{"gene":"SNF8","run_date":"2026-04-28T20:42:08","timeline":{"discoveries":[{"year":1999,"finding":"EAP30 (SNF8) was identified as a subunit of the ELL transcription elongation complex; it interacts directly with ELL and derepresses ELL's inhibitory activity on RNA polymerase II transcription in vitro.","method":"Co-purification/biochemical characterization of the ELL complex; in vitro transcription assay demonstrating derepression of RNA Pol II activity","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 — in vitro transcription assay showing functional derepression, single lab","pmids":["10419521"],"is_preprint":false},{"year":1995,"finding":"In S. cerevisiae, SNF8 is required for derepression of SUC2 (invertase) gene expression in response to glucose limitation; genetic analysis showed that SNF7 and SNF8 are functionally related (double mutant shows no additive phenotype), while snf8 ssn6 double mutants were extremely sick, placing SNF8 genetically distinct from SNF1/SNF4 and SNF2/SNF5/SNF6 groups.","method":"Gene disruption/null mutation, genetic epistasis analysis with suppressors (spt6/ssn20, ssn6), invertase derepression assay","journal":"Yeast (Chichester, England)","confidence":"Medium","confidence_rationale":"Tier 2 — clean KO with defined biochemical phenotype, genetic epistasis in yeast ortholog","pmids":["7785322"],"is_preprint":false},{"year":2007,"finding":"Vps22/EAP30 (SNF8), as an ESCRT-II subunit, is required for lysosomal degradation of ubiquitinated receptors (EGFR and CXCR4); its depletion causes accumulation of EGFR on limiting membranes of early endosomes and aberrantly small multivesicular bodies (MVBs). Notably, depletion of Vps22 — unlike depletion of Hrs or Tsg101 (ESCRT-I) — did not sustain EGF-induced ERK1/2 phosphorylation and nuclear translocation, suggesting EGF signaling termination occurs prior to ESCRT-II engagement.","method":"siRNA knockdown in HeLa cells, immunofluorescence/confocal microscopy, EGFR degradation assay, ERK1/2 phosphorylation and nuclear translocation assay","journal":"Traffic (Copenhagen, Denmark)","confidence":"High","confidence_rationale":"Tier 2 — reciprocal localization, functional KD with multiple orthogonal readouts (degradation, morphology, signaling), moderate evidence","pmids":["17714434"],"is_preprint":false},{"year":2006,"finding":"RILP (Rab-interacting lysosomal protein) interacts with VPS22 (SNF8) and VPS36 of ESCRT-II, and this interaction mediates membrane recruitment of ESCRT-II subunits; overexpression of RILP leads to enlarged and clustered MVBs and retards sorting of internalized EGF to degradation.","method":"Co-immunoprecipitation, yeast two-hybrid (for domain mapping), overexpression with EEA1/LBPA marker analysis, EGF sorting assay","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 — co-IP and functional overexpression, replicated in two independent labs (PMIDs 17010938 and 16857164)","pmids":["17010938","16857164"],"is_preprint":false},{"year":2006,"finding":"RILP interacts specifically with the N-terminal half of VPS22 (SNF8/EAP30); this interaction was validated by co-immunoprecipitation and colocalization of GFP-RILP and HA-VPS22 in endosomes.","method":"Yeast two-hybrid screen (isolation), co-immunoprecipitation, confocal immunofluorescence colocalization","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 3 — yeast two-hybrid plus co-IP plus colocalization, replicated by second lab","pmids":["16857164"],"is_preprint":false},{"year":2009,"finding":"MCM2 binds directly to EAP30 (SNF8) and competes with ELL for binding to EAP30, thereby potentially modulating the stability of the Holo-ELL complex and regulating the balance between RNA Pol II elongation and initiation activities.","method":"Co-immunoprecipitation, competition binding assay","journal":"FEBS letters","confidence":"Low","confidence_rationale":"Tier 3 — single lab, single method (co-IP), no functional transcription readout","pmids":["19819239"],"is_preprint":false},{"year":2012,"finding":"SNF8 (EAP30/VPS22) physically interacts with the amino-terminal cytoplasmic domain (first 107 amino acids) of TRPC6 channel; overexpression of SNF8 enhances TRPC6-mediated whole-cell currents and NFAT-mediated transcription, while RNAi knockdown of SNF8 partially inhibits NFAT activation, without altering cell-surface TRPC6 levels.","method":"Yeast two-hybrid screen (initial discovery), co-immunoprecipitation from mammalian cells, whole-cell patch clamp electrophysiology, NFAT-luciferase reporter assay, surface biotinylation, RNAi knockdown","journal":"BMC cell biology","confidence":"Medium","confidence_rationale":"Tier 2 — multiple orthogonal methods (co-IP, electrophysiology, reporter assay, RNAi), single lab","pmids":["23171048"],"is_preprint":false},{"year":2017,"finding":"EAP30 (SNF8) localizes in part to the nucleus where it forms a virus-inducible complex with IRF3 and its co-activator CBP; this nuclear complex is required for IRF3 binding to target gene promoters (IFN-β, IFN-λ1, ISG56) and induction of type I/III IFNs and ISGs in response to dsRNA or viral infection, acting downstream of IRF3 phosphorylation/activation and independently of NF-κB-driven inflammatory gene induction.","method":"siRNA knockdown with IFN/ISG induction assays, nuclear fractionation, co-immunoprecipitation of EAP30-IRF3-CBP complex, chromatin immunoprecipitation (IRF3 promoter binding), antiviral assays (VSV, HCV)","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods (KD phenotype, nuclear fractionation, co-IP, ChIP) with strong mechanistic resolution, single lab","pmids":["29084253"],"is_preprint":false},{"year":2005,"finding":"In fission yeast, Dot2 (the EAP30/SNF8 homolog) negatively regulates meiotic spindle pole body (SPB) maturation; dot2 mutants accumulate excess electron-dense material near SPBs forming aberrant MTOCs with multipolar spindles. SPB aberrations correlated with elevated Pcp1 (pericentrin ortholog) levels, and reducing pcp1 expression suppressed the dot2 SPB phenotype, placing Dot2 upstream of Pcp1-dependent SPB maturation.","method":"dot2 mutant analysis, electron microscopy, genetic epistasis (pcp1 suppression of dot2), immunofluorescence","journal":"Developmental cell","confidence":"Medium","confidence_rationale":"Tier 2 — genetic epistasis with morphological validation in ortholog, single lab","pmids":["15992541"],"is_preprint":false},{"year":2020,"finding":"In C. elegans, VPS-22/SNF8 acts in the same pathway as DAF-16/FOXO to regulate longevity; knockdown of vps-22/snf8 reduces nuclear localization of DAF-16, and overexpression of daf-16 rescues the short-lived phenotype of vps-22/snf8 knockdown, positioning VPS-22/SNF8 upstream of DAF-16 nuclear activity.","method":"RNAi knockdown lifespan assay, daf-16 null genetic epistasis, DAF-16::GFP nuclear localization imaging, downstream gene expression analysis","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 — genetic epistasis with localization readout in C. elegans ortholog, single lab","pmids":["32829877"],"is_preprint":false}],"current_model":"SNF8/EAP30/VPS22 is a multifunctional ESCRT-II subunit that acts at endosomes to sort ubiquitinated receptors (e.g., EGFR, CXCR4) into MVBs for lysosomal degradation — a process regulated by RILP-mediated ESCRT-II membrane recruitment — and also functions in the nucleus as part of the ELL RNA Pol II elongation complex and as an IRF3/CBP co-activator complex required for antiviral IFN gene induction, while additionally modulating TRPC6 channel activity and, in model organisms, regulating centrosome/SPB maturation and FOXO/DAF-16-dependent longevity."},"narrative":{"teleology":[{"year":1995,"claim":"Establishing SNF8 as a transcriptional regulator: genetic analysis in yeast showed that SNF8 is required for glucose-regulated SUC2 gene derepression and is functionally related to SNF7 but genetically distinct from SNF1/SNF4 and SWI/SNF groups, defining it as part of a separate chromatin/transcription regulatory pathway.","evidence":"Gene disruption, genetic epistasis, and invertase derepression assays in S. cerevisiae","pmids":["7785322"],"confidence":"Medium","gaps":["Molecular mechanism of SNF8-dependent transcriptional derepression in yeast was not defined","Whether SNF8 functions as part of a stable complex in yeast was unknown","Relationship to endosomal sorting function not yet recognized"]},{"year":1999,"claim":"Identifying a direct transcriptional mechanism: EAP30/SNF8 was found as a subunit of the mammalian ELL elongation complex, where it directly binds ELL and derepresses ELL's inhibitory effect on RNA Pol II transcription, providing the first biochemical activity for SNF8 in metazoans.","evidence":"Co-purification of the ELL complex and in vitro transcription assay measuring RNA Pol II derepression","pmids":["10419521"],"confidence":"Medium","gaps":["In vivo relevance of ELL complex derepression not demonstrated","Whether EAP30 has functions independent of ELL was unknown","Target gene specificity of ELL-EAP30 complex not defined"]},{"year":2005,"claim":"Revealing a role in centrosome/SPB maturation: the fission yeast SNF8 ortholog Dot2 negatively regulates spindle pole body maturation, with dot2 mutants showing excess MTOC material and multipolar spindles that are suppressed by reducing Pcp1/pericentrin, placing SNF8 upstream of centrosome component homeostasis.","evidence":"Mutant analysis with electron microscopy and genetic epistasis (pcp1 suppression) in S. pombe","pmids":["15992541"],"confidence":"Medium","gaps":["Whether this reflects an ESCRT-II-dependent or ESCRT-II-independent function was not determined","Mechanism by which Dot2 regulates Pcp1 levels was not identified","Conservation of this SPB role in mammalian cells untested"]},{"year":2006,"claim":"Defining the membrane recruitment mechanism: RILP was shown to interact directly with the N-terminal half of VPS22/SNF8 and to recruit ESCRT-II to endosomal membranes, connecting Rab7-RILP signaling to ESCRT-II-mediated receptor sorting.","evidence":"Yeast two-hybrid, co-immunoprecipitation, confocal colocalization, and EGF sorting assays in mammalian cells; confirmed by two independent labs","pmids":["17010938","16857164"],"confidence":"Medium","gaps":["Whether RILP is the sole recruitment factor for ESCRT-II was not resolved","Structural basis of the RILP–VPS22 interaction not determined","Relative contribution of RILP versus lipid-binding (VPS36) to ESCRT-II membrane association unclear"]},{"year":2007,"claim":"Establishing the core endosomal sorting function: siRNA depletion of VPS22/SNF8 in HeLa cells demonstrated that ESCRT-II is required for lysosomal degradation of EGFR and CXCR4 and for normal MVB biogenesis, while showing that EGF-induced ERK signaling is terminated prior to ESCRT-II engagement — distinguishing ESCRT-II's role from upstream ESCRT-0/I in signal attenuation.","evidence":"siRNA knockdown with EGFR degradation kinetics, confocal microscopy of MVB morphology, and ERK1/2 phosphorylation/translocation assays in HeLa cells","pmids":["17714434"],"confidence":"High","gaps":["Range of cargo receptors requiring ESCRT-II beyond EGFR/CXCR4 not systematically defined","Whether ESCRT-II has cargo-selective versus general roles in MVB formation not resolved"]},{"year":2009,"claim":"Suggesting competition for SNF8 between transcription and replication complexes: MCM2 was found to bind EAP30/SNF8 and compete with ELL for the same binding site, raising the possibility that SNF8 coordinates elongation and initiation activities.","evidence":"Co-immunoprecipitation and competition binding assay","pmids":["19819239"],"confidence":"Low","gaps":["Single-method co-IP without functional transcription or replication readout","Physiological context in which MCM2-EAP30 interaction is relevant not established","No independent confirmation of the competition model"]},{"year":2012,"claim":"Uncovering a non-ESCRT channel-regulatory function: SNF8 was found to bind the TRPC6 cytoplasmic domain and enhance TRPC6-mediated currents and downstream NFAT transcriptional activation without altering channel surface expression, identifying SNF8 as a modulator of ion channel gating.","evidence":"Yeast two-hybrid, co-immunoprecipitation, whole-cell patch clamp electrophysiology, NFAT-luciferase reporter, surface biotinylation, and RNAi in mammalian cells","pmids":["23171048"],"confidence":"Medium","gaps":["Mechanism by which SNF8 enhances TRPC6 currents (gating vs. conductance) not resolved","Whether this is an ESCRT-II-dependent or SNF8-autonomous function unknown","In vivo physiological relevance (e.g., in podocytes) not tested"]},{"year":2017,"claim":"Establishing a nuclear role in innate antiviral immunity: SNF8 was shown to form a virus-inducible nuclear complex with IRF3 and CBP that is required for IRF3 binding to IFN-β/IFN-λ1/ISG56 promoters and for type I/III interferon induction, acting downstream of IRF3 phosphorylation and independently of NF-κB signaling.","evidence":"siRNA knockdown with IFN/ISG induction assays, nuclear fractionation, co-immunoprecipitation of EAP30–IRF3–CBP, ChIP for IRF3 at promoters, and antiviral assays (VSV, HCV)","pmids":["29084253"],"confidence":"High","gaps":["Whether SNF8 recruits chromatin remodeling or additional coactivators to IRF3 targets not determined","Structural basis of the IRF3–EAP30–CBP ternary complex unknown","Whether ESCRT-II partners VPS25/VPS36 participate in the nuclear antiviral function not tested"]},{"year":2020,"claim":"Linking ESCRT-II to longevity regulation: in C. elegans, VPS-22/SNF8 promotes nuclear localization of DAF-16/FOXO and acts in the same genetic pathway, with DAF-16 overexpression rescuing the short-lived phenotype of vps-22 knockdown, positioning SNF8 upstream of FOXO-dependent longevity.","evidence":"RNAi lifespan assay, daf-16 epistasis, DAF-16::GFP nuclear localization, and target gene expression in C. elegans","pmids":["32829877"],"confidence":"Medium","gaps":["Whether the effect on DAF-16 localization is direct or via endosomal signaling regulation not resolved","Conservation in mammalian aging pathways not tested","Which endosomal cargo drives the longevity phenotype is unknown"]},{"year":null,"claim":"How SNF8 partitions between its ESCRT-II endosomal function and its nuclear transcriptional/antiviral roles remains unresolved, as does whether these represent independent or coordinated activities.","evidence":"","pmids":[],"confidence":"Low","gaps":["No regulatory mechanism identified that switches SNF8 between endosomal and nuclear pools","No structural model of full-length human ESCRT-II with SNF8 in a membrane-associated state","Systematic identification of SNF8-dependent cargo in mammalian cells not performed"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,6,7]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[0,7]}],"localization":[{"term_id":"GO:0005768","term_label":"endosome","supporting_discovery_ids":[2,3,4]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[7]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[2,3]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[2,3]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[7]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[0,7]}],"complexes":["ESCRT-II","ELL elongation complex","IRF3-CBP-EAP30 complex"],"partners":["VPS36","VPS25","ELL","RILP","IRF3","CREBBP","TRPC6","MCM2"],"other_free_text":[]},"mechanistic_narrative":"SNF8 (also known as EAP30/VPS22) is a core subunit of the ESCRT-II complex that functions in endosomal sorting of ubiquitinated membrane receptors and also participates in nuclear transcriptional regulation. As an ESCRT-II component, SNF8 is required for sorting ubiquitinated EGFR and CXCR4 into multivesicular bodies for lysosomal degradation, with its membrane recruitment mediated by direct interaction of its N-terminal region with RILP [PMID:17714434, PMID:16857164]. In the nucleus, SNF8 serves dual transcriptional roles: it associates with the ELL elongation complex to derepress RNA Pol II elongation activity [PMID:10419521], and it forms a virus-inducible complex with IRF3 and CBP that is essential for IRF3 binding to IFN-β/IFN-λ1 promoters and antiviral interferon gene induction [PMID:29084253]. SNF8 additionally modulates TRPC6 channel currents and downstream NFAT activation through direct interaction with the TRPC6 cytoplasmic domain, independent of effects on channel surface expression [PMID:23171048]."},"prefetch_data":{"uniprot":{"accession":"Q96H20","full_name":"Vacuolar-sorting protein SNF8","aliases":["ELL-associated protein of 30 kDa","ESCRT-II complex subunit VPS22","hVps22"],"length_aa":258,"mass_kda":28.9,"function":"Component of the endosomal sorting complex required for transport II (ESCRT-II), which is required for multivesicular body (MVB) formation and sorting of endosomal cargo proteins into MVBs, and plays a role in autophagy (PubMed:38423010). The MVB pathway mediates delivery of transmembrane proteins into the lumen of the lysosome for degradation. The ESCRT-II complex is probably involved in the recruitment of the ESCRT-III complex. The ESCRT-II complex may also play a role in transcription regulation by participating in derepression of transcription by RNA polymerase II, possibly via its interaction with ELL. Required for degradation of both endocytosed EGF and EGFR, but not for the EGFR ligand-mediated internalization. It is also required for the degradation of CXCR4. Required for the exosomal release of SDCBP, CD63 and syndecan (PubMed:22660413)","subcellular_location":"Cytoplasm; Endosome membrane; Nucleus; Late endosome membrane","url":"https://www.uniprot.org/uniprotkb/Q96H20/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/SNF8","classification":"Common Essential","n_dependent_lines":1157,"n_total_lines":1208,"dependency_fraction":0.9577814569536424},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000159210","cell_line_id":"CID000783","localizations":[{"compartment":"cytoplasmic","grade":3},{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"VPS36","stoichiometry":10.0},{"gene":"VPS25","stoichiometry":10.0},{"gene":"UBE3B","stoichiometry":0.2},{"gene":"TNPO2","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000783","total_profiled":1310},"omim":[{"mim_id":"620784","title":"NEURODEVELOPMENTAL DISORDER PLUS OPTIC ATROPHY; NEDOA","url":"https://www.omim.org/entry/620784"},{"mim_id":"620783","title":"DEVELOPMENTAL AND EPILEPTIC ENCEPHALOPATHY 115; DEE115","url":"https://www.omim.org/entry/620783"},{"mim_id":"610907","title":"VACUOLAR PROTEIN SORTING 25 HOMOLOG; VPS25","url":"https://www.omim.org/entry/610907"},{"mim_id":"610904","title":"SNF8 SUBUNIT OF ESCRIT-II; SNF8","url":"https://www.omim.org/entry/610904"},{"mim_id":"610903","title":"VACUOLAR PROTEIN SORTING 36 HOMOLOG; VPS36","url":"https://www.omim.org/entry/610903"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Cytosol","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/SNF8"},"hgnc":{"alias_symbol":["EAP30","VPS22","Dot3"],"prev_symbol":[]},"alphafold":{"accession":"Q96H20","domains":[{"cath_id":"1.10.10.10","chopping":"44-155","consensus_level":"medium","plddt":85.2311,"start":44,"end":155},{"cath_id":"1.10.10.10","chopping":"174-246","consensus_level":"medium","plddt":89.8823,"start":174,"end":246}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96H20","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q96H20-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q96H20-F1-predicted_aligned_error_v6.png","plddt_mean":84.94},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=SNF8","jax_strain_url":"https://www.jax.org/strain/search?query=SNF8"},"sequence":{"accession":"Q96H20","fasta_url":"https://rest.uniprot.org/uniprotkb/Q96H20.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q96H20/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96H20"}},"corpus_meta":[{"pmid":"17714434","id":"PMC_17714434","title":"Vps22/EAP30 in ESCRT-II mediates endosomal sorting of growth factor and chemokine receptors destined for lysosomal degradation.","date":"2007","source":"Traffic (Copenhagen, Denmark)","url":"https://pubmed.ncbi.nlm.nih.gov/17714434","citation_count":97,"is_preprint":false},{"pmid":"17010938","id":"PMC_17010938","title":"RILP interacts with VPS22 and VPS36 of ESCRT-II and regulates their membrane recruitment.","date":"2006","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/17010938","citation_count":49,"is_preprint":false},{"pmid":"16857164","id":"PMC_16857164","title":"RILP interacts with the VPS22 component of the ESCRT-II complex.","date":"2006","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/16857164","citation_count":43,"is_preprint":false},{"pmid":"10419521","id":"PMC_10419521","title":"Cloning and characterization of the EAP30 subunit of the ELL complex that confers derepression of transcription by RNA polymerase II.","date":"1999","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/10419521","citation_count":35,"is_preprint":false},{"pmid":"23275199","id":"PMC_23275199","title":"Knockout of the VPS22 component of the ESCRT-II complex in rice (Oryza sativa L.) causes chalky endosperm and early seedling lethality.","date":"2012","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/23275199","citation_count":26,"is_preprint":false},{"pmid":"15992541","id":"PMC_15992541","title":"The fission yeast homolog of the human transcription factor EAP30 blocks meiotic spindle pole body amplification.","date":"2005","source":"Developmental cell","url":"https://pubmed.ncbi.nlm.nih.gov/15992541","citation_count":22,"is_preprint":false},{"pmid":"7785322","id":"PMC_7785322","title":"Molecular analysis of the SNF8 gene of Saccharomyces cerevisiae.","date":"1995","source":"Yeast (Chichester, England)","url":"https://pubmed.ncbi.nlm.nih.gov/7785322","citation_count":20,"is_preprint":false},{"pmid":"29084253","id":"PMC_29084253","title":"Pivotal role for the ESCRT-II complex subunit EAP30/SNF8 in IRF3-dependent innate antiviral defense.","date":"2017","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/29084253","citation_count":19,"is_preprint":false},{"pmid":"23171048","id":"PMC_23171048","title":"SNF8, a member of the ESCRT-II complex, interacts with TRPC6 and enhances its channel activity.","date":"2012","source":"BMC cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/23171048","citation_count":11,"is_preprint":false},{"pmid":"19819239","id":"PMC_19819239","title":"The interaction between EAP30 and ELL is modulated by MCM2.","date":"2009","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/19819239","citation_count":4,"is_preprint":false},{"pmid":"32829877","id":"PMC_32829877","title":"VPS-22/SNF8 regulates longevity via modulating the activity of DAF-16 in C. elegans.","date":"2020","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/32829877","citation_count":3,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":5879,"output_tokens":2687,"usd":0.028971},"stage2":{"model":"claude-opus-4-6","input_tokens":6020,"output_tokens":3072,"usd":0.16035},"total_usd":0.189321,"stage1_batch_id":"msgbatch_011rkcQdVV1RaFq9XhcG6df6","stage2_batch_id":"msgbatch_01RzgrJafdF4EWHhj5mNi6dh","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1999,\n      \"finding\": \"EAP30 (SNF8) was identified as a subunit of the ELL transcription elongation complex; it interacts directly with ELL and derepresses ELL's inhibitory activity on RNA polymerase II transcription in vitro.\",\n      \"method\": \"Co-purification/biochemical characterization of the ELL complex; in vitro transcription assay demonstrating derepression of RNA Pol II activity\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — in vitro transcription assay showing functional derepression, single lab\",\n      \"pmids\": [\"10419521\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1995,\n      \"finding\": \"In S. cerevisiae, SNF8 is required for derepression of SUC2 (invertase) gene expression in response to glucose limitation; genetic analysis showed that SNF7 and SNF8 are functionally related (double mutant shows no additive phenotype), while snf8 ssn6 double mutants were extremely sick, placing SNF8 genetically distinct from SNF1/SNF4 and SNF2/SNF5/SNF6 groups.\",\n      \"method\": \"Gene disruption/null mutation, genetic epistasis analysis with suppressors (spt6/ssn20, ssn6), invertase derepression assay\",\n      \"journal\": \"Yeast (Chichester, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — clean KO with defined biochemical phenotype, genetic epistasis in yeast ortholog\",\n      \"pmids\": [\"7785322\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Vps22/EAP30 (SNF8), as an ESCRT-II subunit, is required for lysosomal degradation of ubiquitinated receptors (EGFR and CXCR4); its depletion causes accumulation of EGFR on limiting membranes of early endosomes and aberrantly small multivesicular bodies (MVBs). Notably, depletion of Vps22 — unlike depletion of Hrs or Tsg101 (ESCRT-I) — did not sustain EGF-induced ERK1/2 phosphorylation and nuclear translocation, suggesting EGF signaling termination occurs prior to ESCRT-II engagement.\",\n      \"method\": \"siRNA knockdown in HeLa cells, immunofluorescence/confocal microscopy, EGFR degradation assay, ERK1/2 phosphorylation and nuclear translocation assay\",\n      \"journal\": \"Traffic (Copenhagen, Denmark)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal localization, functional KD with multiple orthogonal readouts (degradation, morphology, signaling), moderate evidence\",\n      \"pmids\": [\"17714434\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"RILP (Rab-interacting lysosomal protein) interacts with VPS22 (SNF8) and VPS36 of ESCRT-II, and this interaction mediates membrane recruitment of ESCRT-II subunits; overexpression of RILP leads to enlarged and clustered MVBs and retards sorting of internalized EGF to degradation.\",\n      \"method\": \"Co-immunoprecipitation, yeast two-hybrid (for domain mapping), overexpression with EEA1/LBPA marker analysis, EGF sorting assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — co-IP and functional overexpression, replicated in two independent labs (PMIDs 17010938 and 16857164)\",\n      \"pmids\": [\"17010938\", \"16857164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"RILP interacts specifically with the N-terminal half of VPS22 (SNF8/EAP30); this interaction was validated by co-immunoprecipitation and colocalization of GFP-RILP and HA-VPS22 in endosomes.\",\n      \"method\": \"Yeast two-hybrid screen (isolation), co-immunoprecipitation, confocal immunofluorescence colocalization\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — yeast two-hybrid plus co-IP plus colocalization, replicated by second lab\",\n      \"pmids\": [\"16857164\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"MCM2 binds directly to EAP30 (SNF8) and competes with ELL for binding to EAP30, thereby potentially modulating the stability of the Holo-ELL complex and regulating the balance between RNA Pol II elongation and initiation activities.\",\n      \"method\": \"Co-immunoprecipitation, competition binding assay\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — single lab, single method (co-IP), no functional transcription readout\",\n      \"pmids\": [\"19819239\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"SNF8 (EAP30/VPS22) physically interacts with the amino-terminal cytoplasmic domain (first 107 amino acids) of TRPC6 channel; overexpression of SNF8 enhances TRPC6-mediated whole-cell currents and NFAT-mediated transcription, while RNAi knockdown of SNF8 partially inhibits NFAT activation, without altering cell-surface TRPC6 levels.\",\n      \"method\": \"Yeast two-hybrid screen (initial discovery), co-immunoprecipitation from mammalian cells, whole-cell patch clamp electrophysiology, NFAT-luciferase reporter assay, surface biotinylation, RNAi knockdown\",\n      \"journal\": \"BMC cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (co-IP, electrophysiology, reporter assay, RNAi), single lab\",\n      \"pmids\": [\"23171048\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"EAP30 (SNF8) localizes in part to the nucleus where it forms a virus-inducible complex with IRF3 and its co-activator CBP; this nuclear complex is required for IRF3 binding to target gene promoters (IFN-β, IFN-λ1, ISG56) and induction of type I/III IFNs and ISGs in response to dsRNA or viral infection, acting downstream of IRF3 phosphorylation/activation and independently of NF-κB-driven inflammatory gene induction.\",\n      \"method\": \"siRNA knockdown with IFN/ISG induction assays, nuclear fractionation, co-immunoprecipitation of EAP30-IRF3-CBP complex, chromatin immunoprecipitation (IRF3 promoter binding), antiviral assays (VSV, HCV)\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods (KD phenotype, nuclear fractionation, co-IP, ChIP) with strong mechanistic resolution, single lab\",\n      \"pmids\": [\"29084253\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In fission yeast, Dot2 (the EAP30/SNF8 homolog) negatively regulates meiotic spindle pole body (SPB) maturation; dot2 mutants accumulate excess electron-dense material near SPBs forming aberrant MTOCs with multipolar spindles. SPB aberrations correlated with elevated Pcp1 (pericentrin ortholog) levels, and reducing pcp1 expression suppressed the dot2 SPB phenotype, placing Dot2 upstream of Pcp1-dependent SPB maturation.\",\n      \"method\": \"dot2 mutant analysis, electron microscopy, genetic epistasis (pcp1 suppression of dot2), immunofluorescence\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis with morphological validation in ortholog, single lab\",\n      \"pmids\": [\"15992541\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In C. elegans, VPS-22/SNF8 acts in the same pathway as DAF-16/FOXO to regulate longevity; knockdown of vps-22/snf8 reduces nuclear localization of DAF-16, and overexpression of daf-16 rescues the short-lived phenotype of vps-22/snf8 knockdown, positioning VPS-22/SNF8 upstream of DAF-16 nuclear activity.\",\n      \"method\": \"RNAi knockdown lifespan assay, daf-16 null genetic epistasis, DAF-16::GFP nuclear localization imaging, downstream gene expression analysis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis with localization readout in C. elegans ortholog, single lab\",\n      \"pmids\": [\"32829877\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"SNF8/EAP30/VPS22 is a multifunctional ESCRT-II subunit that acts at endosomes to sort ubiquitinated receptors (e.g., EGFR, CXCR4) into MVBs for lysosomal degradation — a process regulated by RILP-mediated ESCRT-II membrane recruitment — and also functions in the nucleus as part of the ELL RNA Pol II elongation complex and as an IRF3/CBP co-activator complex required for antiviral IFN gene induction, while additionally modulating TRPC6 channel activity and, in model organisms, regulating centrosome/SPB maturation and FOXO/DAF-16-dependent longevity.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"SNF8 (also known as EAP30/VPS22) is a core subunit of the ESCRT-II complex that functions in endosomal sorting of ubiquitinated membrane receptors and also participates in nuclear transcriptional regulation. As an ESCRT-II component, SNF8 is required for sorting ubiquitinated EGFR and CXCR4 into multivesicular bodies for lysosomal degradation, with its membrane recruitment mediated by direct interaction of its N-terminal region with RILP [PMID:17714434, PMID:16857164]. In the nucleus, SNF8 serves dual transcriptional roles: it associates with the ELL elongation complex to derepress RNA Pol II elongation activity [PMID:10419521], and it forms a virus-inducible complex with IRF3 and CBP that is essential for IRF3 binding to IFN-β/IFN-λ1 promoters and antiviral interferon gene induction [PMID:29084253]. SNF8 additionally modulates TRPC6 channel currents and downstream NFAT activation through direct interaction with the TRPC6 cytoplasmic domain, independent of effects on channel surface expression [PMID:23171048].\",\n  \"teleology\": [\n    {\n      \"year\": 1995,\n      \"claim\": \"Establishing SNF8 as a transcriptional regulator: genetic analysis in yeast showed that SNF8 is required for glucose-regulated SUC2 gene derepression and is functionally related to SNF7 but genetically distinct from SNF1/SNF4 and SWI/SNF groups, defining it as part of a separate chromatin/transcription regulatory pathway.\",\n      \"evidence\": \"Gene disruption, genetic epistasis, and invertase derepression assays in S. cerevisiae\",\n      \"pmids\": [\"7785322\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Molecular mechanism of SNF8-dependent transcriptional derepression in yeast was not defined\",\n        \"Whether SNF8 functions as part of a stable complex in yeast was unknown\",\n        \"Relationship to endosomal sorting function not yet recognized\"\n      ]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Identifying a direct transcriptional mechanism: EAP30/SNF8 was found as a subunit of the mammalian ELL elongation complex, where it directly binds ELL and derepresses ELL's inhibitory effect on RNA Pol II transcription, providing the first biochemical activity for SNF8 in metazoans.\",\n      \"evidence\": \"Co-purification of the ELL complex and in vitro transcription assay measuring RNA Pol II derepression\",\n      \"pmids\": [\"10419521\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"In vivo relevance of ELL complex derepression not demonstrated\",\n        \"Whether EAP30 has functions independent of ELL was unknown\",\n        \"Target gene specificity of ELL-EAP30 complex not defined\"\n      ]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Revealing a role in centrosome/SPB maturation: the fission yeast SNF8 ortholog Dot2 negatively regulates spindle pole body maturation, with dot2 mutants showing excess MTOC material and multipolar spindles that are suppressed by reducing Pcp1/pericentrin, placing SNF8 upstream of centrosome component homeostasis.\",\n      \"evidence\": \"Mutant analysis with electron microscopy and genetic epistasis (pcp1 suppression) in S. pombe\",\n      \"pmids\": [\"15992541\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Whether this reflects an ESCRT-II-dependent or ESCRT-II-independent function was not determined\",\n        \"Mechanism by which Dot2 regulates Pcp1 levels was not identified\",\n        \"Conservation of this SPB role in mammalian cells untested\"\n      ]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Defining the membrane recruitment mechanism: RILP was shown to interact directly with the N-terminal half of VPS22/SNF8 and to recruit ESCRT-II to endosomal membranes, connecting Rab7-RILP signaling to ESCRT-II-mediated receptor sorting.\",\n      \"evidence\": \"Yeast two-hybrid, co-immunoprecipitation, confocal colocalization, and EGF sorting assays in mammalian cells; confirmed by two independent labs\",\n      \"pmids\": [\"17010938\", \"16857164\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Whether RILP is the sole recruitment factor for ESCRT-II was not resolved\",\n        \"Structural basis of the RILP–VPS22 interaction not determined\",\n        \"Relative contribution of RILP versus lipid-binding (VPS36) to ESCRT-II membrane association unclear\"\n      ]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Establishing the core endosomal sorting function: siRNA depletion of VPS22/SNF8 in HeLa cells demonstrated that ESCRT-II is required for lysosomal degradation of EGFR and CXCR4 and for normal MVB biogenesis, while showing that EGF-induced ERK signaling is terminated prior to ESCRT-II engagement — distinguishing ESCRT-II's role from upstream ESCRT-0/I in signal attenuation.\",\n      \"evidence\": \"siRNA knockdown with EGFR degradation kinetics, confocal microscopy of MVB morphology, and ERK1/2 phosphorylation/translocation assays in HeLa cells\",\n      \"pmids\": [\"17714434\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Range of cargo receptors requiring ESCRT-II beyond EGFR/CXCR4 not systematically defined\",\n        \"Whether ESCRT-II has cargo-selective versus general roles in MVB formation not resolved\"\n      ]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Suggesting competition for SNF8 between transcription and replication complexes: MCM2 was found to bind EAP30/SNF8 and compete with ELL for the same binding site, raising the possibility that SNF8 coordinates elongation and initiation activities.\",\n      \"evidence\": \"Co-immunoprecipitation and competition binding assay\",\n      \"pmids\": [\"19819239\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"Single-method co-IP without functional transcription or replication readout\",\n        \"Physiological context in which MCM2-EAP30 interaction is relevant not established\",\n        \"No independent confirmation of the competition model\"\n      ]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Uncovering a non-ESCRT channel-regulatory function: SNF8 was found to bind the TRPC6 cytoplasmic domain and enhance TRPC6-mediated currents and downstream NFAT transcriptional activation without altering channel surface expression, identifying SNF8 as a modulator of ion channel gating.\",\n      \"evidence\": \"Yeast two-hybrid, co-immunoprecipitation, whole-cell patch clamp electrophysiology, NFAT-luciferase reporter, surface biotinylation, and RNAi in mammalian cells\",\n      \"pmids\": [\"23171048\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Mechanism by which SNF8 enhances TRPC6 currents (gating vs. conductance) not resolved\",\n        \"Whether this is an ESCRT-II-dependent or SNF8-autonomous function unknown\",\n        \"In vivo physiological relevance (e.g., in podocytes) not tested\"\n      ]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Establishing a nuclear role in innate antiviral immunity: SNF8 was shown to form a virus-inducible nuclear complex with IRF3 and CBP that is required for IRF3 binding to IFN-β/IFN-λ1/ISG56 promoters and for type I/III interferon induction, acting downstream of IRF3 phosphorylation and independently of NF-κB signaling.\",\n      \"evidence\": \"siRNA knockdown with IFN/ISG induction assays, nuclear fractionation, co-immunoprecipitation of EAP30–IRF3–CBP, ChIP for IRF3 at promoters, and antiviral assays (VSV, HCV)\",\n      \"pmids\": [\"29084253\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether SNF8 recruits chromatin remodeling or additional coactivators to IRF3 targets not determined\",\n        \"Structural basis of the IRF3–EAP30–CBP ternary complex unknown\",\n        \"Whether ESCRT-II partners VPS25/VPS36 participate in the nuclear antiviral function not tested\"\n      ]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Linking ESCRT-II to longevity regulation: in C. elegans, VPS-22/SNF8 promotes nuclear localization of DAF-16/FOXO and acts in the same genetic pathway, with DAF-16 overexpression rescuing the short-lived phenotype of vps-22 knockdown, positioning SNF8 upstream of FOXO-dependent longevity.\",\n      \"evidence\": \"RNAi lifespan assay, daf-16 epistasis, DAF-16::GFP nuclear localization, and target gene expression in C. elegans\",\n      \"pmids\": [\"32829877\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Whether the effect on DAF-16 localization is direct or via endosomal signaling regulation not resolved\",\n        \"Conservation in mammalian aging pathways not tested\",\n        \"Which endosomal cargo drives the longevity phenotype is unknown\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How SNF8 partitions between its ESCRT-II endosomal function and its nuclear transcriptional/antiviral roles remains unresolved, as does whether these represent independent or coordinated activities.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"No regulatory mechanism identified that switches SNF8 between endosomal and nuclear pools\",\n        \"No structural model of full-length human ESCRT-II with SNF8 in a membrane-associated state\",\n        \"Systematic identification of SNF8-dependent cargo in mammalian cells not performed\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 6, 7]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [0, 7]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005768\", \"supporting_discovery_ids\": [2, 3, 4]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [2, 3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [7]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [0, 7]}\n    ],\n    \"complexes\": [\n      \"ESCRT-II\",\n      \"ELL elongation complex\",\n      \"IRF3-CBP-EAP30 complex\"\n    ],\n    \"partners\": [\n      \"VPS36\",\n      \"VPS25\",\n      \"ELL\",\n      \"RILP\",\n      \"IRF3\",\n      \"CREBBP\",\n      \"TRPC6\",\n      \"MCM2\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}