{"gene":"ITFG2","run_date":"2026-06-10T01:55:23","timeline":{"discoveries":[{"year":2017,"finding":"ITFG2 is a component of the KICSTOR complex (with KPTN, C12orf66, and SZT2) that localizes to lysosomes, binds and recruits GATOR1 (but not GATOR2) to the lysosomal surface, and is necessary for GATOR1 to interact with its substrates the Rag GTPases and with GATOR2; loss of KICSTOR prevents amino acid or glucose deprivation from inhibiting mTORC1.","method":"Co-immunoprecipitation, subcellular fractionation/localization, siRNA knockdown, mTORC1 activity assays in cultured human cells, and SZT2-knockout mouse tissue analysis","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, lysosomal localization with functional consequence, genetic KO in mice, replicated by multiple subsequent studies","pmids":["28199306"],"is_preprint":false},{"year":2025,"finding":"Cryo-EM and computational modeling revealed that within KICSTOR, SZT2 forms a crescent-shaped scaffold that binds the ITFG2–KPTN heterodimer and C12orf66 at its C terminus; GATOR1 binds the SZT2 N-terminal domain via NPRL3; disruption of this GATOR1–KICSTOR interface hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient status. SZT2 and C12orf66 preferentially interact with negatively charged lipids, a requirement for lysosomal localization of the entire complex.","method":"Cryo-electron microscopy, computational modeling, biochemical interaction assays, lipid-binding assays, functional mTORC1 activity and TFE3 localization assays","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with biochemical validation and functional assays in a single rigorous study","pmids":["41198956"],"is_preprint":false},{"year":2025,"finding":"FBXO2 directly interacts with KPTN (a KICSTOR subunit) via its F-box-associated domain and promotes K48- and K63-linked polyubiquitination of KPTN at lysines 49, 67, 262, and 265; this ubiquitination disrupts the KPTN–ITFG2 and KPTN–SZT2 interactions, impairing KICSTOR's ability to recruit GATOR1 to the lysosomal surface and thereby activating mTORC1 signaling.","method":"Co-immunoprecipitation, ubiquitination assays, site-directed mutagenesis of KPTN lysine residues, mTORC1 activity assays, lysosomal localization experiments","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — direct interaction, mutagenesis identifying specific ubiquitination sites, functional mTORC1 readout, multiple orthogonal methods in one study","pmids":["41401028"],"is_preprint":false},{"year":2022,"finding":"Crystal structures of SAMTOR show that it interacts with GATOR1–KICSTOR (which contains ITFG2) through its SAM-dependent methyltransferase domain; conformational change in the SAMTOR helical domain upon SAM binding modulates the interaction of SAMTOR with the GATOR1–KICSTOR complex, thereby regulating mTORC1 activity in response to SAM levels.","method":"Crystal structure determination (apo and SAM-bound), site-directed mutagenesis, functional binding and mTORC1 activity assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — crystal structure with mutagenesis, but ITFG2's specific contribution within KICSTOR is inferred from complex membership rather than directly probed","pmids":["35776786"],"is_preprint":false},{"year":2013,"finding":"ITFG2 (Itfg2) is an intracellular protein in mice that is required for normal B cell differentiation; Itfg2-deficient mice show retention of B cells in the spleen, reduced serum IgG, defective cell migration in vitro, a shift of B cell maturation from germinal centers to extrafollicular regions, and blocked plasma cell deposition in bone marrow; bone marrow transplantation of Itfg2-deficient cells was sufficient to impair germinal center development in wild-type recipients, indicating hematopoietic cell-intrinsic activity.","method":"Itfg2-knockout mice, flow cytometry, bone marrow transplantation, in vitro migration assay, immunization experiments","journal":"Journal of immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KO mouse with defined cellular phenotypes, bone marrow transplantation establishing cell-intrinsic role, multiple readouts in single lab","pmids":["23997217"],"is_preprint":false},{"year":2024,"finding":"ITFG2 forms a complex with NEDD4-2 and ATP5b (mitochondrial ATP synthase β subunit) in cardiomyocytes; ITFG2 inhibits NEDD4-2-mediated ubiquitination of ATP5b, thereby preserving mitochondrial ATP production, reducing ROS, and maintaining mitochondrial membrane potential under hypoxic conditions; overexpression in a mouse MI model reduced infarct size and improved cardiac function.","method":"Co-immunoprecipitation, ubiquitination assay, AAV9-mediated overexpression, shRNA/siRNA knockdown, cardiac-specific transgenic mice, mitochondrial function assays (ATP, ROS, MMP), echocardiography","journal":"Biochemical pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP identifying complex, ubiquitination assay, multiple in vivo and in vitro models, single lab","pmids":["38848780"],"is_preprint":false},{"year":2024,"finding":"ITFG2 binds NEDD4-2 in cardiomyocytes and decreases NEDD4-2's interaction with SERCA2a, preventing NEDD4-2-mediated ubiquitination and degradation of SERCA2a; this preserves calcium homeostasis and cardiac contractility under ischemic conditions.","method":"Co-immunoprecipitation, ubiquitination assay, AAV9 overexpression and shRNA knockdown in MI mouse model, echocardiography, calcium measurement in primary cardiomyocytes","journal":"Biochemical pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assays with functional cardiac readouts, single lab, two orthogonal methods","pmids":["39477020"],"is_preprint":false},{"year":2025,"finding":"ITFG2 reduces the binding affinity between NEDD4-2 and Nav1.5 (voltage-gated sodium channel), inhibiting Nav1.5 ubiquitination; ITFG2 overexpression increased peak Nav1.5 current by ~50%, upregulated Nav1.5 protein expression, increased conduction velocity, and reduced ischemic ventricular arrhythmias in MI mice.","method":"Co-immunoprecipitation, ubiquitination assay, patch-clamp electrophysiology, AAV9 overexpression and shRNA knockdown in MI mouse model, optical mapping of epicardial conduction","journal":"European journal of pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assays with electrophysiological validation, single lab, multiple orthogonal methods","pmids":["39864577"],"is_preprint":false},{"year":2021,"finding":"Loss of ITFG2 (along with other KICSTOR/GATOR1 components) identified in a whole-genome CRISPR/Cas9 knockout screen confers resistance to PI3Kα inhibition in PIK3CA-mutated breast cancer cells by sustaining mTORC1 signaling, placing ITFG2 as a negative regulator of mTORC1 downstream of PI3K-AKT.","method":"Genome-wide CRISPR/Cas9 sgRNA knockout screen, validated by individual gene knockouts, mTORC1 signaling assays, sensitivity assays with PI3Kα inhibitors","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR screen with functional validation, epistatic placement relative to PI3K-AKT and mTORC1, single lab","pmids":["33685991"],"is_preprint":false}],"current_model":"ITFG2 is a subunit of the lysosome-associated KICSTOR complex (with KPTN, C12orf66, and SZT2), where it forms a heterodimer with KPTN on a crescent-shaped SZT2 scaffold; KICSTOR recruits GATOR1 to the lysosomal surface to enable GAP activity toward Rag GTPases and mTORC1 inhibition in response to nutrient deprivation. In cardiomyocytes, ITFG2 additionally functions as a NEDD4-2 inhibitor, blocking ubiquitination of SERCA2a, ATP5b, and Nav1.5 to preserve mitochondrial function, calcium homeostasis, and sodium channel activity under ischemic conditions. In B cells, ITFG2 is required cell-intrinsically for germinal center formation and normal humoral immune responses."},"narrative":{"mechanistic_narrative":"ITFG2 is a subunit of the lysosome-associated KICSTOR complex (with KPTN, C12orf66, and SZT2) that couples nutrient status to mTORC1 signaling by recruiting GATOR1 to the lysosomal surface, where GATOR1 acts on the Rag GTPases to enable amino acid- and glucose-dependent mTORC1 inhibition [PMID:28199306]. Structurally, ITFG2 forms a heterodimer with KPTN docked onto a crescent-shaped SZT2 scaffold, with GATOR1 engaging SZT2's N-terminal domain via NPRL3; the complex localizes to lysosomes through SZT2/C12orf66 binding to negatively charged lipids, and disrupting the GATOR1–KICSTOR interface hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient cues [PMID:41198956]. This assembly is regulated by ubiquitination: FBXO2 ubiquitinates KPTN to dissolve the KPTN–ITFG2 and KPTN–SZT2 interactions and thereby derepress mTORC1 [PMID:41401028], and loss of ITFG2 sustains mTORC1 signaling to confer resistance to PI3Kα inhibition in PIK3CA-mutant breast cancer cells, placing it as a negative regulator of mTORC1 downstream of PI3K-AKT [PMID:33685991]. Independently of KICSTOR, ITFG2 acts in cardiomyocytes as an inhibitor of the E3 ligase NEDD4-2, blocking ubiquitination of ATP5b, SERCA2a, and Nav1.5 to preserve mitochondrial function, calcium homeostasis, and sodium-channel conduction under ischemic stress [PMID:38848780, PMID:39477020, PMID:39864577]. ITFG2 is also required cell-intrinsically in the hematopoietic compartment for germinal center formation and normal humoral immunity [PMID:23997217].","teleology":[{"year":2013,"claim":"Established the first in vivo function for ITFG2, showing it is a hematopoietic cell-intrinsic factor required for B cell differentiation and germinal center responses, before any molecular mechanism was known.","evidence":"Itfg2-knockout mice with flow cytometry, immunization, in vitro migration assays, and bone marrow transplantation","pmids":["23997217"],"confidence":"Medium","gaps":["No molecular partner or biochemical activity identified to explain the B cell phenotype","Connection to the later-defined KICSTOR/mTORC1 role not established","Single lab"]},{"year":2017,"claim":"Defined ITFG2's core molecular function by identifying it as a subunit of the KICSTOR complex that recruits GATOR1 to lysosomes, linking ITFG2 mechanistically to nutrient-dependent mTORC1 inhibition.","evidence":"Co-IP, subcellular fractionation, siRNA knockdown, and mTORC1 activity assays in human cells plus SZT2-KO mouse tissue","pmids":["28199306"],"confidence":"High","gaps":["ITFG2's specific contribution within KICSTOR not separated from other subunits","No structural basis for assembly","Relationship to the B cell phenotype unaddressed"]},{"year":2021,"claim":"Placed ITFG2 epistatically as a negative regulator of mTORC1 downstream of PI3K-AKT, with disease relevance: its loss sustains mTORC1 to confer PI3Kα-inhibitor resistance.","evidence":"Genome-wide CRISPR/Cas9 screen with individual knockout and drug-sensitivity validation in PIK3CA-mutant breast cancer cells","pmids":["33685991"],"confidence":"Medium","gaps":["Does not resolve ITFG2-specific versus complex-wide contribution","Mechanism of resistance beyond mTORC1 reactivation not dissected"]},{"year":2022,"claim":"Showed how upstream methionine/SAM sensing feeds into the GATOR1–KICSTOR module, contextualizing the complex ITFG2 belongs to within SAM-responsive mTORC1 regulation.","evidence":"Apo and SAM-bound crystal structures of SAMTOR with mutagenesis and mTORC1 activity assays","pmids":["35776786"],"confidence":"Medium","gaps":["ITFG2's direct role inferred from complex membership, not probed","No structure of ITFG2 within KICSTOR"]},{"year":2024,"claim":"Revealed a KICSTOR-independent function: in cardiomyocytes ITFG2 binds NEDD4-2 and shields its substrates from ubiquitination, protecting mitochondrial and calcium-handling machinery under ischemia.","evidence":"Co-IP, ubiquitination assays, AAV9 overexpression/knockdown in MI mouse models, mitochondrial and calcium functional assays","pmids":["38848780","39477020"],"confidence":"Medium","gaps":["Structural basis of the ITFG2–NEDD4-2 interaction unknown","Relationship between the NEDD4-2 and KICSTOR functions unresolved","Single lab"]},{"year":2025,"claim":"Provided the structural architecture of KICSTOR and a ubiquitin-based regulatory mechanism, defining how the ITFG2–KPTN heterodimer is positioned on SZT2 and how FBXO2 ubiquitination of KPTN dismantles the complex.","evidence":"Cryo-EM and computational modeling of KICSTOR; Co-IP, lysine-mutagenesis ubiquitination assays, and mTORC1 readouts for FBXO2","pmids":["41198956","41401028"],"confidence":"High","gaps":["High-resolution structure of ITFG2 itself not detailed","Physiological trigger for FBXO2-mediated KPTN ubiquitination unknown"]},{"year":2025,"claim":"Extended the cardiac NEDD4-2 inhibitory role to electrical function, showing ITFG2 preserves Nav1.5 current and conduction velocity to suppress ischemic arrhythmias.","evidence":"Co-IP, ubiquitination assays, patch-clamp, AAV9 overexpression/knockdown in MI mice, optical mapping","pmids":["39864577"],"confidence":"Medium","gaps":["Whether NEDD4-2 substrate selectivity is direct or indirect not resolved","Single lab"]},{"year":null,"claim":"It remains unknown how ITFG2's lysosomal KICSTOR/mTORC1 function relates mechanistically to its NEDD4-2-dependent cardiac role and its B cell-intrinsic requirement.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying model linking mTORC1 regulation, NEDD4-2 inhibition, and germinal center formation","ITFG2-specific catalytic or binding determinants not defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,1]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[5,6,7]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[1]}],"localization":[{"term_id":"GO:0005764","term_label":"lysosome","supporting_discovery_ids":[0,1]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,8]},{"term_id":"R-HSA-8953897","term_label":"Cellular responses to stimuli","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4]}],"complexes":["KICSTOR"],"partners":["KPTN","SZT2","C12ORF66","GATOR1","FBXO2","NEDD4-2","SAMTOR"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q969R8","full_name":"KICSTOR complex protein ITFG2","aliases":["Integrin-alpha FG-GAP repeat-containing protein 2"],"length_aa":447,"mass_kda":49.3,"function":"As part of the KICSTOR complex functions in the amino acid-sensing branch of the TORC1 signaling pathway. Recruits, in an amino acid-independent manner, the GATOR1 complex to the lysosomal membranes and allows its interaction with GATOR2 and the RAG GTPases. Functions upstream of the RAG GTPases and is required to negatively regulate mTORC1 signaling in absence of amino acids. In absence of the KICSTOR complex mTORC1 is constitutively localized to the lysosome and activated. The KICSTOR complex is also probably involved in the regulation of mTORC1 by glucose","subcellular_location":"Lysosome membrane","url":"https://www.uniprot.org/uniprotkb/Q969R8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ITFG2","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/ITFG2","total_profiled":1310},"omim":[{"mim_id":"617421","title":"INTEGRIN-ALPHA FG-GAP REPEAT-CONTAINING PROTEIN 2; ITFG2","url":"https://www.omim.org/entry/617421"},{"mim_id":"617420","title":"KICSTOR SUBUNIT 2; KICS2","url":"https://www.omim.org/entry/617420"},{"mim_id":"615620","title":"KAPTIN; KPTN","url":"https://www.omim.org/entry/615620"},{"mim_id":"615463","title":"SZT2 SUBUNIT OF KICSTOR COMPLEX; SZT2","url":"https://www.omim.org/entry/615463"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Vesicles","reliability":"Supported"},{"location":"Golgi apparatus","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ITFG2"},"hgnc":{"alias_symbol":["MDS028","KICS3"],"prev_symbol":["FGGAP1"]},"alphafold":{"accession":"Q969R8","domains":[{"cath_id":"2.130.10.10","chopping":"1-97_110-214_237-391","consensus_level":"high","plddt":92.0482,"start":1,"end":391},{"cath_id":"-","chopping":"394-425","consensus_level":"high","plddt":89.0428,"start":394,"end":425}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q969R8","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q969R8-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q969R8-F1-predicted_aligned_error_v6.png","plddt_mean":86.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ITFG2","jax_strain_url":"https://www.jax.org/strain/search?query=ITFG2"},"sequence":{"accession":"Q969R8","fasta_url":"https://rest.uniprot.org/uniprotkb/Q969R8.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q969R8/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q969R8"}},"corpus_meta":[{"pmid":"28199306","id":"PMC_28199306","title":"KICSTOR recruits GATOR1 to the lysosome and is necessary for nutrients to regulate mTORC1.","date":"2017","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/28199306","citation_count":272,"is_preprint":false},{"pmid":"35776786","id":"PMC_35776786","title":"Molecular mechanism of S-adenosylmethionine sensing by SAMTOR in mTORC1 signaling.","date":"2022","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/35776786","citation_count":34,"is_preprint":false},{"pmid":"33685991","id":"PMC_33685991","title":"Genomic Alterations in PIK3CA-Mutated Breast Cancer Result in mTORC1 Activation and Limit the Sensitivity to PI3Kα Inhibitors.","date":"2021","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/33685991","citation_count":33,"is_preprint":false},{"pmid":"33083013","id":"PMC_33083013","title":"Genomic testing in 1019 individuals from 349 Pakistani families results in high diagnostic yield and clinical utility.","date":"2020","source":"NPJ genomic medicine","url":"https://pubmed.ncbi.nlm.nih.gov/33083013","citation_count":31,"is_preprint":false},{"pmid":"16734683","id":"PMC_16734683","title":"Radiation hybrid mapping of 18 positional and physiological candidate genes for arthrogryposis multiplex congenita on porcine chromosome 5.","date":"2006","source":"Animal genetics","url":"https://pubmed.ncbi.nlm.nih.gov/16734683","citation_count":15,"is_preprint":false},{"pmid":"23997217","id":"PMC_23997217","title":"Integrin-α FG-GAP repeat-containing protein 2 is critical for normal B cell differentiation and controls disease development in a lupus model.","date":"2013","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/23997217","citation_count":10,"is_preprint":false},{"pmid":"38848780","id":"PMC_38848780","title":"ITFG2, an immune-modulatory protein, targets ATP 5b to maintain mitochondrial function in myocardial infarction.","date":"2024","source":"Biochemical pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/38848780","citation_count":4,"is_preprint":false},{"pmid":"38142287","id":"PMC_38142287","title":"Co-methylation analyses identify CpGs associated with lipid traits in Chinese discordant monozygotic twins.","date":"2024","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38142287","citation_count":3,"is_preprint":false},{"pmid":"41198956","id":"PMC_41198956","title":"Architecture of the human KICSTOR and GATOR1-KICSTOR complexes.","date":"2025","source":"Nature structural & molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/41198956","citation_count":2,"is_preprint":false},{"pmid":"39477020","id":"PMC_39477020","title":"ITFG2 as a NEDD4-2 inhibitor: Preserving calcium homeostasis to prevent myocardial ischemic injury.","date":"2024","source":"Biochemical pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/39477020","citation_count":1,"is_preprint":false},{"pmid":"39864577","id":"PMC_39864577","title":"A New target of ischemic ventricular arrhythmias-ITFG2.","date":"2025","source":"European journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/39864577","citation_count":1,"is_preprint":false},{"pmid":"41401028","id":"PMC_41401028","title":"FBXO2-mediated KPTN ubiquitination promotes amino acid-dependent mTORC1 signaling and tumor growth.","date":"2025","source":"The Journal of clinical investigation","url":"https://pubmed.ncbi.nlm.nih.gov/41401028","citation_count":1,"is_preprint":false},{"pmid":"41041534","id":"PMC_41041534","title":"Genomic relationship between polycystic ovary syndrome and bipolar disorder.","date":"2025","source":"Research square","url":"https://pubmed.ncbi.nlm.nih.gov/41041534","citation_count":0,"is_preprint":false},{"pmid":"41158364","id":"PMC_41158364","title":"The potential of circITFG2 as a therapeutic target in lung squamous cell carcinoma.","date":"2025","source":"Journal of thoracic disease","url":"https://pubmed.ncbi.nlm.nih.gov/41158364","citation_count":0,"is_preprint":false},{"pmid":"42258707","id":"PMC_42258707","title":"Host genetic regulation of rumen 6-hydroxymelatonin reduces methane emissions in dairy cattle.","date":"2026","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/42258707","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":10200,"output_tokens":2713,"usd":0.035647,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9919,"output_tokens":3031,"usd":0.062685,"stage2_stop_reason":"end_turn"},"total_usd":0.098332,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2017,\n      \"finding\": \"ITFG2 is a component of the KICSTOR complex (with KPTN, C12orf66, and SZT2) that localizes to lysosomes, binds and recruits GATOR1 (but not GATOR2) to the lysosomal surface, and is necessary for GATOR1 to interact with its substrates the Rag GTPases and with GATOR2; loss of KICSTOR prevents amino acid or glucose deprivation from inhibiting mTORC1.\",\n      \"method\": \"Co-immunoprecipitation, subcellular fractionation/localization, siRNA knockdown, mTORC1 activity assays in cultured human cells, and SZT2-knockout mouse tissue analysis\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, lysosomal localization with functional consequence, genetic KO in mice, replicated by multiple subsequent studies\",\n      \"pmids\": [\"28199306\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Cryo-EM and computational modeling revealed that within KICSTOR, SZT2 forms a crescent-shaped scaffold that binds the ITFG2–KPTN heterodimer and C12orf66 at its C terminus; GATOR1 binds the SZT2 N-terminal domain via NPRL3; disruption of this GATOR1–KICSTOR interface hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient status. SZT2 and C12orf66 preferentially interact with negatively charged lipids, a requirement for lysosomal localization of the entire complex.\",\n      \"method\": \"Cryo-electron microscopy, computational modeling, biochemical interaction assays, lipid-binding assays, functional mTORC1 activity and TFE3 localization assays\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with biochemical validation and functional assays in a single rigorous study\",\n      \"pmids\": [\"41198956\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FBXO2 directly interacts with KPTN (a KICSTOR subunit) via its F-box-associated domain and promotes K48- and K63-linked polyubiquitination of KPTN at lysines 49, 67, 262, and 265; this ubiquitination disrupts the KPTN–ITFG2 and KPTN–SZT2 interactions, impairing KICSTOR's ability to recruit GATOR1 to the lysosomal surface and thereby activating mTORC1 signaling.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, site-directed mutagenesis of KPTN lysine residues, mTORC1 activity assays, lysosomal localization experiments\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct interaction, mutagenesis identifying specific ubiquitination sites, functional mTORC1 readout, multiple orthogonal methods in one study\",\n      \"pmids\": [\"41401028\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Crystal structures of SAMTOR show that it interacts with GATOR1–KICSTOR (which contains ITFG2) through its SAM-dependent methyltransferase domain; conformational change in the SAMTOR helical domain upon SAM binding modulates the interaction of SAMTOR with the GATOR1–KICSTOR complex, thereby regulating mTORC1 activity in response to SAM levels.\",\n      \"method\": \"Crystal structure determination (apo and SAM-bound), site-directed mutagenesis, functional binding and mTORC1 activity assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — crystal structure with mutagenesis, but ITFG2's specific contribution within KICSTOR is inferred from complex membership rather than directly probed\",\n      \"pmids\": [\"35776786\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ITFG2 (Itfg2) is an intracellular protein in mice that is required for normal B cell differentiation; Itfg2-deficient mice show retention of B cells in the spleen, reduced serum IgG, defective cell migration in vitro, a shift of B cell maturation from germinal centers to extrafollicular regions, and blocked plasma cell deposition in bone marrow; bone marrow transplantation of Itfg2-deficient cells was sufficient to impair germinal center development in wild-type recipients, indicating hematopoietic cell-intrinsic activity.\",\n      \"method\": \"Itfg2-knockout mice, flow cytometry, bone marrow transplantation, in vitro migration assay, immunization experiments\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO mouse with defined cellular phenotypes, bone marrow transplantation establishing cell-intrinsic role, multiple readouts in single lab\",\n      \"pmids\": [\"23997217\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ITFG2 forms a complex with NEDD4-2 and ATP5b (mitochondrial ATP synthase β subunit) in cardiomyocytes; ITFG2 inhibits NEDD4-2-mediated ubiquitination of ATP5b, thereby preserving mitochondrial ATP production, reducing ROS, and maintaining mitochondrial membrane potential under hypoxic conditions; overexpression in a mouse MI model reduced infarct size and improved cardiac function.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, AAV9-mediated overexpression, shRNA/siRNA knockdown, cardiac-specific transgenic mice, mitochondrial function assays (ATP, ROS, MMP), echocardiography\",\n      \"journal\": \"Biochemical pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP identifying complex, ubiquitination assay, multiple in vivo and in vitro models, single lab\",\n      \"pmids\": [\"38848780\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ITFG2 binds NEDD4-2 in cardiomyocytes and decreases NEDD4-2's interaction with SERCA2a, preventing NEDD4-2-mediated ubiquitination and degradation of SERCA2a; this preserves calcium homeostasis and cardiac contractility under ischemic conditions.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, AAV9 overexpression and shRNA knockdown in MI mouse model, echocardiography, calcium measurement in primary cardiomyocytes\",\n      \"journal\": \"Biochemical pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assays with functional cardiac readouts, single lab, two orthogonal methods\",\n      \"pmids\": [\"39477020\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ITFG2 reduces the binding affinity between NEDD4-2 and Nav1.5 (voltage-gated sodium channel), inhibiting Nav1.5 ubiquitination; ITFG2 overexpression increased peak Nav1.5 current by ~50%, upregulated Nav1.5 protein expression, increased conduction velocity, and reduced ischemic ventricular arrhythmias in MI mice.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, patch-clamp electrophysiology, AAV9 overexpression and shRNA knockdown in MI mouse model, optical mapping of epicardial conduction\",\n      \"journal\": \"European journal of pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assays with electrophysiological validation, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"39864577\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Loss of ITFG2 (along with other KICSTOR/GATOR1 components) identified in a whole-genome CRISPR/Cas9 knockout screen confers resistance to PI3Kα inhibition in PIK3CA-mutated breast cancer cells by sustaining mTORC1 signaling, placing ITFG2 as a negative regulator of mTORC1 downstream of PI3K-AKT.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 sgRNA knockout screen, validated by individual gene knockouts, mTORC1 signaling assays, sensitivity assays with PI3Kα inhibitors\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR screen with functional validation, epistatic placement relative to PI3K-AKT and mTORC1, single lab\",\n      \"pmids\": [\"33685991\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ITFG2 is a subunit of the lysosome-associated KICSTOR complex (with KPTN, C12orf66, and SZT2), where it forms a heterodimer with KPTN on a crescent-shaped SZT2 scaffold; KICSTOR recruits GATOR1 to the lysosomal surface to enable GAP activity toward Rag GTPases and mTORC1 inhibition in response to nutrient deprivation. In cardiomyocytes, ITFG2 additionally functions as a NEDD4-2 inhibitor, blocking ubiquitination of SERCA2a, ATP5b, and Nav1.5 to preserve mitochondrial function, calcium homeostasis, and sodium channel activity under ischemic conditions. In B cells, ITFG2 is required cell-intrinsically for germinal center formation and normal humoral immune responses.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ITFG2 is a subunit of the lysosome-associated KICSTOR complex (with KPTN, C12orf66, and SZT2) that couples nutrient status to mTORC1 signaling by recruiting GATOR1 to the lysosomal surface, where GATOR1 acts on the Rag GTPases to enable amino acid- and glucose-dependent mTORC1 inhibition [#0]. Structurally, ITFG2 forms a heterodimer with KPTN docked onto a crescent-shaped SZT2 scaffold, with GATOR1 engaging SZT2's N-terminal domain via NPRL3; the complex localizes to lysosomes through SZT2/C12orf66 binding to negatively charged lipids, and disrupting the GATOR1\\u2013KICSTOR interface hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient cues [#1]. This assembly is regulated by ubiquitination: FBXO2 ubiquitinates KPTN to dissolve the KPTN\\u2013ITFG2 and KPTN\\u2013SZT2 interactions and thereby derepress mTORC1 [#2], and loss of ITFG2 sustains mTORC1 signaling to confer resistance to PI3K\\u03b1 inhibition in PIK3CA-mutant breast cancer cells, placing it as a negative regulator of mTORC1 downstream of PI3K-AKT [#8]. Independently of KICSTOR, ITFG2 acts in cardiomyocytes as an inhibitor of the E3 ligase NEDD4-2, blocking ubiquitination of ATP5b, SERCA2a, and Nav1.5 to preserve mitochondrial function, calcium homeostasis, and sodium-channel conduction under ischemic stress [#5, #6, #7]. ITFG2 is also required cell-intrinsically in the hematopoietic compartment for germinal center formation and normal humoral immunity [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2013,\n      \"claim\": \"Established the first in vivo function for ITFG2, showing it is a hematopoietic cell-intrinsic factor required for B cell differentiation and germinal center responses, before any molecular mechanism was known.\",\n      \"evidence\": \"Itfg2-knockout mice with flow cytometry, immunization, in vitro migration assays, and bone marrow transplantation\",\n      \"pmids\": [\"23997217\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No molecular partner or biochemical activity identified to explain the B cell phenotype\", \"Connection to the later-defined KICSTOR/mTORC1 role not established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined ITFG2's core molecular function by identifying it as a subunit of the KICSTOR complex that recruits GATOR1 to lysosomes, linking ITFG2 mechanistically to nutrient-dependent mTORC1 inhibition.\",\n      \"evidence\": \"Co-IP, subcellular fractionation, siRNA knockdown, and mTORC1 activity assays in human cells plus SZT2-KO mouse tissue\",\n      \"pmids\": [\"28199306\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"ITFG2's specific contribution within KICSTOR not separated from other subunits\", \"No structural basis for assembly\", \"Relationship to the B cell phenotype unaddressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Placed ITFG2 epistatically as a negative regulator of mTORC1 downstream of PI3K-AKT, with disease relevance: its loss sustains mTORC1 to confer PI3K\\u03b1-inhibitor resistance.\",\n      \"evidence\": \"Genome-wide CRISPR/Cas9 screen with individual knockout and drug-sensitivity validation in PIK3CA-mutant breast cancer cells\",\n      \"pmids\": [\"33685991\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Does not resolve ITFG2-specific versus complex-wide contribution\", \"Mechanism of resistance beyond mTORC1 reactivation not dissected\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed how upstream methionine/SAM sensing feeds into the GATOR1\\u2013KICSTOR module, contextualizing the complex ITFG2 belongs to within SAM-responsive mTORC1 regulation.\",\n      \"evidence\": \"Apo and SAM-bound crystal structures of SAMTOR with mutagenesis and mTORC1 activity assays\",\n      \"pmids\": [\"35776786\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"ITFG2's direct role inferred from complex membership, not probed\", \"No structure of ITFG2 within KICSTOR\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Revealed a KICSTOR-independent function: in cardiomyocytes ITFG2 binds NEDD4-2 and shields its substrates from ubiquitination, protecting mitochondrial and calcium-handling machinery under ischemia.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, AAV9 overexpression/knockdown in MI mouse models, mitochondrial and calcium functional assays\",\n      \"pmids\": [\"38848780\", \"39477020\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of the ITFG2\\u2013NEDD4-2 interaction unknown\", \"Relationship between the NEDD4-2 and KICSTOR functions unresolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Provided the structural architecture of KICSTOR and a ubiquitin-based regulatory mechanism, defining how the ITFG2\\u2013KPTN heterodimer is positioned on SZT2 and how FBXO2 ubiquitination of KPTN dismantles the complex.\",\n      \"evidence\": \"Cryo-EM and computational modeling of KICSTOR; Co-IP, lysine-mutagenesis ubiquitination assays, and mTORC1 readouts for FBXO2\",\n      \"pmids\": [\"41198956\", \"41401028\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"High-resolution structure of ITFG2 itself not detailed\", \"Physiological trigger for FBXO2-mediated KPTN ubiquitination unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended the cardiac NEDD4-2 inhibitory role to electrical function, showing ITFG2 preserves Nav1.5 current and conduction velocity to suppress ischemic arrhythmias.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, patch-clamp, AAV9 overexpression/knockdown in MI mice, optical mapping\",\n      \"pmids\": [\"39864577\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether NEDD4-2 substrate selectivity is direct or indirect not resolved\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown how ITFG2's lysosomal KICSTOR/mTORC1 function relates mechanistically to its NEDD4-2-dependent cardiac role and its B cell-intrinsic requirement.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying model linking mTORC1 regulation, NEDD4-2 inhibition, and germinal center formation\", \"ITFG2-specific catalytic or binding determinants not defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [5, 6, 7]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005764\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 8]},\n      {\"term_id\": \"R-HSA-8953897\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"complexes\": [\"KICSTOR\"],\n    \"partners\": [\"KPTN\", \"SZT2\", \"C12orf66\", \"GATOR1\", \"FBXO2\", \"NEDD4-2\", \"SAMTOR\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}