{"gene":"MEI4","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":1989,"finding":"MEI4 in S. cerevisiae is required for meiotic DNA double-strand break formation and acts upstream of RAD52 in the meiotic recombination pathway, as established by genetic epistasis: mei4 spo13 rad52 mutants produce viable spores, placing MEI4 before RAD52.","method":"Genetic epistasis analysis (double/triple mutant analysis, spo13 suppression assay)","journal":"Genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean loss-of-function with defined pathway position via epistasis, single lab","pmids":["2693205"],"is_preprint":false},{"year":1992,"finding":"S. cerevisiae MEI4 encodes a 450-amino-acid protein with meiosis-specific transcription; mei4 mutants form long axial elements but fail to undergo chromosome synapsis, establishing MEI4 as required for synaptonemal complex formation.","method":"DNA sequencing, cytological analysis of meiotic chromosomes in mei4 deletion strains, Northern blot","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function with defined cytological phenotype (axial elements without synapsis), single lab","pmids":["1545815"],"is_preprint":false},{"year":1998,"finding":"S. pombe mei4+ encodes a meiosis-specific transcription factor with an N-terminal forkhead/HNF3 DNA-binding domain that binds a GTAAAYA consensus sequence in the spo6+ promoter; a C-terminal 140-amino-acid region acts as a transcriptional activation domain.","method":"Functional complementation cloning, gel mobility shift assay (GST-Mei4 forkhead domain fusion), one-hybrid assay, promoter deletion analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct DNA-binding assay with recombinant protein, deletion analysis of activation domain, promoter element deletion validating functional consequence; multiple orthogonal methods","pmids":["9528784"],"is_preprint":false},{"year":2000,"finding":"S. pombe Mei4 binds a cis-acting element called FLEX in the promoter of its own gene and of multiple meiotic target genes (mde1–mde9), and mei4 transcription is positively autoregulated.","method":"Genome-wide promoter scanning, Northern blotting, forkhead-domain binding to FLEX element, reporter gene assay with ectopic Mei4 overproduction","journal":"Genetics","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — direct binding to FLEX element, Northern blot confirmation of target regulation, autoregulation confirmed by reporter assay; multiple orthogonal methods in single lab","pmids":["10747048"],"is_preprint":false},{"year":2006,"finding":"S. cerevisiae Mei4, Rec114, and Mer2 form a distinct protein complex required for meiotic DSB formation; all three proteins co-immunoprecipitate and co-localize in foci on meiotic chromosomes, partially overlapping each other but distinct from Mre11 and Rec102 foci.","method":"Co-immunoprecipitation, cytological co-localization (fluorescence microscopy) on meiotic chromosomes","journal":"Genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP and cytological co-localization, replicated by subsequent independent studies","pmids":["16783010"],"is_preprint":false},{"year":2005,"finding":"In S. pombe, the Rad3-Cds1 checkpoint pathway suppresses mei4+ transcription when premeiotic DNA replication is blocked, thereby coupling initiation of meiotic recombination (DSB formation) with DNA replication completion.","method":"Hydroxyurea block of replication, analysis in rad3, cds1, chk1, mek1 deletion mutants, Northern blot of mei4+ mRNA","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with defined checkpoint mutants, Northern blot, single lab","pmids":["16286472"],"is_preprint":false},{"year":2007,"finding":"In S. cerevisiae, Mei4 chromosome localization is strongly dependent on Rec114 and Mer2; systematic two-hybrid and deletion analyses identified protein regions required for Mei4–Rec114 interaction; Rec102 N-terminal sequences mediate interactions with Rec114 and Mei4, connecting the Mei4-Rec114-Mer2 subgroup to Spo11.","method":"Two-hybrid analysis, deletion mapping, cytological localization in various deletion backgrounds","journal":"Chromosoma","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — two-hybrid plus cytological localization dependencies, single lab with multiple mutants","pmids":["17558514"],"is_preprint":false},{"year":2010,"finding":"Mouse MEI4 is functionally conserved for meiotic DSB formation: Mei4−/− mice are deficient in meiotic DSBs, MEI4 localizes to discrete foci on meiotic chromosome axes (not overlapping DMC1 or RPA foci), and mouse MEI4 and REC114 interact directly with conserved motifs required for this interaction.","method":"Knockout mouse analysis, cytological localization (immunofluorescence), direct protein interaction assay (pulldown/co-IP), mutational analysis of conserved motifs","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 2 / Strong — KO mouse with defined phenotype, cytological localization, direct interaction assay with mutagenesis; multiple orthogonal methods replicated in mouse system","pmids":["20551173"],"is_preprint":false},{"year":2011,"finding":"S. pombe Rec24 (ortholog of mouse Mei4) is required genome-wide for crossing-over, localizes to linear elements on meiotic chromosomes in a Rec12 (Spo11)-independent manner, and forms complexes in vivo with Rec7; Rec7 is required for stabilization of Rec24 on linear elements.","method":"Gene disruption analysis, cytological localization, co-immunoprecipitation","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO phenotype analysis, cytological localization, co-IP; single lab","pmids":["21429938"],"is_preprint":false},{"year":2011,"finding":"S. pombe Mei4 (forkhead transcription factor) is required for telomere dispersal from the spindle pole body at meiosis I onset; mei4Δ cells arrest with clustered telomeres, and a temperature-sensitive mei4-N136A allele in the forkhead DNA-binding domain shows the same phenotype reversibly.","method":"Gene deletion, temperature-sensitive point mutant in forkhead domain, live-cell imaging of telomere position","journal":"Yeast (Chichester, England)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — defined genetic perturbation with cytological readout, single lab","pmids":["21449049"],"is_preprint":false},{"year":2014,"finding":"In S. pombe, Mei4 (transcription factor) controls termination of meiotic nuclear movements; loss of Mei4 prolongs nuclear movements, while Mei4 overproduction accelerates their termination; the Cds1 replication checkpoint represses mei4+ expression to link DNA replication to nuclear movement timing.","method":"Gene deletion/overproduction experiments, live-cell imaging of nuclear movement duration, checkpoint mutant analysis","journal":"Genes to cells","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — loss-of-function and gain-of-function with defined cellular phenotype (nuclear movement duration), single lab","pmids":["25492408"],"is_preprint":false},{"year":2015,"finding":"Mouse HORMAD1 (a chromosome axis component) is required for MEI4 localization on meiotic chromosome axes; MEI1, REC8, and RAD21L also contribute to proper MEI4 localization; quantitative correlation between axis-associated MEI4 levels and DSB formation suggests MEI4 is a limiting factor for DSB formation.","method":"Knockout mouse analyses (HORMAD1 KO, MEI1 KO, REC8 KO, RAD21L KO), immunofluorescence cytology, quantitative analysis of MEI4 foci and DSB markers","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple independent KO models tested with quantitative cytological readout, multiple orthogonal genetic perturbations","pmids":["25795304"],"is_preprint":false},{"year":2016,"finding":"S. pombe Cuf2 is a transcriptional co-regulator that physically interacts with Mei4 in the nucleus; Cuf2 requires Mei4 for its function, and Cuf2 chromatin occupancy at target gene promoters depends on FLEX-like elements bound by Mei4.","method":"Co-immunoprecipitation, bimolecular fluorescence complementation (BiFC), chromatin immunoprecipitation (ChIP), promoter deletion analysis","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — co-IP plus BiFC plus ChIP; multiple methods in single lab","pmids":["26986212"],"is_preprint":false},{"year":2018,"finding":"Mouse REC114 is essential for meiotic DSB formation and forms a stable complex with MEI4 and IHO1 (Mer2 ortholog) in spermatocytes; in vitro reconstitution showed a stable complex between the REC114 C-terminal domain and the MEI4 N-terminal domain; the REC114 N-terminal domain has a structure similar to Pleckstrin homology (PH) domains.","method":"Mouse knockout analysis, co-immunoprecipitation from spermatocytes, in vitro reconstitution of REC114–MEI4 domain complex, X-ray crystallography of REC114 N-terminal domain","journal":"Life science alliance","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution of domain complex, structural determination by crystallography, KO mouse phenotype, co-IP from endogenous tissue; multiple orthogonal methods","pmids":["30569039"],"is_preprint":false},{"year":2020,"finding":"In S. cerevisiae, PAF1C histone modification complex components (Rtf1, Cdc73) show synthetic DSB formation defects when combined with tagged (but not untagged) alleles of REC114 or MER2, and SET1 deletion similarly affects tagged REC114 in a manner independent of SPP1, suggesting histone modification machinery influences the Rec114-Mer2-Mei4 complex in DSB formation.","method":"Genetic analysis, meiotic DSB assay in double mutants","journal":"International journal of molecular sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — genetic interaction using tagged alleles with unclear relevance; single lab, indirect evidence","pmids":["32290544"],"is_preprint":false},{"year":2023,"finding":"The S. cerevisiae Rec114–Mei4 complex forms a 2:1 heterotrimer (two Rec114 : one Mei4) in which Rec114 C-terminal dimers cup an N-terminal Mei4 α-helix; this minimal complex binds DNA cooperatively and forms large condensates; single-molecule experiments show the complex bridges multiple DNA duplexes and can condense DNA through long-range interactions. AlphaFold2 structural models are conserved across eukaryotes.","method":"NMR spectroscopy, AlphaFold2 structural modeling, in vitro reconstitution of heterotrimeric complex, single-molecule DNA condensation assay, mutagenesis","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1 / Strong — NMR-supported structural model with mutagenesis validation, in vitro reconstitution, single-molecule functional assay; multiple orthogonal methods","pmids":["37442580"],"is_preprint":false},{"year":2023,"finding":"AlphaFold2 modeling supported by NMR and SAXS shows that Rec114–Mei4 forms a 2:1 heterotrimer where Rec114 C-terminal dimers form α-helical chains that cup the N-terminal Mei4 α-helix; the interaction domain contains two DNA-binding sites pointing in opposite directions driving DNA-dependent condensation; Mer2 forms a parallel homotetrameric coiled coil that bridges coaligned DNA duplexes; both structures are conserved across eukaryotes though DNA-binding properties vary.","method":"AlphaFold2 modeling, NMR spectroscopy, SAXS, mutagenesis, in vitro DNA condensation assay","journal":"Genes & development","confidence":"High","confidence_rationale":"Tier 1 / Strong — structural determination by NMR and SAXS with mutagenesis and functional assay; multiple orthogonal methods; replicated across two independent papers same year","pmids":["37442581"],"is_preprint":false},{"year":2023,"finding":"Mouse REC114 forms homodimers and associates with MEI4 as a 2:1 REC114:MEI4 heterotrimer that further dimerizes; IHO1 forms coiled-coil-based tetramers; IHO1 directly interacts with the PH domain of REC114 via the same surface recognized by TOPOVIBL and ANKRD31, suggesting REC114 acts as a regulatory platform for mutually exclusive interactions.","method":"AlphaFold2 modeling, biochemical characterization (SEC, native MS), co-IP, mutagenesis","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — structural modeling validated by biochemical methods, direct interaction mapping, multiple orthogonal approaches; replicated architecture consistent with yeast studies","pmids":["37431931"],"is_preprint":false},{"year":2024,"finding":"Bi-allelic missense variants in human MEI4 (in the N-terminal region, amino acids 98–305) reduce MEI4–DNA interaction in vitro, and a knock-in mouse model with one variant causes female infertility with oogenesis developmental defects, demonstrating that MEI4's DNA-binding activity is required for its reproductive function.","method":"In vitro DNA interaction assay with variant MEI4 proteins, knock-in mouse model generation, oogenesis phenotype analysis","journal":"Human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro DNA interaction assay plus knock-in mouse model; single lab","pmids":["38252283"],"is_preprint":false},{"year":2025,"finding":"The MEI4 C-terminus is required for stabilizing the MEI4–REC114 subcomplex on chromosome axes; Mei4Arg356* (C-terminal truncation) mice exhibit severe DSB formation defects, massive oocyte apoptosis via a HORMAD1-dependent synapsis checkpoint, and surviving oocytes show aneuploidy-driven complete embryonic arrest; heterozygous mice show intermediate follicle depletion consistent with haploinsufficiency.","method":"Exome sequencing of POI patients, in vitro DSB formation assay, knock-in mouse model (Mei4Arg356*), cytological analysis, embryonic development assay","journal":"Journal of genetics and genomics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — knock-in mouse model with defined molecular and cellular phenotypes, in vitro functional assay; single lab","pmids":["41419020"],"is_preprint":false},{"year":2025,"finding":"Mre11 is recruited to meiotic DSB sites via its C-terminal IDR, which contains a short α-helix that binds a conserved region of Mer2; this interaction is required for Mre11 foci formation during meiosis and DSB formation; the Mer2 interaction is part of the RMM (Rec114-Mei4-Mer2) condensation-dependent recruitment mechanism for Mre11.","method":"In vitro condensate assay, mutagenesis of Mer2-binding helix in Mre11, in vivo foci analysis","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro assay plus in vivo foci with mutagenesis; preprint, not yet peer-reviewed; Mei4 is part of the RMM complex implicated but focus is on Mre11-Mer2 interaction","pmids":["bio_10.1101_2025.07.08.663703"],"is_preprint":true}],"current_model":"MEI4 (and its orthologs across eukaryotes) is a conserved component of the meiotic DSB machinery that forms a 2:1 heterotrimeric complex with REC114 (two REC114 per one MEI4 N-terminal helix), localizes to chromosome axes in a manner dependent on HORMAD1, REC8, and IHO1/Mer2, and promotes SPO11-mediated DSB formation by cooperatively binding DNA and driving condensate formation that assembles the DSB machinery; in S. pombe and other fungi, the Mei4 ortholog additionally functions as a forkhead transcription factor activating middle-phase meiotic genes via FLEX-element binding."},"narrative":{"mechanistic_narrative":"MEI4 is a conserved component of the meiotic recombination machinery that promotes SPO11/Spo11-mediated DNA double-strand break (DSB) formation, acting genetically upstream of strand-exchange and repair factors such as RAD52 [PMID:2693205, PMID:20551173]. MEI4 functions as the core of a stable, conserved complex with REC114 and the Mer2 ortholog IHO1, in which two REC114 C-terminal dimers cup an N-terminal MEI4 α-helix to form a 2:1 REC114:MEI4 heterotrimer that can further oligomerize [PMID:30569039, PMID:37442580, PMID:37431931]. This minimal REC114–MEI4 unit binds DNA cooperatively through two oppositely oriented DNA-binding sites and drives DNA-dependent condensate formation, bridging and condensing multiple DNA duplexes — the biophysical basis for assembling the DSB machinery [PMID:37442580, PMID:37442581]. MEI4 concentrates in discrete foci on meiotic chromosome axes distinct from downstream recombination markers, and its axis loading depends on the chromosome-axis factor HORMAD1 together with the cohesins REC8/RAD21L, with axis-associated MEI4 levels quantitatively limiting DSB formation [PMID:20551173, PMID:25795304]. MEI4's DNA-binding activity, residing in its N-terminal region, and the C-terminal segment that stabilizes the MEI4–REC114 subcomplex on axes are both required for fertility: bi-allelic and truncating MEI4 variants impair DNA binding or axis stabilization and cause oogenesis defects and premature ovarian insufficiency, modeled in knock-in mice [PMID:38252283, PMID:41419020]. In the fission yeast and budding yeast lineages, the Mei4 ortholog has an additional, distinct role as a forkhead/HNF3 transcription factor that binds the FLEX cis-element to activate middle-meiotic genes and autoregulate its own expression, coordinating recombination timing with premeiotic DNA replication [PMID:9528784, PMID:10747048, PMID:16286472].","teleology":[{"year":1989,"claim":"Established where MEI4 acts in meiotic recombination, placing it upstream of the strand-exchange/repair machinery rather than within it.","evidence":"Genetic epistasis with spo13 suppression in S. cerevisiae mei4 spo13 rad52 triple mutants","pmids":["2693205"],"confidence":"Medium","gaps":["Did not define a molecular activity for MEI4","No protein partners identified","Mechanism of DSB promotion unknown"]},{"year":1992,"claim":"Defined MEI4 as a meiosis-specific gene whose product is required for synaptonemal complex formation, linking it to chromosome morphogenesis.","evidence":"Sequencing, Northern blot, and cytology of meiotic chromosomes in S. cerevisiae mei4 deletion strains","pmids":["1545815"],"confidence":"Medium","gaps":["Whether synapsis failure is a direct consequence of failed DSB formation not resolved","No biochemical activity assigned"]},{"year":1998,"claim":"Revealed that the S. pombe Mei4 ortholog is a sequence-specific forkhead transcription factor, defining a transcriptional function distinct from the DSB role in budding yeast/mouse.","evidence":"Complementation cloning, gel-shift with recombinant forkhead domain, one-hybrid and promoter deletion in S. pombe","pmids":["9528784"],"confidence":"High","gaps":["Full target-gene repertoire not yet mapped","Relationship between fission-yeast transcriptional role and DSB-machinery role of orthologs unaddressed"]},{"year":2000,"claim":"Identified the FLEX cis-element as the genomic recognition site through which S. pombe Mei4 activates middle-meiotic genes and autoregulates itself, building the transcriptional regulatory logic.","evidence":"Genome-wide promoter scanning, Northern blots, forkhead-FLEX binding, and reporter assays in S. pombe","pmids":["10747048"],"confidence":"High","gaps":["Co-factors required for activation not yet identified","How FLEX targets are selected genome-wide unclear"]},{"year":2005,"claim":"Showed that mei4+ transcription is gated by the replication checkpoint, coupling onset of meiotic recombination to completion of premeiotic DNA replication.","evidence":"Hydroxyurea replication block with rad3/cds1/chk1/mek1 mutants and Northern blot in S. pombe","pmids":["16286472"],"confidence":"Medium","gaps":["Direct checkpoint target within the mei4 promoter not defined","Applies to the transcriptional role; checkpoint control of the metazoan DSB role separate"]},{"year":2006,"claim":"Defined the Mei4–Rec114–Mer2 complex as a discrete physical and cytological entity required for DSB formation, distinct from the Mre11/Rec102 machinery.","evidence":"Co-immunoprecipitation and cytological co-localization on meiotic chromosomes in S. cerevisiae","pmids":["16783010"],"confidence":"High","gaps":["Stoichiometry and direct binding interfaces not resolved","Biochemical activity of the complex unknown"]},{"year":2007,"claim":"Mapped the interaction hierarchy showing Mei4 axis localization depends on Rec114/Mer2 and connecting the subcomplex to Spo11 via Rec102.","evidence":"Two-hybrid, deletion mapping, and cytological localization in deletion backgrounds in S. cerevisiae","pmids":["17558514"],"confidence":"Medium","gaps":["Direct vs indirect nature of Mei4-Spo11 link not established","Interaction regions not resolved structurally"]},{"year":2010,"claim":"Demonstrated functional conservation of MEI4 to mammals, showing it is essential for DSB formation, localizes to axis foci, and directly binds REC114 via conserved motifs.","evidence":"Mei4-/- mouse phenotyping, immunofluorescence, and pulldown/co-IP with motif mutagenesis","pmids":["20551173"],"confidence":"High","gaps":["Complex stoichiometry and structure not yet defined","How axis foci promote SPO11 cleavage unknown"]},{"year":2011,"claim":"Showed the S. pombe DSB-pathway ortholog Rec24 is required genome-wide for crossing-over and is stabilized on linear elements by Rec7, separating axis loading from Spo11 activity.","evidence":"Gene disruption, cytological localization, and co-IP in S. pombe","pmids":["21429938"],"confidence":"Medium","gaps":["Molecular basis of linear-element recruitment unresolved","Direct DNA-binding not tested here"]},{"year":2011,"claim":"Extended the fission-yeast transcription-factor role to meiotic chromosome dynamics, showing forkhead-domain-dependent control of telomere dispersal at meiosis I.","evidence":"Deletion and temperature-sensitive forkhead-domain point mutant with live-cell telomere imaging in S. pombe","pmids":["21449049"],"confidence":"Medium","gaps":["Direct transcriptional targets driving telomere dispersal not identified"]},{"year":2014,"claim":"Linked Mei4 transcriptional output to timing of meiotic nuclear movements under checkpoint control, reinforcing replication-coupled regulation of meiotic progression.","evidence":"Deletion/overproduction with live-cell imaging and checkpoint-mutant analysis in S. pombe","pmids":["25492408"],"confidence":"Medium","gaps":["Effector genes mediating nuclear-movement termination not defined"]},{"year":2015,"claim":"Identified the chromosome-axis determinants of MEI4 localization and established that axis-bound MEI4 is a quantitative limiting factor for DSB formation.","evidence":"HORMAD1, MEI1, REC8, RAD21L knockout mice with quantitative immunofluorescence of MEI4 and DSB markers","pmids":["25795304"],"confidence":"High","gaps":["Direct vs indirect dependence of MEI4 on each axis factor not dissected","Mechanism converting axis loading into cleavage unknown"]},{"year":2018,"claim":"Resolved the mammalian REC114–MEI4–IHO1 complex biochemically and structurally, reconstituting the REC114-CTD:MEI4-NTD interaction and solving the REC114 N-terminal PH-like domain.","evidence":"REC114 KO mouse, co-IP from spermatocytes, in vitro domain reconstitution, X-ray crystallography","pmids":["30569039"],"confidence":"High","gaps":["Complex stoichiometry not yet quantified","DNA-binding/condensation activity not yet characterized"]},{"year":2023,"claim":"Defined the molecular architecture and biochemical activity of the complex: a 2:1 REC114:MEI4 heterotrimer with opposed DNA-binding sites that drives cooperative DNA binding and condensation, providing the physical mechanism for DSB-machinery assembly.","evidence":"NMR/SAXS, AlphaFold2 modeling, in vitro reconstitution, single-molecule DNA condensation, and mutagenesis (S. cerevisiae and conserved models)","pmids":["37442580","37442581"],"confidence":"High","gaps":["How condensates direct SPO11 to specific sites not resolved","In vivo relevance of condensation vs other functions"]},{"year":2023,"claim":"Established REC114 as a regulatory platform: the same PH-domain surface mediates mutually exclusive interactions with IHO1, TOPOVIBL, and ANKRD31, organizing the higher-order DSB machinery around the MEI4 heterotrimer.","evidence":"AlphaFold2 modeling with SEC, native MS, co-IP, and mutagenesis in the mouse system","pmids":["37431931"],"confidence":"High","gaps":["Temporal ordering of competing interactions in vivo unknown","Functional consequence of heterotrimer dimerization not defined"]},{"year":2024,"claim":"Connected MEI4 to human disease by showing N-terminal variants reduce DNA binding and cause oogenesis defects, establishing DNA-binding activity as required for reproductive function.","evidence":"In vitro DNA interaction assays with variant proteins and a knock-in mouse model with oogenesis phenotyping","pmids":["38252283"],"confidence":"Medium","gaps":["Single lab; allelic spectrum in patients limited","Whether variants affect condensation as well as binding not tested"]},{"year":2025,"claim":"Showed the MEI4 C-terminus stabilizes the MEI4–REC114 subcomplex on axes and that its truncation causes severe DSB defects, oocyte apoptosis via the HORMAD1-dependent synapsis checkpoint, and premature ovarian insufficiency, with haploinsufficiency.","evidence":"POI exome sequencing, in vitro DSB assay, Mei4Arg356* knock-in mouse with cytology and embryonic-development analysis","pmids":["41419020"],"confidence":"Medium","gaps":["Structural basis of C-terminal axis stabilization not defined","Single lab"]},{"year":null,"claim":"How the RMM condensate is converted into site-specific SPO11 cleavage — i.e. how DNA condensation and axis tethering are mechanistically coupled to catalytic DSB formation — remains unresolved.","evidence":"No direct evidence in the timeline reconstitutes SPO11 cleavage from the assembled RMM condensate","pmids":[],"confidence":"Low","gaps":["No reconstitution of SPO11 cleavage from the RMM complex","Mechanism coupling condensation to catalysis unknown","Mre11 recruitment via Mer2 only shown in a preprint (idx 20)"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[2,3,15,16,18]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[2,3]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[13,15,17]}],"localization":[{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[4,7,11]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[2,12]}],"pathway":[{"term_id":"R-HSA-1474165","term_label":"Reproduction","supporting_discovery_ids":[7,18,19]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[0,7,11]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[2,3]}],"complexes":["REC114-MEI4-Mer2/IHO1 (RMM) complex"],"partners":["REC114","IHO1","MER2","REC102","HORMAD1","CUF2","REC7"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"A8MW99","full_name":"Meiosis-specific protein MEI4","aliases":[],"length_aa":385,"mass_kda":44.0,"function":"Required for DNA double-strand breaks (DSBs) formation in unsynapsed regions during meiotic recombination. Probably acts by forming a complex with IHO1 and REC114, which activates DSBs formation in unsynapsed regions, an essential step to ensure completion of synapsis","subcellular_location":"Chromosome","url":"https://www.uniprot.org/uniprotkb/A8MW99/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MEI4","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/MEI4","total_profiled":1310},"omim":[{"mim_id":"619190","title":"INTERACTOR OF HORMAD1 1; IHO1","url":"https://www.omim.org/entry/619190"},{"mim_id":"618423","title":"ANKYRIN REPEAT DOMAIN-CONTAINING PROTEIN 31; ANKRD31","url":"https://www.omim.org/entry/618423"},{"mim_id":"618421","title":"REC114 MEIOTIC RECOMBINATION PROTEIN; REC114","url":"https://www.omim.org/entry/618421"},{"mim_id":"618417","title":"MEIOTIC DOUBLE-STRANDED BREAK FORMATION PROTEIN 4; MEI4","url":"https://www.omim.org/entry/618417"},{"mim_id":"609824","title":"HORMA DOMAIN-CONTAINING PROTEIN 1; HORMAD1","url":"https://www.omim.org/entry/609824"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"kidney","ntpm":3.9},{"tissue":"pancreas","ntpm":1.9},{"tissue":"testis","ntpm":3.2},{"tissue":"thyroid gland","ntpm":2.0}],"url":"https://www.proteinatlas.org/search/MEI4"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"A8MW99","domains":[{"cath_id":"-","chopping":"148-165_175-273","consensus_level":"medium","plddt":78.2194,"start":148,"end":273},{"cath_id":"-","chopping":"276-385","consensus_level":"medium","plddt":80.0506,"start":276,"end":385}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/A8MW99","model_url":"https://alphafold.ebi.ac.uk/files/AF-A8MW99-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-A8MW99-F1-predicted_aligned_error_v6.png","plddt_mean":70.19},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MEI4","jax_strain_url":"https://www.jax.org/strain/search?query=MEI4"},"sequence":{"accession":"A8MW99","fasta_url":"https://rest.uniprot.org/uniprotkb/A8MW99.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/A8MW99/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/A8MW99"}},"corpus_meta":[{"pmid":"20551173","id":"PMC_20551173","title":"Functional conservation of Mei4 for meiotic DNA double-strand break formation from yeasts to mice.","date":"2010","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/20551173","citation_count":164,"is_preprint":false},{"pmid":"9528784","id":"PMC_9528784","title":"The Schizosaccharomyces pombe mei4+ gene encodes a meiosis-specific transcription factor containing a forkhead DNA-binding domain.","date":"1998","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/9528784","citation_count":118,"is_preprint":false},{"pmid":"17558514","id":"PMC_17558514","title":"Interactions between Mei4, Rec114, and other proteins required for meiotic DNA double-strand break formation in Saccharomyces cerevisiae.","date":"2007","source":"Chromosoma","url":"https://pubmed.ncbi.nlm.nih.gov/17558514","citation_count":111,"is_preprint":false},{"pmid":"16783010","id":"PMC_16783010","title":"Saccharomyces cerevisiae Mer2, Mei4 and Rec114 form a complex required for meiotic double-strand break formation.","date":"2006","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/16783010","citation_count":92,"is_preprint":false},{"pmid":"10747048","id":"PMC_10747048","title":"Autoregulated expression of Schizosaccharomyces pombe meiosis-specific transcription factor Mei4 and a genome-wide search for its target genes.","date":"2000","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/10747048","citation_count":88,"is_preprint":false},{"pmid":"30569039","id":"PMC_30569039","title":"Mouse REC114 is essential for meiotic DNA double-strand break formation and forms a complex with MEI4.","date":"2018","source":"Life science alliance","url":"https://pubmed.ncbi.nlm.nih.gov/30569039","citation_count":80,"is_preprint":false},{"pmid":"2693205","id":"PMC_2693205","title":"MEI4, a yeast gene required for meiotic recombination.","date":"1989","source":"Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/2693205","citation_count":64,"is_preprint":false},{"pmid":"25795304","id":"PMC_25795304","title":"MEI4 – a central player in the regulation of meiotic DNA double-strand break formation in the mouse.","date":"2015","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/25795304","citation_count":58,"is_preprint":false},{"pmid":"1545815","id":"PMC_1545815","title":"MEI4, a meiosis-specific yeast gene required for chromosome synapsis.","date":"1992","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/1545815","citation_count":51,"is_preprint":false},{"pmid":"16286472","id":"PMC_16286472","title":"Rad3-Cds1 mediates coupling of initiation of meiotic recombination with DNA replication. Mei4-dependent transcription as a potential target of meiotic checkpoint.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16286472","citation_count":27,"is_preprint":false},{"pmid":"21429938","id":"PMC_21429938","title":"Functional interactions of Rec24, the fission yeast ortholog of mouse Mei4, with the meiotic recombination-initiation complex.","date":"2011","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/21429938","citation_count":21,"is_preprint":false},{"pmid":"37442581","id":"PMC_37442581","title":"Evolutionary conservation of the structure and function of meiotic Rec114-Mei4 and Mer2 complexes.","date":"2023","source":"Genes & development","url":"https://pubmed.ncbi.nlm.nih.gov/37442581","citation_count":19,"is_preprint":false},{"pmid":"37431931","id":"PMC_37431931","title":"Characterization of the REC114-MEI4-IHO1 complex regulating meiotic DNA double-strand break 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genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38252283","citation_count":10,"is_preprint":false},{"pmid":"25492408","id":"PMC_25492408","title":"Meiotic nuclear movements in fission yeast are regulated by the transcription factor Mei4 downstream of a Cds1-dependent replication checkpoint pathway.","date":"2014","source":"Genes to cells : devoted to molecular & cellular mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/25492408","citation_count":10,"is_preprint":false},{"pmid":"32290544","id":"PMC_32290544","title":"Genetic Interactions of Histone Modification Machinery Set1 and PAF1C with the Recombination Complex Rec114-Mer2-Mei4 in the Formation of Meiotic DNA Double-Strand Breaks.","date":"2020","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/32290544","citation_count":6,"is_preprint":false},{"pmid":"26986212","id":"PMC_26986212","title":"Cuf2 Is a Transcriptional Co-Regulator that Interacts with Mei4 for Timely Expression of Middle-Phase Meiotic Genes.","date":"2016","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/26986212","citation_count":3,"is_preprint":false},{"pmid":"36711595","id":"PMC_36711595","title":"Structure and DNA bridging activity of the essential Rec114â€\"Mei4 trimer interface.","date":"2023","source":"bioRxiv : the preprint server for biology","url":"https://pubmed.ncbi.nlm.nih.gov/36711595","citation_count":1,"is_preprint":false},{"pmid":"41419020","id":"PMC_41419020","title":"MEI4 variations drive female reproductive disorders via impaired oocyte abundance and developmental potential.","date":"2025","source":"Journal of genetics and genomics = Yi chuan xue bao","url":"https://pubmed.ncbi.nlm.nih.gov/41419020","citation_count":0,"is_preprint":false},{"pmid":"42011758","id":"PMC_42011758","title":"Mei4 deficiency induces sexual dimorphism in early meiosis and promotes production of unreduced eggs in zebrafish.","date":"2026","source":"Zoological 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pigs","date":"2024-06-24","source":"bioRxiv","url":"https://doi.org/10.1101/2024.06.20.599484","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":13077,"output_tokens":5029,"usd":0.057333,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13423,"output_tokens":5064,"usd":0.096857,"stage2_stop_reason":"end_turn"},"total_usd":0.15419,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 1989,\n      \"finding\": \"MEI4 in S. cerevisiae is required for meiotic DNA double-strand break formation and acts upstream of RAD52 in the meiotic recombination pathway, as established by genetic epistasis: mei4 spo13 rad52 mutants produce viable spores, placing MEI4 before RAD52.\",\n      \"method\": \"Genetic epistasis analysis (double/triple mutant analysis, spo13 suppression assay)\",\n      \"journal\": \"Genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean loss-of-function with defined pathway position via epistasis, single lab\",\n      \"pmids\": [\"2693205\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1992,\n      \"finding\": \"S. cerevisiae MEI4 encodes a 450-amino-acid protein with meiosis-specific transcription; mei4 mutants form long axial elements but fail to undergo chromosome synapsis, establishing MEI4 as required for synaptonemal complex formation.\",\n      \"method\": \"DNA sequencing, cytological analysis of meiotic chromosomes in mei4 deletion strains, Northern blot\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function with defined cytological phenotype (axial elements without synapsis), single lab\",\n      \"pmids\": [\"1545815\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1998,\n      \"finding\": \"S. pombe mei4+ encodes a meiosis-specific transcription factor with an N-terminal forkhead/HNF3 DNA-binding domain that binds a GTAAAYA consensus sequence in the spo6+ promoter; a C-terminal 140-amino-acid region acts as a transcriptional activation domain.\",\n      \"method\": \"Functional complementation cloning, gel mobility shift assay (GST-Mei4 forkhead domain fusion), one-hybrid assay, promoter deletion analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct DNA-binding assay with recombinant protein, deletion analysis of activation domain, promoter element deletion validating functional consequence; multiple orthogonal methods\",\n      \"pmids\": [\"9528784\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2000,\n      \"finding\": \"S. pombe Mei4 binds a cis-acting element called FLEX in the promoter of its own gene and of multiple meiotic target genes (mde1–mde9), and mei4 transcription is positively autoregulated.\",\n      \"method\": \"Genome-wide promoter scanning, Northern blotting, forkhead-domain binding to FLEX element, reporter gene assay with ectopic Mei4 overproduction\",\n      \"journal\": \"Genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — direct binding to FLEX element, Northern blot confirmation of target regulation, autoregulation confirmed by reporter assay; multiple orthogonal methods in single lab\",\n      \"pmids\": [\"10747048\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"S. cerevisiae Mei4, Rec114, and Mer2 form a distinct protein complex required for meiotic DSB formation; all three proteins co-immunoprecipitate and co-localize in foci on meiotic chromosomes, partially overlapping each other but distinct from Mre11 and Rec102 foci.\",\n      \"method\": \"Co-immunoprecipitation, cytological co-localization (fluorescence microscopy) on meiotic chromosomes\",\n      \"journal\": \"Genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP and cytological co-localization, replicated by subsequent independent studies\",\n      \"pmids\": [\"16783010\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"In S. pombe, the Rad3-Cds1 checkpoint pathway suppresses mei4+ transcription when premeiotic DNA replication is blocked, thereby coupling initiation of meiotic recombination (DSB formation) with DNA replication completion.\",\n      \"method\": \"Hydroxyurea block of replication, analysis in rad3, cds1, chk1, mek1 deletion mutants, Northern blot of mei4+ mRNA\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with defined checkpoint mutants, Northern blot, single lab\",\n      \"pmids\": [\"16286472\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"In S. cerevisiae, Mei4 chromosome localization is strongly dependent on Rec114 and Mer2; systematic two-hybrid and deletion analyses identified protein regions required for Mei4–Rec114 interaction; Rec102 N-terminal sequences mediate interactions with Rec114 and Mei4, connecting the Mei4-Rec114-Mer2 subgroup to Spo11.\",\n      \"method\": \"Two-hybrid analysis, deletion mapping, cytological localization in various deletion backgrounds\",\n      \"journal\": \"Chromosoma\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — two-hybrid plus cytological localization dependencies, single lab with multiple mutants\",\n      \"pmids\": [\"17558514\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Mouse MEI4 is functionally conserved for meiotic DSB formation: Mei4−/− mice are deficient in meiotic DSBs, MEI4 localizes to discrete foci on meiotic chromosome axes (not overlapping DMC1 or RPA foci), and mouse MEI4 and REC114 interact directly with conserved motifs required for this interaction.\",\n      \"method\": \"Knockout mouse analysis, cytological localization (immunofluorescence), direct protein interaction assay (pulldown/co-IP), mutational analysis of conserved motifs\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — KO mouse with defined phenotype, cytological localization, direct interaction assay with mutagenesis; multiple orthogonal methods replicated in mouse system\",\n      \"pmids\": [\"20551173\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"S. pombe Rec24 (ortholog of mouse Mei4) is required genome-wide for crossing-over, localizes to linear elements on meiotic chromosomes in a Rec12 (Spo11)-independent manner, and forms complexes in vivo with Rec7; Rec7 is required for stabilization of Rec24 on linear elements.\",\n      \"method\": \"Gene disruption analysis, cytological localization, co-immunoprecipitation\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO phenotype analysis, cytological localization, co-IP; single lab\",\n      \"pmids\": [\"21429938\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"S. pombe Mei4 (forkhead transcription factor) is required for telomere dispersal from the spindle pole body at meiosis I onset; mei4Δ cells arrest with clustered telomeres, and a temperature-sensitive mei4-N136A allele in the forkhead DNA-binding domain shows the same phenotype reversibly.\",\n      \"method\": \"Gene deletion, temperature-sensitive point mutant in forkhead domain, live-cell imaging of telomere position\",\n      \"journal\": \"Yeast (Chichester, England)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — defined genetic perturbation with cytological readout, single lab\",\n      \"pmids\": [\"21449049\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"In S. pombe, Mei4 (transcription factor) controls termination of meiotic nuclear movements; loss of Mei4 prolongs nuclear movements, while Mei4 overproduction accelerates their termination; the Cds1 replication checkpoint represses mei4+ expression to link DNA replication to nuclear movement timing.\",\n      \"method\": \"Gene deletion/overproduction experiments, live-cell imaging of nuclear movement duration, checkpoint mutant analysis\",\n      \"journal\": \"Genes to cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — loss-of-function and gain-of-function with defined cellular phenotype (nuclear movement duration), single lab\",\n      \"pmids\": [\"25492408\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Mouse HORMAD1 (a chromosome axis component) is required for MEI4 localization on meiotic chromosome axes; MEI1, REC8, and RAD21L also contribute to proper MEI4 localization; quantitative correlation between axis-associated MEI4 levels and DSB formation suggests MEI4 is a limiting factor for DSB formation.\",\n      \"method\": \"Knockout mouse analyses (HORMAD1 KO, MEI1 KO, REC8 KO, RAD21L KO), immunofluorescence cytology, quantitative analysis of MEI4 foci and DSB markers\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple independent KO models tested with quantitative cytological readout, multiple orthogonal genetic perturbations\",\n      \"pmids\": [\"25795304\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"S. pombe Cuf2 is a transcriptional co-regulator that physically interacts with Mei4 in the nucleus; Cuf2 requires Mei4 for its function, and Cuf2 chromatin occupancy at target gene promoters depends on FLEX-like elements bound by Mei4.\",\n      \"method\": \"Co-immunoprecipitation, bimolecular fluorescence complementation (BiFC), chromatin immunoprecipitation (ChIP), promoter deletion analysis\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — co-IP plus BiFC plus ChIP; multiple methods in single lab\",\n      \"pmids\": [\"26986212\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Mouse REC114 is essential for meiotic DSB formation and forms a stable complex with MEI4 and IHO1 (Mer2 ortholog) in spermatocytes; in vitro reconstitution showed a stable complex between the REC114 C-terminal domain and the MEI4 N-terminal domain; the REC114 N-terminal domain has a structure similar to Pleckstrin homology (PH) domains.\",\n      \"method\": \"Mouse knockout analysis, co-immunoprecipitation from spermatocytes, in vitro reconstitution of REC114–MEI4 domain complex, X-ray crystallography of REC114 N-terminal domain\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution of domain complex, structural determination by crystallography, KO mouse phenotype, co-IP from endogenous tissue; multiple orthogonal methods\",\n      \"pmids\": [\"30569039\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In S. cerevisiae, PAF1C histone modification complex components (Rtf1, Cdc73) show synthetic DSB formation defects when combined with tagged (but not untagged) alleles of REC114 or MER2, and SET1 deletion similarly affects tagged REC114 in a manner independent of SPP1, suggesting histone modification machinery influences the Rec114-Mer2-Mei4 complex in DSB formation.\",\n      \"method\": \"Genetic analysis, meiotic DSB assay in double mutants\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — genetic interaction using tagged alleles with unclear relevance; single lab, indirect evidence\",\n      \"pmids\": [\"32290544\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The S. cerevisiae Rec114–Mei4 complex forms a 2:1 heterotrimer (two Rec114 : one Mei4) in which Rec114 C-terminal dimers cup an N-terminal Mei4 α-helix; this minimal complex binds DNA cooperatively and forms large condensates; single-molecule experiments show the complex bridges multiple DNA duplexes and can condense DNA through long-range interactions. AlphaFold2 structural models are conserved across eukaryotes.\",\n      \"method\": \"NMR spectroscopy, AlphaFold2 structural modeling, in vitro reconstitution of heterotrimeric complex, single-molecule DNA condensation assay, mutagenesis\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — NMR-supported structural model with mutagenesis validation, in vitro reconstitution, single-molecule functional assay; multiple orthogonal methods\",\n      \"pmids\": [\"37442580\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"AlphaFold2 modeling supported by NMR and SAXS shows that Rec114–Mei4 forms a 2:1 heterotrimer where Rec114 C-terminal dimers form α-helical chains that cup the N-terminal Mei4 α-helix; the interaction domain contains two DNA-binding sites pointing in opposite directions driving DNA-dependent condensation; Mer2 forms a parallel homotetrameric coiled coil that bridges coaligned DNA duplexes; both structures are conserved across eukaryotes though DNA-binding properties vary.\",\n      \"method\": \"AlphaFold2 modeling, NMR spectroscopy, SAXS, mutagenesis, in vitro DNA condensation assay\",\n      \"journal\": \"Genes & development\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — structural determination by NMR and SAXS with mutagenesis and functional assay; multiple orthogonal methods; replicated across two independent papers same year\",\n      \"pmids\": [\"37442581\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Mouse REC114 forms homodimers and associates with MEI4 as a 2:1 REC114:MEI4 heterotrimer that further dimerizes; IHO1 forms coiled-coil-based tetramers; IHO1 directly interacts with the PH domain of REC114 via the same surface recognized by TOPOVIBL and ANKRD31, suggesting REC114 acts as a regulatory platform for mutually exclusive interactions.\",\n      \"method\": \"AlphaFold2 modeling, biochemical characterization (SEC, native MS), co-IP, mutagenesis\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — structural modeling validated by biochemical methods, direct interaction mapping, multiple orthogonal approaches; replicated architecture consistent with yeast studies\",\n      \"pmids\": [\"37431931\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Bi-allelic missense variants in human MEI4 (in the N-terminal region, amino acids 98–305) reduce MEI4–DNA interaction in vitro, and a knock-in mouse model with one variant causes female infertility with oogenesis developmental defects, demonstrating that MEI4's DNA-binding activity is required for its reproductive function.\",\n      \"method\": \"In vitro DNA interaction assay with variant MEI4 proteins, knock-in mouse model generation, oogenesis phenotype analysis\",\n      \"journal\": \"Human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro DNA interaction assay plus knock-in mouse model; single lab\",\n      \"pmids\": [\"38252283\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"The MEI4 C-terminus is required for stabilizing the MEI4–REC114 subcomplex on chromosome axes; Mei4Arg356* (C-terminal truncation) mice exhibit severe DSB formation defects, massive oocyte apoptosis via a HORMAD1-dependent synapsis checkpoint, and surviving oocytes show aneuploidy-driven complete embryonic arrest; heterozygous mice show intermediate follicle depletion consistent with haploinsufficiency.\",\n      \"method\": \"Exome sequencing of POI patients, in vitro DSB formation assay, knock-in mouse model (Mei4Arg356*), cytological analysis, embryonic development assay\",\n      \"journal\": \"Journal of genetics and genomics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — knock-in mouse model with defined molecular and cellular phenotypes, in vitro functional assay; single lab\",\n      \"pmids\": [\"41419020\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Mre11 is recruited to meiotic DSB sites via its C-terminal IDR, which contains a short α-helix that binds a conserved region of Mer2; this interaction is required for Mre11 foci formation during meiosis and DSB formation; the Mer2 interaction is part of the RMM (Rec114-Mei4-Mer2) condensation-dependent recruitment mechanism for Mre11.\",\n      \"method\": \"In vitro condensate assay, mutagenesis of Mer2-binding helix in Mre11, in vivo foci analysis\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro assay plus in vivo foci with mutagenesis; preprint, not yet peer-reviewed; Mei4 is part of the RMM complex implicated but focus is on Mre11-Mer2 interaction\",\n      \"pmids\": [\"bio_10.1101_2025.07.08.663703\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"MEI4 (and its orthologs across eukaryotes) is a conserved component of the meiotic DSB machinery that forms a 2:1 heterotrimeric complex with REC114 (two REC114 per one MEI4 N-terminal helix), localizes to chromosome axes in a manner dependent on HORMAD1, REC8, and IHO1/Mer2, and promotes SPO11-mediated DSB formation by cooperatively binding DNA and driving condensate formation that assembles the DSB machinery; in S. pombe and other fungi, the Mei4 ortholog additionally functions as a forkhead transcription factor activating middle-phase meiotic genes via FLEX-element binding.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MEI4 is a conserved component of the meiotic recombination machinery that promotes SPO11/Spo11-mediated DNA double-strand break (DSB) formation, acting genetically upstream of strand-exchange and repair factors such as RAD52 [#0, #7]. MEI4 functions as the core of a stable, conserved complex with REC114 and the Mer2 ortholog IHO1, in which two REC114 C-terminal dimers cup an N-terminal MEI4 α-helix to form a 2:1 REC114:MEI4 heterotrimer that can further oligomerize [#13, #15, #17]. This minimal REC114–MEI4 unit binds DNA cooperatively through two oppositely oriented DNA-binding sites and drives DNA-dependent condensate formation, bridging and condensing multiple DNA duplexes — the biophysical basis for assembling the DSB machinery [#15, #16]. MEI4 concentrates in discrete foci on meiotic chromosome axes distinct from downstream recombination markers, and its axis loading depends on the chromosome-axis factor HORMAD1 together with the cohesins REC8/RAD21L, with axis-associated MEI4 levels quantitatively limiting DSB formation [#7, #11]. MEI4's DNA-binding activity, residing in its N-terminal region, and the C-terminal segment that stabilizes the MEI4–REC114 subcomplex on axes are both required for fertility: bi-allelic and truncating MEI4 variants impair DNA binding or axis stabilization and cause oogenesis defects and premature ovarian insufficiency, modeled in knock-in mice [#18, #19]. In the fission yeast and budding yeast lineages, the Mei4 ortholog has an additional, distinct role as a forkhead/HNF3 transcription factor that binds the FLEX cis-element to activate middle-meiotic genes and autoregulate its own expression, coordinating recombination timing with premeiotic DNA replication [#2, #3, #5].\",\n  \"teleology\": [\n    {\n      \"year\": 1989,\n      \"claim\": \"Established where MEI4 acts in meiotic recombination, placing it upstream of the strand-exchange/repair machinery rather than within it.\",\n      \"evidence\": \"Genetic epistasis with spo13 suppression in S. cerevisiae mei4 spo13 rad52 triple mutants\",\n      \"pmids\": [\"2693205\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not define a molecular activity for MEI4\", \"No protein partners identified\", \"Mechanism of DSB promotion unknown\"]\n    },\n    {\n      \"year\": 1992,\n      \"claim\": \"Defined MEI4 as a meiosis-specific gene whose product is required for synaptonemal complex formation, linking it to chromosome morphogenesis.\",\n      \"evidence\": \"Sequencing, Northern blot, and cytology of meiotic chromosomes in S. cerevisiae mei4 deletion strains\",\n      \"pmids\": [\"1545815\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether synapsis failure is a direct consequence of failed DSB formation not resolved\", \"No biochemical activity assigned\"]\n    },\n    {\n      \"year\": 1998,\n      \"claim\": \"Revealed that the S. pombe Mei4 ortholog is a sequence-specific forkhead transcription factor, defining a transcriptional function distinct from the DSB role in budding yeast/mouse.\",\n      \"evidence\": \"Complementation cloning, gel-shift with recombinant forkhead domain, one-hybrid and promoter deletion in S. pombe\",\n      \"pmids\": [\"9528784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full target-gene repertoire not yet mapped\", \"Relationship between fission-yeast transcriptional role and DSB-machinery role of orthologs unaddressed\"]\n    },\n    {\n      \"year\": 2000,\n      \"claim\": \"Identified the FLEX cis-element as the genomic recognition site through which S. pombe Mei4 activates middle-meiotic genes and autoregulates itself, building the transcriptional regulatory logic.\",\n      \"evidence\": \"Genome-wide promoter scanning, Northern blots, forkhead-FLEX binding, and reporter assays in S. pombe\",\n      \"pmids\": [\"10747048\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Co-factors required for activation not yet identified\", \"How FLEX targets are selected genome-wide unclear\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Showed that mei4+ transcription is gated by the replication checkpoint, coupling onset of meiotic recombination to completion of premeiotic DNA replication.\",\n      \"evidence\": \"Hydroxyurea replication block with rad3/cds1/chk1/mek1 mutants and Northern blot in S. pombe\",\n      \"pmids\": [\"16286472\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct checkpoint target within the mei4 promoter not defined\", \"Applies to the transcriptional role; checkpoint control of the metazoan DSB role separate\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Defined the Mei4–Rec114–Mer2 complex as a discrete physical and cytological entity required for DSB formation, distinct from the Mre11/Rec102 machinery.\",\n      \"evidence\": \"Co-immunoprecipitation and cytological co-localization on meiotic chromosomes in S. cerevisiae\",\n      \"pmids\": [\"16783010\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Stoichiometry and direct binding interfaces not resolved\", \"Biochemical activity of the complex unknown\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Mapped the interaction hierarchy showing Mei4 axis localization depends on Rec114/Mer2 and connecting the subcomplex to Spo11 via Rec102.\",\n      \"evidence\": \"Two-hybrid, deletion mapping, and cytological localization in deletion backgrounds in S. cerevisiae\",\n      \"pmids\": [\"17558514\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct vs indirect nature of Mei4-Spo11 link not established\", \"Interaction regions not resolved structurally\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Demonstrated functional conservation of MEI4 to mammals, showing it is essential for DSB formation, localizes to axis foci, and directly binds REC114 via conserved motifs.\",\n      \"evidence\": \"Mei4-/- mouse phenotyping, immunofluorescence, and pulldown/co-IP with motif mutagenesis\",\n      \"pmids\": [\"20551173\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Complex stoichiometry and structure not yet defined\", \"How axis foci promote SPO11 cleavage unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Showed the S. pombe DSB-pathway ortholog Rec24 is required genome-wide for crossing-over and is stabilized on linear elements by Rec7, separating axis loading from Spo11 activity.\",\n      \"evidence\": \"Gene disruption, cytological localization, and co-IP in S. pombe\",\n      \"pmids\": [\"21429938\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of linear-element recruitment unresolved\", \"Direct DNA-binding not tested here\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Extended the fission-yeast transcription-factor role to meiotic chromosome dynamics, showing forkhead-domain-dependent control of telomere dispersal at meiosis I.\",\n      \"evidence\": \"Deletion and temperature-sensitive forkhead-domain point mutant with live-cell telomere imaging in S. pombe\",\n      \"pmids\": [\"21449049\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct transcriptional targets driving telomere dispersal not identified\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Linked Mei4 transcriptional output to timing of meiotic nuclear movements under checkpoint control, reinforcing replication-coupled regulation of meiotic progression.\",\n      \"evidence\": \"Deletion/overproduction with live-cell imaging and checkpoint-mutant analysis in S. pombe\",\n      \"pmids\": [\"25492408\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Effector genes mediating nuclear-movement termination not defined\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified the chromosome-axis determinants of MEI4 localization and established that axis-bound MEI4 is a quantitative limiting factor for DSB formation.\",\n      \"evidence\": \"HORMAD1, MEI1, REC8, RAD21L knockout mice with quantitative immunofluorescence of MEI4 and DSB markers\",\n      \"pmids\": [\"25795304\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct vs indirect dependence of MEI4 on each axis factor not dissected\", \"Mechanism converting axis loading into cleavage unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Resolved the mammalian REC114–MEI4–IHO1 complex biochemically and structurally, reconstituting the REC114-CTD:MEI4-NTD interaction and solving the REC114 N-terminal PH-like domain.\",\n      \"evidence\": \"REC114 KO mouse, co-IP from spermatocytes, in vitro domain reconstitution, X-ray crystallography\",\n      \"pmids\": [\"30569039\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Complex stoichiometry not yet quantified\", \"DNA-binding/condensation activity not yet characterized\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined the molecular architecture and biochemical activity of the complex: a 2:1 REC114:MEI4 heterotrimer with opposed DNA-binding sites that drives cooperative DNA binding and condensation, providing the physical mechanism for DSB-machinery assembly.\",\n      \"evidence\": \"NMR/SAXS, AlphaFold2 modeling, in vitro reconstitution, single-molecule DNA condensation, and mutagenesis (S. cerevisiae and conserved models)\",\n      \"pmids\": [\"37442580\", \"37442581\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How condensates direct SPO11 to specific sites not resolved\", \"In vivo relevance of condensation vs other functions\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established REC114 as a regulatory platform: the same PH-domain surface mediates mutually exclusive interactions with IHO1, TOPOVIBL, and ANKRD31, organizing the higher-order DSB machinery around the MEI4 heterotrimer.\",\n      \"evidence\": \"AlphaFold2 modeling with SEC, native MS, co-IP, and mutagenesis in the mouse system\",\n      \"pmids\": [\"37431931\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Temporal ordering of competing interactions in vivo unknown\", \"Functional consequence of heterotrimer dimerization not defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Connected MEI4 to human disease by showing N-terminal variants reduce DNA binding and cause oogenesis defects, establishing DNA-binding activity as required for reproductive function.\",\n      \"evidence\": \"In vitro DNA interaction assays with variant proteins and a knock-in mouse model with oogenesis phenotyping\",\n      \"pmids\": [\"38252283\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single lab; allelic spectrum in patients limited\", \"Whether variants affect condensation as well as binding not tested\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Showed the MEI4 C-terminus stabilizes the MEI4–REC114 subcomplex on axes and that its truncation causes severe DSB defects, oocyte apoptosis via the HORMAD1-dependent synapsis checkpoint, and premature ovarian insufficiency, with haploinsufficiency.\",\n      \"evidence\": \"POI exome sequencing, in vitro DSB assay, Mei4Arg356* knock-in mouse with cytology and embryonic-development analysis\",\n      \"pmids\": [\"41419020\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of C-terminal axis stabilization not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the RMM condensate is converted into site-specific SPO11 cleavage — i.e. how DNA condensation and axis tethering are mechanistically coupled to catalytic DSB formation — remains unresolved.\",\n      \"evidence\": \"No direct evidence in the timeline reconstitutes SPO11 cleavage from the assembled RMM condensate\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No reconstitution of SPO11 cleavage from the RMM complex\", \"Mechanism coupling condensation to catalysis unknown\", \"Mre11 recruitment via Mer2 only shown in a preprint (idx 20)\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [2, 3, 15, 16, 18]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [13, 15, 17]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [4, 7, 11]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [2, 12]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1474165\", \"supporting_discovery_ids\": [7, 18, 19]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [0, 7, 11]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [2, 3]}\n    ],\n    \"complexes\": [\"REC114-MEI4-Mer2/IHO1 (RMM) complex\"],\n    \"partners\": [\"REC114\", \"IHO1\", \"MER2\", \"REC102\", \"HORMAD1\", \"CUF2\", \"REC7\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}