{"gene":"CENPI","run_date":"2026-06-09T22:57:18","timeline":{"discoveries":[{"year":1997,"finding":"Mis6 (CENP-I ortholog in S. pombe) localizes to centromeres throughout the cell cycle and is required during G1/S phase to establish specialized inner centromere chromatin (disrupted micrococcal nuclease pattern in mutants) necessary for faithful sister chromatid segregation.","method":"Temperature-sensitive mutant analysis, minichromosome loss assay, micrococcal nuclease chromatin analysis, fluorescence microscopy","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (genetic, chromatin biochemistry, microscopy) in a foundational study, independently replicated by subsequent work","pmids":["9230309"],"is_preprint":false},{"year":2000,"finding":"Fission yeast Mis6 is required for loading SpCENP-A onto inner centromere chromatin; in mis6 mutants SpCENP-A fails to localize to centromeres, establishing Mis6 as upstream of CENP-A deposition.","method":"Genetic epistasis, immunofluorescence of SpCENP-A localization in mis6 mutants, cell cycle analysis","journal":"Science","confidence":"High","confidence_rationale":"Tier 2 / Strong — epistasis combined with fluorescence localization, replicated across multiple subsequent studies","pmids":["10864871"],"is_preprint":false},{"year":1999,"finding":"Fission yeast Mis6 (together with Mis12) is required for correct metaphase spindle length; mis6 mutants show 35–60% extension of metaphase spindle length, indicating a role in spindle morphogenesis through proper sister centromere connection.","method":"Fluorescence microscopy of spindle length in mis6 mutants, suppressor analysis","journal":"Genes & development","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean loss-of-function with defined cellular phenotype, single lab","pmids":["10398680"],"is_preprint":false},{"year":2002,"finding":"Human CENP-I is a constitutive kinetochore component that co-localizes with CENP-A, -C, and -H throughout the cell cycle; conditional knockout in chicken DT40 cells shows that CENP-I (and CENP-H) is required for centromeric localization of CENP-C but not CENP-A.","method":"Conditional gene knockout (DT40), immunocytochemistry, co-localization studies","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean conditional KO with defined molecular phenotype (CENP-C delocalization), multiple orthogonal methods","pmids":["11970896"],"is_preprint":false},{"year":2002,"finding":"Budding yeast Ctf3p (CENP-I ortholog) interacts with Mcm22p and Mcm16p and binds centromere DNA in a Ctf19p-dependent manner; unlike fission yeast Mis6, Ctf3p is not required for loading of Cse4p (CENP-A homolog), but Ctf3p and Ctf19p require Cse4p for proper centromere binding.","method":"Two-hybrid screen, chromatin immunoprecipitation (ChIP), genetic synthetic dosage lethality screen, fluorescence microscopy","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — ChIP demonstrating direct centromere binding, genetic epistasis, protein interaction assays","pmids":["11782448"],"is_preprint":false},{"year":2003,"finding":"Human CENP-I depletion from kinetochores causes loss of CENP-F, MAD1, and MAD2 localization at kinetochores, a G2 delay, and failure to arrest mitosis despite unattached kinetochores; MAD2-dependent mitotic delay requires a collective threshold from many unattached kinetochores.","method":"Antibody microinjection/depletion, immunofluorescence, time-lapse microscopy, spindle checkpoint assays","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean depletion with multiple defined molecular phenotypes, replicated independently","pmids":["12640463"],"is_preprint":false},{"year":2005,"finding":"The fission yeast Mis6-complex physically interacts with Mad2 when the spindle checkpoint is activated, and is required (along with the Nuf2-complex) for Mad2 accumulation at unattached kinetochores; N-terminal fragments of Mis6 localize along the mitotic spindle, suggesting microtubule-binding capacity.","method":"Co-immunoprecipitation, fluorescence microscopy of Mad2/Bub1 localization in mis6 mutants, ectopic expression of Mis6 fragments","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — reciprocal co-IP plus loss-of-function localization assay, single lab","pmids":["15930132"],"is_preprint":false},{"year":2011,"finding":"The Mis6-Mal2-Sim4 complex in fission yeast contains 12 subunits including Mis17, which acts as a regulatory module; Mis17 is hyperphosphorylated by multiple kinases (AMPK, Yak1, Ark1, Ssk2, P-TEFb), and its overproduction causes dominant-negative missegregation and disrupts CenH3/CENP-A recruitment without delocalizing existing Mis6 or CenH3.","method":"Mass spectrometry, FLAG/TAP pulldowns, kinase-deletion mutant analysis, dominant-negative overexpression, chromosome segregation assays","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-identified complex composition, genetic and biochemical follow-up, single lab","pmids":["21445296"],"is_preprint":false},{"year":2014,"finding":"Human CENP-I is required for stable association of the RZZ complex and Mad1 with kinetochores and inhibits their dynein-mediated removal; Aurora B regulates RZZ/Mad1 association while CENP-I inhibits dissociation, together forming a molecular switch that maintains spindle checkpoint signal at prometaphase kinetochores.","method":"siRNA depletion, immunofluorescence, live-cell imaging, epistasis analysis with Aurora B inhibition","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — two orthogonal loss-of-function approaches (CENP-I depletion + Aurora B inhibition) with defined molecular phenotypes, clear epistasis established","pmids":["24862574"],"is_preprint":false},{"year":2014,"finding":"Fission yeast Eic1 links the Mis18 complex with the CCAN/Mis6/Ctf19 complex by interacting with Fta7 (CENP-Q/Okp1) and other CCAN subunits, thereby enabling temporally regulated recruitment of Mis18/Scm3(HJURP) CENP-A loading factors to centromeres.","method":"Co-immunoprecipitation, mass spectrometry, conditional depletion, localization assays","journal":"Open biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS and co-IP identifying complex connections, single lab","pmids":["24789708"],"is_preprint":false},{"year":2014,"finding":"Fission yeast Kis1/Eic1/Mis19 is required to maintain Mis6/CENP-I and Cnp1/CENP-A at kinetochores; loss of Kis1 causes delocalization of Mis6 and CENP-A and defective kinetochore-microtubule attachment with spindle defects.","method":"Forward genetic screen, fluorescence microscopy, conditional depletion","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic screen with defined localization phenotype for Mis6, single lab","pmids":["25375240"],"is_preprint":false},{"year":2015,"finding":"Human CENP-I can recruit M18BP1 to centromeres (independently of CENP-C) and thereby enhance CENP-A assembly; tethering experiments showed CENP-I induces de novo CENP-A assembly at ectopic sites by first recruiting CENP-C and then M18BP1.","method":"Tethering of tetR-fusion proteins to synthetic alphoid(tetO) HAC array, immunofluorescence, CENP-A ChIP","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — tethering/ectopic recruitment assay with defined downstream factors, single lab","pmids":["26527398"],"is_preprint":false},{"year":2019,"finding":"The mRNA decay pathway (exo2/pan2 ribonucleases) negatively regulates Mis17-Mis6 complex levels (affecting Mis17 protein stability), while the EKC/KEOPS complex negatively regulates centromeric localization of Mis6 and CENP-A independently of Mis17 protein levels, through mechanisms involving kinase activity.","method":"Whole-genome suppressor sequencing, double mutant analysis, Western blot of Mis17 levels, centromere localization assays","journal":"G3 (Bethesda, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — suppressor screen with mechanistic follow-up (protein levels + localization), single lab","pmids":["30967422"],"is_preprint":false},{"year":2020,"finding":"Human CENP-I contains a conserved helix (α11) that forms intramolecular interactions with N-terminal HEAT repeats; deletion of this helix causes protein aggregation in vitro and dramatically reduces interaction with CENP-H and CENP-M; mutations in conserved residues on this helix specifically weaken binding to CENP-M but not CENP-H in HeLa cells.","method":"In vitro protein aggregation assay, co-immunoprecipitation in HeLa cells, mutagenesis, structural analysis of fungal CENP-I","journal":"Journal of molecular recognition","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro reconstitution with mutagenesis but single lab and limited structural validation","pmids":["32017295"],"is_preprint":false},{"year":2021,"finding":"Budding yeast Ctf3/CENP-I provides a docking site for the desumoylase Ulp2 at the kinetochore; a conserved surface of Ctf3 binds Ulp2, and Ctf3 mutations that disable Ulp2 recruitment cause elevated inner kinetochore sumoylation and defective chromosome segregation.","method":"Protein interaction mapping, Ctf3 surface mutations, sumoylation assays, chromosome segregation assays","journal":"The Journal of cell biology","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — direct binding site identified by mutagenesis, functional consequence (sumoylation + segregation defect), single lab","pmids":["34081091"],"is_preprint":false},{"year":2021,"finding":"Loss of human CENP-I impairs homologous recombination (HR) DNA double-strand break repair while having no effect on non-homologous end-joining (NHEJ); CENP-I loss increases endogenous 53BP1 foci and R-loop formation, and RNaseH1 expression restores HR capacity in CENP-I-deficient cells.","method":"siRNA knockdown, HR/NHEJ reporter assays, 53BP1 foci quantification, ionizing radiation survival, RNaseH1 rescue experiment","journal":"Cancers","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal functional assays (HR reporters, foci, rescue), single lab","pmids":["34206916"],"is_preprint":false},{"year":2022,"finding":"Fission yeast Mis6 (CENP-I) and Mis15 (CENP-N) are required during mitosis to retain pre-existing CENP-A at centromeres by suppressing RNA Pol II-dependent non-coding transcription at the central core; inhibition of RNA Pol II rescues CENP-A loss in mis6 mutant cells.","method":"Conditional depletion during mitosis, CENP-A localization assays, non-coding RNA quantification, RNA Pol II inhibition rescue experiment","journal":"Communications biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional depletion with mechanistic rescue by Pol II inhibition, single lab","pmids":["35970865"],"is_preprint":false},{"year":2023,"finding":"Human CENP-I directly interacts with centromeric DNA, preferentially recognizing AT-rich elements via a consecutive DNA-binding surface formed by conserved charged residues at the end of N-terminal HEAT repeats; DNA binding-deficient mutants retain interaction with CENP-H/K and CENP-M but show severely diminished centromeric localization, impaired chromosome alignment, and failure to load newly synthesized CENP-A; CENP-I stabilizes CENP-A nucleosomes by binding nucleosomal DNA rather than histones.","method":"In vitro DNA binding assay, mutagenesis of DNA-binding surface, co-immunoprecipitation, CENP-A loading assay, chromosome alignment assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with mutagenesis, multiple functional readouts (localization, CENP-A loading, chromosome alignment), single lab","pmids":["36888657"],"is_preprint":false},{"year":2025,"finding":"FOXM1 acts as a transcriptional activator of CENPI in glioblastoma cells, as demonstrated by JASPAR prediction, dual luciferase reporter assay, and ChIP; CENPI overexpression or exogenous L-Arg/L-Pro rescues the reduced proline and arginine metabolism caused by FOXM1 knockdown.","method":"Dual luciferase reporter assay, ChIP, siRNA knockdown, metabolic assays","journal":"Journal of neuropathology and experimental neurology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, limited mechanistic follow-up linking FOXM1 to CENPI transcription, no direct protein-level mechanistic insight into CENPI function","pmids":["41206587"],"is_preprint":false}],"current_model":"CENP-I (Mis6/Ctf3) is a constitutive inner kinetochore HEAT-repeat protein that directly binds AT-rich centromeric DNA to stabilize CENP-A nucleosomes and promote new CENP-A deposition; it acts upstream of CENP-C localization (but not CENP-A), recruits M18BP1 to facilitate HJURP-mediated CENP-A loading, maintains CENP-A against mitotic non-coding transcription, provides a docking site for the desumoylase Ulp2 to control inner kinetochore sumoylation, and serves as a platform for spindle checkpoint proteins (MAD1, MAD2, RZZ) at unattached kinetochores while antagonizing dynein-mediated checkpoint protein stripping—together with Aurora B forming a molecular switch for robust spindle assembly checkpoint signaling; additionally, CENP-I participates in homologous recombination DNA repair by suppressing R-loop accumulation."},"narrative":{"mechanistic_narrative":"CENP-I (Mis6/Ctf3) is a constitutive inner kinetochore protein that builds and maintains the centromeric chromatin platform required for faithful chromosome segregation [PMID:9230309, PMID:11970896]. It directly binds AT-rich centromeric DNA through a consecutive surface formed by conserved charged residues at the end of its N-terminal HEAT repeats, and this DNA contact—rather than histone binding—drives its own centromeric localization, stabilizes CENP-A nucleosomes, and enables loading of newly synthesized CENP-A [PMID:36888657]. CENP-I acts upstream of CENP-A deposition in fission yeast and of CENP-C localization in vertebrate cells, while being dispensable for CENP-A localization itself in vertebrates [PMID:10864871, PMID:11970896], and it promotes CENP-A assembly by recruiting CENP-C and M18BP1 to enable HJURP/Mis18-mediated loading [PMID:26527398]. It also protects pre-existing CENP-A during mitosis by suppressing RNA Pol II-dependent non-coding transcription at the central core [PMID:35970865]. Within the kinetochore, CENP-I assembles through conserved HEAT-repeat contacts with CENP-H, CENP-K, and CENP-M [PMID:32017295, PMID:36888657] and provides a docking site for the desumoylase Ulp2 that controls inner kinetochore sumoylation [PMID:34081091]. Beyond architecture, CENP-I sustains spindle assembly checkpoint signaling: it is required for kinetochore association of MAD1, MAD2, CENP-F, and the RZZ complex, and—together with Aurora B—forms a molecular switch that retains checkpoint proteins by antagonizing their dynein-mediated stripping at unattached kinetochores [PMID:12640463, PMID:24862574]. Independently of its kinetochore role, human CENP-I promotes homologous-recombination repair of DNA double-strand breaks by suppressing R-loop accumulation, as RNaseH1 expression restores HR in CENP-I-deficient cells [PMID:34206916].","teleology":[{"year":1997,"claim":"Established that the CENP-I ortholog Mis6 is a constitutive centromere component acting in G1/S to build specialized inner centromere chromatin, identifying it as a foundational determinant of centromere identity rather than a transient mitotic factor.","evidence":"Temperature-sensitive mutant analysis with minichromosome loss and micrococcal nuclease chromatin profiling in fission yeast","pmids":["9230309"],"confidence":"High","gaps":["Molecular nature of the chromatin alteration undefined","No direct DNA- or protein-binding activity demonstrated","Relationship to CENP-A not yet tested"]},{"year":1999,"claim":"Linked Mis6 function to mitotic spindle morphogenesis, showing centromere defects translate into aberrant metaphase spindle geometry.","evidence":"Fluorescence microscopy of spindle length and suppressor analysis in mis6 mutants","pmids":["10398680"],"confidence":"Medium","gaps":["Spindle defect likely indirect via centromere structure","Mechanism connecting centromere to spindle length unresolved"]},{"year":2000,"claim":"Placed Mis6 upstream of CENP-A deposition, defining its role in establishing the epigenetic centromere mark.","evidence":"Genetic epistasis and immunofluorescence of SpCENP-A in mis6 mutants","pmids":["10864871"],"confidence":"High","gaps":["Did not distinguish establishment versus maintenance of CENP-A","Direct loading mechanism unknown"]},{"year":2002,"claim":"Defined the vertebrate kinetochore hierarchy, showing human/chicken CENP-I is required for CENP-C but not CENP-A localization, revealing species differences in the CENP-A dependency relationship.","evidence":"Conditional knockout in DT40 cells with co-localization immunocytochemistry","pmids":["11970896"],"confidence":"High","gaps":["Direct versus indirect requirement for CENP-C not separated","DNA-binding basis of localization not addressed"]},{"year":2002,"claim":"Identified the budding yeast Ctf3 sub-complex (with Mcm22/Mcm16) and showed Cse4/CENP-A-dependent, Ctf19-dependent centromere binding, mapping CENP-I into the CCAN interaction network.","evidence":"Two-hybrid, ChIP, synthetic dosage lethality, and microscopy in budding yeast","pmids":["11782448"],"confidence":"High","gaps":["Ctf3 dispensable for Cse4 loading, contrasting fission yeast — unifying mechanism unclear","Direct DNA contact versus indirect recruitment not resolved"]},{"year":2003,"claim":"Connected CENP-I to spindle checkpoint competence, showing its depletion abolishes kinetochore MAD1/MAD2/CENP-F and impairs mitotic arrest at unattached kinetochores.","evidence":"Antibody depletion, immunofluorescence, time-lapse imaging and checkpoint assays in human cells","pmids":["12640463"],"confidence":"High","gaps":["Whether checkpoint defect is architectural or signaling-specific not separated","Mechanism of MAD1/MAD2 recruitment unknown"]},{"year":2005,"claim":"Provided physical evidence that the Mis6 complex engages Mad2 upon checkpoint activation and is required for Mad2 accumulation at unattached kinetochores.","evidence":"Reciprocal co-IP and Mad2/Bub1 localization in mis6 mutants, plus Mis6 fragment localization in fission yeast","pmids":["15930132"],"confidence":"Medium","gaps":["Microtubule-binding by N-terminal fragments not functionally validated","Direct versus indirect Mad2 interaction unresolved","Single lab"]},{"year":2014,"claim":"Resolved the CENP-I checkpoint mechanism, showing it retains RZZ/Mad1 by inhibiting dynein-mediated stripping while Aurora B regulates their association — a two-input molecular switch for robust SAC signaling.","evidence":"siRNA depletion with Aurora B inhibition epistasis, immunofluorescence and live imaging in human cells","pmids":["24862574"],"confidence":"High","gaps":["Direct molecular target of CENP-I in blocking stripping unknown","Structural basis of the switch undefined"]},{"year":2014,"claim":"Connected CENP-I/CCAN to the CENP-A loading machinery via Eic1/Mis19, showing physical bridging of Mis18/HJURP factors to the Mis6/Ctf19 complex enables temporally regulated CENP-A deposition.","evidence":"Co-IP, mass spectrometry, conditional depletion and localization in fission yeast","pmids":["24789708","25375240"],"confidence":"Medium","gaps":["Direct CENP-I contact with Eic1 versus other CCAN subunits unmapped","Maintenance versus establishment role of the bridge unclear"]},{"year":2015,"claim":"Demonstrated CENP-I can drive de novo CENP-A assembly by recruiting CENP-C and then M18BP1, defining a CENP-C-independent route to M18BP1 recruitment.","evidence":"Tethering to synthetic alphoid(tetO) HAC array with immunofluorescence and CENP-A ChIP","pmids":["26527398"],"confidence":"Medium","gaps":["Ectopic tethering may not reflect endogenous order of events","Direct M18BP1 binding not shown","Single lab"]},{"year":2020,"claim":"Mapped CENP-I intramolecular architecture, showing a conserved α11 helix folds against N-terminal HEAT repeats to maintain folding and CENP-H/CENP-M binding.","evidence":"In vitro aggregation assays, co-IP, mutagenesis and fungal structural analysis","pmids":["32017295"],"confidence":"Medium","gaps":["Human full-length structure not solved","Functional consequence of α11 mutations in cells limited"]},{"year":2021,"claim":"Identified a non-kinetochore function: CENP-I promotes homologous-recombination DSB repair by limiting R-loop formation, with RNaseH1 rescuing HR in its absence.","evidence":"siRNA, HR/NHEJ reporters, 53BP1 foci, IR survival and RNaseH1 rescue in human cells","pmids":["34206916"],"confidence":"Medium","gaps":["Whether CENP-I acts directly at break sites or via centromere transcription unclear","Mechanism of R-loop suppression undefined","Single lab"]},{"year":2021,"claim":"Revealed CENP-I as a desumoylase docking platform, with a conserved Ctf3 surface binding Ulp2 to control inner kinetochore sumoylation and segregation fidelity.","evidence":"Interaction mapping, surface mutagenesis, sumoylation and segregation assays in budding yeast","pmids":["34081091"],"confidence":"High","gaps":["Conservation of the Ulp2-docking role in humans untested","Sumoylation substrates at the kinetochore not enumerated"]},{"year":2022,"claim":"Showed CENP-I maintains pre-existing CENP-A during mitosis by suppressing RNA Pol II non-coding transcription, since Pol II inhibition rescues CENP-A loss in mis6 mutants.","evidence":"Conditional mitotic depletion, CENP-A localization, ncRNA quantification and Pol II inhibition rescue in fission yeast","pmids":["35970865"],"confidence":"Medium","gaps":["Direct mechanism of transcriptional suppression by CENP-I unknown","Link to its DNA-binding activity not tested"]},{"year":2023,"claim":"Established the central biochemical activity: CENP-I directly binds AT-rich centromeric DNA via charged residues at its N-terminal HEAT repeats, and this DNA contact—not histone binding—drives localization, CENP-A nucleosome stabilization, CENP-A loading and chromosome alignment.","evidence":"In vitro DNA-binding assays, DNA-surface mutagenesis, co-IP, CENP-A loading and chromosome alignment assays in human cells","pmids":["36888657"],"confidence":"High","gaps":["Structure of the CENP-I–DNA complex not solved","How DNA binding integrates with CCAN assembly and transcription suppression unresolved"]},{"year":2025,"claim":"Placed CENPI under FOXM1 transcriptional control with links to proline/arginine metabolism in glioblastoma, extending it into a cancer regulatory context.","evidence":"JASPAR prediction, dual luciferase reporter, ChIP, siRNA and metabolic assays in glioblastoma cells","pmids":["41206587"],"confidence":"Low","gaps":["No direct protein-level mechanism linking CENPI to metabolism","Single lab, correlative","Generality beyond glioblastoma untested"]},{"year":null,"claim":"How CENP-I's direct DNA-binding activity is mechanistically integrated with CCAN assembly, transcriptional suppression of the centromere core, checkpoint protein retention, and its R-loop-suppressing DNA-repair role remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No high-resolution structure of human CENP-I bound to centromeric DNA or nucleosome","Mechanistic basis of dynein-stripping antagonism unknown","Whether the HR/R-loop function is separable from the kinetochore role untested"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[4,17]}],"localization":[{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[0,3,17]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[3,11]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[3,5,8]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[15]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[1,11,16,17]}],"complexes":["CCAN/Mis6-Ctf19 complex","kinetochore"],"partners":["CENP-C","CENP-H","CENP-M","CENP-K","M18BP1","MAD2","ULP2","MCM22"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q92674","full_name":"Centromere protein I","aliases":["FSH primary response protein 1","Follicle-stimulating hormone primary response protein","Interphase centromere complex protein 19","Leucine-rich primary response protein 1"],"length_aa":756,"mass_kda":86.7,"function":"Component of the CENPA-CAD (nucleosome distal) complex, a complex recruited to centromeres which is involved in assembly of kinetochore proteins, mitotic progression and chromosome segregation. May be involved in incorporation of newly synthesized CENPA into centromeres via its interaction with the CENPA-NAC complex. Required for the localization of CENPF, MAD1L1 and MAD2 (MAD2L1 or MAD2L2) to kinetochores. Involved in the response of gonadal tissues to follicle-stimulating hormone","subcellular_location":"Nucleus; Chromosome, centromere","url":"https://www.uniprot.org/uniprotkb/Q92674/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/CENPI","classification":"Common Essential","n_dependent_lines":852,"n_total_lines":1208,"dependency_fraction":0.7052980132450332},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/CENPI","total_profiled":1310},"omim":[{"mim_id":"611511","title":"MLF1-INTERACTING PROTEIN; MLF1IP","url":"https://www.omim.org/entry/611511"},{"mim_id":"611506","title":"CENTROMERIC PROTEIN Q; CENPQ","url":"https://www.omim.org/entry/611506"},{"mim_id":"611505","title":"CENTROMERIC PROTEIN P; CENPP","url":"https://www.omim.org/entry/611505"},{"mim_id":"611504","title":"CENTROMERIC PROTEIN O; CENPO","url":"https://www.omim.org/entry/611504"},{"mim_id":"611503","title":"CENTROMERIC PROTEIN L; CENPL","url":"https://www.omim.org/entry/611503"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nuclear bodies","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"bone marrow","ntpm":5.2},{"tissue":"lymphoid tissue","ntpm":4.9}],"url":"https://www.proteinatlas.org/search/CENPI"},"hgnc":{"alias_symbol":["LRPR1","CENP-I","Mis6"],"prev_symbol":["FSHPRH1"]},"alphafold":{"accession":"Q92674","domains":[{"cath_id":"-","chopping":"687-737","consensus_level":"medium","plddt":82.6702,"start":687,"end":737},{"cath_id":"1.25.40","chopping":"85-248","consensus_level":"medium","plddt":83.1731,"start":85,"end":248}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q92674","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q92674-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q92674-F1-predicted_aligned_error_v6.png","plddt_mean":73.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CENPI","jax_strain_url":"https://www.jax.org/strain/search?query=CENPI"},"sequence":{"accession":"Q92674","fasta_url":"https://rest.uniprot.org/uniprotkb/Q92674.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q92674/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q92674"}},"corpus_meta":[{"pmid":"10864871","id":"PMC_10864871","title":"Requirement of Mis6 centromere connector for localizing a CENP-A-like protein in fission yeast.","date":"2000","source":"Science (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/10864871","citation_count":341,"is_preprint":false},{"pmid":"9230309","id":"PMC_9230309","title":"Mis6, a fission yeast inner centromere protein, acts during G1/S and forms specialized chromatin required for equal segregation.","date":"1997","source":"Cell","url":"https://pubmed.ncbi.nlm.nih.gov/9230309","citation_count":198,"is_preprint":false},{"pmid":"10398680","id":"PMC_10398680","title":"Proper metaphase spindle length is determined by centromere proteins Mis12 and Mis6 required for faithful chromosome segregation.","date":"1999","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/10398680","citation_count":179,"is_preprint":false},{"pmid":"12640463","id":"PMC_12640463","title":"Human CENP-I specifies localization of CENP-F, MAD1 and MAD2 to kinetochores and is essential for mitosis.","date":"2003","source":"Nature cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/12640463","citation_count":129,"is_preprint":false},{"pmid":"11970896","id":"PMC_11970896","title":"CENP-I is essential for centromere function in vertebrate cells.","date":"2002","source":"Developmental cell","url":"https://pubmed.ncbi.nlm.nih.gov/11970896","citation_count":118,"is_preprint":false},{"pmid":"11782448","id":"PMC_11782448","title":"Ctf3p, the Mis6 budding yeast homolog, interacts with Mcm22p and Mcm16p at the yeast outer kinetochore.","date":"2002","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/11782448","citation_count":108,"is_preprint":false},{"pmid":"26527398","id":"PMC_26527398","title":"CENP-C and CENP-I are key connecting factors for kinetochore and CENP-A assembly.","date":"2015","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/26527398","citation_count":54,"is_preprint":false},{"pmid":"24862574","id":"PMC_24862574","title":"CENP-I and Aurora B act as a molecular switch that ties RZZ/Mad1 recruitment to kinetochore attachment status.","date":"2014","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/24862574","citation_count":42,"is_preprint":false},{"pmid":"24789708","id":"PMC_24789708","title":"Eic1 links Mis18 with the CCAN/Mis6/Ctf19 complex to promote CENP-A assembly.","date":"2014","source":"Open biology","url":"https://pubmed.ncbi.nlm.nih.gov/24789708","citation_count":37,"is_preprint":false},{"pmid":"15930132","id":"PMC_15930132","title":"Spindle checkpoint signaling requires the mis6 kinetochore subcomplex, which interacts with mad2 and mitotic spindles.","date":"2005","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/15930132","citation_count":31,"is_preprint":false},{"pmid":"29936263","id":"PMC_29936263","title":"CENPI is overexpressed in colorectal cancer and regulates cell migration and invasion.","date":"2018","source":"Gene","url":"https://pubmed.ncbi.nlm.nih.gov/29936263","citation_count":25,"is_preprint":false},{"pmid":"21445296","id":"PMC_21445296","title":"Mis17 is a regulatory module of the Mis6-Mal2-Sim4 centromere complex that is required for the recruitment of CenH3/CENP-A in fission yeast.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/21445296","citation_count":21,"is_preprint":false},{"pmid":"25375240","id":"PMC_25375240","title":"The kinetochore protein Kis1/Eic1/Mis19 ensures the integrity of mitotic spindles through maintenance of kinetochore factors Mis6/CENP-I and CENP-A.","date":"2014","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/25375240","citation_count":16,"is_preprint":false},{"pmid":"21888900","id":"PMC_21888900","title":"Anti-CENPI autoantibodies in scleroderma patients with features of autoimmune liver diseases.","date":"2011","source":"Clinica chimica acta; international journal of clinical chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/21888900","citation_count":16,"is_preprint":false},{"pmid":"8921378","id":"PMC_8921378","title":"Sequence and chromosomal location of a human homologue of LRPR1, an FSH primary response gene.","date":"1996","source":"Genomics","url":"https://pubmed.ncbi.nlm.nih.gov/8921378","citation_count":15,"is_preprint":false},{"pmid":"36888657","id":"PMC_36888657","title":"CENP-I directly targets centromeric DNA to support CENP-A deposition and centromere maintenance.","date":"2023","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/36888657","citation_count":9,"is_preprint":false},{"pmid":"31912435","id":"PMC_31912435","title":"Centromere protein I (CENPI) is a candidate gene for X-linked steroid sensitive nephrotic syndrome.","date":"2020","source":"Journal of nephrology","url":"https://pubmed.ncbi.nlm.nih.gov/31912435","citation_count":8,"is_preprint":false},{"pmid":"37465344","id":"PMC_37465344","title":"Pan-Cancer Analysis Reveals CENPI as a Potential Biomarker and Therapeutic Target in Adrenocortical Carcinoma.","date":"2023","source":"Journal of inflammation research","url":"https://pubmed.ncbi.nlm.nih.gov/37465344","citation_count":8,"is_preprint":false},{"pmid":"34081091","id":"PMC_34081091","title":"Ctf3/CENP-I provides a docking site for the desumoylase Ulp2 at the kinetochore.","date":"2021","source":"The Journal of cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/34081091","citation_count":6,"is_preprint":false},{"pmid":"34206916","id":"PMC_34206916","title":"Loss of CENP-I Impairs Homologous Recombination and Sensitizes Cells to PARP1 Inhibition.","date":"2021","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/34206916","citation_count":5,"is_preprint":false},{"pmid":"9701787","id":"PMC_9701787","title":"Differential regulation of leucine-rich primary response gene 1 (LRPR1) mRNA expression in rat testis and ovary.","date":"1998","source":"Molecular human reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/9701787","citation_count":4,"is_preprint":false},{"pmid":"40406242","id":"PMC_40406242","title":"The m6A reader YTHDF3 promotes TNBC progression by regulating CENPI stabilization.","date":"2025","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/40406242","citation_count":4,"is_preprint":false},{"pmid":"35970865","id":"PMC_35970865","title":"The Mis6 inner kinetochore subcomplex maintains CENP-A nucleosomes against centromeric non-coding transcription during mitosis.","date":"2022","source":"Communications biology","url":"https://pubmed.ncbi.nlm.nih.gov/35970865","citation_count":3,"is_preprint":false},{"pmid":"30967422","id":"PMC_30967422","title":"Negative Regulation of the Mis17-Mis6 Centromere Complex by mRNA Decay Pathway and EKC/KEOPS Complex in Schizosaccharomyces pombe.","date":"2019","source":"G3 (Bethesda, Md.)","url":"https://pubmed.ncbi.nlm.nih.gov/30967422","citation_count":2,"is_preprint":false},{"pmid":"32017295","id":"PMC_32017295","title":"Structural insights into the intramolecular interactions of centromere protein CENP-I.","date":"2020","source":"Journal of molecular recognition : JMR","url":"https://pubmed.ncbi.nlm.nih.gov/32017295","citation_count":2,"is_preprint":false},{"pmid":"40784804","id":"PMC_40784804","title":"[CENPI promotes the migration of liver cancer cells and the epithelial-mesenchymal transition process by activating the RAS/MEK/ERK signaling axis].","date":"2025","source":"Zhonghua gan zang bing za zhi = Zhonghua ganzangbing zazhi = Chinese journal of hepatology","url":"https://pubmed.ncbi.nlm.nih.gov/40784804","citation_count":1,"is_preprint":false},{"pmid":"41206587","id":"PMC_41206587","title":"FOXM1/CENPI axis regulation of proline and arginine metabolism in glioblastoma cells.","date":"2026","source":"Journal of neuropathology and experimental neurology","url":"https://pubmed.ncbi.nlm.nih.gov/41206587","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":14406,"output_tokens":4528,"usd":0.055569,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":12510,"output_tokens":4901,"usd":0.092537,"stage2_stop_reason":"end_turn"},"total_usd":0.148106,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1997,\n      \"finding\": \"Mis6 (CENP-I ortholog in S. pombe) localizes to centromeres throughout the cell cycle and is required during G1/S phase to establish specialized inner centromere chromatin (disrupted micrococcal nuclease pattern in mutants) necessary for faithful sister chromatid segregation.\",\n      \"method\": \"Temperature-sensitive mutant analysis, minichromosome loss assay, micrococcal nuclease chromatin analysis, fluorescence microscopy\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (genetic, chromatin biochemistry, microscopy) in a foundational study, independently replicated by subsequent work\",\n      \"pmids\": [\"9230309\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"Fission yeast Mis6 is required for loading SpCENP-A onto inner centromere chromatin; in mis6 mutants SpCENP-A fails to localize to centromeres, establishing Mis6 as upstream of CENP-A deposition.\",\n      \"method\": \"Genetic epistasis, immunofluorescence of SpCENP-A localization in mis6 mutants, cell cycle analysis\",\n      \"journal\": \"Science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — epistasis combined with fluorescence localization, replicated across multiple subsequent studies\",\n      \"pmids\": [\"10864871\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1999,\n      \"finding\": \"Fission yeast Mis6 (together with Mis12) is required for correct metaphase spindle length; mis6 mutants show 35–60% extension of metaphase spindle length, indicating a role in spindle morphogenesis through proper sister centromere connection.\",\n      \"method\": \"Fluorescence microscopy of spindle length in mis6 mutants, suppressor analysis\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean loss-of-function with defined cellular phenotype, single lab\",\n      \"pmids\": [\"10398680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Human CENP-I is a constitutive kinetochore component that co-localizes with CENP-A, -C, and -H throughout the cell cycle; conditional knockout in chicken DT40 cells shows that CENP-I (and CENP-H) is required for centromeric localization of CENP-C but not CENP-A.\",\n      \"method\": \"Conditional gene knockout (DT40), immunocytochemistry, co-localization studies\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean conditional KO with defined molecular phenotype (CENP-C delocalization), multiple orthogonal methods\",\n      \"pmids\": [\"11970896\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Budding yeast Ctf3p (CENP-I ortholog) interacts with Mcm22p and Mcm16p and binds centromere DNA in a Ctf19p-dependent manner; unlike fission yeast Mis6, Ctf3p is not required for loading of Cse4p (CENP-A homolog), but Ctf3p and Ctf19p require Cse4p for proper centromere binding.\",\n      \"method\": \"Two-hybrid screen, chromatin immunoprecipitation (ChIP), genetic synthetic dosage lethality screen, fluorescence microscopy\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — ChIP demonstrating direct centromere binding, genetic epistasis, protein interaction assays\",\n      \"pmids\": [\"11782448\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Human CENP-I depletion from kinetochores causes loss of CENP-F, MAD1, and MAD2 localization at kinetochores, a G2 delay, and failure to arrest mitosis despite unattached kinetochores; MAD2-dependent mitotic delay requires a collective threshold from many unattached kinetochores.\",\n      \"method\": \"Antibody microinjection/depletion, immunofluorescence, time-lapse microscopy, spindle checkpoint assays\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean depletion with multiple defined molecular phenotypes, replicated independently\",\n      \"pmids\": [\"12640463\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The fission yeast Mis6-complex physically interacts with Mad2 when the spindle checkpoint is activated, and is required (along with the Nuf2-complex) for Mad2 accumulation at unattached kinetochores; N-terminal fragments of Mis6 localize along the mitotic spindle, suggesting microtubule-binding capacity.\",\n      \"method\": \"Co-immunoprecipitation, fluorescence microscopy of Mad2/Bub1 localization in mis6 mutants, ectopic expression of Mis6 fragments\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal co-IP plus loss-of-function localization assay, single lab\",\n      \"pmids\": [\"15930132\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"The Mis6-Mal2-Sim4 complex in fission yeast contains 12 subunits including Mis17, which acts as a regulatory module; Mis17 is hyperphosphorylated by multiple kinases (AMPK, Yak1, Ark1, Ssk2, P-TEFb), and its overproduction causes dominant-negative missegregation and disrupts CenH3/CENP-A recruitment without delocalizing existing Mis6 or CenH3.\",\n      \"method\": \"Mass spectrometry, FLAG/TAP pulldowns, kinase-deletion mutant analysis, dominant-negative overexpression, chromosome segregation assays\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-identified complex composition, genetic and biochemical follow-up, single lab\",\n      \"pmids\": [\"21445296\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Human CENP-I is required for stable association of the RZZ complex and Mad1 with kinetochores and inhibits their dynein-mediated removal; Aurora B regulates RZZ/Mad1 association while CENP-I inhibits dissociation, together forming a molecular switch that maintains spindle checkpoint signal at prometaphase kinetochores.\",\n      \"method\": \"siRNA depletion, immunofluorescence, live-cell imaging, epistasis analysis with Aurora B inhibition\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — two orthogonal loss-of-function approaches (CENP-I depletion + Aurora B inhibition) with defined molecular phenotypes, clear epistasis established\",\n      \"pmids\": [\"24862574\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Fission yeast Eic1 links the Mis18 complex with the CCAN/Mis6/Ctf19 complex by interacting with Fta7 (CENP-Q/Okp1) and other CCAN subunits, thereby enabling temporally regulated recruitment of Mis18/Scm3(HJURP) CENP-A loading factors to centromeres.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, conditional depletion, localization assays\",\n      \"journal\": \"Open biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS and co-IP identifying complex connections, single lab\",\n      \"pmids\": [\"24789708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Fission yeast Kis1/Eic1/Mis19 is required to maintain Mis6/CENP-I and Cnp1/CENP-A at kinetochores; loss of Kis1 causes delocalization of Mis6 and CENP-A and defective kinetochore-microtubule attachment with spindle defects.\",\n      \"method\": \"Forward genetic screen, fluorescence microscopy, conditional depletion\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic screen with defined localization phenotype for Mis6, single lab\",\n      \"pmids\": [\"25375240\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Human CENP-I can recruit M18BP1 to centromeres (independently of CENP-C) and thereby enhance CENP-A assembly; tethering experiments showed CENP-I induces de novo CENP-A assembly at ectopic sites by first recruiting CENP-C and then M18BP1.\",\n      \"method\": \"Tethering of tetR-fusion proteins to synthetic alphoid(tetO) HAC array, immunofluorescence, CENP-A ChIP\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — tethering/ectopic recruitment assay with defined downstream factors, single lab\",\n      \"pmids\": [\"26527398\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"The mRNA decay pathway (exo2/pan2 ribonucleases) negatively regulates Mis17-Mis6 complex levels (affecting Mis17 protein stability), while the EKC/KEOPS complex negatively regulates centromeric localization of Mis6 and CENP-A independently of Mis17 protein levels, through mechanisms involving kinase activity.\",\n      \"method\": \"Whole-genome suppressor sequencing, double mutant analysis, Western blot of Mis17 levels, centromere localization assays\",\n      \"journal\": \"G3 (Bethesda, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — suppressor screen with mechanistic follow-up (protein levels + localization), single lab\",\n      \"pmids\": [\"30967422\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Human CENP-I contains a conserved helix (α11) that forms intramolecular interactions with N-terminal HEAT repeats; deletion of this helix causes protein aggregation in vitro and dramatically reduces interaction with CENP-H and CENP-M; mutations in conserved residues on this helix specifically weaken binding to CENP-M but not CENP-H in HeLa cells.\",\n      \"method\": \"In vitro protein aggregation assay, co-immunoprecipitation in HeLa cells, mutagenesis, structural analysis of fungal CENP-I\",\n      \"journal\": \"Journal of molecular recognition\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro reconstitution with mutagenesis but single lab and limited structural validation\",\n      \"pmids\": [\"32017295\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Budding yeast Ctf3/CENP-I provides a docking site for the desumoylase Ulp2 at the kinetochore; a conserved surface of Ctf3 binds Ulp2, and Ctf3 mutations that disable Ulp2 recruitment cause elevated inner kinetochore sumoylation and defective chromosome segregation.\",\n      \"method\": \"Protein interaction mapping, Ctf3 surface mutations, sumoylation assays, chromosome segregation assays\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct binding site identified by mutagenesis, functional consequence (sumoylation + segregation defect), single lab\",\n      \"pmids\": [\"34081091\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Loss of human CENP-I impairs homologous recombination (HR) DNA double-strand break repair while having no effect on non-homologous end-joining (NHEJ); CENP-I loss increases endogenous 53BP1 foci and R-loop formation, and RNaseH1 expression restores HR capacity in CENP-I-deficient cells.\",\n      \"method\": \"siRNA knockdown, HR/NHEJ reporter assays, 53BP1 foci quantification, ionizing radiation survival, RNaseH1 rescue experiment\",\n      \"journal\": \"Cancers\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal functional assays (HR reporters, foci, rescue), single lab\",\n      \"pmids\": [\"34206916\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Fission yeast Mis6 (CENP-I) and Mis15 (CENP-N) are required during mitosis to retain pre-existing CENP-A at centromeres by suppressing RNA Pol II-dependent non-coding transcription at the central core; inhibition of RNA Pol II rescues CENP-A loss in mis6 mutant cells.\",\n      \"method\": \"Conditional depletion during mitosis, CENP-A localization assays, non-coding RNA quantification, RNA Pol II inhibition rescue experiment\",\n      \"journal\": \"Communications biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional depletion with mechanistic rescue by Pol II inhibition, single lab\",\n      \"pmids\": [\"35970865\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Human CENP-I directly interacts with centromeric DNA, preferentially recognizing AT-rich elements via a consecutive DNA-binding surface formed by conserved charged residues at the end of N-terminal HEAT repeats; DNA binding-deficient mutants retain interaction with CENP-H/K and CENP-M but show severely diminished centromeric localization, impaired chromosome alignment, and failure to load newly synthesized CENP-A; CENP-I stabilizes CENP-A nucleosomes by binding nucleosomal DNA rather than histones.\",\n      \"method\": \"In vitro DNA binding assay, mutagenesis of DNA-binding surface, co-immunoprecipitation, CENP-A loading assay, chromosome alignment assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with mutagenesis, multiple functional readouts (localization, CENP-A loading, chromosome alignment), single lab\",\n      \"pmids\": [\"36888657\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FOXM1 acts as a transcriptional activator of CENPI in glioblastoma cells, as demonstrated by JASPAR prediction, dual luciferase reporter assay, and ChIP; CENPI overexpression or exogenous L-Arg/L-Pro rescues the reduced proline and arginine metabolism caused by FOXM1 knockdown.\",\n      \"method\": \"Dual luciferase reporter assay, ChIP, siRNA knockdown, metabolic assays\",\n      \"journal\": \"Journal of neuropathology and experimental neurology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, limited mechanistic follow-up linking FOXM1 to CENPI transcription, no direct protein-level mechanistic insight into CENPI function\",\n      \"pmids\": [\"41206587\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CENP-I (Mis6/Ctf3) is a constitutive inner kinetochore HEAT-repeat protein that directly binds AT-rich centromeric DNA to stabilize CENP-A nucleosomes and promote new CENP-A deposition; it acts upstream of CENP-C localization (but not CENP-A), recruits M18BP1 to facilitate HJURP-mediated CENP-A loading, maintains CENP-A against mitotic non-coding transcription, provides a docking site for the desumoylase Ulp2 to control inner kinetochore sumoylation, and serves as a platform for spindle checkpoint proteins (MAD1, MAD2, RZZ) at unattached kinetochores while antagonizing dynein-mediated checkpoint protein stripping—together with Aurora B forming a molecular switch for robust spindle assembly checkpoint signaling; additionally, CENP-I participates in homologous recombination DNA repair by suppressing R-loop accumulation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CENP-I (Mis6/Ctf3) is a constitutive inner kinetochore protein that builds and maintains the centromeric chromatin platform required for faithful chromosome segregation [#0, #3]. It directly binds AT-rich centromeric DNA through a consecutive surface formed by conserved charged residues at the end of its N-terminal HEAT repeats, and this DNA contact—rather than histone binding—drives its own centromeric localization, stabilizes CENP-A nucleosomes, and enables loading of newly synthesized CENP-A [#17]. CENP-I acts upstream of CENP-A deposition in fission yeast and of CENP-C localization in vertebrate cells, while being dispensable for CENP-A localization itself in vertebrates [#1, #3], and it promotes CENP-A assembly by recruiting CENP-C and M18BP1 to enable HJURP/Mis18-mediated loading [#11]. It also protects pre-existing CENP-A during mitosis by suppressing RNA Pol II-dependent non-coding transcription at the central core [#16]. Within the kinetochore, CENP-I assembles through conserved HEAT-repeat contacts with CENP-H, CENP-K, and CENP-M [#13, #17] and provides a docking site for the desumoylase Ulp2 that controls inner kinetochore sumoylation [#14]. Beyond architecture, CENP-I sustains spindle assembly checkpoint signaling: it is required for kinetochore association of MAD1, MAD2, CENP-F, and the RZZ complex, and—together with Aurora B—forms a molecular switch that retains checkpoint proteins by antagonizing their dynein-mediated stripping at unattached kinetochores [#5, #8]. Independently of its kinetochore role, human CENP-I promotes homologous-recombination repair of DNA double-strand breaks by suppressing R-loop accumulation, as RNaseH1 expression restores HR in CENP-I-deficient cells [#15].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Established that the CENP-I ortholog Mis6 is a constitutive centromere component acting in G1/S to build specialized inner centromere chromatin, identifying it as a foundational determinant of centromere identity rather than a transient mitotic factor.\",\n      \"evidence\": \"Temperature-sensitive mutant analysis with minichromosome loss and micrococcal nuclease chromatin profiling in fission yeast\",\n      \"pmids\": [\"9230309\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular nature of the chromatin alteration undefined\", \"No direct DNA- or protein-binding activity demonstrated\", \"Relationship to CENP-A not yet tested\"]\n    },\n    {\n      \"year\": 1999,\n      \"claim\": \"Linked Mis6 function to mitotic spindle morphogenesis, showing centromere defects translate into aberrant metaphase spindle geometry.\",\n      \"evidence\": \"Fluorescence microscopy of spindle length and suppressor analysis in mis6 mutants\",\n      \"pmids\": [\"10398680\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Spindle defect likely indirect via centromere structure\", \"Mechanism connecting centromere to spindle length unresolved\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Placed Mis6 upstream of CENP-A deposition, defining its role in establishing the epigenetic centromere mark.\",\n      \"evidence\": \"Genetic epistasis and immunofluorescence of SpCENP-A in mis6 mutants\",\n      \"pmids\": [\"10864871\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not distinguish establishment versus maintenance of CENP-A\", \"Direct loading mechanism unknown\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Defined the vertebrate kinetochore hierarchy, showing human/chicken CENP-I is required for CENP-C but not CENP-A localization, revealing species differences in the CENP-A dependency relationship.\",\n      \"evidence\": \"Conditional knockout in DT40 cells with co-localization immunocytochemistry\",\n      \"pmids\": [\"11970896\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct versus indirect requirement for CENP-C not separated\", \"DNA-binding basis of localization not addressed\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Identified the budding yeast Ctf3 sub-complex (with Mcm22/Mcm16) and showed Cse4/CENP-A-dependent, Ctf19-dependent centromere binding, mapping CENP-I into the CCAN interaction network.\",\n      \"evidence\": \"Two-hybrid, ChIP, synthetic dosage lethality, and microscopy in budding yeast\",\n      \"pmids\": [\"11782448\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ctf3 dispensable for Cse4 loading, contrasting fission yeast — unifying mechanism unclear\", \"Direct DNA contact versus indirect recruitment not resolved\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Connected CENP-I to spindle checkpoint competence, showing its depletion abolishes kinetochore MAD1/MAD2/CENP-F and impairs mitotic arrest at unattached kinetochores.\",\n      \"evidence\": \"Antibody depletion, immunofluorescence, time-lapse imaging and checkpoint assays in human cells\",\n      \"pmids\": [\"12640463\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether checkpoint defect is architectural or signaling-specific not separated\", \"Mechanism of MAD1/MAD2 recruitment unknown\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Provided physical evidence that the Mis6 complex engages Mad2 upon checkpoint activation and is required for Mad2 accumulation at unattached kinetochores.\",\n      \"evidence\": \"Reciprocal co-IP and Mad2/Bub1 localization in mis6 mutants, plus Mis6 fragment localization in fission yeast\",\n      \"pmids\": [\"15930132\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Microtubule-binding by N-terminal fragments not functionally validated\", \"Direct versus indirect Mad2 interaction unresolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Resolved the CENP-I checkpoint mechanism, showing it retains RZZ/Mad1 by inhibiting dynein-mediated stripping while Aurora B regulates their association — a two-input molecular switch for robust SAC signaling.\",\n      \"evidence\": \"siRNA depletion with Aurora B inhibition epistasis, immunofluorescence and live imaging in human cells\",\n      \"pmids\": [\"24862574\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct molecular target of CENP-I in blocking stripping unknown\", \"Structural basis of the switch undefined\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected CENP-I/CCAN to the CENP-A loading machinery via Eic1/Mis19, showing physical bridging of Mis18/HJURP factors to the Mis6/Ctf19 complex enables temporally regulated CENP-A deposition.\",\n      \"evidence\": \"Co-IP, mass spectrometry, conditional depletion and localization in fission yeast\",\n      \"pmids\": [\"24789708\", \"25375240\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct CENP-I contact with Eic1 versus other CCAN subunits unmapped\", \"Maintenance versus establishment role of the bridge unclear\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Demonstrated CENP-I can drive de novo CENP-A assembly by recruiting CENP-C and then M18BP1, defining a CENP-C-independent route to M18BP1 recruitment.\",\n      \"evidence\": \"Tethering to synthetic alphoid(tetO) HAC array with immunofluorescence and CENP-A ChIP\",\n      \"pmids\": [\"26527398\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ectopic tethering may not reflect endogenous order of events\", \"Direct M18BP1 binding not shown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Mapped CENP-I intramolecular architecture, showing a conserved α11 helix folds against N-terminal HEAT repeats to maintain folding and CENP-H/CENP-M binding.\",\n      \"evidence\": \"In vitro aggregation assays, co-IP, mutagenesis and fungal structural analysis\",\n      \"pmids\": [\"32017295\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Human full-length structure not solved\", \"Functional consequence of α11 mutations in cells limited\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Identified a non-kinetochore function: CENP-I promotes homologous-recombination DSB repair by limiting R-loop formation, with RNaseH1 rescuing HR in its absence.\",\n      \"evidence\": \"siRNA, HR/NHEJ reporters, 53BP1 foci, IR survival and RNaseH1 rescue in human cells\",\n      \"pmids\": [\"34206916\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether CENP-I acts directly at break sites or via centromere transcription unclear\", \"Mechanism of R-loop suppression undefined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Revealed CENP-I as a desumoylase docking platform, with a conserved Ctf3 surface binding Ulp2 to control inner kinetochore sumoylation and segregation fidelity.\",\n      \"evidence\": \"Interaction mapping, surface mutagenesis, sumoylation and segregation assays in budding yeast\",\n      \"pmids\": [\"34081091\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conservation of the Ulp2-docking role in humans untested\", \"Sumoylation substrates at the kinetochore not enumerated\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Showed CENP-I maintains pre-existing CENP-A during mitosis by suppressing RNA Pol II non-coding transcription, since Pol II inhibition rescues CENP-A loss in mis6 mutants.\",\n      \"evidence\": \"Conditional mitotic depletion, CENP-A localization, ncRNA quantification and Pol II inhibition rescue in fission yeast\",\n      \"pmids\": [\"35970865\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct mechanism of transcriptional suppression by CENP-I unknown\", \"Link to its DNA-binding activity not tested\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established the central biochemical activity: CENP-I directly binds AT-rich centromeric DNA via charged residues at its N-terminal HEAT repeats, and this DNA contact—not histone binding—drives localization, CENP-A nucleosome stabilization, CENP-A loading and chromosome alignment.\",\n      \"evidence\": \"In vitro DNA-binding assays, DNA-surface mutagenesis, co-IP, CENP-A loading and chromosome alignment assays in human cells\",\n      \"pmids\": [\"36888657\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the CENP-I–DNA complex not solved\", \"How DNA binding integrates with CCAN assembly and transcription suppression unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Placed CENPI under FOXM1 transcriptional control with links to proline/arginine metabolism in glioblastoma, extending it into a cancer regulatory context.\",\n      \"evidence\": \"JASPAR prediction, dual luciferase reporter, ChIP, siRNA and metabolic assays in glioblastoma cells\",\n      \"pmids\": [\"41206587\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No direct protein-level mechanism linking CENPI to metabolism\", \"Single lab, correlative\", \"Generality beyond glioblastoma untested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CENP-I's direct DNA-binding activity is mechanistically integrated with CCAN assembly, transcriptional suppression of the centromere core, checkpoint protein retention, and its R-loop-suppressing DNA-repair role remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No high-resolution structure of human CENP-I bound to centromeric DNA or nucleosome\", \"Mechanistic basis of dynein-stripping antagonism unknown\", \"Whether the HR/R-loop function is separable from the kinetochore role untested\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [4, 17]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [0, 3, 17]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [3, 11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [3, 5, 8]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [15]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [1, 11, 16, 17]}\n    ],\n    \"complexes\": [\n      \"CCAN/Mis6-Ctf19 complex\",\n      \"kinetochore\"\n    ],\n    \"partners\": [\n      \"CENP-C\",\n      \"CENP-H\",\n      \"CENP-M\",\n      \"CENP-K\",\n      \"M18BP1\",\n      \"MAD2\",\n      \"Ulp2\",\n      \"Mcm22\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}