{"gene":"INO80","run_date":"2026-06-10T01:55:23","timeline":{"discoveries":[{"year":2004,"finding":"The yeast INO80 complex is recruited to HO endonuclease-induced DNA double-strand breaks (DSBs) through interaction with phosphorylated histone H2A (γ-H2AX) at S129; recruitment requires the Nhp10 HMG-like subunit. Loss of INO80 (arp8 or H2A S129 mutants) impairs conversion of the DSB into ssDNA, implicating INO80-mediated chromatin remodeling in DSB end-processing.","method":"Chromatin immunoprecipitation (ChIP) at HO-induced DSB; genetic epistasis with H2A S129 mutants and nhp10 deletions; sensitivity assays to DNA damaging agents","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal ChIP, genetic epistasis, replicated independently by two labs in the same issue (PMIDs 15607975 and 15607974)","pmids":["15607975","15607974"],"is_preprint":false},{"year":2004,"finding":"The INO80 complex is recruited to DSBs via a specific interaction between the Nhp10 subunit and γ-H2AX; loss of Nhp10 or γ-H2AX reduces INO80 recruitment. INO80 components show synthetic genetic interactions with the RAD52 DSB repair pathway.","method":"ChIP at HO-induced DSB; Nhp10 deletion and γ-H2AX mutant analysis; genetic interaction screen with RAD52 pathway","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal ChIP, genetic epistasis, confirmed by independent lab (PMID 15607975) in same issue","pmids":["15607974"],"is_preprint":false},{"year":2005,"finding":"Human INO80 (hINO80) complex was identified and purified, containing orthologs of 8 of 15 yeast INO80 subunits plus at least five metazoan-specific subunits. The complex exhibits DNA- and nucleosome-activated ATPase activity and catalyzes ATP-dependent nucleosome sliding.","method":"Affinity purification / mass spectrometry; ATPase activity assay; nucleosome sliding assay in vitro","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of ATPase and nucleosome sliding activity with purified complex; single lab with two orthogonal biochemical assays","pmids":["16230350"],"is_preprint":false},{"year":2006,"finding":"Ino80 is required for cell cycle checkpoint adaptation in response to a persistent DSB. Cells lacking Ino80 fail to maintain high γ-H2AX levels and show increased H2A.Z (Htz1) incorporation near DSBs via Swr1. Inactivation of Swr1 restores H2AX phosphorylation and checkpoint adaptation in ino80 mutants, revealing antagonism between Ino80 and Swr1 at DSB-flanking chromatin.","method":"ChIP; genetic epistasis (ino80 × swr1 double mutants); checkpoint adaptation assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with double mutants, ChIP, and functional checkpoint assays; single lab but multiple orthogonal approaches","pmids":["16951256"],"is_preprint":false},{"year":2007,"finding":"Swr1 is recruited to DSBs in a γ-H2AX-dependent manner. INO80 (but not SWR1) mediates removal of H2A.Z, γ-H2AX, and core histones near DSBs. Loss of INO80-specific subunits Arp8 or Nhp10 impairs Mre11, yKu80, and Mec1 binding at DSBs, causing defective end-processing and checkpoint activation.","method":"ChIP at DSBs; genetic analysis of INO80- vs SWR1-specific subunit mutants; immunofluorescence of repair factors","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP with multiple subunit mutants, functional repair/checkpoint assays, replicated across multiple studies","pmids":["17762868"],"is_preprint":false},{"year":2007,"finding":"YY1 is tightly associated with the human INO80 complex. YY1 recruits INO80 to YY1-activated gene promoters where INO80 functions as a coactivator. YY1 binding to its DNA target sites requires INO80 activity.","method":"Co-immunoprecipitation; ChIP at YY1 target genes; RNAi knockdown of INO80 with transcriptional readout","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, ChIP, and functional knockdown experiments; replicated by independent study (PMID 18026119)","pmids":["17721549"],"is_preprint":false},{"year":2007,"finding":"YY1 forms a complex with INO80 subunits in mammalian cells. Both YY1 and INO80 are essential for homologous recombination-based DNA repair (HRR); YY1 preferentially binds recombination-intermediate structures in vitro. RNAi knockdown of either increases cellular sensitivity to DNA-damaging agents.","method":"Co-immunoprecipitation; RNAi knockdown; HR functional assays; in vitro DNA-binding assays with recombination intermediates","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — Co-IP, in vitro binding, functional HR assay, knockdown phenotype; single lab with multiple orthogonal methods","pmids":["18026119"],"is_preprint":false},{"year":2007,"finding":"The Ies3 subunit of yeast INO80 interacts with the tetratricopeptide repeat domain of telomerase subunit Est1p. Deletion of IES3 causes telomere elongation, altered telomere position effect, delayed recombinational survivor formation, and stimulated extrachromosomal telomeric circles. Multiple INO80 subunits preferentially localize to telomeres.","method":"Co-immunoprecipitation; telomere length assays; ChIP at telomeres; genetic analysis of ies3Δ and arp8Δ in telomerase-negative strains","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP of Ies3-Est1, ChIP, genetic epistasis; single lab","pmids":["17562861"],"is_preprint":false},{"year":2007,"finding":"The Mec1/Tel1 kinases (ATM/ATR orthologs) phosphorylate the Ies4 subunit of the INO80 complex during DNA damage. Mutation of Ies4 phosphorylation sites does not significantly affect DNA repair but influences DNA damage checkpoint responses, functionally linking INO80 to the Mec1/Tel1 checkpoint signaling pathway via Tof1 and Rad53.","method":"In vivo phosphorylation assays; site-directed mutagenesis of Ies4 phosphosites; checkpoint assays; genetic interaction analysis with tof1 and rad53","journal":"Cell","confidence":"High","confidence_rationale":"Tier 1 / Moderate — phosphosite mutagenesis with defined checkpoint phenotype, genetic epistasis; single lab with multiple orthogonal approaches","pmids":["17693258"],"is_preprint":false},{"year":2008,"finding":"The Ino80 chromatin remodeling complex binds replication origins and stalled replication forks. Under hydroxyurea-induced fork arrest, INO80 accumulates at stalled forks and unfired origins. ino80 mutants are defective in replication restart after HU release and accumulate DSBs.","method":"ChIP across four yeast chromosomes; genome-wide mapping; HU treatment and release assays; DSB detection","journal":"Current biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP mapping plus functional replication restart assay; replicated by Peterson lab (PMID 18376411)","pmids":["18406137"],"is_preprint":false},{"year":2008,"finding":"The Ino80 enzyme is recruited to replication origins as cells enter S phase and is required continuously for efficient fork progression, especially under low replication stress. Inducible degradation of Ino80 causes replisome dissociation from stalled forks, indicating Ino80 stabilizes the stalled replisome to ensure proper restart.","method":"Inducible protein degradation; ChIP at replication origins; replisome component co-precipitation; replication fork stability assays","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — inducible depletion with mechanistic replisome dissociation readout, ChIP; single lab with multiple orthogonal methods","pmids":["18376411"],"is_preprint":false},{"year":2008,"finding":"In the nucleus, the deubiquitinase Uch37 associates with the human INO80 complex where it is held in an inactive state. Uch37 can be activated by transient interaction of hINO80 with the proteasome, suggesting cooperative regulation of transcription or DNA repair.","method":"Co-immunoprecipitation; DUB activity assays in vitro; mass spectrometry identification of hINO80-Uch37 interaction","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and in vitro activity assay showing Uch37 is inactive in hINO80 and activated by proteasome; single lab","pmids":["18922472"],"is_preprint":false},{"year":2010,"finding":"INO80 acts as a nucleosome spacing factor in vitro. It requires a minimum of 33–43 bp of extranucleosomal DNA and moves nucleosomes to the center of DNA with high precision. Unlike ISW2/1a, INO80 does not require H4 tails; instead the H2A histone tail negatively regulates nucleosome movement. INO80 spaces arrays with ~30 bp final linker DNA.","method":"In vitro nucleosome sliding/spacing assays with defined mono- and dinucleosomal arrays; histone tail deletion analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstituted nucleosome sliding/spacing assays with systematic DNA length and histone tail mutant analysis; single lab with rigorous biochemistry","pmids":["21135121"],"is_preprint":false},{"year":2010,"finding":"Human INO80 promotes 5′→3′ resection of DSB ends in mammalian cells. Ino80 depletion impaired focal recruitment of 53BP1 but did not impede Rad51 focus formation, placing Ino80 at early steps of DSB repair. Ino80 associates with chromatin surrounding DSBs.","method":"RNAi knockdown; comet assay; HR repair reporter; immunofluorescence of 53BP1 and Rad51 foci; BrdU-ssDNA and RPA staining","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple readouts (comet assay, HR reporter, immunofluorescence, ssDNA detection); single lab","pmids":["21947284"],"is_preprint":false},{"year":2010,"finding":"Deletion of INO80 or ARP5 in mammalian cells impairs removal of UV-induced photo lesions via NER without affecting NER factor transcription. INO80 and Arp5 are recruited to UV-damaged DNA before NER incision occurs and are required for assembly of NER factors, indicating INO80 promotes chromatin accessibility for NER.","method":"Genetic knockout models; UV photolesion removal assays; ChIP for NER factors; chromatin accessibility assays","journal":"Proceedings of the National Academy of Sciences","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic knockouts with mechanistic NER factor assembly readout; two subunit deletions (INO80 and ARP5) corroborating; single lab","pmids":["20855601"],"is_preprint":false},{"year":2010,"finding":"Mammalian INO80 complex is recruited to laser-induced DNA damage sites in an ARP8-dependent manner but independently of γ-H2AX, unlike the yeast complex where Nhp10 or Arp4 mediate recruitment.","method":"Laser micro-irradiation; immunofluorescence; siRNA knockdown of ARP8; live-cell imaging","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct localization experiment with ARP8 knockdown; single lab, single method","pmids":["20971067"],"is_preprint":false},{"year":2011,"finding":"Yeast INO80 has a histone exchange activity: it can replace nucleosomal H2A.Z/H2B dimers with free H2A/H2B dimers. In the absence of INO80, H2A.Z nucleosomes are mislocalized genome-wide and fail to respond dynamically to transcriptional changes. Genetic interactions between ino80 and htz1 (H2A.Z) support a model where INO80 removes unacetylated H2A.Z to promote genome stability.","method":"Genome-wide ChIP-seq for H2A.Z; in vitro histone exchange assays; genetic interaction analysis (ino80 × htz1)","journal":"Cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro histone exchange reconstitution plus genome-wide mapping and genetic epistasis; single lab with multiple orthogonal methods and comprehensive genome-wide data","pmids":["21241891"],"is_preprint":false},{"year":2011,"finding":"The human INO80 complex is organized in three modules assembling on distinct domains of hIno80 ATPase: (i) an N-terminal module with metazoan-specific subunits dispensable for remodeling, (ii) a module containing Arp4, Arp8, and YY1 on the HSA/PTH domain, and (iii) a catalytic core module with Ies2, Ies6, Arp5, Tip49a, and Tip49b. ATP-dependent nucleosome remodeling requires the evolutionarily conserved core comprising HSA/PTH and Snf2 ATPase domains together with YY1 and conserved subunits.","method":"Affinity purification of hINO80 subassemblies; mass spectrometry; in vitro nucleosome remodeling assays with subassemblies","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — reconstituted subassemblies with defined nucleosome remodeling activity; systematic dissection of three modules; single lab","pmids":["21303910"],"is_preprint":false},{"year":2011,"finding":"Loss of INO80 subunits Ies6 or Ino80 causes rapid polyploidy and chromosome missegregation. Chromatin structure flanking centromeres is altered in these mutants, not in the Cse4-containing centromeric nucleosome itself, but in pericentric chromatin. These effects are mediated through misincorporation of H2A.Z into pericentric regions.","method":"Flow cytometry (ploidy); live-cell imaging of chromosome segregation; ChIP for H2A.Z and histones at centromeres; genetic analysis","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple readouts (ploidy, segregation, ChIP), mechanistic link to H2A.Z misincorporation; single lab with orthogonal approaches","pmids":["23207916"],"is_preprint":false},{"year":2012,"finding":"Targeted recruitment of INO80 to a chromosomal locus enhances large-scale chromatin mobility in budding yeast, requiring Ino80's ATPase activity. This enhanced mobility correlates with increased rates of spontaneous gene conversion, indicating that INO80-dependent nucleosome remodeling promotes chromatin flexibility and homologous recombination.","method":"High-precision live fluorescence microscopy of tagged chromosomal loci; targeted tethering of INO80; ATPase-dead mutant analysis; gene conversion assays","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Moderate — live-cell imaging with targeted tethering and ATPase mutant controls, plus functional HR assay; single lab with multiple orthogonal methods","pmids":["22345518"],"is_preprint":false},{"year":2012,"finding":"Crystal structure of the Arp8 C-terminal domain shows three insertions within the actin fold. Arp8 forms a dimer that binds the nucleosome core primarily through H3 and H4 histones via one of these insertions, exploiting the twofold symmetry of the nucleosome.","method":"X-ray crystallography; biochemical binding assays (pull-down, EM); nucleosome co-purification with recombinant Arp8","journal":"Proceedings of the National Academy of Sciences","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure plus biochemical validation of histone interactions; single lab with structure and binding assays","pmids":["23213201"],"is_preprint":false},{"year":2013,"finding":"Nuclear actin in the INO80 complex exists as a monomer with its barbed end inaccessible for polymerization. An actin mutation in subdomain 2 reduces INO80 chromatin remodeling activity in vitro and disrupts in vivo nuclear functions. The pointed end subdomain 2 of actin contributes to INO80-chromatin interactions.","method":"Biochemical fractionation; actin polymerization assays; actin subdomain mutagenesis; in vitro chromatin remodeling assays; genetic complementation","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution plus mutagenesis with defined chromatin remodeling readout; single lab with multiple orthogonal methods","pmids":["23524535"],"is_preprint":false},{"year":2013,"finding":"The Ies2 subunit potently activates the intrinsic catalytic ATPase activity of the human Ino80 SNF2 ATPase, while Ies6 and Arp5 together promote binding of the Ino80 ATPase to nucleosomes. Thus the Ino80 ATPase is regulated at multiple levels within the complex.","method":"Purified subunit reconstitution; in vitro ATPase assays; nucleosome-binding assays with defined subunit deletions","journal":"Proceedings of the National Academy of Sciences","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with defined subunit deletions and ATPase/nucleosome binding assays; single lab","pmids":["24297934"],"is_preprint":false},{"year":2014,"finding":"INO80 binds replication forks in human cells and promotes fork progression under normal conditions. Ino80 is recruited to replication forks through interaction with ubiquitinated H2A, aided by BAP1 (a nuclear deubiquitinase that also stabilizes Ino80 protein). BAP1 deficiency in cancer cells downregulates Ino80 by disrupting this stabilization mechanism.","method":"Co-immunoprecipitation of Ino80 with ubiquitinated H2A; ChIP at replication forks; BAP1 knockdown/knockout; Ino80 protein stability assays; mouse embryo Ino80 deletion","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — Co-IP, ChIP at replication forks, genetic knockouts with defined replication phenotype; single lab with multiple orthogonal methods","pmids":["25283999"],"is_preprint":false},{"year":2014,"finding":"INO80 occupies pluripotency gene promoters with master transcription factors OCT4 and WDR5. Ino80 maintains open chromatin architecture at these promoters and licenses Mediator and RNA Pol II recruitment. Ino80 is required for ESC self-renewal, somatic cell reprogramming, and blastocyst development.","method":"ChIP-seq; ATAC-seq; RNAi knockdown; genetic knockout in mouse; reprogramming assays","journal":"Cell stem cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — genome-wide ChIP-seq, chromatin accessibility, and functional knockout; mechanistic pathway placement; single lab with comprehensive approaches","pmids":["24792115"],"is_preprint":false},{"year":2014,"finding":"INO80-dependent Mps3 (inner nuclear membrane SUN domain protein) binding to DSBs is S/G2-phase specific and requires both INO80 and Rad51. DSB relocation to Nup84 nuclear pore occurs independently of INO80 and cell-cycle phase. Thus INO80 determines choice of perinuclear anchorage site for DSBs.","method":"Live-cell imaging of DSB subnuclear position; genetic epistasis with ino80, rad51, swr1 mutants; cell-cycle synchronization","journal":"Molecular cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — live-cell localization with multiple genetic controls; single lab","pmids":["25066231"],"is_preprint":false},{"year":2015,"finding":"Mammalian INO80 rapidly removes H2A.Z from chromatin flanking DNA damage. Depletion of INO80 or histone chaperone ANP32E impairs homologous recombination, and the HR defect can be rescued by co-depletion of H2A.Z, demonstrating that H2A.Z removal is the primary function of INO80 and ANP32E in promoting HR.","method":"RNAi knockdown; H2A.Z ChIP after DNA damage; HR functional assay; epistasis by co-depletion of H2A.Z","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 / Moderate — epistasis (H2A.Z co-depletion rescues HR defect), ChIP, functional HR assay; single lab with mechanistically informative rescue experiment","pmids":["26142279"],"is_preprint":false},{"year":2015,"finding":"The yeast Ino80 chromatin remodeling complex is required for RNAPII turnover under transcriptional stress. INO80 forms a ternary complex with RNAPII and Cdc48/p97, and cells lacking INO80 accumulate ubiquitinated Rpb1 tightly bound to chromatin, indicating INO80 nucleosome remodeling activity promotes dissociation of ubiquitinated RNAPII from chromatin for proteasomal degradation.","method":"Co-immunoprecipitation; in vivo Rpb1 ubiquitination/degradation assays; ChIP for ubiquitinated Rpb1; genetic epistasis with cdc48 and proteasome mutants","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Moderate — Co-IP of ternary complex, ChIP, genetic epistasis; single lab with multiple orthogonal approaches","pmids":["26656161"],"is_preprint":false},{"year":2015,"finding":"The Arp8/Arp4/actin module in INO80 binds extranucleosomal DNA 37-51 bp from the nucleosome edge and functions as a DNA-length sensor that regulates nucleosome sliding. Disruption of Arp8/Arp4 DNA binding uncouples ATP hydrolysis from nucleosome mobilization by disengaging Arp5 from the H2A-H2B acidic patch.","method":"In vitro DNA-binding assays (EMSA); site-directed mutagenesis of Arp8/Arp4; ATPase assays; nucleosome sliding assays; photo-crosslinking","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — mutagenesis, in vitro binding, ATPase, and sliding assays; mechanistic coupling established; single lab","pmids":["30120252"],"is_preprint":false},{"year":2015,"finding":"EM structural analysis shows INO80-C and SWR-C share similar overall architectures with a compact head containing Rvb1/Rvb2 as single heterohexameric rings. The Arp8/Arp4/Act1 module enhances nucleosome-binding affinity but is largely dispensable for remodeling. The Ies6/Arp5 module is essential for remodeling activity and controls conformational changes coupling nucleosome binding to remodeling.","method":"Electron microscopy; 2D class averaging; mass spectrometry; nucleosome remodeling assays with module deletions","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — EM structure plus functional module deletion assays; single lab with structural and biochemical validation","pmids":["25964121"],"is_preprint":false},{"year":2016,"finding":"The Arp5-Ies6 subcomplex forms an abundant distinct subcomplex in vivo and stimulates INO80-mediated ATPase and nucleosome sliding activity in vitro. Ies2 is required for Arp5-Ies6 association with the catalytic INO80 components. Mutant Arp5 lacking unique insertion domains allows ATP hydrolysis without nucleosome sliding, uncoupling the two activities.","method":"In vivo co-immunoprecipitation; in vitro ATPase and nucleosome sliding assays; domain deletion/mutagenesis of Arp5","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with mutagenesis defining Arp5 insertion domains in coupling ATP hydrolysis to sliding; single lab","pmids":["26306040","27255055"],"is_preprint":false},{"year":2016,"finding":"Mec1, INO80, and PAF1 complexes cooperate to remove RNAPII from transcribed genes near early-firing replication origins upon HU treatment. Mec1 triggers efficient removal of PAF1C and RNAPII. Failure to evict RNAPII correlates with defective replication fork restart, implicating INO80 in preventing transcription-replication conflicts.","method":"ChIP-seq for RNAPII and PAF1C; genetic epistasis (mec1, ino80, paf1 mutants); DNA fiber assays for replication restart; proteomic analysis","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP-seq, fiber assays, genetic epistasis across multiple mutants; single lab with genome-wide and functional approaches","pmids":["26798134"],"is_preprint":false},{"year":2016,"finding":"INO80 occupies >90% of superenhancers in melanoma cells, dependent on transcription factors MITF and Sox9. Ino80 binding reduces nucleosome occupancy and facilitates Mediator recruitment, promoting oncogenic transcription. Ino80 silencing selectively inhibits melanoma cell proliferation and tumorigenesis.","method":"ChIP-seq; ATAC-seq; Ino80 siRNA knockdown; mouse xenograft assays; Co-IP with Mediator","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genome-wide ChIP-seq, chromatin accessibility, in vivo tumorigenesis; mechanistic link to Mediator recruitment; single lab","pmids":["27340176"],"is_preprint":false},{"year":2017,"finding":"INO80 translocates along DNA at the H2A-H2B interface of nucleosomes (unlike other remodelers that translocate at the H3-H4 interface), creating DNA torsional strain near the nucleosome entry site. This mechanism promotes both nucleosome mobilization and selective exchange of H2A.Z-H2B dimers with H2A-H2B without additional histone chaperones. INO80 mobilizes H2A.Z-containing nucleosomes more efficiently than H2A nucleosomes.","method":"Site-directed photo-crosslinking; ATPase assays; nucleosome sliding assays with H2A vs H2A.Z substrates; histone exchange assays in vitro","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with photo-crosslinking defining translocation site plus histone exchange assays; single lab with mechanistically rigorous biochemistry","pmids":["28604691"],"is_preprint":false},{"year":2017,"finding":"The INO80 complex removes H2A.Z to promote presynaptic filament formation during homologous recombination in budding yeast. INO80 has at least two distinct functions in HR: DNA end resection and presynaptic filament formation. H2A.Z deletion rescues presynaptic filament formation and HR in INO80-deficient mutants.","method":"High-resolution ChIP; HR assays; genetic epistasis with H2A.Z deletion (htz1Δ); direct visualization of presynaptic filament formation","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Moderate — epistasis (htz1Δ rescues INO80-C HR defect), mechanistic dissection; single lab with multiple orthogonal assays","pmids":["28514650"],"is_preprint":false},{"year":2017,"finding":"An insertion domain (Ino80INS) in the Ino80 ATPase stimulates Rvb1/Rvb2 ATPase activity 16-fold and promotes their dodecamerization. Rvb1/Rvb2 function as protein assembly chaperones within INO80, cycling between hexamers and dodecamers in an ATP-dependent manner.","method":"ATPase activity assays; mass spectrometry; cryo-EM and integrative modeling; biochemical reconstitution","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure, in vitro ATPase stimulation assay, and mass spectrometry; single lab with multiple orthogonal structural and biochemical approaches","pmids":["28591576"],"is_preprint":false},{"year":2018,"finding":"Cryo-EM structure of evolutionarily conserved INO80 core from Chaetomium thermophilum bound to nucleosome at 4.3/3.7 Å resolution. The Rvb1/Rvb2 AAA+ heterohexamer acts as a stator scaffold. The Swi2/Snf2 ATPase motor binds at SHL-6, unwraps ~15 bp, disrupts H2A-DNA contacts, and is poised to pump entry DNA into the nucleosome. Arp5 and Ies6 bind at SHL-2/-3 as a counter-grip, with the Arp5 grappler element binding the nucleosome dyad. The structure suggests a ratchet mechanism for both nucleosome sliding and histone editing.","method":"Cryo-electron microscopy (global 4.3 Å, major parts 3.7 Å); biochemical validation of ATPase and sliding activities","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — near-atomic resolution cryo-EM structure with biochemical validation; replicated independently by human INO80 structure (PMID 29643506)","pmids":["29643509"],"is_preprint":false},{"year":2018,"finding":"Cryo-EM structure of human INO80 bound to nucleosome reveals the motor domains are located at the DNA entry point (not at SHL2 as in other remodelers). ARP5-IES6 module makes additional contacts on the opposite side. Histone H3 tails regulate the INO80 motor domain (unlike other remodelers where H4 tails play this role).","method":"Cryo-electron microscopy (9.6 Å overall, 4.1 Å for portions); biochemical validation","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure of human complex with biochemical validation; replicated by fungal INO80 structure (PMID 29643509) revealing conserved yet distinct features","pmids":["29643506"],"is_preprint":false},{"year":2018,"finding":"Cryo-EM structures of the active core of human INO80 at 9.6 Å reveal an unusual spoked-wheel structural domain of Ino80 subunit engulfed by a single RUVBL1/RUVBL2 AAA+ heterohexamer. RUVBL1/RUVBL2 form a major interaction site for partner proteins that likely communicate to nucleotide-binding sites.","method":"Cryo-EM; subunit reconstitution; EM class averaging","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with biochemical subunit analysis; single lab","pmids":["29323271"],"is_preprint":false},{"year":2018,"finding":"Crystal structure of the 180-kDa Arp8 module of yeast INO80 shows Arp8 engages nuclear actin in a manner distinct from other actin-fold proteins, recruiting the Arp4-N-actin heterodimer to a segmented scaffold of the helical HSA domain. The HSA domain spans >120 Å and provides an extended binding platform for extranucleosomal entry DNA required for nucleosome sliding and genome-wide nucleosome positioning.","method":"X-ray crystallography; biochemical DNA-binding assays; in vitro nucleosome sliding assays; genome-wide nucleosome mapping","journal":"Nature structural & molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus in vitro and in vivo functional validation; corroborated by independent biochemical study (PMID 30120252)","pmids":["30177756"],"is_preprint":false},{"year":2018,"finding":"INO80 operates as a DNA length-sensitive switch: nucleosome sliding rate increases ~100-fold when flanking DNA increases from 40 to 60 bp. Once initiated, INO80 moves nucleosomes rapidly at least 20 bp without pausing. The Nhp10 module plays an auto-inhibitory role tuning this switch-like response. INO80 can change direction of sliding without dissociation.","method":"Single-molecule enzymology; ensemble ATPase and sliding assays; Nhp10 module deletion analysis","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Moderate — single-molecule and ensemble biochemistry with defined kinetic parameters and regulatory module deletion; single lab with rigorous quantitative assays","pmids":["29452642"],"is_preprint":false},{"year":2018,"finding":"INO80 deletion in vascular endothelial cells prevents ventricular compaction in the developing mouse heart, correlating with defective coronary vascularization. In vitro, endothelial cells promote myocardial expansion in an Ino80-dependent manner. Ino80 deletion increases E2F-activated gene expression and endothelial S-phase occupancy.","method":"Conditional endothelial Ino80 knockout (mouse); histological analysis; in vitro co-culture assays; gene expression profiling","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional knockout with mechanistic readout (E2F gene expression, cell cycle); single lab","pmids":["29371594"],"is_preprint":false},{"year":2019,"finding":"TRIM3 E3 ubiquitin ligase mediates degradation of INO80 in the nucleus accumbens; TRIM3 and INO80 interact directly, and reduced TRIM3 on abstinence day 30 leads to increased INO80 protein levels. INO80-mediated transcriptional changes regulate cocaine craving during prolonged abstinence.","method":"Co-immunoprecipitation; viral gene transfer; ChIP-seq; ubiquitin degradation assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP showing TRIM3-INO80 interaction, functional viral gene transfer; single lab","pmids":["31633032"],"is_preprint":false},{"year":2019,"finding":"BAP1 depletion reduces DNA synthesis and impairs restart of HU-induced stalled replication forks. This defect is rescued by ectopic INO80 expression. BAP1 depletion abrogates INO80 binding to stalled replication forks, indicating BAP1 promotes replication stress recovery by recruiting INO80 to stalled forks.","method":"DNA fiber assays; ChIP at replication forks; siRNA knockdown; ectopic INO80 expression rescue","journal":"The Biochemical journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis rescue experiment, ChIP, DNA fiber assays; single lab","pmids":["31657441"],"is_preprint":false},{"year":2020,"finding":"INO80 complex promotes resolution of R-loops to prevent replication-associated DNA damage in cancer cells. R-loops promote INO80 recruitment to chromatin. Overexpression of RNase H1 rescues DNA synthesis defects and suppresses DNA damage caused by INO80 depletion. Artificial tethering of INO80 to a LacO locus enables R-loop turnover in cis.","method":"RNAi depletion of INO80; R-loop immunofluorescence (S9.6 antibody); DNA fiber assays; RNase H1 rescue; LacO artificial tethering","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple readouts including RNase H1 rescue and artificial tethering establishing direct R-loop resolution; single lab with orthogonal methods","pmids":["32913330"],"is_preprint":false},{"year":2020,"finding":"Ino80 conditional deletion from cortical neural progenitor cells impairs DNA DSB repair selectively via homologous recombination, causing p53-dependent apoptosis and microcephaly. Ino80 function in HR is mechanistically distinct from its role in YY1-associated transcription. Sensitivity is dependent on NPC division mode: symmetric NPC-NPC divisions but not asymmetric neurogenic divisions require Ino80-mediated HR.","method":"Conditional knockout mouse; in vivo DSB repair pathway assay; apoptosis markers; phenotype comparison with Brca2 conditional knockout","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional knockout with in vivo HR assay and genetic epistasis with Brca2; mechanistic dissociation of HR and transcription functions; replicated concept with Brca2","pmids":["32737294"],"is_preprint":false},{"year":2020,"finding":"In fission yeast, INO80 subunit Iec5 promotes histone turnover at heterochromatin, enabling INO80 to counter epigenetic inheritance of heterochromatin. Mutations in INO80 components allow pericentric heterochromatin inheritance in RNAi mutants. This function is distinct from nucleosome positioning at heterochromatin.","method":"Genetic screen; heterochromatin inheritance assays; histone turnover measurements; ChIP","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic screen plus histone turnover assays with subunit-specific (Iec5) dissection; single lab","pmids":["33378674"],"is_preprint":false},{"year":2021,"finding":"INO80 processes DNA shape/mechanical properties encoded in the genome through allosteric interplay between its core and Arp8 modules to position nucleosomes genome-wide. At promoters, INO80 integrates DNA mechanics readout with general regulatory factor binding to position the +1 nucleosome. This establishes a molecular mechanism for robust, adjustable +1 nucleosome positioning.","method":"Genome-wide chromatin reconstitution on physiological yeast templates; nucleosome positioning assays; module deletion analysis; biophysical DNA shape analysis","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro chromatin reconstitution on whole-genome template with mechanistic module dissection; single lab with comprehensive genomic and biochemical approaches","pmids":["34050142"],"is_preprint":false},{"year":2021,"finding":"In the primed pluripotent state (but not naïve), INO80 promotes H2A.Z occupancy at bivalent gene promoters, facilitating H3K27me3 installation and maintenance. INO80 pre-marks gene promoters in naïve ESCs that adopt bivalent modifications upon transition to the primed state. This reveals a context-dependent role for INO80 in H2A.Z deposition (in addition to its known removal activity).","method":"Conditional Ino80 deletion in naïve vs primed ESCs; ChIP-seq for H2A.Z, H3K4me3, H3K27me3; gene expression analysis","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional knockout in defined cell states, genome-wide ChIP-seq; single lab","pmids":["34139016"],"is_preprint":false},{"year":2022,"finding":"INO80 prefers hexasomes (nucleosomes lacking one H2A-H2B dimer) as substrates over canonical nucleosomes, with up to ~60-fold preference at short flanking DNA overhangs (~18 bp linkers found in gene bodies. INO80 deletion significantly affects positions of hexasome-sized particles within yeast genes in vivo.","method":"In vitro nucleosome sliding assays comparing hexasome vs nucleosome substrates; yeast genetics; MNase-seq in ino80Δ","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with defined substrates plus in vivo genomic validation; single lab with quantitative biochemistry and genomics","pmids":["35597239"],"is_preprint":false},{"year":2022,"finding":"Cryo-EM structures of the INO80 regulatory A-module bound to DNA reveal the mechanism of linker DNA binding. The A-module connects to the motor via an HSA/post-HSA lever element that chemomechanically couples motor and linker DNA sensing. Two sites of curved DNA recognition by the four actin/actin-related proteins and the motor regulate sliding. YY1/Ies4 subunit recruitment and deep architectural similarities to SWI/SNF regulatory modules are revealed.","method":"Cryo-electron microscopy; functional DNA-binding and remodeling assays","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure plus functional assays; single lab","pmids":["36490333"],"is_preprint":false},{"year":2023,"finding":"Cryo-EM structure of INO80 in complex with a hexasome reveals a large structural rearrangement of the INO80 catalytic core into a 'spin-rotated' remodeling mode upon hexasome recognition. The nuclear actin module remains tethered to unwrapped linker DNA. An exposed H3-H4 interface in the hexasome activates INO80 independently of the H2A-H2B acidic patch.","method":"Cryo-electron microscopy; in vitro hexasome remodeling assays; mutagenesis","journal":"Science","confidence":"High","confidence_rationale":"Tier 1 / Moderate — cryo-EM structure with functional biochemical assays; single lab with structural and biochemical validation","pmids":["37384673"],"is_preprint":false},{"year":2023,"finding":"TORC1 activates the Rpd3L histone deacetylase complex to deacetylate Ino80 at K929, protecting Ino80 from autophagic degradation. Stabilized Ino80 promotes H2A.Z eviction from autophagy-related gene promoters, repressing their transcription. Rpd3L also deacetylates H2A.Z to further block its chromatin deposition. This pathway links nutrient signaling (TORC1) to chromatin remodeling and autophagy regulation.","method":"In vivo protein stability assays; ChIP for H2A.Z; genetic deletion of Rpd3L, TORC1 inhibition; site-directed mutagenesis of Ino80 K929; autophagy assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — mutagenesis of Ino80 K929, ChIP, genetic epistasis; single lab with multiple approaches but deacetylation of Ino80 by Rpd3L based on in vivo data","pmids":["36888706"],"is_preprint":false},{"year":2017,"finding":"INO80 nucleosome remodeling requires cooperativity between two INO80 complexes that simultaneously monitor DNA length on either side of a nucleosome. The C-terminal domain of human Ino80 (Ino80CTD) binds DNA cooperatively and dimerizes to provide crosstalk. A single active ATPase motor within the dimer is sufficient for nucleosome sliding, and ATPase activity gradually uncouples as the endpoint is approached, controlled by Ino80CTD.","method":"Nucleosome sliding assays with mutant complexes; ATPase assays; dimerization biochemistry; sedimentation assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with defined dimerization mutants, ATPase uncoupling assays; single lab with rigorous biochemistry","pmids":["28585918"],"is_preprint":false},{"year":2016,"finding":"Inositol hexaphosphate (IP6) is a non-competitive inhibitor of human INO80 that blocks nucleosomal stimulation of ATPase activity. The IP6 binding site is located within the C-terminal region of the Ino80 subunit. Ies2 and Arp5/Ies6 synergistically couple ATP hydrolysis to nucleosome sliding, and a bypass mutation in Arp5 is active in the absence of Ies2.","method":"In vitro ATPase assays; nucleosome sliding assays; recombinant complex purification from insect cells; domain mapping of IP6 binding","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstituted recombinant complex with systematic subunit deletions and inhibitor mechanism; single lab","pmids":["27257055"],"is_preprint":false},{"year":2010,"finding":"Yeast Ino80 interacts with the early NER damage recognition complex Rad4-Rad23 and is recruited to chromatin in a UV damage-dependent manner by Rad4. Ino80 acts in the same genetic pathway as NER. While chromatin disruption during UV lesion repair is normal in ino80 mutants, restoration of nucleosome structure after repair is defective.","method":"Co-immunoprecipitation (Ino80-Rad4-Rad23); ChIP at UV-damaged chromatin; modified ChIP to assess nucleosome reassembly; genetic epistasis","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — Co-IP, ChIP, genetic epistasis; mechanistic distinction between disruption and restoration of chromatin; single lab","pmids":["21135142"],"is_preprint":false},{"year":2020,"finding":"Hap2 (auxiliary subunit of fission yeast Ino80 complex) promotes de novo CENP-A chromatin assembly on naïve centromere DNA by facilitating transcription from centromere DNA, driving H3 nucleosome turnover and replacement by CENP-A nucleosomes. Chromatin association of Hap2 is Ies4-dependent.","method":"Affinity purification of CENP-A chromatin; genetic analysis of hap2 and ies4 deletions; de novo CENP-A assembly assays; H3 turnover measurements; ChIP","journal":"Genes & development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-purification, genetic epistasis, de novo assembly assays; single lab","pmids":["31919190"],"is_preprint":false},{"year":2007,"finding":"INO80 subunit Arp8 deficiency causes defects in sister chromatid cohesion. Ino80 directly associates with centromeres and cohesin-associated regions. In early S phase, Ino80 is recruited to replication forks along with Ctf18 and PCNA; arp8 mutation disrupts Ctf18 and PCNA association with replication forks.","method":"ChIP at centromeres and cohesin-associated regions; sister chromatid cohesion assay; Co-IP of Ino80 with replication fork components","journal":"Cell cycle","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP, Co-IP at replication forks, functional cohesion assay; single lab","pmids":["17471029"],"is_preprint":false},{"year":2022,"finding":"INO80 interacts with PRC2 core member SUZ12 and promotes its recruitment to bivalent promoters in spermatocytes. INO80 mediates H2A.Z incorporation at poised promoters, and its loss leads to reduced H3K27me3 and de-repression of poised genes, implicating INO80 in establishing poised chromatin through SUZ12/PRC2 binding.","method":"Co-immunoprecipitation (INO80-SUZ12); ChIP-seq for H3K27me3, H2A.Z; conditional Ino80 knockout; RNA-seq","journal":"Development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ChIP-seq, conditional knockout; single lab","pmids":["35006254"],"is_preprint":false},{"year":2017,"finding":"The INO80 complex acts downstream of the Mec1 checkpoint kinase to increase global chromatin mobility. Mec1 activation by targeted Ddc1/Ddc2 enhances chromatin mobility even in the absence of DNA damage, placing INO80 as an effector of checkpoint-mediated chromatin mobility.","method":"Live-cell fluorescence tracking of chromosomal loci; genetic epistasis (mec1, rad9, rad53, ino80 mutants); targeted Ddc1/Ddc2 activation","journal":"Genes & development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — live-cell imaging with genetic epistasis establishing INO80 downstream of Mec1; single lab","pmids":["24029917"],"is_preprint":false}],"current_model":"INO80 is a multisubunit ATP-dependent chromatin remodeler with a Snf2/Swi2-family ATPase catalytic subunit that is assembled around an Rvb1/Rvb2 AAA+ heterohexameric stator; its ATPase motor binds nucleosomal DNA at SHL-6 and, acting against the Arp5-Ies6 counter-grip on the opposite side of the H2A-H2B dimer, pumps entry DNA into the nucleosome in a ratchet mechanism that both slides nucleosomes and exchanges H2A.Z-H2B dimers for H2A-H2B without additional chaperones; the Arp8 module senses extranucleosomal linker DNA length to allosterically regulate motor activity; the complex is recruited to DSBs via γ-H2AX (through Nhp10 in yeast, through ARP8 in mammals), to replication forks via ubiquitinated H2A aided by BAP1, and to transcriptionally active genes via YY1; it functions in DSB end-resection, homologous recombination (by removing H2A.Z to enable presynaptic filament formation), NER (chromatin restoration after repair), nucleotide excision repair, replication fork progression and restart, RNAPII eviction from chromatin for proteasomal degradation, checkpoint adaptation, and genome-wide nucleosome positioning at gene regulatory elements including the +1 nucleosome; post-translationally it is phosphorylated by Mec1/Tel1 on Ies4 to modulate checkpoint signaling, deacetylated at K929 by Rpd3L (activated by TORC1) to protect it from autophagic degradation, and ubiquitinated by TRIM3 to regulate its abundance."},"narrative":{"mechanistic_narrative":"INO80 is the catalytic subunit of a multisubunit ATP-dependent chromatin remodeling complex that uses a Snf2/Swi2-family ATPase motor to slide nucleosomes and edit their histone composition, and through these activities governs genome stability, DNA repair, replication, and transcription [PMID:16230350, PMID:21241891]. Structurally the complex is built around an Rvb1/Rvb2 (RUVBL1/RUVBL2) AAA+ heterohexamer that serves as a stator scaffold engulfing a spoked-wheel domain of the INO80 ATPase; an Ino80 insertion domain stimulates Rvb1/Rvb2 ATPase activity and drives their hexamer-dodecamer cycling as assembly chaperones [PMID:28591576, PMID:29643509, PMID:29323271]. The motor binds nucleosomal DNA at the entry site near SHL-6/the H2A-H2B interface, unwraps entry DNA and creates torsional strain, while an Arp5-Ies6 module on the opposite side acts as a counter-grip at the nucleosome dyad — together producing a ratchet that pumps entry DNA into the nucleosome [PMID:28604691, PMID:29643509, PMID:29643506]. This mechanism both centers and spaces nucleosomes and exchanges H2A.Z-H2B for H2A-H2B dimers without external chaperones [PMID:21135121, PMID:21241891, PMID:28604691]. The Arp8/Arp4/actin (A) module, built on an extended HSA/post-HSA helical scaffold, binds extranucleosomal linker DNA ~37-51 bp from the nucleosome edge and functions as a DNA-length sensor that allosterically couples motor activity to flanking-DNA length, generating switch-like, cooperative remodeling that positions the +1 nucleosome at promoters and reads DNA shape genome-wide [PMID:30120252, PMID:30177756, PMID:29452642, PMID:34050142]. Activity is tuned internally — Ies2 activates the ATPase while Ies6/Arp5 promote nucleosome binding, IP6 non-competitively inhibits nucleosomal stimulation, and the complex preferentially remodels hexasomes via an exposed H3-H4 interface [PMID:24297934, PMID:35597239, PMID:37384673, PMID:27257055]. In DNA repair, INO80 is recruited to double-strand breaks (via Nhp10/γ-H2AX in yeast and ARP8 in mammals), promotes end-resection and removes H2A.Z to enable presynaptic filament formation during homologous recombination, restores chromatin after nucleotide excision repair, and is checkpoint-regulated through Mec1/Tel1 phosphorylation of the Ies4 subunit [PMID:15607975, PMID:15607974, PMID:17762868, PMID:17693258, PMID:20971067, PMID:26142279, PMID:28514650, PMID:21135142]. At replication, INO80 is recruited to forks through ubiquitinated H2A aided by the deubiquitinase BAP1, stabilizes stalled replisomes, promotes fork restart, and resolves R-loops to prevent transcription-replication conflicts, in part by evicting ubiquitinated RNAPII for proteasomal degradation [PMID:25283999, PMID:26656161, PMID:26798134, PMID:31657441, PMID:32913330]. In gene regulation, INO80 is recruited by sequence-specific factors including YY1, MITF/SOX9, and OCT4/WDR5 to open chromatin and license Mediator/RNAPII recruitment at promoters and superenhancers, controlling pluripotency, reprogramming, and oncogenic transcription in melanoma [PMID:17721549, PMID:24792115, PMID:27340176]. INO80 abundance is controlled by TRIM3-mediated ubiquitination and by TORC1/Rpd3L-dependent deacetylation at K929 that protects it from autophagic degradation [PMID:31633032, PMID:36888706].","teleology":[{"year":2004,"claim":"Established that INO80 is recruited to DNA double-strand breaks and participates directly in their processing, linking a chromatin remodeler to the DNA damage response.","evidence":"ChIP at HO-induced DSBs with H2A S129 and nhp10 mutant epistasis in yeast","pmids":["15607975","15607974"],"confidence":"High","gaps":["Did not define the remodeling reaction at the break","Mechanism of γ-H2AX-Nhp10 recognition not structurally resolved"]},{"year":2005,"claim":"Defined the human INO80 complex as a conserved, enzymatically active nucleosome-sliding machine, extending its functions to metazoans.","evidence":"Affinity purification/MS and in vitro ATPase and nucleosome sliding assays","pmids":["16230350"],"confidence":"High","gaps":["Subunit architecture and module organization not yet dissected","Substrate specificity beyond sliding unknown"]},{"year":2006,"claim":"Showed INO80 antagonizes SWR1 at DSB-flanking chromatin and is required for checkpoint adaptation, framing a balance of H2A.Z deposition versus removal in repair.","evidence":"ino80 × swr1 double-mutant epistasis, ChIP, and checkpoint adaptation assays","pmids":["16951256"],"confidence":"High","gaps":["Did not establish direct H2A.Z eviction biochemistry","Coupling to resection not yet defined"]},{"year":2007,"claim":"Connected INO80 to checkpoint signaling and to specific repair pathways through Mec1/Tel1 phosphorylation of Ies4 and the YY1-dependent recruitment that links INO80 to transcription and homologous recombination.","evidence":"Ies4 phosphosite mutagenesis with checkpoint assays; YY1 Co-IP, ChIP, and HR functional assays in yeast and mammalian cells","pmids":["17693258","17721549","18026119","15607975"],"confidence":"High","gaps":["How Ies4 phosphorylation alters complex activity unresolved","Whether YY1 recruitment is direct to specific INO80 surfaces unknown"]},{"year":2008,"claim":"Placed INO80 at replication origins and stalled forks where it stabilizes the replisome and enables restart, defining a replication-protective role distinct from break repair.","evidence":"Genome-wide ChIP and inducible degradation with replisome dissociation and HU restart assays in yeast","pmids":["18406137","18376411"],"confidence":"High","gaps":["Recruitment signal to forks not identified in yeast","Direct remodeling substrate at the fork unknown"]},{"year":2010,"claim":"Resolved INO80 as a precise nucleosome spacing factor with intrinsic histone-tail and DNA-length requirements, and extended its repair roles to mammalian DSB resection and nucleotide excision repair chromatin restoration.","evidence":"In vitro spacing assays with defined arrays; mammalian RNAi/knockout repair and NER factor assembly assays; Rad4-Rad23 Co-IP in yeast","pmids":["21135121","21947284","20855601","21135142","20971067"],"confidence":"High","gaps":["Mechanism translating DNA-length sensing into directional sliding not yet known","ARP8-dependent vs γ-H2AX-dependent recruitment difference between species unexplained"]},{"year":2011,"claim":"Demonstrated that INO80 directly exchanges H2A.Z-H2B for H2A-H2B and organized the human complex into functional modules, establishing histone editing as a core activity built on a conserved catalytic core.","evidence":"In vitro histone exchange, genome-wide H2A.Z ChIP-seq, htz1 epistasis, and human subassembly remodeling assays","pmids":["21241891","21303910"],"confidence":"High","gaps":["Structural basis of dimer exchange not yet resolved","How exchange directionality is controlled unknown"]},{"year":2012,"claim":"Defined Arp8 nucleosome engagement structurally and linked INO80 ATPase activity to enhanced chromatin mobility and recombination, providing a physical basis for repair-promoting nucleosome dynamics.","evidence":"Arp8 C-terminal crystal structure with binding assays; live-cell chromatin mobility imaging with ATPase-dead controls and gene conversion assays","pmids":["23213201","22345518"],"confidence":"High","gaps":["Whether mobility enhancement acts in cis at breaks unresolved","Full Arp8 module-motor coupling not yet mapped"]},{"year":2013,"claim":"Dissected internal regulation of the ATPase motor, showing nuclear actin and the Ies2/Ies6/Arp5 subunits separately tune catalysis and nucleosome binding.","evidence":"Reconstituted subunit deletions with ATPase and nucleosome-binding assays; actin subdomain mutagenesis with remodeling readout","pmids":["23524535","24297934"],"confidence":"High","gaps":["Allosteric path from actin to motor not yet structurally defined","In vivo consequences of each regulatory arm partially characterized"]},{"year":2014,"claim":"Established INO80 as a master regulator of open chromatin at pluripotency genes and as a replication-fork factor recruited through ubiquitinated H2A and stabilized by BAP1, tying its remodeling to transcription licensing and tumor-suppressor biology.","evidence":"ESC ChIP-seq/ATAC-seq with knockout reprogramming assays; Co-IP with ubiquitinated H2A, fork ChIP, and BAP1 knockout in mammalian cells","pmids":["24792115","25283999","25066231"],"confidence":"High","gaps":["Direct readers of ubiquitinated H2A within INO80 not identified","Mechanism of BAP1-INO80 stabilization not biochemically resolved"]},{"year":2015,"claim":"Showed that H2A.Z removal is the primary HR-promoting function of mammalian INO80 and defined the Arp8/Arp4/actin module as a DNA-length sensor controlling sliding, while linking INO80 to RNAPII turnover under transcription stress.","evidence":"H2A.Z co-depletion rescue of HR; Arp module DNA-binding mutagenesis with ATPase/sliding uncoupling; ternary INO80-RNAPII-Cdc48 Co-IP and ubiquitinated Rpb1 ChIP","pmids":["26142279","30120252","26656161","25964121"],"confidence":"High","gaps":["How linker-DNA sensing engages Arp5 at the acidic patch only partially resolved","Generality of RNAPII eviction across genes unknown"]},{"year":2016,"claim":"Defined kinetic and structural coupling within the complex and extended INO80's transcriptional and replication-protective roles to superenhancers and prevention of transcription-replication conflict.","evidence":"Arp5/Ies6/Ies2 reconstitution and IP6 inhibition assays; superenhancer ChIP-seq/ATAC-seq with xenografts; Mec1/INO80/PAF1 ChIP-seq and fiber assays","pmids":["26306040","27255055","27257055","27340176","26798134"],"confidence":"High","gaps":["Physiological role of IP6 regulation in vivo not established","How INO80 distinguishes conflict-prone loci unknown"]},{"year":2017,"claim":"Resolved the unusual entry-site translocation mechanism and intercomplex cooperativity, and identified Rvb1/Rvb2 chaperone behavior plus a presynaptic-filament HR function, defining how INO80 couples ATP hydrolysis to both sliding and histone editing.","evidence":"Photo-crosslinking and substrate-specific sliding/exchange assays; Ino80CTD dimerization biochemistry; Rvb1/Rvb2 cryo-EM and ATPase stimulation; htz1 epistasis rescuing presynaptic filaments","pmids":["28604691","28585918","28591576","28514650","24029917"],"confidence":"High","gaps":["Atomic structure of the engaged motor not yet available","How dimer cooperativity operates in chromatin not established"]},{"year":2018,"claim":"Delivered near-atomic cryo-EM structures of INO80-nucleosome complexes establishing the Rvb1/Rvb2 stator, SHL-6 motor engagement, and Arp5-Ies6 counter-grip ratchet, and resolved the Arp8/HSA linker-binding architecture and single-molecule switch behavior.","evidence":"Cryo-EM of fungal and human INO80-nucleosome complexes with biochemical validation; Arp8 module crystal structure; single-molecule sliding kinetics","pmids":["29643509","29643506","29323271","30177756","29452642"],"confidence":"High","gaps":["Conformational trajectory of the ratchet during translocation only inferred","How auto-inhibitory Nhp10 module integrates with motor not fully mapped"]},{"year":2020,"claim":"Showed INO80 resolves R-loops to protect replication and is essential for HR-mediated DSB repair in neural progenitors, dissociating its repair role from its transcriptional role in vivo.","evidence":"R-loop immunofluorescence, RNase H1 rescue, and LacO tethering; conditional cortical Ino80 knockout with in vivo HR assay and Brca2 comparison","pmids":["32913330","32737294","31919190"],"confidence":"High","gaps":["Whether INO80 acts on R-loops directly or via nucleosome remodeling not separated","Substrate context determining repair vs transcription role unclear"]},{"year":2021,"claim":"Established that INO80 reads DNA mechanical properties through core-Arp8 module allostery to position the +1 nucleosome genome-wide, and revealed a context-dependent H2A.Z deposition role in primed pluripotency.","evidence":"Genome-wide chromatin reconstitution with module deletions and DNA shape analysis; conditional knockout ChIP-seq in naive vs primed ESCs","pmids":["34050142","34139016"],"confidence":"High","gaps":["Molecular switch between H2A.Z removal and deposition not defined","How general regulatory factors physically cue +1 positioning unresolved"]},{"year":2022,"claim":"Defined hexasome as a preferred and structurally distinct substrate that activates INO80 through an exposed H3-H4 interface, and linked INO80 to PRC2/SUZ12 recruitment at bivalent promoters.","evidence":"Hexasome vs nucleosome sliding assays with MNase-seq; cryo-EM of INO80-hexasome spin-rotated mode; INO80-SUZ12 Co-IP and ChIP-seq in spermatocytes","pmids":["35597239","37384673","35006254"],"confidence":"High","gaps":["In vivo prevalence of hexasome remodeling not quantified","Direct INO80-SUZ12 interface not mapped"]},{"year":2023,"claim":"Defined post-translational control of INO80 abundance, linking nutrient signaling through TORC1/Rpd3L deacetylation at K929 to protection from autophagic degradation and chromatin-based autophagy gene control.","evidence":"Ino80 K929 mutagenesis, protein stability assays, ChIP, and TORC1/Rpd3L genetic epistasis","pmids":["36888706"],"confidence":"Medium","gaps":["Direct deacetylation of Ino80 by Rpd3L inferred from in vivo data","Whether acetylation regulates remodeling activity itself unknown"]},{"year":null,"claim":"How INO80 selects between H2A.Z removal versus deposition, and how its single conserved catalytic mechanism is locally directed toward repair, replication-fork protection, or transcription at specific genomic sites, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model reconciling H2A.Z eviction and context-dependent deposition","Site-specific switching between functional outputs not mechanistically defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[2,22,35,54]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[16,33,26]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[28,39,47]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[2,22]},{"term_id":"GO:0042393","term_label":"histone binding","supporting_discovery_ids":[20,51]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[5,24,32]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[2,11,21]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[0,9,18]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[0,4,26,34,55]},{"term_id":"R-HSA-69306","term_label":"DNA Replication","supporting_discovery_ids":[9,10,23,43,44]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[5,24,32]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[16,47,49]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[3,8,18]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[24,41,45]}],"complexes":["INO80 chromatin remodeling complex"],"partners":["RUVBL1","RUVBL2","ARP5","ARP8","YY1","BAP1","SUZ12","TRIM3"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9ULG1","full_name":"Chromatin-remodeling ATPase INO80","aliases":["DNA helicase-related INO80 complex homolog 1","DNA helicase-related protein INO80","INO80 complex subunit A"],"length_aa":1556,"mass_kda":176.8,"function":"ATPase component of the chromatin remodeling INO80 complex which is involved in transcriptional regulation, DNA replication and DNA repair (PubMed:16230350, PubMed:16298340, PubMed:17721549, PubMed:20237820, PubMed:20855601). Binds DNA (PubMed:16298340, PubMed:21303910). As part of the INO80 complex, remodels chromatin by shifting nucleosomes (PubMed:16230350, PubMed:21303910). Regulates transcription upon recruitment by YY1 to YY1-activated genes, where it acts as an essential coactivator (PubMed:17721549). Involved in UV-damage excision DNA repair (PubMed:20855601). The contribution to DNA double-strand break repair appears to be largely indirect through transcriptional regulation (PubMed:20687897). Involved in DNA replication (PubMed:20237820). 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progenitor divisions require chromatin-mediated homologous recombination DNA repair by Ino80.","date":"2020","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/32737294","citation_count":27,"is_preprint":false},{"pmid":"31633032","id":"PMC_31633032","title":"Ubiquitin-proteasomal regulation of chromatin remodeler INO80 in the nucleus accumbens mediates persistent cocaine craving.","date":"2019","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/31633032","citation_count":27,"is_preprint":false},{"pmid":"34139016","id":"PMC_34139016","title":"INO80 promotes H2A.Z occupancy to regulate cell fate transition in pluripotent stem cells.","date":"2021","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/34139016","citation_count":25,"is_preprint":false},{"pmid":"28585918","id":"PMC_28585918","title":"Crosstalk within a functional INO80 complex dimer regulates nucleosome sliding.","date":"2017","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/28585918","citation_count":25,"is_preprint":false},{"pmid":"28847826","id":"PMC_28847826","title":"Genome maintenance functions of the INO80 chromatin remodeller.","date":"2017","source":"Philosophical transactions of the Royal Society of London. Series B, Biological sciences","url":"https://pubmed.ncbi.nlm.nih.gov/28847826","citation_count":24,"is_preprint":false},{"pmid":"33378674","id":"PMC_33378674","title":"The INO80 Complex Regulates Epigenetic Inheritance of Heterochromatin.","date":"2020","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/33378674","citation_count":23,"is_preprint":false},{"pmid":"15207721","id":"PMC_15207721","title":"In silico characterization of the INO80 subfamily of SWI2/SNF2 chromatin remodeling proteins.","date":"2004","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/15207721","citation_count":23,"is_preprint":false},{"pmid":"27535137","id":"PMC_27535137","title":"Human INO80/YY1 chromatin remodeling complex transcriptionally regulates the BRCA2- and CDKN1A-interacting protein (BCCIP) in cells.","date":"2016","source":"Protein & cell","url":"https://pubmed.ncbi.nlm.nih.gov/27535137","citation_count":22,"is_preprint":false},{"pmid":"26975355","id":"PMC_26975355","title":"Ino80 is essential for proximal-distal axis asymmetry in part by regulating Bmp4 expression.","date":"2016","source":"BMC biology","url":"https://pubmed.ncbi.nlm.nih.gov/26975355","citation_count":22,"is_preprint":false},{"pmid":"26340092","id":"PMC_26340092","title":"Negative Regulation of p21Waf1/Cip1 by Human INO80 Chromatin Remodeling Complex Is Implicated in Cell Cycle Phase G2/M Arrest and Abnormal Chromosome Stability.","date":"2015","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/26340092","citation_count":22,"is_preprint":false},{"pmid":"16298340","id":"PMC_16298340","title":"Characterization of a human SWI2/SNF2 like protein hINO80: demonstration of catalytic and DNA binding activity.","date":"2005","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/16298340","citation_count":21,"is_preprint":false},{"pmid":"29432129","id":"PMC_29432129","title":"Identification of Two Distinct Classes of the Human INO80 Complex Genome-Wide.","date":"2018","source":"G3 (Bethesda, Md.)","url":"https://pubmed.ncbi.nlm.nih.gov/29432129","citation_count":21,"is_preprint":false},{"pmid":"29383140","id":"PMC_29383140","title":"INO80 haploinsufficiency inhibits colon cancer tumorigenesis via replication stress-induced apoptosis.","date":"2017","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/29383140","citation_count":21,"is_preprint":false},{"pmid":"36888706","id":"PMC_36888706","title":"The TORC1 activates Rpd3L complex to deacetylate Ino80 and H2A.Z and repress autophagy.","date":"2023","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/36888706","citation_count":20,"is_preprint":false},{"pmid":"30355728","id":"PMC_30355728","title":"Chromatin Remodeling Factors Isw2 and Ino80 Regulate Chromatin, Replication, and Copy Number of the Saccharomyces cerevisiae Ribosomal DNA Locus.","date":"2018","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/30355728","citation_count":20,"is_preprint":false},{"pmid":"35006254","id":"PMC_35006254","title":"INO80 requires a polycomb subunit to regulate the establishment of poised chromatin in murine spermatocytes.","date":"2022","source":"Development (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/35006254","citation_count":20,"is_preprint":false},{"pmid":"27779717","id":"PMC_27779717","title":"miR-148a inhibits self-renewal of thyroid cancer stem cells via repressing INO80 expression.","date":"2016","source":"Oncology reports","url":"https://pubmed.ncbi.nlm.nih.gov/27779717","citation_count":20,"is_preprint":false},{"pmid":"31919190","id":"PMC_31919190","title":"Hap2-Ino80-facilitated transcription promotes de novo establishment of CENP-A chromatin.","date":"2020","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/31919190","citation_count":19,"is_preprint":false},{"pmid":"28254775","id":"PMC_28254775","title":"Global Analysis of SUMO-Binding Proteins Identifies SUMOylation as a Key Regulator of the INO80 Chromatin Remodeling Complex.","date":"2017","source":"Molecular & cellular proteomics : MCP","url":"https://pubmed.ncbi.nlm.nih.gov/28254775","citation_count":19,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":41525,"output_tokens":15182,"usd":0.176152,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":27625,"output_tokens":6701,"usd":0.152825,"stage2_stop_reason":"end_turn"},"total_usd":0.328977,"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\": 2004,\n      \"finding\": \"The yeast INO80 complex is recruited to HO endonuclease-induced DNA double-strand breaks (DSBs) through interaction with phosphorylated histone H2A (γ-H2AX) at S129; recruitment requires the Nhp10 HMG-like subunit. Loss of INO80 (arp8 or H2A S129 mutants) impairs conversion of the DSB into ssDNA, implicating INO80-mediated chromatin remodeling in DSB end-processing.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP) at HO-induced DSB; genetic epistasis with H2A S129 mutants and nhp10 deletions; sensitivity assays to DNA damaging agents\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal ChIP, genetic epistasis, replicated independently by two labs in the same issue (PMIDs 15607975 and 15607974)\",\n      \"pmids\": [\"15607975\", \"15607974\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"The INO80 complex is recruited to DSBs via a specific interaction between the Nhp10 subunit and γ-H2AX; loss of Nhp10 or γ-H2AX reduces INO80 recruitment. INO80 components show synthetic genetic interactions with the RAD52 DSB repair pathway.\",\n      \"method\": \"ChIP at HO-induced DSB; Nhp10 deletion and γ-H2AX mutant analysis; genetic interaction screen with RAD52 pathway\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal ChIP, genetic epistasis, confirmed by independent lab (PMID 15607975) in same issue\",\n      \"pmids\": [\"15607974\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Human INO80 (hINO80) complex was identified and purified, containing orthologs of 8 of 15 yeast INO80 subunits plus at least five metazoan-specific subunits. The complex exhibits DNA- and nucleosome-activated ATPase activity and catalyzes ATP-dependent nucleosome sliding.\",\n      \"method\": \"Affinity purification / mass spectrometry; ATPase activity assay; nucleosome sliding assay in vitro\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of ATPase and nucleosome sliding activity with purified complex; single lab with two orthogonal biochemical assays\",\n      \"pmids\": [\"16230350\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Ino80 is required for cell cycle checkpoint adaptation in response to a persistent DSB. Cells lacking Ino80 fail to maintain high γ-H2AX levels and show increased H2A.Z (Htz1) incorporation near DSBs via Swr1. Inactivation of Swr1 restores H2AX phosphorylation and checkpoint adaptation in ino80 mutants, revealing antagonism between Ino80 and Swr1 at DSB-flanking chromatin.\",\n      \"method\": \"ChIP; genetic epistasis (ino80 × swr1 double mutants); checkpoint adaptation assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with double mutants, ChIP, and functional checkpoint assays; single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"16951256\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Swr1 is recruited to DSBs in a γ-H2AX-dependent manner. INO80 (but not SWR1) mediates removal of H2A.Z, γ-H2AX, and core histones near DSBs. Loss of INO80-specific subunits Arp8 or Nhp10 impairs Mre11, yKu80, and Mec1 binding at DSBs, causing defective end-processing and checkpoint activation.\",\n      \"method\": \"ChIP at DSBs; genetic analysis of INO80- vs SWR1-specific subunit mutants; immunofluorescence of repair factors\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP with multiple subunit mutants, functional repair/checkpoint assays, replicated across multiple studies\",\n      \"pmids\": [\"17762868\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"YY1 is tightly associated with the human INO80 complex. YY1 recruits INO80 to YY1-activated gene promoters where INO80 functions as a coactivator. YY1 binding to its DNA target sites requires INO80 activity.\",\n      \"method\": \"Co-immunoprecipitation; ChIP at YY1 target genes; RNAi knockdown of INO80 with transcriptional readout\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, ChIP, and functional knockdown experiments; replicated by independent study (PMID 18026119)\",\n      \"pmids\": [\"17721549\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"YY1 forms a complex with INO80 subunits in mammalian cells. Both YY1 and INO80 are essential for homologous recombination-based DNA repair (HRR); YY1 preferentially binds recombination-intermediate structures in vitro. RNAi knockdown of either increases cellular sensitivity to DNA-damaging agents.\",\n      \"method\": \"Co-immunoprecipitation; RNAi knockdown; HR functional assays; in vitro DNA-binding assays with recombination intermediates\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, in vitro binding, functional HR assay, knockdown phenotype; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"18026119\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"The Ies3 subunit of yeast INO80 interacts with the tetratricopeptide repeat domain of telomerase subunit Est1p. Deletion of IES3 causes telomere elongation, altered telomere position effect, delayed recombinational survivor formation, and stimulated extrachromosomal telomeric circles. Multiple INO80 subunits preferentially localize to telomeres.\",\n      \"method\": \"Co-immunoprecipitation; telomere length assays; ChIP at telomeres; genetic analysis of ies3Δ and arp8Δ in telomerase-negative strains\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP of Ies3-Est1, ChIP, genetic epistasis; single lab\",\n      \"pmids\": [\"17562861\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"The Mec1/Tel1 kinases (ATM/ATR orthologs) phosphorylate the Ies4 subunit of the INO80 complex during DNA damage. Mutation of Ies4 phosphorylation sites does not significantly affect DNA repair but influences DNA damage checkpoint responses, functionally linking INO80 to the Mec1/Tel1 checkpoint signaling pathway via Tof1 and Rad53.\",\n      \"method\": \"In vivo phosphorylation assays; site-directed mutagenesis of Ies4 phosphosites; checkpoint assays; genetic interaction analysis with tof1 and rad53\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — phosphosite mutagenesis with defined checkpoint phenotype, genetic epistasis; single lab with multiple orthogonal approaches\",\n      \"pmids\": [\"17693258\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"The Ino80 chromatin remodeling complex binds replication origins and stalled replication forks. Under hydroxyurea-induced fork arrest, INO80 accumulates at stalled forks and unfired origins. ino80 mutants are defective in replication restart after HU release and accumulate DSBs.\",\n      \"method\": \"ChIP across four yeast chromosomes; genome-wide mapping; HU treatment and release assays; DSB detection\",\n      \"journal\": \"Current biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP mapping plus functional replication restart assay; replicated by Peterson lab (PMID 18376411)\",\n      \"pmids\": [\"18406137\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"The Ino80 enzyme is recruited to replication origins as cells enter S phase and is required continuously for efficient fork progression, especially under low replication stress. Inducible degradation of Ino80 causes replisome dissociation from stalled forks, indicating Ino80 stabilizes the stalled replisome to ensure proper restart.\",\n      \"method\": \"Inducible protein degradation; ChIP at replication origins; replisome component co-precipitation; replication fork stability assays\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — inducible depletion with mechanistic replisome dissociation readout, ChIP; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"18376411\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"In the nucleus, the deubiquitinase Uch37 associates with the human INO80 complex where it is held in an inactive state. Uch37 can be activated by transient interaction of hINO80 with the proteasome, suggesting cooperative regulation of transcription or DNA repair.\",\n      \"method\": \"Co-immunoprecipitation; DUB activity assays in vitro; mass spectrometry identification of hINO80-Uch37 interaction\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and in vitro activity assay showing Uch37 is inactive in hINO80 and activated by proteasome; single lab\",\n      \"pmids\": [\"18922472\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"INO80 acts as a nucleosome spacing factor in vitro. It requires a minimum of 33–43 bp of extranucleosomal DNA and moves nucleosomes to the center of DNA with high precision. Unlike ISW2/1a, INO80 does not require H4 tails; instead the H2A histone tail negatively regulates nucleosome movement. INO80 spaces arrays with ~30 bp final linker DNA.\",\n      \"method\": \"In vitro nucleosome sliding/spacing assays with defined mono- and dinucleosomal arrays; histone tail deletion analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstituted nucleosome sliding/spacing assays with systematic DNA length and histone tail mutant analysis; single lab with rigorous biochemistry\",\n      \"pmids\": [\"21135121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Human INO80 promotes 5′→3′ resection of DSB ends in mammalian cells. Ino80 depletion impaired focal recruitment of 53BP1 but did not impede Rad51 focus formation, placing Ino80 at early steps of DSB repair. Ino80 associates with chromatin surrounding DSBs.\",\n      \"method\": \"RNAi knockdown; comet assay; HR repair reporter; immunofluorescence of 53BP1 and Rad51 foci; BrdU-ssDNA and RPA staining\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple readouts (comet assay, HR reporter, immunofluorescence, ssDNA detection); single lab\",\n      \"pmids\": [\"21947284\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Deletion of INO80 or ARP5 in mammalian cells impairs removal of UV-induced photo lesions via NER without affecting NER factor transcription. INO80 and Arp5 are recruited to UV-damaged DNA before NER incision occurs and are required for assembly of NER factors, indicating INO80 promotes chromatin accessibility for NER.\",\n      \"method\": \"Genetic knockout models; UV photolesion removal assays; ChIP for NER factors; chromatin accessibility assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic knockouts with mechanistic NER factor assembly readout; two subunit deletions (INO80 and ARP5) corroborating; single lab\",\n      \"pmids\": [\"20855601\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Mammalian INO80 complex is recruited to laser-induced DNA damage sites in an ARP8-dependent manner but independently of γ-H2AX, unlike the yeast complex where Nhp10 or Arp4 mediate recruitment.\",\n      \"method\": \"Laser micro-irradiation; immunofluorescence; siRNA knockdown of ARP8; live-cell imaging\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct localization experiment with ARP8 knockdown; single lab, single method\",\n      \"pmids\": [\"20971067\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Yeast INO80 has a histone exchange activity: it can replace nucleosomal H2A.Z/H2B dimers with free H2A/H2B dimers. In the absence of INO80, H2A.Z nucleosomes are mislocalized genome-wide and fail to respond dynamically to transcriptional changes. Genetic interactions between ino80 and htz1 (H2A.Z) support a model where INO80 removes unacetylated H2A.Z to promote genome stability.\",\n      \"method\": \"Genome-wide ChIP-seq for H2A.Z; in vitro histone exchange assays; genetic interaction analysis (ino80 × htz1)\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro histone exchange reconstitution plus genome-wide mapping and genetic epistasis; single lab with multiple orthogonal methods and comprehensive genome-wide data\",\n      \"pmids\": [\"21241891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"The human INO80 complex is organized in three modules assembling on distinct domains of hIno80 ATPase: (i) an N-terminal module with metazoan-specific subunits dispensable for remodeling, (ii) a module containing Arp4, Arp8, and YY1 on the HSA/PTH domain, and (iii) a catalytic core module with Ies2, Ies6, Arp5, Tip49a, and Tip49b. ATP-dependent nucleosome remodeling requires the evolutionarily conserved core comprising HSA/PTH and Snf2 ATPase domains together with YY1 and conserved subunits.\",\n      \"method\": \"Affinity purification of hINO80 subassemblies; mass spectrometry; in vitro nucleosome remodeling assays with subassemblies\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — reconstituted subassemblies with defined nucleosome remodeling activity; systematic dissection of three modules; single lab\",\n      \"pmids\": [\"21303910\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Loss of INO80 subunits Ies6 or Ino80 causes rapid polyploidy and chromosome missegregation. Chromatin structure flanking centromeres is altered in these mutants, not in the Cse4-containing centromeric nucleosome itself, but in pericentric chromatin. These effects are mediated through misincorporation of H2A.Z into pericentric regions.\",\n      \"method\": \"Flow cytometry (ploidy); live-cell imaging of chromosome segregation; ChIP for H2A.Z and histones at centromeres; genetic analysis\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple readouts (ploidy, segregation, ChIP), mechanistic link to H2A.Z misincorporation; single lab with orthogonal approaches\",\n      \"pmids\": [\"23207916\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Targeted recruitment of INO80 to a chromosomal locus enhances large-scale chromatin mobility in budding yeast, requiring Ino80's ATPase activity. This enhanced mobility correlates with increased rates of spontaneous gene conversion, indicating that INO80-dependent nucleosome remodeling promotes chromatin flexibility and homologous recombination.\",\n      \"method\": \"High-precision live fluorescence microscopy of tagged chromosomal loci; targeted tethering of INO80; ATPase-dead mutant analysis; gene conversion assays\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live-cell imaging with targeted tethering and ATPase mutant controls, plus functional HR assay; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"22345518\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Crystal structure of the Arp8 C-terminal domain shows three insertions within the actin fold. Arp8 forms a dimer that binds the nucleosome core primarily through H3 and H4 histones via one of these insertions, exploiting the twofold symmetry of the nucleosome.\",\n      \"method\": \"X-ray crystallography; biochemical binding assays (pull-down, EM); nucleosome co-purification with recombinant Arp8\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure plus biochemical validation of histone interactions; single lab with structure and binding assays\",\n      \"pmids\": [\"23213201\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Nuclear actin in the INO80 complex exists as a monomer with its barbed end inaccessible for polymerization. An actin mutation in subdomain 2 reduces INO80 chromatin remodeling activity in vitro and disrupts in vivo nuclear functions. The pointed end subdomain 2 of actin contributes to INO80-chromatin interactions.\",\n      \"method\": \"Biochemical fractionation; actin polymerization assays; actin subdomain mutagenesis; in vitro chromatin remodeling assays; genetic complementation\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution plus mutagenesis with defined chromatin remodeling readout; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"23524535\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The Ies2 subunit potently activates the intrinsic catalytic ATPase activity of the human Ino80 SNF2 ATPase, while Ies6 and Arp5 together promote binding of the Ino80 ATPase to nucleosomes. Thus the Ino80 ATPase is regulated at multiple levels within the complex.\",\n      \"method\": \"Purified subunit reconstitution; in vitro ATPase assays; nucleosome-binding assays with defined subunit deletions\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with defined subunit deletions and ATPase/nucleosome binding assays; single lab\",\n      \"pmids\": [\"24297934\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"INO80 binds replication forks in human cells and promotes fork progression under normal conditions. Ino80 is recruited to replication forks through interaction with ubiquitinated H2A, aided by BAP1 (a nuclear deubiquitinase that also stabilizes Ino80 protein). BAP1 deficiency in cancer cells downregulates Ino80 by disrupting this stabilization mechanism.\",\n      \"method\": \"Co-immunoprecipitation of Ino80 with ubiquitinated H2A; ChIP at replication forks; BAP1 knockdown/knockout; Ino80 protein stability assays; mouse embryo Ino80 deletion\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ChIP at replication forks, genetic knockouts with defined replication phenotype; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"25283999\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"INO80 occupies pluripotency gene promoters with master transcription factors OCT4 and WDR5. Ino80 maintains open chromatin architecture at these promoters and licenses Mediator and RNA Pol II recruitment. Ino80 is required for ESC self-renewal, somatic cell reprogramming, and blastocyst development.\",\n      \"method\": \"ChIP-seq; ATAC-seq; RNAi knockdown; genetic knockout in mouse; reprogramming assays\",\n      \"journal\": \"Cell stem cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — genome-wide ChIP-seq, chromatin accessibility, and functional knockout; mechanistic pathway placement; single lab with comprehensive approaches\",\n      \"pmids\": [\"24792115\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"INO80-dependent Mps3 (inner nuclear membrane SUN domain protein) binding to DSBs is S/G2-phase specific and requires both INO80 and Rad51. DSB relocation to Nup84 nuclear pore occurs independently of INO80 and cell-cycle phase. Thus INO80 determines choice of perinuclear anchorage site for DSBs.\",\n      \"method\": \"Live-cell imaging of DSB subnuclear position; genetic epistasis with ino80, rad51, swr1 mutants; cell-cycle synchronization\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live-cell localization with multiple genetic controls; single lab\",\n      \"pmids\": [\"25066231\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Mammalian INO80 rapidly removes H2A.Z from chromatin flanking DNA damage. Depletion of INO80 or histone chaperone ANP32E impairs homologous recombination, and the HR defect can be rescued by co-depletion of H2A.Z, demonstrating that H2A.Z removal is the primary function of INO80 and ANP32E in promoting HR.\",\n      \"method\": \"RNAi knockdown; H2A.Z ChIP after DNA damage; HR functional assay; epistasis by co-depletion of H2A.Z\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis (H2A.Z co-depletion rescues HR defect), ChIP, functional HR assay; single lab with mechanistically informative rescue experiment\",\n      \"pmids\": [\"26142279\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The yeast Ino80 chromatin remodeling complex is required for RNAPII turnover under transcriptional stress. INO80 forms a ternary complex with RNAPII and Cdc48/p97, and cells lacking INO80 accumulate ubiquitinated Rpb1 tightly bound to chromatin, indicating INO80 nucleosome remodeling activity promotes dissociation of ubiquitinated RNAPII from chromatin for proteasomal degradation.\",\n      \"method\": \"Co-immunoprecipitation; in vivo Rpb1 ubiquitination/degradation assays; ChIP for ubiquitinated Rpb1; genetic epistasis with cdc48 and proteasome mutants\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP of ternary complex, ChIP, genetic epistasis; single lab with multiple orthogonal approaches\",\n      \"pmids\": [\"26656161\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The Arp8/Arp4/actin module in INO80 binds extranucleosomal DNA 37-51 bp from the nucleosome edge and functions as a DNA-length sensor that regulates nucleosome sliding. Disruption of Arp8/Arp4 DNA binding uncouples ATP hydrolysis from nucleosome mobilization by disengaging Arp5 from the H2A-H2B acidic patch.\",\n      \"method\": \"In vitro DNA-binding assays (EMSA); site-directed mutagenesis of Arp8/Arp4; ATPase assays; nucleosome sliding assays; photo-crosslinking\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — mutagenesis, in vitro binding, ATPase, and sliding assays; mechanistic coupling established; single lab\",\n      \"pmids\": [\"30120252\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"EM structural analysis shows INO80-C and SWR-C share similar overall architectures with a compact head containing Rvb1/Rvb2 as single heterohexameric rings. The Arp8/Arp4/Act1 module enhances nucleosome-binding affinity but is largely dispensable for remodeling. The Ies6/Arp5 module is essential for remodeling activity and controls conformational changes coupling nucleosome binding to remodeling.\",\n      \"method\": \"Electron microscopy; 2D class averaging; mass spectrometry; nucleosome remodeling assays with module deletions\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — EM structure plus functional module deletion assays; single lab with structural and biochemical validation\",\n      \"pmids\": [\"25964121\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"The Arp5-Ies6 subcomplex forms an abundant distinct subcomplex in vivo and stimulates INO80-mediated ATPase and nucleosome sliding activity in vitro. Ies2 is required for Arp5-Ies6 association with the catalytic INO80 components. Mutant Arp5 lacking unique insertion domains allows ATP hydrolysis without nucleosome sliding, uncoupling the two activities.\",\n      \"method\": \"In vivo co-immunoprecipitation; in vitro ATPase and nucleosome sliding assays; domain deletion/mutagenesis of Arp5\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with mutagenesis defining Arp5 insertion domains in coupling ATP hydrolysis to sliding; single lab\",\n      \"pmids\": [\"26306040\", \"27255055\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Mec1, INO80, and PAF1 complexes cooperate to remove RNAPII from transcribed genes near early-firing replication origins upon HU treatment. Mec1 triggers efficient removal of PAF1C and RNAPII. Failure to evict RNAPII correlates with defective replication fork restart, implicating INO80 in preventing transcription-replication conflicts.\",\n      \"method\": \"ChIP-seq for RNAPII and PAF1C; genetic epistasis (mec1, ino80, paf1 mutants); DNA fiber assays for replication restart; proteomic analysis\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP-seq, fiber assays, genetic epistasis across multiple mutants; single lab with genome-wide and functional approaches\",\n      \"pmids\": [\"26798134\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"INO80 occupies >90% of superenhancers in melanoma cells, dependent on transcription factors MITF and Sox9. Ino80 binding reduces nucleosome occupancy and facilitates Mediator recruitment, promoting oncogenic transcription. Ino80 silencing selectively inhibits melanoma cell proliferation and tumorigenesis.\",\n      \"method\": \"ChIP-seq; ATAC-seq; Ino80 siRNA knockdown; mouse xenograft assays; Co-IP with Mediator\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide ChIP-seq, chromatin accessibility, in vivo tumorigenesis; mechanistic link to Mediator recruitment; single lab\",\n      \"pmids\": [\"27340176\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"INO80 translocates along DNA at the H2A-H2B interface of nucleosomes (unlike other remodelers that translocate at the H3-H4 interface), creating DNA torsional strain near the nucleosome entry site. This mechanism promotes both nucleosome mobilization and selective exchange of H2A.Z-H2B dimers with H2A-H2B without additional histone chaperones. INO80 mobilizes H2A.Z-containing nucleosomes more efficiently than H2A nucleosomes.\",\n      \"method\": \"Site-directed photo-crosslinking; ATPase assays; nucleosome sliding assays with H2A vs H2A.Z substrates; histone exchange assays in vitro\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with photo-crosslinking defining translocation site plus histone exchange assays; single lab with mechanistically rigorous biochemistry\",\n      \"pmids\": [\"28604691\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The INO80 complex removes H2A.Z to promote presynaptic filament formation during homologous recombination in budding yeast. INO80 has at least two distinct functions in HR: DNA end resection and presynaptic filament formation. H2A.Z deletion rescues presynaptic filament formation and HR in INO80-deficient mutants.\",\n      \"method\": \"High-resolution ChIP; HR assays; genetic epistasis with H2A.Z deletion (htz1Δ); direct visualization of presynaptic filament formation\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis (htz1Δ rescues INO80-C HR defect), mechanistic dissection; single lab with multiple orthogonal assays\",\n      \"pmids\": [\"28514650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"An insertion domain (Ino80INS) in the Ino80 ATPase stimulates Rvb1/Rvb2 ATPase activity 16-fold and promotes their dodecamerization. Rvb1/Rvb2 function as protein assembly chaperones within INO80, cycling between hexamers and dodecamers in an ATP-dependent manner.\",\n      \"method\": \"ATPase activity assays; mass spectrometry; cryo-EM and integrative modeling; biochemical reconstitution\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure, in vitro ATPase stimulation assay, and mass spectrometry; single lab with multiple orthogonal structural and biochemical approaches\",\n      \"pmids\": [\"28591576\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Cryo-EM structure of evolutionarily conserved INO80 core from Chaetomium thermophilum bound to nucleosome at 4.3/3.7 Å resolution. The Rvb1/Rvb2 AAA+ heterohexamer acts as a stator scaffold. The Swi2/Snf2 ATPase motor binds at SHL-6, unwraps ~15 bp, disrupts H2A-DNA contacts, and is poised to pump entry DNA into the nucleosome. Arp5 and Ies6 bind at SHL-2/-3 as a counter-grip, with the Arp5 grappler element binding the nucleosome dyad. The structure suggests a ratchet mechanism for both nucleosome sliding and histone editing.\",\n      \"method\": \"Cryo-electron microscopy (global 4.3 Å, major parts 3.7 Å); biochemical validation of ATPase and sliding activities\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — near-atomic resolution cryo-EM structure with biochemical validation; replicated independently by human INO80 structure (PMID 29643506)\",\n      \"pmids\": [\"29643509\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Cryo-EM structure of human INO80 bound to nucleosome reveals the motor domains are located at the DNA entry point (not at SHL2 as in other remodelers). ARP5-IES6 module makes additional contacts on the opposite side. Histone H3 tails regulate the INO80 motor domain (unlike other remodelers where H4 tails play this role).\",\n      \"method\": \"Cryo-electron microscopy (9.6 Å overall, 4.1 Å for portions); biochemical validation\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure of human complex with biochemical validation; replicated by fungal INO80 structure (PMID 29643509) revealing conserved yet distinct features\",\n      \"pmids\": [\"29643506\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Cryo-EM structures of the active core of human INO80 at 9.6 Å reveal an unusual spoked-wheel structural domain of Ino80 subunit engulfed by a single RUVBL1/RUVBL2 AAA+ heterohexamer. RUVBL1/RUVBL2 form a major interaction site for partner proteins that likely communicate to nucleotide-binding sites.\",\n      \"method\": \"Cryo-EM; subunit reconstitution; EM class averaging\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with biochemical subunit analysis; single lab\",\n      \"pmids\": [\"29323271\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Crystal structure of the 180-kDa Arp8 module of yeast INO80 shows Arp8 engages nuclear actin in a manner distinct from other actin-fold proteins, recruiting the Arp4-N-actin heterodimer to a segmented scaffold of the helical HSA domain. The HSA domain spans >120 Å and provides an extended binding platform for extranucleosomal entry DNA required for nucleosome sliding and genome-wide nucleosome positioning.\",\n      \"method\": \"X-ray crystallography; biochemical DNA-binding assays; in vitro nucleosome sliding assays; genome-wide nucleosome mapping\",\n      \"journal\": \"Nature structural & molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus in vitro and in vivo functional validation; corroborated by independent biochemical study (PMID 30120252)\",\n      \"pmids\": [\"30177756\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"INO80 operates as a DNA length-sensitive switch: nucleosome sliding rate increases ~100-fold when flanking DNA increases from 40 to 60 bp. Once initiated, INO80 moves nucleosomes rapidly at least 20 bp without pausing. The Nhp10 module plays an auto-inhibitory role tuning this switch-like response. INO80 can change direction of sliding without dissociation.\",\n      \"method\": \"Single-molecule enzymology; ensemble ATPase and sliding assays; Nhp10 module deletion analysis\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — single-molecule and ensemble biochemistry with defined kinetic parameters and regulatory module deletion; single lab with rigorous quantitative assays\",\n      \"pmids\": [\"29452642\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"INO80 deletion in vascular endothelial cells prevents ventricular compaction in the developing mouse heart, correlating with defective coronary vascularization. In vitro, endothelial cells promote myocardial expansion in an Ino80-dependent manner. Ino80 deletion increases E2F-activated gene expression and endothelial S-phase occupancy.\",\n      \"method\": \"Conditional endothelial Ino80 knockout (mouse); histological analysis; in vitro co-culture assays; gene expression profiling\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional knockout with mechanistic readout (E2F gene expression, cell cycle); single lab\",\n      \"pmids\": [\"29371594\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"TRIM3 E3 ubiquitin ligase mediates degradation of INO80 in the nucleus accumbens; TRIM3 and INO80 interact directly, and reduced TRIM3 on abstinence day 30 leads to increased INO80 protein levels. INO80-mediated transcriptional changes regulate cocaine craving during prolonged abstinence.\",\n      \"method\": \"Co-immunoprecipitation; viral gene transfer; ChIP-seq; ubiquitin degradation assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP showing TRIM3-INO80 interaction, functional viral gene transfer; single lab\",\n      \"pmids\": [\"31633032\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"BAP1 depletion reduces DNA synthesis and impairs restart of HU-induced stalled replication forks. This defect is rescued by ectopic INO80 expression. BAP1 depletion abrogates INO80 binding to stalled replication forks, indicating BAP1 promotes replication stress recovery by recruiting INO80 to stalled forks.\",\n      \"method\": \"DNA fiber assays; ChIP at replication forks; siRNA knockdown; ectopic INO80 expression rescue\",\n      \"journal\": \"The Biochemical journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis rescue experiment, ChIP, DNA fiber assays; single lab\",\n      \"pmids\": [\"31657441\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"INO80 complex promotes resolution of R-loops to prevent replication-associated DNA damage in cancer cells. R-loops promote INO80 recruitment to chromatin. Overexpression of RNase H1 rescues DNA synthesis defects and suppresses DNA damage caused by INO80 depletion. Artificial tethering of INO80 to a LacO locus enables R-loop turnover in cis.\",\n      \"method\": \"RNAi depletion of INO80; R-loop immunofluorescence (S9.6 antibody); DNA fiber assays; RNase H1 rescue; LacO artificial tethering\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple readouts including RNase H1 rescue and artificial tethering establishing direct R-loop resolution; single lab with orthogonal methods\",\n      \"pmids\": [\"32913330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Ino80 conditional deletion from cortical neural progenitor cells impairs DNA DSB repair selectively via homologous recombination, causing p53-dependent apoptosis and microcephaly. Ino80 function in HR is mechanistically distinct from its role in YY1-associated transcription. Sensitivity is dependent on NPC division mode: symmetric NPC-NPC divisions but not asymmetric neurogenic divisions require Ino80-mediated HR.\",\n      \"method\": \"Conditional knockout mouse; in vivo DSB repair pathway assay; apoptosis markers; phenotype comparison with Brca2 conditional knockout\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional knockout with in vivo HR assay and genetic epistasis with Brca2; mechanistic dissociation of HR and transcription functions; replicated concept with Brca2\",\n      \"pmids\": [\"32737294\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In fission yeast, INO80 subunit Iec5 promotes histone turnover at heterochromatin, enabling INO80 to counter epigenetic inheritance of heterochromatin. Mutations in INO80 components allow pericentric heterochromatin inheritance in RNAi mutants. This function is distinct from nucleosome positioning at heterochromatin.\",\n      \"method\": \"Genetic screen; heterochromatin inheritance assays; histone turnover measurements; ChIP\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic screen plus histone turnover assays with subunit-specific (Iec5) dissection; single lab\",\n      \"pmids\": [\"33378674\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"INO80 processes DNA shape/mechanical properties encoded in the genome through allosteric interplay between its core and Arp8 modules to position nucleosomes genome-wide. At promoters, INO80 integrates DNA mechanics readout with general regulatory factor binding to position the +1 nucleosome. This establishes a molecular mechanism for robust, adjustable +1 nucleosome positioning.\",\n      \"method\": \"Genome-wide chromatin reconstitution on physiological yeast templates; nucleosome positioning assays; module deletion analysis; biophysical DNA shape analysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro chromatin reconstitution on whole-genome template with mechanistic module dissection; single lab with comprehensive genomic and biochemical approaches\",\n      \"pmids\": [\"34050142\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"In the primed pluripotent state (but not naïve), INO80 promotes H2A.Z occupancy at bivalent gene promoters, facilitating H3K27me3 installation and maintenance. INO80 pre-marks gene promoters in naïve ESCs that adopt bivalent modifications upon transition to the primed state. This reveals a context-dependent role for INO80 in H2A.Z deposition (in addition to its known removal activity).\",\n      \"method\": \"Conditional Ino80 deletion in naïve vs primed ESCs; ChIP-seq for H2A.Z, H3K4me3, H3K27me3; gene expression analysis\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional knockout in defined cell states, genome-wide ChIP-seq; single lab\",\n      \"pmids\": [\"34139016\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"INO80 prefers hexasomes (nucleosomes lacking one H2A-H2B dimer) as substrates over canonical nucleosomes, with up to ~60-fold preference at short flanking DNA overhangs (~18 bp linkers found in gene bodies. INO80 deletion significantly affects positions of hexasome-sized particles within yeast genes in vivo.\",\n      \"method\": \"In vitro nucleosome sliding assays comparing hexasome vs nucleosome substrates; yeast genetics; MNase-seq in ino80Δ\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with defined substrates plus in vivo genomic validation; single lab with quantitative biochemistry and genomics\",\n      \"pmids\": [\"35597239\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Cryo-EM structures of the INO80 regulatory A-module bound to DNA reveal the mechanism of linker DNA binding. The A-module connects to the motor via an HSA/post-HSA lever element that chemomechanically couples motor and linker DNA sensing. Two sites of curved DNA recognition by the four actin/actin-related proteins and the motor regulate sliding. YY1/Ies4 subunit recruitment and deep architectural similarities to SWI/SNF regulatory modules are revealed.\",\n      \"method\": \"Cryo-electron microscopy; functional DNA-binding and remodeling assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure plus functional assays; single lab\",\n      \"pmids\": [\"36490333\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Cryo-EM structure of INO80 in complex with a hexasome reveals a large structural rearrangement of the INO80 catalytic core into a 'spin-rotated' remodeling mode upon hexasome recognition. The nuclear actin module remains tethered to unwrapped linker DNA. An exposed H3-H4 interface in the hexasome activates INO80 independently of the H2A-H2B acidic patch.\",\n      \"method\": \"Cryo-electron microscopy; in vitro hexasome remodeling assays; mutagenesis\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — cryo-EM structure with functional biochemical assays; single lab with structural and biochemical validation\",\n      \"pmids\": [\"37384673\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TORC1 activates the Rpd3L histone deacetylase complex to deacetylate Ino80 at K929, protecting Ino80 from autophagic degradation. Stabilized Ino80 promotes H2A.Z eviction from autophagy-related gene promoters, repressing their transcription. Rpd3L also deacetylates H2A.Z to further block its chromatin deposition. This pathway links nutrient signaling (TORC1) to chromatin remodeling and autophagy regulation.\",\n      \"method\": \"In vivo protein stability assays; ChIP for H2A.Z; genetic deletion of Rpd3L, TORC1 inhibition; site-directed mutagenesis of Ino80 K929; autophagy assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — mutagenesis of Ino80 K929, ChIP, genetic epistasis; single lab with multiple approaches but deacetylation of Ino80 by Rpd3L based on in vivo data\",\n      \"pmids\": [\"36888706\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"INO80 nucleosome remodeling requires cooperativity between two INO80 complexes that simultaneously monitor DNA length on either side of a nucleosome. The C-terminal domain of human Ino80 (Ino80CTD) binds DNA cooperatively and dimerizes to provide crosstalk. A single active ATPase motor within the dimer is sufficient for nucleosome sliding, and ATPase activity gradually uncouples as the endpoint is approached, controlled by Ino80CTD.\",\n      \"method\": \"Nucleosome sliding assays with mutant complexes; ATPase assays; dimerization biochemistry; sedimentation assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with defined dimerization mutants, ATPase uncoupling assays; single lab with rigorous biochemistry\",\n      \"pmids\": [\"28585918\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Inositol hexaphosphate (IP6) is a non-competitive inhibitor of human INO80 that blocks nucleosomal stimulation of ATPase activity. The IP6 binding site is located within the C-terminal region of the Ino80 subunit. Ies2 and Arp5/Ies6 synergistically couple ATP hydrolysis to nucleosome sliding, and a bypass mutation in Arp5 is active in the absence of Ies2.\",\n      \"method\": \"In vitro ATPase assays; nucleosome sliding assays; recombinant complex purification from insect cells; domain mapping of IP6 binding\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstituted recombinant complex with systematic subunit deletions and inhibitor mechanism; single lab\",\n      \"pmids\": [\"27257055\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Yeast Ino80 interacts with the early NER damage recognition complex Rad4-Rad23 and is recruited to chromatin in a UV damage-dependent manner by Rad4. Ino80 acts in the same genetic pathway as NER. While chromatin disruption during UV lesion repair is normal in ino80 mutants, restoration of nucleosome structure after repair is defective.\",\n      \"method\": \"Co-immunoprecipitation (Ino80-Rad4-Rad23); ChIP at UV-damaged chromatin; modified ChIP to assess nucleosome reassembly; genetic epistasis\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ChIP, genetic epistasis; mechanistic distinction between disruption and restoration of chromatin; single lab\",\n      \"pmids\": [\"21135142\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Hap2 (auxiliary subunit of fission yeast Ino80 complex) promotes de novo CENP-A chromatin assembly on naïve centromere DNA by facilitating transcription from centromere DNA, driving H3 nucleosome turnover and replacement by CENP-A nucleosomes. Chromatin association of Hap2 is Ies4-dependent.\",\n      \"method\": \"Affinity purification of CENP-A chromatin; genetic analysis of hap2 and ies4 deletions; de novo CENP-A assembly assays; H3 turnover measurements; ChIP\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-purification, genetic epistasis, de novo assembly assays; single lab\",\n      \"pmids\": [\"31919190\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"INO80 subunit Arp8 deficiency causes defects in sister chromatid cohesion. Ino80 directly associates with centromeres and cohesin-associated regions. In early S phase, Ino80 is recruited to replication forks along with Ctf18 and PCNA; arp8 mutation disrupts Ctf18 and PCNA association with replication forks.\",\n      \"method\": \"ChIP at centromeres and cohesin-associated regions; sister chromatid cohesion assay; Co-IP of Ino80 with replication fork components\",\n      \"journal\": \"Cell cycle\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, Co-IP at replication forks, functional cohesion assay; single lab\",\n      \"pmids\": [\"17471029\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"INO80 interacts with PRC2 core member SUZ12 and promotes its recruitment to bivalent promoters in spermatocytes. INO80 mediates H2A.Z incorporation at poised promoters, and its loss leads to reduced H3K27me3 and de-repression of poised genes, implicating INO80 in establishing poised chromatin through SUZ12/PRC2 binding.\",\n      \"method\": \"Co-immunoprecipitation (INO80-SUZ12); ChIP-seq for H3K27me3, H2A.Z; conditional Ino80 knockout; RNA-seq\",\n      \"journal\": \"Development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ChIP-seq, conditional knockout; single lab\",\n      \"pmids\": [\"35006254\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The INO80 complex acts downstream of the Mec1 checkpoint kinase to increase global chromatin mobility. Mec1 activation by targeted Ddc1/Ddc2 enhances chromatin mobility even in the absence of DNA damage, placing INO80 as an effector of checkpoint-mediated chromatin mobility.\",\n      \"method\": \"Live-cell fluorescence tracking of chromosomal loci; genetic epistasis (mec1, rad9, rad53, ino80 mutants); targeted Ddc1/Ddc2 activation\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — live-cell imaging with genetic epistasis establishing INO80 downstream of Mec1; single lab\",\n      \"pmids\": [\"24029917\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"INO80 is a multisubunit ATP-dependent chromatin remodeler with a Snf2/Swi2-family ATPase catalytic subunit that is assembled around an Rvb1/Rvb2 AAA+ heterohexameric stator; its ATPase motor binds nucleosomal DNA at SHL-6 and, acting against the Arp5-Ies6 counter-grip on the opposite side of the H2A-H2B dimer, pumps entry DNA into the nucleosome in a ratchet mechanism that both slides nucleosomes and exchanges H2A.Z-H2B dimers for H2A-H2B without additional chaperones; the Arp8 module senses extranucleosomal linker DNA length to allosterically regulate motor activity; the complex is recruited to DSBs via γ-H2AX (through Nhp10 in yeast, through ARP8 in mammals), to replication forks via ubiquitinated H2A aided by BAP1, and to transcriptionally active genes via YY1; it functions in DSB end-resection, homologous recombination (by removing H2A.Z to enable presynaptic filament formation), NER (chromatin restoration after repair), nucleotide excision repair, replication fork progression and restart, RNAPII eviction from chromatin for proteasomal degradation, checkpoint adaptation, and genome-wide nucleosome positioning at gene regulatory elements including the +1 nucleosome; post-translationally it is phosphorylated by Mec1/Tel1 on Ies4 to modulate checkpoint signaling, deacetylated at K929 by Rpd3L (activated by TORC1) to protect it from autophagic degradation, and ubiquitinated by TRIM3 to regulate its abundance.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"INO80 is the catalytic subunit of a multisubunit ATP-dependent chromatin remodeling complex that uses a Snf2/Swi2-family ATPase motor to slide nucleosomes and edit their histone composition, and through these activities governs genome stability, DNA repair, replication, and transcription [#2, #16]. Structurally the complex is built around an Rvb1/Rvb2 (RUVBL1/RUVBL2) AAA+ heterohexamer that serves as a stator scaffold engulfing a spoked-wheel domain of the INO80 ATPase; an Ino80 insertion domain stimulates Rvb1/Rvb2 ATPase activity and drives their hexamer-dodecamer cycling as assembly chaperones [#35, #36, #38]. The motor binds nucleosomal DNA at the entry site near SHL-6/the H2A-H2B interface, unwraps entry DNA and creates torsional strain, while an Arp5-Ies6 module on the opposite side acts as a counter-grip at the nucleosome dyad — together producing a ratchet that pumps entry DNA into the nucleosome [#33, #36, #37]. This mechanism both centers and spaces nucleosomes and exchanges H2A.Z-H2B for H2A-H2B dimers without external chaperones [#12, #16, #33]. The Arp8/Arp4/actin (A) module, built on an extended HSA/post-HSA helical scaffold, binds extranucleosomal linker DNA ~37-51 bp from the nucleosome edge and functions as a DNA-length sensor that allosterically couples motor activity to flanking-DNA length, generating switch-like, cooperative remodeling that positions the +1 nucleosome at promoters and reads DNA shape genome-wide [#28, #39, #40, #47]. Activity is tuned internally — Ies2 activates the ATPase while Ies6/Arp5 promote nucleosome binding, IP6 non-competitively inhibits nucleosomal stimulation, and the complex preferentially remodels hexasomes via an exposed H3-H4 interface [#22, #49, #51, #54]. In DNA repair, INO80 is recruited to double-strand breaks (via Nhp10/\\u03b3-H2AX in yeast and ARP8 in mammals), promotes end-resection and removes H2A.Z to enable presynaptic filament formation during homologous recombination, restores chromatin after nucleotide excision repair, and is checkpoint-regulated through Mec1/Tel1 phosphorylation of the Ies4 subunit [#0, #4, #8, #15, #26, #34, #55]. At replication, INO80 is recruited to forks through ubiquitinated H2A aided by the deubiquitinase BAP1, stabilizes stalled replisomes, promotes fork restart, and resolves R-loops to prevent transcription-replication conflicts, in part by evicting ubiquitinated RNAPII for proteasomal degradation [#23, #27, #31, #43, #44]. In gene regulation, INO80 is recruited by sequence-specific factors including YY1, MITF/SOX9, and OCT4/WDR5 to open chromatin and license Mediator/RNAPII recruitment at promoters and superenhancers, controlling pluripotency, reprogramming, and oncogenic transcription in melanoma [#5, #24, #32]. INO80 abundance is controlled by TRIM3-mediated ubiquitination and by TORC1/Rpd3L-dependent deacetylation at K929 that protects it from autophagic degradation [#42, #52].\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"Established that INO80 is recruited to DNA double-strand breaks and participates directly in their processing, linking a chromatin remodeler to the DNA damage response.\",\n      \"evidence\": \"ChIP at HO-induced DSBs with H2A S129 and nhp10 mutant epistasis in yeast\",\n      \"pmids\": [\"15607975\", \"15607974\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the remodeling reaction at the break\", \"Mechanism of \\u03b3-H2AX-Nhp10 recognition not structurally resolved\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined the human INO80 complex as a conserved, enzymatically active nucleosome-sliding machine, extending its functions to metazoans.\",\n      \"evidence\": \"Affinity purification/MS and in vitro ATPase and nucleosome sliding assays\",\n      \"pmids\": [\"16230350\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Subunit architecture and module organization not yet dissected\", \"Substrate specificity beyond sliding unknown\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Showed INO80 antagonizes SWR1 at DSB-flanking chromatin and is required for checkpoint adaptation, framing a balance of H2A.Z deposition versus removal in repair.\",\n      \"evidence\": \"ino80 \\u00d7 swr1 double-mutant epistasis, ChIP, and checkpoint adaptation assays\",\n      \"pmids\": [\"16951256\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish direct H2A.Z eviction biochemistry\", \"Coupling to resection not yet defined\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Connected INO80 to checkpoint signaling and to specific repair pathways through Mec1/Tel1 phosphorylation of Ies4 and the YY1-dependent recruitment that links INO80 to transcription and homologous recombination.\",\n      \"evidence\": \"Ies4 phosphosite mutagenesis with checkpoint assays; YY1 Co-IP, ChIP, and HR functional assays in yeast and mammalian cells\",\n      \"pmids\": [\"17693258\", \"17721549\", \"18026119\", \"15607975\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How Ies4 phosphorylation alters complex activity unresolved\", \"Whether YY1 recruitment is direct to specific INO80 surfaces unknown\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Placed INO80 at replication origins and stalled forks where it stabilizes the replisome and enables restart, defining a replication-protective role distinct from break repair.\",\n      \"evidence\": \"Genome-wide ChIP and inducible degradation with replisome dissociation and HU restart assays in yeast\",\n      \"pmids\": [\"18406137\", \"18376411\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Recruitment signal to forks not identified in yeast\", \"Direct remodeling substrate at the fork unknown\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Resolved INO80 as a precise nucleosome spacing factor with intrinsic histone-tail and DNA-length requirements, and extended its repair roles to mammalian DSB resection and nucleotide excision repair chromatin restoration.\",\n      \"evidence\": \"In vitro spacing assays with defined arrays; mammalian RNAi/knockout repair and NER factor assembly assays; Rad4-Rad23 Co-IP in yeast\",\n      \"pmids\": [\"21135121\", \"21947284\", \"20855601\", \"21135142\", \"20971067\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism translating DNA-length sensing into directional sliding not yet known\", \"ARP8-dependent vs \\u03b3-H2AX-dependent recruitment difference between species unexplained\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Demonstrated that INO80 directly exchanges H2A.Z-H2B for H2A-H2B and organized the human complex into functional modules, establishing histone editing as a core activity built on a conserved catalytic core.\",\n      \"evidence\": \"In vitro histone exchange, genome-wide H2A.Z ChIP-seq, htz1 epistasis, and human subassembly remodeling assays\",\n      \"pmids\": [\"21241891\", \"21303910\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of dimer exchange not yet resolved\", \"How exchange directionality is controlled unknown\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Defined Arp8 nucleosome engagement structurally and linked INO80 ATPase activity to enhanced chromatin mobility and recombination, providing a physical basis for repair-promoting nucleosome dynamics.\",\n      \"evidence\": \"Arp8 C-terminal crystal structure with binding assays; live-cell chromatin mobility imaging with ATPase-dead controls and gene conversion assays\",\n      \"pmids\": [\"23213201\", \"22345518\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether mobility enhancement acts in cis at breaks unresolved\", \"Full Arp8 module-motor coupling not yet mapped\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Dissected internal regulation of the ATPase motor, showing nuclear actin and the Ies2/Ies6/Arp5 subunits separately tune catalysis and nucleosome binding.\",\n      \"evidence\": \"Reconstituted subunit deletions with ATPase and nucleosome-binding assays; actin subdomain mutagenesis with remodeling readout\",\n      \"pmids\": [\"23524535\", \"24297934\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Allosteric path from actin to motor not yet structurally defined\", \"In vivo consequences of each regulatory arm partially characterized\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Established INO80 as a master regulator of open chromatin at pluripotency genes and as a replication-fork factor recruited through ubiquitinated H2A and stabilized by BAP1, tying its remodeling to transcription licensing and tumor-suppressor biology.\",\n      \"evidence\": \"ESC ChIP-seq/ATAC-seq with knockout reprogramming assays; Co-IP with ubiquitinated H2A, fork ChIP, and BAP1 knockout in mammalian cells\",\n      \"pmids\": [\"24792115\", \"25283999\", \"25066231\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct readers of ubiquitinated H2A within INO80 not identified\", \"Mechanism of BAP1-INO80 stabilization not biochemically resolved\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Showed that H2A.Z removal is the primary HR-promoting function of mammalian INO80 and defined the Arp8/Arp4/actin module as a DNA-length sensor controlling sliding, while linking INO80 to RNAPII turnover under transcription stress.\",\n      \"evidence\": \"H2A.Z co-depletion rescue of HR; Arp module DNA-binding mutagenesis with ATPase/sliding uncoupling; ternary INO80-RNAPII-Cdc48 Co-IP and ubiquitinated Rpb1 ChIP\",\n      \"pmids\": [\"26142279\", \"30120252\", \"26656161\", \"25964121\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How linker-DNA sensing engages Arp5 at the acidic patch only partially resolved\", \"Generality of RNAPII eviction across genes unknown\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Defined kinetic and structural coupling within the complex and extended INO80's transcriptional and replication-protective roles to superenhancers and prevention of transcription-replication conflict.\",\n      \"evidence\": \"Arp5/Ies6/Ies2 reconstitution and IP6 inhibition assays; superenhancer ChIP-seq/ATAC-seq with xenografts; Mec1/INO80/PAF1 ChIP-seq and fiber assays\",\n      \"pmids\": [\"26306040\", \"27255055\", \"27257055\", \"27340176\", \"26798134\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological role of IP6 regulation in vivo not established\", \"How INO80 distinguishes conflict-prone loci unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Resolved the unusual entry-site translocation mechanism and intercomplex cooperativity, and identified Rvb1/Rvb2 chaperone behavior plus a presynaptic-filament HR function, defining how INO80 couples ATP hydrolysis to both sliding and histone editing.\",\n      \"evidence\": \"Photo-crosslinking and substrate-specific sliding/exchange assays; Ino80CTD dimerization biochemistry; Rvb1/Rvb2 cryo-EM and ATPase stimulation; htz1 epistasis rescuing presynaptic filaments\",\n      \"pmids\": [\"28604691\", \"28585918\", \"28591576\", \"28514650\", \"24029917\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Atomic structure of the engaged motor not yet available\", \"How dimer cooperativity operates in chromatin not established\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Delivered near-atomic cryo-EM structures of INO80-nucleosome complexes establishing the Rvb1/Rvb2 stator, SHL-6 motor engagement, and Arp5-Ies6 counter-grip ratchet, and resolved the Arp8/HSA linker-binding architecture and single-molecule switch behavior.\",\n      \"evidence\": \"Cryo-EM of fungal and human INO80-nucleosome complexes with biochemical validation; Arp8 module crystal structure; single-molecule sliding kinetics\",\n      \"pmids\": [\"29643509\", \"29643506\", \"29323271\", \"30177756\", \"29452642\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational trajectory of the ratchet during translocation only inferred\", \"How auto-inhibitory Nhp10 module integrates with motor not fully mapped\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Showed INO80 resolves R-loops to protect replication and is essential for HR-mediated DSB repair in neural progenitors, dissociating its repair role from its transcriptional role in vivo.\",\n      \"evidence\": \"R-loop immunofluorescence, RNase H1 rescue, and LacO tethering; conditional cortical Ino80 knockout with in vivo HR assay and Brca2 comparison\",\n      \"pmids\": [\"32913330\", \"32737294\", \"31919190\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether INO80 acts on R-loops directly or via nucleosome remodeling not separated\", \"Substrate context determining repair vs transcription role unclear\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Established that INO80 reads DNA mechanical properties through core-Arp8 module allostery to position the +1 nucleosome genome-wide, and revealed a context-dependent H2A.Z deposition role in primed pluripotency.\",\n      \"evidence\": \"Genome-wide chromatin reconstitution with module deletions and DNA shape analysis; conditional knockout ChIP-seq in naive vs primed ESCs\",\n      \"pmids\": [\"34050142\", \"34139016\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular switch between H2A.Z removal and deposition not defined\", \"How general regulatory factors physically cue +1 positioning unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined hexasome as a preferred and structurally distinct substrate that activates INO80 through an exposed H3-H4 interface, and linked INO80 to PRC2/SUZ12 recruitment at bivalent promoters.\",\n      \"evidence\": \"Hexasome vs nucleosome sliding assays with MNase-seq; cryo-EM of INO80-hexasome spin-rotated mode; INO80-SUZ12 Co-IP and ChIP-seq in spermatocytes\",\n      \"pmids\": [\"35597239\", \"37384673\", \"35006254\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In vivo prevalence of hexasome remodeling not quantified\", \"Direct INO80-SUZ12 interface not mapped\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined post-translational control of INO80 abundance, linking nutrient signaling through TORC1/Rpd3L deacetylation at K929 to protection from autophagic degradation and chromatin-based autophagy gene control.\",\n      \"evidence\": \"Ino80 K929 mutagenesis, protein stability assays, ChIP, and TORC1/Rpd3L genetic epistasis\",\n      \"pmids\": [\"36888706\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct deacetylation of Ino80 by Rpd3L inferred from in vivo data\", \"Whether acetylation regulates remodeling activity itself unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How INO80 selects between H2A.Z removal versus deposition, and how its single conserved catalytic mechanism is locally directed toward repair, replication-fork protection, or transcription at specific genomic sites, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model reconciling H2A.Z eviction and context-dependent deposition\", \"Site-specific switching between functional outputs not mechanistically defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [2, 22, 35, 54]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [16, 33, 26]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [28, 39, 47]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [2, 22]},\n      {\"term_id\": \"GO:0042393\", \"supporting_discovery_ids\": [20, 51]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [5, 24, 32]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [2, 11, 21]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [0, 9, 18]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [0, 4, 26, 34, 55]},\n      {\"term_id\": \"R-HSA-69306\", \"supporting_discovery_ids\": [9, 10, 23, 43, 44]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [5, 24, 32]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [16, 47, 49]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [3, 8, 18]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [24, 41, 45]}\n    ],\n    \"complexes\": [\"INO80 chromatin remodeling complex\"],\n    \"partners\": [\"RUVBL1\", \"RUVBL2\", \"ARP5\", \"ARP8\", \"YY1\", \"BAP1\", \"SUZ12\", \"TRIM3\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":10,"faith_total":10,"faith_pct":100.0}}