protein expression and translation assay, in vivo xenograft model","journal":"Advanced science","confidence":"Medium","confidence_rationale":"Tier 2 — chemoproteomic target ID combined with mechanistic RNA-binding and translational regulation assays","pmids":["37401167"],"is_preprint":false},{"year":2023,"finding":"Under PFOS exposure, ATP5F1B (ATP5B) redistributes from the plasma membrane to mitochondria and interacts with transferrin receptor 2 (TFR2), facilitating TFR2 translocation to mitochondria, leading to mitochondrial iron overload that precedes and causes insulin resistance; stabilizing ATP5B on the plasma membrane or knockdown of ATP5B blocked TFR2 translocation and prevented insulin resistance.","method":"Subcellular fractionation, Co-IP, pharmacological inhibition of ectopic ATP synthase, ATP5B knockdown, mitochondrial iron measurement, insulin resistance assays in hepatocytes and mouse liver","journal":"Ecotoxicology and environmental safety","confidence":"Medium","confidence_rationale":"Tier 2 — multiple orthogonal approaches (Co-IP, fractionation, KD, pharmacological) in vitro and in vivo","pmids":["36801541"],"is_preprint":false},{"year":2024,"finding":"Mitochondria-localized acetyltransferase MOF directly acetylates ATP5F1B (ATP5B) at lysine K201; this acetylation, co-regulated by the deacetylase SIRT3, impairs mitochondrial respiration and energy metabolism; overexpression of mitochondria-targeted MOF in mice causes mitochondrial dysfunction, cardiac remodeling, and heart failure, and SIRT3 knockout aggravates these effects.","method":"Quantitative lysine acetylome mass spectrometry, mitochondria-targeted MOF overexpression mouse model, SIRT3 knockout, mitochondrial respiration assay, cardiac phenotyping","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1-2 — site-specific PTM identified by acetylome MS, validated by in vivo mouse models with functional mitochondrial and cardiac readouts","pmids":["39392752"],"is_preprint":false},{"year":2024,"finding":"Under PFOS exposure, ATP5F1B (ATP5B) interacts with TRPML1 (lysosomal calcium channel) and VDAC1 (mitochondrial calcium channel), facilitating calcium transmission from lysosomes to mitochondria; inhibiting ATP5B expression or retaining ATP5B on the plasma membrane disrupts TRPML1-VDAC1 interaction and reverses mitochondrial calcium overload and insulin resistance.","method":"Co-IP, subcellular fractionation, siRNA knockdown, calcium imaging, pharmacological inhibition, mouse liver experiments","journal":"Ecotoxicology and environmental safety","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP confirms ternary complex, multiple functional interventions validate the pathway","pmids":["38626609"],"is_preprint":false},{"year":2024,"finding":"Under PFOS exposure, ATP5F1B (ATP5B) interacts with VDAC1 and promotes VDAC1 translocation from the plasma membrane to mitochondria, where it undergoes oligomerization; this VDAC1 oligomerization activates the NLRP3 inflammasome; knockdown of ATP5B or immobilization of ATP5B on the plasma membrane prevents VDAC1 oligomerization and NLRP3 activation.","method":"Co-IP, VDAC1 oligomerization assay, siRNA knockdown, plasma membrane ATP5B stabilization, NLRP3 inflammasome activation assay in hepatocytes and mouse liver","journal":"Ecotoxicology and environmental safety","confidence":"Medium","confidence_rationale":"Tier 2 — protein interaction confirmed by Co-IP with mechanistic functional readout (NLRP3 activation) and genetic/pharmacological rescue","pmids":["38944014"],"is_preprint":false},{"year":2024,"finding":"ATP5F1B (ATP5B) interacts with TGEV (transmissible gastroenteritis coronavirus) nonstructural protein Nsp2; downregulation of ATP5B promotes TGEV replication, indicating that ATP5B functions as a negative regulator of TGEV replication.","method":"Immunoprecipitation/LC-MS/MS, Co-IP, indirect immunofluorescence, siRNA knockdown with viral replication assay","journal":"Virulence","confidence":"Medium","confidence_rationale":"Tier 2-3 — Co-IP and IFA confirm interaction; functional RNAi establishes antiviral role","pmids":["39239724"],"is_preprint":false},{"year":2025,"finding":"ATP5F1B (Atp5f1b) interacts with the mitochondrial protein Cend1; Cend1 deficiency in knockout mice exacerbates cerebral ischemia/reperfusion injury with impaired mitochondrial membrane potential, mPTP opening, ATP content reduction, and decreased Complex V activity; Cend1 dimerization via GXXXA motifs is required for ATP synthesis enhancement; the small molecule Tianeptine stabilizes Cend1 dimers, elevates ATP, and confers neuroprotection in a Cend1-dependent manner.","method":"Cend1 knockout mouse model, Co-IP (Atp5f1b-Cend1 interaction), mitochondrial function assays, mutagenesis (G130P), small molecule treatment, neurological phenotyping","journal":"Communications biology","confidence":"Medium","confidence_rationale":"Tier 2 — Co-IP confirms interaction, in vivo KO and mutagenesis establish mechanistic link to Complex V activity","pmids":["41469760"],"is_preprint":false},{"year":2023,"finding":"The Pseudomonas aeruginosa flagellar hook protein FlgE directly interacts with cell-surface ATP5F1B (ATP5B) on macrophages, and blocking ATP5B attenuates FlgE-induced NF-κB/MAPK signaling, inflammatory responses, SR-A1 upregulation, and foam cell formation, indicating ATP5B mediates pathogen-induced pro-atherogenic macrophage activation.","method":"Pull-down assay, Western blotting, blocking experiments with anti-ATP5B, in vitro macrophage assays, ApoE-/- mouse atherosclerosis model","journal":"Atherosclerosis","confidence":"Low","confidence_rationale":"Tier 3 — pull-down confirms interaction; functional blocking is partial; single-lab study with limited mechanistic depth on ATP5B's molecular role","pmids":["38278062"],"is_preprint":false},{"year":2020,"finding":"The bovine ATP5F1B (ATP5B) promoter contains two transcriptional start sites; the transcription factors MyoD and GATA1 bind to specific sites in the proximal promoter region (-539/+220 relative to TSS) and drive ATP5B basal transcription, as demonstrated by 5'-RACE, deletion analysis, site-directed mutagenesis, and ChIP assays.","method":"5'-RACE, luciferase reporter deletion assay, site-directed mutagenesis, chromatin immunoprecipitation (ChIP)","journal":"Animal biotechnology","confidence":"Medium","confidence_rationale":"Tier 2 — multiple complementary methods (reporter assay, mutagenesis, ChIP) establish transcriptional regulation mechanism","pmids":["33124493"],"is_preprint":false}],"current_model":"ATP5F1B encodes the catalytic β-subunit of the mitochondrial F1-ATPase (Complex V); its crystal structure established a rotary catalytic mechanism in which three β-subunits cycle through distinct nucleotide-bound states; beyond mitochondrial ATP synthesis, ATP5F1B is ectopically present at the plasma membrane where it acts as an HDL/apoA-I receptor on hepatocytes and an angiostatin-binding protein on endothelial cells; its activity is regulated by MOF-catalyzed acetylation at K201 (counteracted by SIRT3), and pathogenic dominant-negative missense variants cause Complex V deficiency and autosomal dominant dystonia; ATP5F1B also interacts with viral RNAs and proteins (serving as a pro-viral factor for HSV-1, rotavirus, and HBV, but an antiviral factor for TGEV), and participates in calcium and iron homeostasis through interactions with TRPML1, VDAC1, and TFR2 at mitochondria-associated membranes."},"narrative":{"teleology":[{"year":1994,"claim":"The fundamental question of how the three β-subunits of F1-ATPase collaborate during catalysis was resolved by determining that each β-subunit simultaneously adopts a distinct nucleotide-bound conformation, providing the structural basis for the rotary catalytic mechanism.","evidence":"X-ray crystallography of bovine mitochondrial F1-ATPase at 2.8 Å resolution","pmids":["8065448"],"confidence":"High","gaps":["Rotation was structurally inferred but not yet directly observed","No human-specific structure at the time","Post-translational regulatory mechanisms unknown"]},{"year":1999,"claim":"The discovery that ATP5F1B is ectopically expressed on the endothelial cell surface and serves as the angiostatin receptor established that ATP synthase subunits have non-canonical extramitochondrial functions.","evidence":"Ligand blot, N-terminal sequencing, flow cytometry, and antibody blocking of angiostatin's antiproliferative effect on endothelial cells","pmids":["10077593"],"confidence":"High","gaps":["Mechanism of ATP5F1B trafficking to the plasma membrane unresolved","Stoichiometry and assembly state of ectopic complex unknown"]},{"year":2003,"claim":"Ectopic ATP5F1B on hepatocytes was shown to function as a high-affinity apoA-I receptor whose ATP hydrolase activity generates ADP to trigger HDL endocytosis, establishing a physiological role for cell-surface ATP synthase in lipoprotein metabolism.","evidence":"Receptor isolation from hepatocyte membranes, surface activity assays, and ex vivo perfused rat liver endocytosis experiments","pmids":["12511957"],"confidence":"High","gaps":["Structural basis for apoA-I recognition by ectopic β-subunit undetermined","In vivo contribution to systemic HDL clearance not quantified"]},{"year":2011,"claim":"ATP5F1B was identified as a pro-viral host factor for HSV-1 (regulated via miR-101 targeting of its 3′UTR) and as a plasma membrane signaling partner of the calcium channel α2/δ1 subunit in developing myotubes, broadening its functional repertoire beyond energy metabolism.","evidence":"3′UTR luciferase reporter plus siRNA/plaque assays for HSV-1; FRET, Co-IP, and calcium transient imaging in myotubes","pmids":["21291913","21490313"],"confidence":"Medium","gaps":["Molecular mechanism by which ATP5F1B promotes HSV-1 replication unclear","Whether the calcium channel interaction occurs independently of ectopic ATP synthase complex is unknown"]},{"year":2019,"claim":"ATP5F1B was shown to bind rotavirus RNA 3′UTR consensus sequences and promote late-stage viral particle maturation without affecting RNA replication, pinpointing its pro-viral contribution to a specific assembly step.","evidence":"RaPID RNA–protein interaction screen, siRNA knockdown, chemical inhibition in cell lines and human intestinal enteroids","pmids":["30770472"],"confidence":"High","gaps":["Whether ATP5F1B's RNA-binding and catalytic functions are separable in the viral context is untested","No structural data on the ATP5F1B–RV RNA interface"]},{"year":2022,"claim":"Cell-surface ATP5F1B on hepatocytes was identified as a receptor for the myristoylated preS1 domain of HBV, with knockdown reducing cccDNA formation, establishing ATP5F1B as an entry co-factor for HBV alongside NTCP.","evidence":"Flow cytometry, myristoylated peptide binding assay, siRNA knockdown with cccDNA quantification in NTCP-expressing HepG2 cells","pmids":["36076968"],"confidence":"Medium","gaps":["Relative contribution of ATP5F1B versus NTCP to HBV entry kinetics not delineated","Whether ATP hydrolase activity is required for HBV internalization is unknown"]},{"year":2023,"claim":"Heterozygous missense variants in ATP5F1B were shown to cause autosomal dominant early-onset dystonia through a dominant-negative mechanism that severely impairs Complex V activity while preserving protein levels, establishing ATP5F1B as a Mendelian disease gene.","evidence":"Genetic segregation in families, Complex V enzymatic activity and mitochondrial membrane potential assays in patient fibroblasts","pmids":["36860166"],"confidence":"High","gaps":["No structural explanation for how p.Thr334Pro and p.Val482Ala exert dominant-negative effects","Penetrance modifiers not identified","No animal model recapitulating the dystonia phenotype"]},{"year":2023,"claim":"ATP5F1B was found to bind HIF-1α mRNA and mediate translational inhibition when engaged by adamantaniline-containing small molecules, revealing an unexpected role as an RNA-binding translational regulator.","evidence":"Chemoproteomic target identification (affinity-based protein profiling), mRNA-binding assay, HIF-1α translation assay, in vivo xenograft model","pmids":["37401167"],"confidence":"Medium","gaps":["Endogenous regulation of HIF-1α translation by ATP5F1B in the absence of small molecules not demonstrated","RNA-binding site on ATP5F1B not mapped"]},{"year":2023,"claim":"Under PFOS-induced stress, redistribution of ATP5F1B from the plasma membrane to mitochondria was shown to drive pathological iron and calcium overload by chaperoning TFR2 and bridging TRPML1–VDAC1, linking ectopic ATP5F1B trafficking to mitochondria-associated membrane signaling.","evidence":"Co-IP, subcellular fractionation, calcium and iron imaging, siRNA knockdown, and pharmacological stabilization of surface ATP5F1B in hepatocytes and mouse liver","pmids":["36801541","38626609","38944014"],"confidence":"Medium","gaps":["Whether ATP5F1B redistribution occurs in physiological (non-PFOS) contexts is unknown","Direct biophysical evidence for a ternary ATP5F1B–TRPML1–VDAC1 complex is lacking","Mechanism by which ATP5F1B promotes VDAC1 oligomerization is undefined"]},{"year":2024,"claim":"MOF-mediated acetylation of ATP5F1B at K201, counteracted by SIRT3, was established as a regulatory switch for mitochondrial respiration; hyperacetylation impairs Complex V activity and causes cardiac remodeling and heart failure in vivo.","evidence":"Quantitative acetylome mass spectrometry, mitochondria-targeted MOF overexpression and SIRT3 knockout mouse models, mitochondrial respiration and cardiac phenotyping","pmids":["39392752"],"confidence":"High","gaps":["Whether K201 acetylation affects rotary mechanism or assembly is not structurally resolved","Tissue-specific acetylation dynamics beyond heart are unexplored"]},{"year":2025,"claim":"ATP5F1B was shown to interact with Cend1, whose dimerization is required for full Complex V activity and neuroprotection during cerebral ischemia/reperfusion, revealing a regulatory partner that modulates ATP synthase function in the brain.","evidence":"Cend1 knockout mice, Co-IP, mutagenesis of Cend1 GXXXA dimerization motifs, small molecule (Tianeptine) stabilization of Cend1 dimers","pmids":["41469760"],"confidence":"Medium","gaps":["Direct binding interface between ATP5F1B and Cend1 not mapped","Whether Cend1 is a stoichiometric complex subunit or a transient regulator is unclear"]},{"year":null,"claim":"Key unresolved questions include the structural mechanism of ATP5F1B trafficking to the plasma membrane, the molecular basis for its multi-target RNA-binding capacity, how dominant-negative dystonia variants structurally disrupt the F1 hexamer, and whether ectopic versus mitochondrial pools of ATP5F1B are independently regulated.","evidence":"","pmids":[],"confidence":"Low","gaps":["No trafficking mechanism for ectopic ATP5F1B established","No cryo-EM or crystal structure of disease-mutant F1 complex","Relative physiological importance of RNA-binding versus catalytic functions undetermined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[0,2,12,15]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,2]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[7,13]},{"term_id":"GO:0001618","term_label":"virus receptor activity","supporting_discovery_ids":[11]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[0,6,12,14,15,16,17,19]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[1,2,4,10,11,14,16,17,20]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[0,2,6,15,19]},{"term_id":"R-HSA-382551","term_label":"Transport of small molecules","supporting_discovery_ids":[0,2,15]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[3,7,11,12,18]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[17,20]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,8]}],"complexes":["F1-ATPase (mitochondrial ATP synthase F1 sector)","Ectopic cell-surface ATP synthase complex"],"partners":["ATP5F1A","VDAC1","CAV1","TRPML1","TFR2","SIRT3","KAT8","CEND1"],"other_free_text":[]},"mechanistic_narrative":"ATP5F1B is the catalytic β-subunit of mitochondrial F1-ATPase (Complex V), whose three copies cycle through distinct nucleotide-bound conformations to drive rotary ATP synthesis, as established by the landmark bovine F1 crystal structure [PMID:8065448]. Beyond its canonical mitochondrial role, ATP5F1B is ectopically displayed on the plasma membrane of hepatocytes, endothelial cells, and cancer cells, where it functions as a high-affinity receptor for apoA-I mediating HDL endocytosis [PMID:12511957], a binding site for angiostatin mediating anti-angiogenic signaling [PMID:10077593], and a receptor for the HBV preS1 peptide facilitating viral entry [PMID:36076968]. Catalytic activity is regulated by MOF-mediated acetylation at K201 (counteracted by SIRT3), with hyperacetylation impairing mitochondrial respiration and causing cardiac remodeling in vivo [PMID:39392752]. Heterozygous dominant-negative missense variants in ATP5F1B (p.Thr334Pro, p.Val482Ala) cause Complex V deficiency and autosomal dominant early-onset dystonia [PMID:36860166]."},"prefetch_data":{"uniprot":{"accession":"P06576","full_name":"ATP synthase F(1) complex subunit beta, mitochondrial","aliases":["ATP synthase F1 subunit beta"],"length_aa":529,"mass_kda":56.6,"function":"Catalytic subunit beta, of the mitochondrial membrane ATP synthase complex (F(1)F(0) ATP synthase or Complex V) that produces ATP from ADP in the presence of a proton gradient across the membrane which is generated by electron transport complexes of the respiratory chain (Probable) (PubMed:37244256). ATP synthase complex consist of a soluble F(1) head domain - the catalytic core - and a membrane F(1) domain - the membrane proton channel (PubMed:37244256). These two domains are linked by a central stalk rotating inside the F(1) region and a stationary peripheral stalk (PubMed:37244256). During catalysis, ATP synthesis in the catalytic domain of F(1) is coupled via a rotary mechanism of the central stalk subunits to proton translocation (Probable). In vivo, can only synthesize ATP although its ATP hydrolase activity can be activated artificially in vitro (By similarity). With the subunit alpha (ATP5F1A), forms the catalytic core in the F(1) domain (PubMed:37244256)","subcellular_location":"Mitochondrion inner membrane","url":"https://www.uniprot.org/uniprotkb/P06576/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/ATP5F1B","classification":"Common Essential","n_dependent_lines":995,"n_total_lines":1208,"dependency_fraction":0.8236754966887417},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CAPZB","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/ATP5F1B","total_profiled":1310},"omim":[{"mim_id":"621502","title":"DYSTONIA 38, SUSCEPTIBILITY TO; DYT38","url":"https://www.omim.org/entry/621502"},{"mim_id":"620085","title":"HYPERMETABOLISM DUE TO UNCOUPLED MITOCHONDRIAL OXIDATIVE PHOSPHORYLATION 2; HUMOP2","url":"https://www.omim.org/entry/620085"},{"mim_id":"603281","title":"2-PRIME,5-PRIME-@OLIGOADENYLATE SYNTHETASE-LIKE; OASL","url":"https://www.omim.org/entry/603281"},{"mim_id":"603150","title":"ATP SYNTHASE F1, SUBUNIT DELTA; ATP5F1D","url":"https://www.omim.org/entry/603150"},{"mim_id":"300111","title":"PRICKLE PLANAR CELL POLARITY PROTEIN 3; PRICKLE3","url":"https://www.omim.org/entry/300111"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Mitochondria","reliability":"Enhanced"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal 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factor since its knockdown inhibits HSV-1 replication, and re-expression of ATP5B lacking the 3'UTR rescues viral replication despite miR-101 overexpression.\",\n      \"method\": \"Luciferase reporter assay (3'UTR binding), siRNA knockdown, ectopic overexpression of 3'UTR-lacking ATP5B, plaque assay\",\n      \"journal\": \"Antiviral research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (reporter assay, KD, rescue experiment) in a single study with clear functional readout\",\n      \"pmids\": [\"21291913\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"ATP5B (ATP5F1B) interacts with the calcium channel α2/δ1 subunit at the plasma membrane of developing myotubes, forming a functional signaling complex that accelerates the rate of decline of calcium transients, particularly during repetitive stimulation.\",\n      \"method\": \"FRET, co-immunoprecipitation, calcium imaging in developing myotubes\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — reciprocal Co-IP confirmed by FRET and functional calcium transient readout\",\n      \"pmids\": [\"21490313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Deimination (arginine-to-citrulline conversion) of the mRNA carrier REF is required for efficient ATP5B mRNA transport to mitochondria; loss of REF deimination impairs ATP5B mRNA binding and transport, reducing mitochondrial ATP synthase activity.\",\n      \"method\": \"mRNA binding assays, mRNA transport assays in ND4 transgenic mice (MS model), pharmacological inhibition of deimination in PC12 cells with ATP synthase activity measurement\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — functional assays in cell and mouse model, but single lab study\",\n      \"pmids\": [\"22261716\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ATP5B (ATP5F1B) binds with high affinity to the conserved 3'UTR consensus sequences of rotavirus (RV) RNA, co-localizes with viral RNA and viroplasm during infection, and is required for late-stage RV virion maturation/assembly; siRNA depletion of ATP5B or chemical inhibition of ATP synthase reduces infectious viral progeny without affecting intracellular viral RNA levels or translation.\",\n      \"method\": \"RaPID proteomics screen, siRNA knockdown, chemical inhibition, plaque assay, co-localization imaging, human intestinal enteroid model\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (proteomics, genetic KD, chemical inhibition, primary tissue model) with defined mechanistic step\",\n      \"pmids\": [\"30770472\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ATP5B (ATP5F1B) overexpression in gastric cancer cells increases intracellular and extracellular ATP levels and activates the FAK/AKT/MMP2 signaling pathway via the plasma membrane P2X7 receptor, promoting cell migration, invasion, and proliferation; inhibitors of P2X7, FAK, AKT, and MMP2 suppress these effects.\",\n      \"method\": \"Overexpression, siRNA knockdown, pharmacological inhibition, ATP measurement, western blot for pathway components, migration/invasion assays\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — pathway placement by pharmacological epistasis and overexpression, single lab\",\n      \"pmids\": [\"33715234\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ATP5B (ATP5F1B) is required for osteoclast differentiation and bone resorption; its inhibition impairs F-actin ring formation, reduces osteoclast-related gene/protein expression, impairs vacuolar proton secretion and MMP9 secretion, causes mitochondrial dysfunction, and protects against bone erosion in collagen-induced arthritis mice.\",\n      \"method\": \"Lentiviral siRNA knockdown in bone marrow macrophage-derived osteoclasts, in vivo local administration to arthritic joints, bone resorption assays, F-actin staining, gene/protein expression analysis\",\n      \"journal\": \"Pharmacological research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — loss-of-function both in vitro and in vivo with multiple mechanistic readouts\",\n      \"pmids\": [\"33515708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Ectopic (plasma membrane) ATP5B co-localizes and physically interacts with caveolin-1 (Cav-1) in MDA-MB-231 breast cancer cells; Cav-1 knockdown reduces cell migration and invasion induced by cholesterol-stimulated ectopic ATP5B upregulation, and cholesterol cannot rescue this effect after Cav-1 knockdown.\",\n      \"method\": \"Double immunofluorescence, co-immunoprecipitation, siRNA knockdown, migration/invasion assays, cholesterol loading\",\n      \"journal\": \"Medical oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP plus functional KD readout, single lab\",\n      \"pmids\": [\"34009483\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ATP5B (ATP5F1B) is expressed on the cell surface of hepatocellular carcinoma cells, binds myristoylated (but not non-myristoylated) HBV preS1 peptide (residues 2-47), and is required for HBV entry; knockdown of ATP5B in NTCP-expressing HepG2 cells reduces HBV infectivity and cccDNA formation.\",\n      \"method\": \"Cell surface expression assay, peptide binding assay, siRNA knockdown, HBV infection assay, cccDNA measurement\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — binding specificity demonstrated plus functional KD with defined viral entry readout, single lab\",\n      \"pmids\": [\"36076968\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Two heterozygous missense variants in ATP5F1B (p.Thr334Pro and p.Val482Ala) cause autosomal dominant isolated dystonia with incomplete penetrance through a dominant-negative mechanism; functional studies in patient fibroblasts show severe reduction of complex V (ATP synthase) activity and impaired mitochondrial membrane potential without reduction of ATP5F1B protein levels.\",\n      \"method\": \"Patient genetics (family segregation), functional assays in mutant fibroblasts (complex V activity, mitochondrial membrane potential measurement)\",\n      \"journal\": \"Brain\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — human genetics combined with direct enzymatic and bioenergetic functional validation in patient cells\",\n      \"pmids\": [\"36860166\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ATP5B (ATP5F1B) binds HIF-1α mRNA; the small molecule HI-101 (adamantaniline derivative) promotes this interaction, thereby inhibiting HIF-1α mRNA translation and downstream transcriptional activity, leading to antitumor effects in xenograft models.\",\n      \"method\": \"High-throughput screening, affinity-based protein profiling (probe HI-102), RNA-protein interaction assay, HIF-1α protein/activity assays, xenograft mouse model\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — chemical probe-based target ID plus mechanistic RNA-binding and functional translation assays, single lab\",\n      \"pmids\": [\"37401167\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Under PFOS exposure, plasma membrane ATP5B interacts with transferrin receptor 2 (TFR2) and both proteins co-translocate from the plasma membrane to mitochondria, causing mitochondrial iron overload that triggers hepatic insulin resistance; stabilizing ATP5B at the plasma membrane or knocking down ATP5B prevents TFR2 translocation and reverses iron overload and insulin resistance.\",\n      \"method\": \"Co-immunoprecipitation, subcellular fractionation/localization, siRNA knockdown, pharmacological manipulation of plasma membrane ATP synthase activity, mitochondrial iron measurement, insulin signaling assays in hepatocytes and mouse liver\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP, localization, genetic and pharmacological manipulation with defined mechanistic pathway, single lab\",\n      \"pmids\": [\"36801541\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Flagellar hook protein FlgE from Pseudomonas aeruginosa directly interacts with cell surface ATP5B; blocking ATP5B attenuates FlgE-induced NF-κB/MAPK signaling, SR-A1 upregulation, foam cell formation, and inflammatory responses in macrophages.\",\n      \"method\": \"Pull-down assay, western blot for signaling components, ATP5B blocking experiments, lipid uptake assays, atherosclerosis mouse model\",\n      \"journal\": \"Atherosclerosis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — pull-down plus functional blocking, single lab\",\n      \"pmids\": [\"38278062\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Mitochondria-localized lysine acetyltransferase MOF directly acetylates ATP5B (ATP5F1B) at lysine K201; this acetylation (counteracted by SIRT3 deacetylase) impairs mitochondrial respiration and energy metabolism. Overexpression of mitochondria-targeted MOF causes ATP5B hyperacetylation, mitochondrial dysfunction, cardiac remodeling, and heart failure in mice; SIRT3 knockout aggravates these effects.\",\n      \"method\": \"Quantitative lysine acetylome mass spectrometry, in vitro acetyltransferase assay, mouse cardiac overexpression models, SIRT3 KO, mitochondrial respiration assays\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — substrate identified by acetylome MS, writer (MOF) and eraser (SIRT3) characterized, specific site (K201) defined, validated in vitro and in vivo\",\n      \"pmids\": [\"39392752\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Under PFOS exposure, ATP5B interacts with TRPML1 (lysosomal calcium channel) and VDAC1 (mitochondrial calcium channel), bridging lysosome-to-mitochondria calcium transfer; knockdown of ATP5B or immobilization of ATP5B at the plasma membrane disrupts the TRPML1-VDAC1 interaction, reverses mitochondrial calcium overload, and worsens lysosomal calcium accumulation.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, pharmacological inhibition of mitochondrial calcium uptake, calcium imaging in hepatocytes and mouse liver\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP with functional rescue experiments, single lab\",\n      \"pmids\": [\"38626609\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ATP5B interacts with VDAC1 under PFOS exposure; ATP5B knockdown or plasma membrane immobilization of ATP5B prevents VDAC1 translocation from plasma membrane to mitochondria and reduces VDAC1 oligomerization, thereby attenuating NLRP3 inflammasome activation.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, plasma membrane ATP5B stabilization, VDAC1 oligomerization assay, NLRP3 inflammasome activation assays in hepatocytes and mouse liver\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — interaction and functional pathway placement supported by genetic and biochemical approaches, single lab\",\n      \"pmids\": [\"38944014\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TGEV coronavirus Nsp2 directly interacts with ATP5B; downregulation of ATP5B promotes TGEV replication, indicating ATP5B functions as a negative regulator of TGEV replication.\",\n      \"method\": \"Co-immunoprecipitation (Co-IP), indirect immunofluorescence (IFA), siRNA knockdown, viral replication assays\",\n      \"journal\": \"Virulence\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP confirmed by IFA with functional KD readout, single lab\",\n      \"pmids\": [\"39239724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cend1 protein interacts with ATP5F1B (Atp5f1b) at mitochondria; Cend1 forms dimers via conserved GXXXA motifs to enhance ATP synthesis, and disruption of Cend1 dimerization (G130P mutation) abolishes ATP-enhancing effects and worsens cerebral ischemia/reperfusion injury. The small molecule Tianeptine stabilizes Cend1 dimers and confers neuroprotection in a Cend1-dependent manner.\",\n      \"method\": \"Cend1 KO mice, co-immunoprecipitation (Cend1-Atp5f1b), mutagenesis (G130P), ATP/mitochondrial functional assays, infarct volume measurement, small-molecule rescue\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — genetic KO with mechanistic mutagenesis, interaction confirmed by Co-IP, functional ATP synthase readout, single lab\",\n      \"pmids\": [\"41469760\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The transcription factors MyoD and GATA1 bind to the proximal promoter region of bovine ATP5B (within -539/+220 relative to the TSS) and drive its basal transcription, as demonstrated by site-directed mutagenesis of their binding sites and chromatin immunoprecipitation.\",\n      \"method\": \"5'-RACE, luciferase reporter assays with 5'-deletion constructs, site-directed mutagenesis, chromatin immunoprecipitation (ChIP)\",\n      \"journal\": \"Animal biotechnology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP plus mutagenesis in reporter assay, but bovine ortholog study\",\n      \"pmids\": [\"33124493\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ATP5F1B (ATP5B) is the catalytic β-subunit of the mitochondrial F1FO-ATP synthase that synthesizes ATP; its activity is regulated by MOF-catalyzed acetylation at K201 (reversed by SIRT3), its mRNA transport to mitochondria depends on deimination of the carrier protein REF, it forms functional complexes at both mitochondrial and ectopic plasma membrane locations (interacting with α2/δ1, Cav-1, TFR2, TRPML1, and VDAC1) to regulate calcium signaling, iron homeostasis, and inflammasome activation, and it participates in viral replication cycles (HSV-1, rotavirus, HBV, TGEV) as a pro- or anti-viral host factor, while dominant-negative missense variants cause complex V deficiency and autosomal dominant isolated dystonia.\"\n}\n```\n\n--- ROUND 2 (R2/R3, augmented corpus) ---\n\n```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1994,\n      \"finding\": \"Crystal structure of bovine mitochondrial F1-ATPase at 2.8 Å resolution revealed that the three catalytic β-subunits (ATP5F1B orthologs) differ in conformation and bound nucleotide, supporting a rotary catalytic mechanism in which the three catalytic subunits are in different states of the catalytic cycle at any instant, with interconversion achieved by rotation of the α3β3 subassembly relative to the γ-subunit.\",\n      \"method\": \"X-ray crystallography at 2.8 Å resolution\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — landmark crystal structure, foundational paper with >2500 citations providing direct structural evidence for rotary catalysis\",\n      \"pmids\": [\"8065448\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"The β-subunit of ATP synthase (ATP5F1B) is present on the surface of human endothelial cells and functions as a binding protein for angiostatin; angiostatin binding to cell-surface ATP synthase α/β-subunits mediates its antiproliferative and antiangiogenic effects on endothelial cells.\",\n      \"method\": \"Ligand blot analysis, amino-terminal sequencing, peptide mass fingerprinting, immunologic analysis, flow cytometry, immunofluorescence, anti-subunit antibody inhibition of angiostatin antiproliferative effect\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (ligand blot, sequencing, flow cytometry, functional antibody inhibition) in a highly cited study\",\n      \"pmids\": [\"10077593\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The β-chain of ATP synthase (ATP5F1B) is ectopically expressed on the surface of hepatocytes and functions as a high-affinity receptor for apolipoprotein A-I (apoA-I), mediating HDL endocytosis; receptor stimulation by apoA-I triggers holo-HDL endocytosis via a mechanism strictly dependent on ADP generation by cell-surface ATP hydrolase activity.\",\n      \"method\": \"Biochemical receptor isolation, immunologic confirmation of ectopic localization, cell-surface ATP hydrolase activity assay, endocytosis assay with ATP synthase inhibitor in perfused rat liver ex vivo\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — multiple orthogonal biochemical and functional approaches including ex vivo perfused liver experiments, >389 citations\",\n      \"pmids\": [\"12511957\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"miR-101 directly binds to the 3'UTR of ATP5F1B (ATP5B) mRNA and negatively regulates ATP5B protein expression; knockdown of ATP5B significantly inhibits HSV-1 replication, demonstrating that ATP5B functions as a pro-viral host factor for HSV-1.\",\n      \"method\": \"Luciferase 3'UTR reporter assay, siRNA knockdown, plaque assay, real-time PCR\",\n      \"journal\": \"Antiviral research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple methods (reporter, RNAi, functional viral assay) in single study\",\n      \"pmids\": [\"21291913\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"ATP5F1B (ATP5B) interacts with the calcium channel α2/δ1 subunit at the plasma membrane of developing myotubes, forming a functional signaling complex that accelerates the rate of decline of calcium transients, particularly during trains of stimulation pulses.\",\n      \"method\": \"FRET, coimmunoprecipitation, fluorescence imaging, calcium transient measurements in myotubes\",\n      \"journal\": \"American journal of physiology. Cell physiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal Co-IP and FRET confirming interaction, with functional readout (calcium transient kinetics)\",\n      \"pmids\": [\"21490313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"In a mouse model of multiple sclerosis (ND4 mice), the mRNA export factor REF undergoes loss of deimination (arginine-to-citrulline conversion), which impairs its binding strength to ATP5F1B mRNA and reduces ATP5B mRNA transport to mitochondria; pharmacological inhibition of deimination in PC12 cells reduced mitochondrial ATP synthase activity.\",\n      \"method\": \"mRNA binding assay, mRNA transport assay, ATP synthase activity assay after deimination inhibition\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal assays linking REF deimination to ATP5B mRNA transport and ATP synthase activity\",\n      \"pmids\": [\"22261716\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Knockdown of Atp5b in mouse spermatocytes (GC2 cells) decreased mitochondrial membrane potential and induced apoptosis, demonstrating that Atp5b is required for maintaining mitochondrial integrity in spermatocytes.\",\n      \"method\": \"siRNA knockdown, flow cytometry (mitochondrial membrane potential, apoptosis assay)\",\n      \"journal\": \"Environmental toxicology and pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct knockdown with defined mechanistic phenotype (MMP loss, apoptosis)\",\n      \"pmids\": [\"27525561\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ATP5F1B (ATP5B) binds to the 3'UTR consensus sequences of rotavirus (RV) RNA with high affinity; during RV infection, ATP5B co-localizes with viral RNA and viroplasm; siRNA-mediated depletion of ATP5B (or other ATP synthase subunits) reduces production of infectious viral progeny without altering intracellular viral RNA levels or translation, placing ATP5B as a positive regulator of late-stage RV maturation/particle formation.\",\n      \"method\": \"RaPID proteomics-based RNA-protein interaction screen, siRNA knockdown, viral plaque assay, chemical inhibition in human intestinal enteroids\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — RNA-protein interaction screen, confirmed by siRNA and chemical inhibition in both cell culture and primary enteroids with mechanistic specificity (late-stage maturation, not RNA replication)\",\n      \"pmids\": [\"30770472\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ATP5F1B (ATP5B) overexpression in gastric cancer cells elevates intracellular ATP content, increases extracellular ATP secretion, activates the P2X7 purinergic receptor, and thereby activates the FAK/AKT/MMP2 signaling pathway to promote cell migration, invasion, and proliferation; inhibitors of P2X7, FAK, AKT, and MMP2 suppress these effects.\",\n      \"method\": \"Overexpression and knockdown in gastric cancer cell lines, ATP content assay, migration/invasion assays, Western blotting for pathway components, pharmacological inhibitor experiments\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — gain- and loss-of-function with multiple downstream pathway readouts in a single study\",\n      \"pmids\": [\"33715234\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Inhibition of ATP5F1B (ATP5B) by siRNA lentivirus impairs osteoclast differentiation, suppresses osteoclast-related gene and protein expression, significantly impairs F-actin ring formation, decreases adhesion-associated proteins, causes mitochondrial dysfunction, and impairs vacuolar proton secretion and MMP9 secretion, thereby protecting arthritic mouse joints from bone erosion.\",\n      \"method\": \"Lentiviral siRNA delivery in vitro and intra-articular in vivo, gene/protein expression analysis, F-actin staining, bone resorption pit assay, mitochondrial function assay\",\n      \"journal\": \"Pharmacological research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vitro and in vivo loss-of-function with multiple mechanistic readouts\",\n      \"pmids\": [\"33515708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Ectopic ATP5F1B (ATP5B) at the plasma membrane co-localizes and physically interacts with caveolin-1 (Cav-1) in MDA-MB-231 breast cancer cells; Cav-1 knockdown reduces migration and invasion abilities and is required for the pro-migratory/invasive function of ectopic ATP5B.\",\n      \"method\": \"Coimmunoprecipitation, double immunofluorescence staining, siRNA knockdown, migration/invasion assays\",\n      \"journal\": \"Medical oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — Co-IP confirmed interaction with functional siRNA data but single-lab study\",\n      \"pmids\": [\"34009483\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cell-surface ATP5F1B (ATP5B) on hepatocellular carcinoma cells binds the myristoylated (but not non-myristoylated) preS1 2-47 peptide of hepatitis B virus; knockdown of ATP5B in NTCP-expressing HepG2 cells reduces HBV infectivity with less cccDNA formation, establishing ATP5B as an essential factor for HBV cell entry.\",\n      \"method\": \"Flow cytometry (cell surface expression), binding assay with myristoylated preS1 peptide, siRNA knockdown, HBV infection assay (cccDNA quantification)\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — binding specificity established plus functional RNAi with mechanistic endpoint (cccDNA)\",\n      \"pmids\": [\"36076968\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Two heterozygous missense variants in ATP5F1B (p.Thr334Pro and p.Val482Ala) segregate with autosomal dominant early-onset isolated dystonia with incomplete penetrance; functional studies in patient fibroblasts showed preserved ATP5F1B protein levels but severe reduction of complex V (ATP synthase) activity and impaired mitochondrial membrane potential, consistent with a dominant-negative mechanism.\",\n      \"method\": \"Genetic segregation analysis, protein quantification, complex V enzymatic activity assay, mitochondrial membrane potential measurement in patient fibroblasts\",\n      \"journal\": \"Brain\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — disease variants with functional validation using multiple mitochondrial assays in patient-derived cells, establishing dominant-negative mechanism\",\n      \"pmids\": [\"36860166\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ATP5F1B (ATP5B) binds HIF-1α mRNA; small molecules (HI-derivatives containing an adamantaniline moiety) promote this binding and thereby inhibit HIF-1α translation without affecting mRNA levels; target identification was achieved via affinity-based protein profiling of the probe HI-102.\",\n      \"method\": \"Affinity-based protein profiling (chemoproteomic target ID), mRNA-binding assay, HIF-1α protein expression and translation assay, in vivo xenograft model\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — chemoproteomic target ID combined with mechanistic RNA-binding and translational regulation assays\",\n      \"pmids\": [\"37401167\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Under PFOS exposure, ATP5F1B (ATP5B) redistributes from the plasma membrane to mitochondria and interacts with transferrin receptor 2 (TFR2), facilitating TFR2 translocation to mitochondria, leading to mitochondrial iron overload that precedes and causes insulin resistance; stabilizing ATP5B on the plasma membrane or knockdown of ATP5B blocked TFR2 translocation and prevented insulin resistance.\",\n      \"method\": \"Subcellular fractionation, Co-IP, pharmacological inhibition of ectopic ATP synthase, ATP5B knockdown, mitochondrial iron measurement, insulin resistance assays in hepatocytes and mouse liver\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal approaches (Co-IP, fractionation, KD, pharmacological) in vitro and in vivo\",\n      \"pmids\": [\"36801541\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Mitochondria-localized acetyltransferase MOF directly acetylates ATP5F1B (ATP5B) at lysine K201; this acetylation, co-regulated by the deacetylase SIRT3, impairs mitochondrial respiration and energy metabolism; overexpression of mitochondria-targeted MOF in mice causes mitochondrial dysfunction, cardiac remodeling, and heart failure, and SIRT3 knockout aggravates these effects.\",\n      \"method\": \"Quantitative lysine acetylome mass spectrometry, mitochondria-targeted MOF overexpression mouse model, SIRT3 knockout, mitochondrial respiration assay, cardiac phenotyping\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 — site-specific PTM identified by acetylome MS, validated by in vivo mouse models with functional mitochondrial and cardiac readouts\",\n      \"pmids\": [\"39392752\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Under PFOS exposure, ATP5F1B (ATP5B) interacts with TRPML1 (lysosomal calcium channel) and VDAC1 (mitochondrial calcium channel), facilitating calcium transmission from lysosomes to mitochondria; inhibiting ATP5B expression or retaining ATP5B on the plasma membrane disrupts TRPML1-VDAC1 interaction and reverses mitochondrial calcium overload and insulin resistance.\",\n      \"method\": \"Co-IP, subcellular fractionation, siRNA knockdown, calcium imaging, pharmacological inhibition, mouse liver experiments\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP confirms ternary complex, multiple functional interventions validate the pathway\",\n      \"pmids\": [\"38626609\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Under PFOS exposure, ATP5F1B (ATP5B) interacts with VDAC1 and promotes VDAC1 translocation from the plasma membrane to mitochondria, where it undergoes oligomerization; this VDAC1 oligomerization activates the NLRP3 inflammasome; knockdown of ATP5B or immobilization of ATP5B on the plasma membrane prevents VDAC1 oligomerization and NLRP3 activation.\",\n      \"method\": \"Co-IP, VDAC1 oligomerization assay, siRNA knockdown, plasma membrane ATP5B stabilization, NLRP3 inflammasome activation assay in hepatocytes and mouse liver\",\n      \"journal\": \"Ecotoxicology and environmental safety\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — protein interaction confirmed by Co-IP with mechanistic functional readout (NLRP3 activation) and genetic/pharmacological rescue\",\n      \"pmids\": [\"38944014\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ATP5F1B (ATP5B) interacts with TGEV (transmissible gastroenteritis coronavirus) nonstructural protein Nsp2; downregulation of ATP5B promotes TGEV replication, indicating that ATP5B functions as a negative regulator of TGEV replication.\",\n      \"method\": \"Immunoprecipitation/LC-MS/MS, Co-IP, indirect immunofluorescence, siRNA knockdown with viral replication assay\",\n      \"journal\": \"Virulence\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 — Co-IP and IFA confirm interaction; functional RNAi establishes antiviral role\",\n      \"pmids\": [\"39239724\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ATP5F1B (Atp5f1b) interacts with the mitochondrial protein Cend1; Cend1 deficiency in knockout mice exacerbates cerebral ischemia/reperfusion injury with impaired mitochondrial membrane potential, mPTP opening, ATP content reduction, and decreased Complex V activity; Cend1 dimerization via GXXXA motifs is required for ATP synthesis enhancement; the small molecule Tianeptine stabilizes Cend1 dimers, elevates ATP, and confers neuroprotection in a Cend1-dependent manner.\",\n      \"method\": \"Cend1 knockout mouse model, Co-IP (Atp5f1b-Cend1 interaction), mitochondrial function assays, mutagenesis (G130P), small molecule treatment, neurological phenotyping\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — Co-IP confirms interaction, in vivo KO and mutagenesis establish mechanistic link to Complex V activity\",\n      \"pmids\": [\"41469760\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The Pseudomonas aeruginosa flagellar hook protein FlgE directly interacts with cell-surface ATP5F1B (ATP5B) on macrophages, and blocking ATP5B attenuates FlgE-induced NF-κB/MAPK signaling, inflammatory responses, SR-A1 upregulation, and foam cell formation, indicating ATP5B mediates pathogen-induced pro-atherogenic macrophage activation.\",\n      \"method\": \"Pull-down assay, Western blotting, blocking experiments with anti-ATP5B, in vitro macrophage assays, ApoE-/- mouse atherosclerosis model\",\n      \"journal\": \"Atherosclerosis\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — pull-down confirms interaction; functional blocking is partial; single-lab study with limited mechanistic depth on ATP5B's molecular role\",\n      \"pmids\": [\"38278062\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The bovine ATP5F1B (ATP5B) promoter contains two transcriptional start sites; the transcription factors MyoD and GATA1 bind to specific sites in the proximal promoter region (-539/+220 relative to TSS) and drive ATP5B basal transcription, as demonstrated by 5'-RACE, deletion analysis, site-directed mutagenesis, and ChIP assays.\",\n      \"method\": \"5'-RACE, luciferase reporter deletion assay, site-directed mutagenesis, chromatin immunoprecipitation (ChIP)\",\n      \"journal\": \"Animal biotechnology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple complementary methods (reporter assay, mutagenesis, ChIP) establish transcriptional regulation mechanism\",\n      \"pmids\": [\"33124493\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ATP5F1B encodes the catalytic β-subunit of the mitochondrial F1-ATPase (Complex V); its crystal structure established a rotary catalytic mechanism in which three β-subunits cycle through distinct nucleotide-bound states; beyond mitochondrial ATP synthesis, ATP5F1B is ectopically present at the plasma membrane where it acts as an HDL/apoA-I receptor on hepatocytes and an angiostatin-binding protein on endothelial cells; its activity is regulated by MOF-catalyzed acetylation at K201 (counteracted by SIRT3), and pathogenic dominant-negative missense variants cause Complex V deficiency and autosomal dominant dystonia; ATP5F1B also interacts with viral RNAs and proteins (serving as a pro-viral factor for HSV-1, rotavirus, and HBV, but an antiviral factor for TGEV), and participates in calcium and iron homeostasis through interactions with TRPML1, VDAC1, and TFR2 at mitochondria-associated membranes.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"ATP5F1B encodes the catalytic β-subunit of the mitochondrial F1FO-ATP synthase and is central to oxidative phosphorylation, but it also functions at ectopic plasma membrane locations and as an RNA-binding protein with roles in calcium signaling, iron homeostasis, inflammasome regulation, and viral infection. Its catalytic activity is regulated by MOF-mediated acetylation at K201, reversed by SIRT3, and hyperacetylation impairs mitochondrial respiration and causes cardiac remodeling in mice [PMID:39392752]. At the plasma membrane, ATP5F1B interacts with calcium channel subunits, TFR2, VDAC1, TRPML1, and caveolin-1, mediating calcium transient modulation, lysosome-to-mitochondria calcium transfer, iron translocation, and NLRP3 inflammasome activation [PMID:21490313, PMID:38626609, PMID:38944014, PMID:36801541]. Heterozygous dominant-negative missense variants (p.Thr334Pro, p.Val482Ala) cause autosomal dominant isolated dystonia with severe reduction of complex V activity [PMID:36860166].\",\n  \"teleology\": [\n    {\n      \"year\": 2011,\n      \"claim\": \"Establishing that ATP5F1B is co-opted by viruses: its expression is required for HSV-1 replication, with miR-101 acting as an antiviral factor by suppressing ATP5B mRNA via its 3ʹUTR, thereby defining ATP5F1B as a pro-viral host factor.\",\n      \"evidence\": \"Luciferase 3ʹUTR reporter, siRNA knockdown, rescue with 3ʹUTR-lacking construct, plaque assay in mammalian cells\",\n      \"pmids\": [\"21291913\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism by which ATP5B promotes HSV-1 replication is undefined\", \"Whether the effect is through mitochondrial ATP synthesis or an alternative function is unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Revealing an ectopic plasma membrane function: ATP5F1B interacts with the calcium channel α2/δ1 subunit at the surface of developing myotubes and modulates calcium transient kinetics, establishing that the protein operates outside mitochondria in signaling.\",\n      \"evidence\": \"FRET, co-immunoprecipitation, and calcium imaging in developing myotubes\",\n      \"pmids\": [\"21490313\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How ATP5F1B reaches the plasma membrane is unresolved\", \"Whether catalytic activity is required for calcium modulation is not tested\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defining a post-translational mechanism for ATP5B mRNA delivery: deimination of the mRNA carrier REF is required for efficient transport of ATP5B mRNA to mitochondria, linking citrullination of a carrier protein to ATP synthase biogenesis.\",\n      \"evidence\": \"mRNA binding/transport assays, pharmacological inhibition of deimination in PC12 cells, ND4 transgenic mouse model\",\n      \"pmids\": [\"22261716\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab finding not independently replicated\", \"Whether deimination of REF selectively controls ATP5B versus other mitochondrial mRNAs is unclear\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Extending the viral role to rotavirus: ATP5F1B binds conserved 3ʹUTR sequences of rotavirus RNA and is required specifically for late-stage virion maturation/assembly, separating its function from viral RNA replication or translation.\",\n      \"evidence\": \"RaPID proteomics, siRNA knockdown, chemical inhibition, co-localization imaging, human intestinal enteroid model\",\n      \"pmids\": [\"30770472\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of RNA binding by ATP5F1B is unknown\", \"Whether RNA-binding and ATPase activities are functionally separable is not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrating that ectopic plasma membrane ATP5F1B drives oncogenic signaling: overexpression increases extracellular ATP and activates a P2X7→FAK/AKT/MMP2 cascade promoting gastric cancer invasion, while interaction with caveolin-1 mediates cholesterol-stimulated breast cancer migration.\",\n      \"evidence\": \"Overexpression/knockdown, pharmacological inhibition of P2X7/FAK/AKT/MMP2, Co-IP of ATP5B–Cav-1, migration/invasion assays\",\n      \"pmids\": [\"33715234\", \"34009483\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"All from single labs without independent replication\", \"Relative contribution of ATP synthesis versus signaling scaffolding at plasma membrane is unresolved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identifying a requirement for ATP5F1B in osteoclast biology: its inhibition impairs F-actin ring formation, proton secretion, MMP9 release, and bone resorption in vitro and protects against bone erosion in arthritic mice.\",\n      \"evidence\": \"Lentiviral shRNA knockdown in bone marrow macrophage-derived osteoclasts, in vivo intra-articular delivery in collagen-induced arthritis model\",\n      \"pmids\": [\"33515708\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether the effect reflects general mitochondrial dysfunction versus a specific ATP5F1B function is not dissected\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Connecting cell-surface ATP5F1B to HBV entry: surface-expressed ATP5F1B binds myristoylated (but not unmodified) HBV preS1, and its knockdown reduces HBV infection and cccDNA formation, implicating it as a co-receptor for hepatitis B virus.\",\n      \"evidence\": \"Cell surface expression assay, peptide binding assay, siRNA knockdown, HBV infection/cccDNA quantification in NTCP-expressing HepG2 cells\",\n      \"pmids\": [\"36076968\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab study\", \"Whether ATP5F1B is sufficient or acts in complex with NTCP for entry is not resolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Establishing human disease causality: dominant-negative ATP5F1B missense variants (Thr334Pro, Val482Ala) cause autosomal dominant isolated dystonia by severely reducing complex V activity without lowering protein levels, proving the β-subunit is essential for neuronal bioenergetics.\",\n      \"evidence\": \"Family segregation studies, complex V enzymatic assays and mitochondrial membrane potential measurement in patient fibroblasts\",\n      \"pmids\": [\"36860166\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Neuronal-specific pathomechanism not characterized\", \"Structural basis of dominant-negative effect not modeled\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Revealing a plasma membrane-to-mitochondria trafficking axis: under PFOS exposure, ATP5F1B co-translocates with TFR2 from the plasma membrane to mitochondria, causing mitochondrial iron overload and hepatic insulin resistance, and also bridges TRPML1–VDAC1 for lysosome-to-mitochondria calcium transfer, positioning ATP5F1B as a dynamic scaffold linking organellar ion fluxes.\",\n      \"evidence\": \"Co-IP, subcellular fractionation, siRNA knockdown, plasma membrane immobilization, calcium/iron imaging in hepatocytes and mouse liver\",\n      \"pmids\": [\"36801541\", \"38626609\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"All from a single research group studying PFOS; generality beyond toxicant exposure is unconfirmed\", \"Mechanism controlling ATP5F1B plasma membrane retention versus internalization is unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Adding RNA-binding breadth: ATP5F1B binds HIF-1α mRNA, and a small molecule (HI-101) that enhances this interaction inhibits HIF-1α translation and tumor growth, demonstrating that ATP5F1B's RNA-binding function extends beyond viral RNA to endogenous mRNAs with translational regulatory consequences.\",\n      \"evidence\": \"High-throughput screen, affinity probe profiling, RNA–protein interaction assay, xenograft model\",\n      \"pmids\": [\"37401167\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab; specificity of ATP5F1B-HIF-1α mRNA interaction versus general RNA binding is not established\", \"Endogenous physiological relevance of this RNA interaction is not demonstrated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identifying the writer-eraser pair controlling ATP5F1B catalytic activity: MOF directly acetylates K201 of ATP5F1B, SIRT3 removes this mark, and the balance determines mitochondrial respiration and cardiac function, establishing a specific post-translational regulatory circuit for ATP synthase.\",\n      \"evidence\": \"Quantitative lysine acetylome MS, in vitro acetyltransferase assay, cardiac-specific MOF overexpression and SIRT3 KO mice, mitochondrial respiration assays\",\n      \"pmids\": [\"39392752\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether K201 acetylation alters catalytic rate, assembly, or both is not dissected\", \"Relevance of this regulatory axis beyond cardiomyocytes is untested\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Expanding the inflammasome connection: ATP5F1B interacts with VDAC1, and its translocation from the plasma membrane promotes VDAC1 oligomerization and NLRP3 inflammasome activation, while ATP5F1B also serves as a receptor for Pseudomonas FlgE-driven NF-κB/MAPK inflammatory signaling.\",\n      \"evidence\": \"Co-IP, VDAC1 oligomerization assay, NLRP3 inflammasome readouts, pull-down with FlgE, blocking experiments in macrophages and mouse liver\",\n      \"pmids\": [\"38944014\", \"38278062\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"FlgE–ATP5B interaction based on pull-down without reciprocal validation\", \"Whether ATP5B directly activates inflammasome or acts indirectly via VDAC1 is not resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defining a mitochondrial partner that enhances ATP synthase: Cend1 interacts with ATP5F1B at mitochondria and dimerizes via GXXXA motifs to stimulate ATP synthesis; disruption worsens cerebral ischemia, and Tianeptine rescues by stabilizing Cend1 dimers.\",\n      \"evidence\": \"Cend1 KO mice, Co-IP, GXXXA mutagenesis, ATP/mitochondrial functional assays, ischemia/reperfusion injury model\",\n      \"pmids\": [\"41469760\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab study\", \"Whether Cend1 modulates ATP5F1B catalysis directly or affects complex V assembly is unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Major open questions include: how ATP5F1B is trafficked to the plasma membrane, the structural basis and specificity of its RNA-binding activity, whether its catalytic and RNA-binding functions are separable, and the neuronal pathomechanism underlying ATP5F1B-linked dystonia.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural model of ATP5F1B in its plasma membrane or RNA-bound states\", \"Mechanism of ectopic localization entirely uncharacterized\", \"Genotype-phenotype correlation for dystonia variants lacks neuronal cell models\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [5, 8, 12, 16]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [3, 9]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [1, 13, 14]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [2, 8, 12, 16]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1, 6, 7, 10, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [5, 8, 12, 16]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [11, 14]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [0, 3, 7, 8, 15]}\n    ],\n    \"complexes\": [\n      \"F1FO-ATP synthase (complex V)\"\n    ],\n    \"partners\": [\n      \"SIRT3\",\n      \"KAT8\",\n      \"VDAC1\",\n      \"TRPML1\",\n      \"TFR2\",\n      \"CAV1\",\n      \"CACNA2D1\",\n      \"CEND1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```\n\n--- ROUND 2 (R2/R3, augmented corpus) ---\n\n```json\n{\n  \"mechanistic_narrative\": \"ATP5F1B is the catalytic β-subunit of mitochondrial F1-ATPase (Complex V), whose three copies cycle through distinct nucleotide-bound conformations to drive rotary ATP synthesis, as established by the landmark bovine F1 crystal structure [PMID:8065448]. Beyond its canonical mitochondrial role, ATP5F1B is ectopically displayed on the plasma membrane of hepatocytes, endothelial cells, and cancer cells, where it functions as a high-affinity receptor for apoA-I mediating HDL endocytosis [PMID:12511957], a binding site for angiostatin mediating anti-angiogenic signaling [PMID:10077593], and a receptor for the HBV preS1 peptide facilitating viral entry [PMID:36076968]. Catalytic activity is regulated by MOF-mediated acetylation at K201 (counteracted by SIRT3), with hyperacetylation impairing mitochondrial respiration and causing cardiac remodeling in vivo [PMID:39392752]. Heterozygous dominant-negative missense variants in ATP5F1B (p.Thr334Pro, p.Val482Ala) cause Complex V deficiency and autosomal dominant early-onset dystonia [PMID:36860166].\",\n  \"teleology\": [\n    {\n      \"year\": 1994,\n      \"claim\": \"The fundamental question of how the three β-subunits of F1-ATPase collaborate during catalysis was resolved by determining that each β-subunit simultaneously adopts a distinct nucleotide-bound conformation, providing the structural basis for the rotary catalytic mechanism.\",\n      \"evidence\": \"X-ray crystallography of bovine mitochondrial F1-ATPase at 2.8 Å resolution\",\n      \"pmids\": [\"8065448\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Rotation was structurally inferred but not yet directly observed\", \"No human-specific structure at the time\", \"Post-translational regulatory mechanisms unknown\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"The discovery that ATP5F1B is ectopically expressed on the endothelial cell surface and serves as the angiostatin receptor established that ATP synthase subunits have non-canonical extramitochondrial functions.\",\n      \"evidence\": \"Ligand blot, N-terminal sequencing, flow cytometry, and antibody blocking of angiostatin's antiproliferative effect on endothelial cells\",\n      \"pmids\": [\"10077593\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of ATP5F1B trafficking to the plasma membrane unresolved\", \"Stoichiometry and assembly state of ectopic complex unknown\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Ectopic ATP5F1B on hepatocytes was shown to function as a high-affinity apoA-I receptor whose ATP hydrolase activity generates ADP to trigger HDL endocytosis, establishing a physiological role for cell-surface ATP synthase in lipoprotein metabolism.\",\n      \"evidence\": \"Receptor isolation from hepatocyte membranes, surface activity assays, and ex vivo perfused rat liver endocytosis experiments\",\n      \"pmids\": [\"12511957\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis for apoA-I recognition by ectopic β-subunit undetermined\", \"In vivo contribution to systemic HDL clearance not quantified\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"ATP5F1B was identified as a pro-viral host factor for HSV-1 (regulated via miR-101 targeting of its 3′UTR) and as a plasma membrane signaling partner of the calcium channel α2/δ1 subunit in developing myotubes, broadening its functional repertoire beyond energy metabolism.\",\n      \"evidence\": \"3′UTR luciferase reporter plus siRNA/plaque assays for HSV-1; FRET, Co-IP, and calcium transient imaging in myotubes\",\n      \"pmids\": [\"21291913\", \"21490313\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular mechanism by which ATP5F1B promotes HSV-1 replication unclear\", \"Whether the calcium channel interaction occurs independently of ectopic ATP synthase complex is unknown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"ATP5F1B was shown to bind rotavirus RNA 3′UTR consensus sequences and promote late-stage viral particle maturation without affecting RNA replication, pinpointing its pro-viral contribution to a specific assembly step.\",\n      \"evidence\": \"RaPID RNA–protein interaction screen, siRNA knockdown, chemical inhibition in cell lines and human intestinal enteroids\",\n      \"pmids\": [\"30770472\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether ATP5F1B's RNA-binding and catalytic functions are separable in the viral context is untested\", \"No structural data on the ATP5F1B–RV RNA interface\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Cell-surface ATP5F1B on hepatocytes was identified as a receptor for the myristoylated preS1 domain of HBV, with knockdown reducing cccDNA formation, establishing ATP5F1B as an entry co-factor for HBV alongside NTCP.\",\n      \"evidence\": \"Flow cytometry, myristoylated peptide binding assay, siRNA knockdown with cccDNA quantification in NTCP-expressing HepG2 cells\",\n      \"pmids\": [\"36076968\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Relative contribution of ATP5F1B versus NTCP to HBV entry kinetics not delineated\", \"Whether ATP hydrolase activity is required for HBV internalization is unknown\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Heterozygous missense variants in ATP5F1B were shown to cause autosomal dominant early-onset dystonia through a dominant-negative mechanism that severely impairs Complex V activity while preserving protein levels, establishing ATP5F1B as a Mendelian disease gene.\",\n      \"evidence\": \"Genetic segregation in families, Complex V enzymatic activity and mitochondrial membrane potential assays in patient fibroblasts\",\n      \"pmids\": [\"36860166\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No structural explanation for how p.Thr334Pro and p.Val482Ala exert dominant-negative effects\", \"Penetrance modifiers not identified\", \"No animal model recapitulating the dystonia phenotype\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"ATP5F1B was found to bind HIF-1α mRNA and mediate translational inhibition when engaged by adamantaniline-containing small molecules, revealing an unexpected role as an RNA-binding translational regulator.\",\n      \"evidence\": \"Chemoproteomic target identification (affinity-based protein profiling), mRNA-binding assay, HIF-1α translation assay, in vivo xenograft model\",\n      \"pmids\": [\"37401167\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Endogenous regulation of HIF-1α translation by ATP5F1B in the absence of small molecules not demonstrated\", \"RNA-binding site on ATP5F1B not mapped\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Under PFOS-induced stress, redistribution of ATP5F1B from the plasma membrane to mitochondria was shown to drive pathological iron and calcium overload by chaperoning TFR2 and bridging TRPML1–VDAC1, linking ectopic ATP5F1B trafficking to mitochondria-associated membrane signaling.\",\n      \"evidence\": \"Co-IP, subcellular fractionation, calcium and iron imaging, siRNA knockdown, and pharmacological stabilization of surface ATP5F1B in hepatocytes and mouse liver\",\n      \"pmids\": [\"36801541\", \"38626609\", \"38944014\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether ATP5F1B redistribution occurs in physiological (non-PFOS) contexts is unknown\", \"Direct biophysical evidence for a ternary ATP5F1B–TRPML1–VDAC1 complex is lacking\", \"Mechanism by which ATP5F1B promotes VDAC1 oligomerization is undefined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"MOF-mediated acetylation of ATP5F1B at K201, counteracted by SIRT3, was established as a regulatory switch for mitochondrial respiration; hyperacetylation impairs Complex V activity and causes cardiac remodeling and heart failure in vivo.\",\n      \"evidence\": \"Quantitative acetylome mass spectrometry, mitochondria-targeted MOF overexpression and SIRT3 knockout mouse models, mitochondrial respiration and cardiac phenotyping\",\n      \"pmids\": [\"39392752\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether K201 acetylation affects rotary mechanism or assembly is not structurally resolved\", \"Tissue-specific acetylation dynamics beyond heart are unexplored\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"ATP5F1B was shown to interact with Cend1, whose dimerization is required for full Complex V activity and neuroprotection during cerebral ischemia/reperfusion, revealing a regulatory partner that modulates ATP synthase function in the brain.\",\n      \"evidence\": \"Cend1 knockout mice, Co-IP, mutagenesis of Cend1 GXXXA dimerization motifs, small molecule (Tianeptine) stabilization of Cend1 dimers\",\n      \"pmids\": [\"41469760\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct binding interface between ATP5F1B and Cend1 not mapped\", \"Whether Cend1 is a stoichiometric complex subunit or a transient regulator is unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the structural mechanism of ATP5F1B trafficking to the plasma membrane, the molecular basis for its multi-target RNA-binding capacity, how dominant-negative dystonia variants structurally disrupt the F1 hexamer, and whether ectopic versus mitochondrial pools of ATP5F1B are independently regulated.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No trafficking mechanism for ectopic ATP5F1B established\", \"No cryo-EM or crystal structure of disease-mutant F1 complex\", \"Relative physiological importance of RNA-binding versus catalytic functions undetermined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [0, 2, 12, 15]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [7, 13]},\n      {\"term_id\": \"GO:0001618\", \"supporting_discovery_ids\": [11]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [0, 6, 12, 14, 15, 16, 17, 19]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1, 2, 4, 10, 11, 14, 16, 17, 20]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [0, 2, 6, 15, 19]},\n      {\"term_id\": \"R-HSA-382551\", \"supporting_discovery_ids\": [0, 2, 15]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [3, 7, 11, 12, 18]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [17, 20]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 8]}\n    ],\n    \"complexes\": [\n      \"F1-ATPase (mitochondrial ATP synthase F1 sector)\",\n      \"Ectopic cell-surface ATP synthase complex\"\n    ],\n    \"partners\": [\n      \"ATP5F1A\",\n      \"VDAC1\",\n      \"CAV1\",\n      \"TRPML1\",\n      \"TFR2\",\n      \"SIRT3\",\n      \"KAT8\",\n      \"CEND1\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}