{"gene":"M1AP","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":2013,"finding":"M1AP (mouse M1ap) is expressed exclusively in germ cells from spermatogonia to secondary spermatocytes, and its knockout in male mice causes meiotic arrest predominantly at metaphase I with failure to properly align chromosomes at the metaphase plate due to abnormal chromosome synapses and failure to form crossover foci, leading to severe oligozoospermia and infertility; female knockouts have histologically normal ovaries and are fertile.","method":"Knockout mouse model (M1ap-deficient mice); histological analysis, immunostaining for meiotic markers, chromosome synapsis analysis","journal":"Biology of reproduction","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean KO mouse with defined cellular phenotype, multiple orthogonal analyses (histology, chromosome synapsis, crossover foci), replicated by subsequent studies","pmids":["23269666"],"is_preprint":false},{"year":2006,"finding":"D6Mm5e (M1AP) mRNA expression is highly restricted to germ cells: expressed in female embryonic germ cells upon entry into meiosis and in male germ cells during late spermatogenesis. Two transcripts arise from alternative splicing of exon 8 without frameshift, using the same stop codon. The protein does not belong to any known protein family and lacks known signature motifs.","method":"RT-PCR, Northern blot, in situ hybridization, sequence analysis","journal":"Developmental dynamics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal expression methods (RT-PCR, Northern blot, ISH) in single study establishing tissue-specific expression pattern","pmids":["16881047"],"is_preprint":false},{"year":2022,"finding":"M1AP physically interacts with the mammalian ZZS complex components SHOC1, TEX11, and SPO16; M1AP localizes to chromosomal axes in a SPO16-dependent manner and colocalizes with TEX11. Ablation of M1AP does not alter SHOC1 localization but reduces recruitment of TEX11 to recombination intermediates, resulting in decreased recombination intermediates and crossovers.","method":"Co-immunoprecipitation/interaction assays identifying M1AP-ZZS binding; immunostaining of chromosome spreads in M1AP-mutant mouse models; analysis of recombination intermediate and crossover numbers in patient-derived and mouse models","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal interaction assays, genetic epistasis via mouse models, multiple orthogonal methods (co-IP, immunostaining, crossover quantification) in single study","pmids":["36440627"],"is_preprint":false},{"year":2022,"finding":"A homozygous M1AP splicing mutation (c.1074+2T>C) abolishes M1AP foci on chromosome axes and leads to metaphase I arrest with decreased crossovers in male mice; M1AP is dispensable for crossover formation and fertility in female mice, showing cytoplasmic (not axial) localization in fetal oocytes.","method":"Patient genetic analysis; mouse model of splicing mutation; immunostaining of chromosome spreads for M1AP foci and crossover markers","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — combination of human patient variant, mouse model, and cytological analysis with multiple markers in same study","pmids":["36440627"],"is_preprint":false},{"year":2025,"finding":"In human males, loss-of-function of M1AP leads to a predominant metaphase I arrest with rare haploid spermatids, caused by reduced recombination intermediates and class I crossover failure (but not incorrect synapsis or unrepaired DNA double-strand breaks, which distinguish ZZS deficiency). This establishes M1AP as an important but non-essential functional enhancer of meiotic recombination, downstream of ZZS-mediated synapsis.","method":"Testicular biopsy phenotyping of men with biallelic/hemizygous M1AP loss-of-function variants; immunostaining for recombination and synapsis markers; genetic epistasis comparison with ZZS-deficient patients","journal":"EMBO molecular medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — in-depth human testicular phenotyping with multiple cytological markers, genetic epistasis with ZZS pathway, replicated across 24 individuals","pmids":["40374915"],"is_preprint":false},{"year":2020,"finding":"A homozygous M1AP frameshift variant (c.676dup, p.Trp226LeufsTer4) results in a truncated protein as demonstrated by heterologous in vitro expression of mutant M1AP, and causes non-obstructive azoospermia in humans.","method":"In vitro heterologous expression of mutant M1AP; whole-exome sequencing; segregation analysis in consanguineous family","journal":"American journal of human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro expression confirms truncation, multiple independent patients, but mechanistic detail limited to protein truncation confirmation","pmids":["32673564"],"is_preprint":false},{"year":2020,"finding":"A homozygous splice-site mutation in M1AP (c.1435-1G>A) leads to complete absence of M1AP protein in patient spermatozoa and is associated with severe oligozoospermia and highly aberrant swollen mitochondrial sheaths with normal axonemal structures.","method":"Protein immunostaining of patient spermatozoa; electron microscopy of sperm ultrastructure; segregation analysis","journal":"Clinical genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct protein absence confirmed by immunostaining in patient tissue, ultrastructural phenotype characterized, single family/patient","pmids":["32017041"],"is_preprint":false},{"year":2020,"finding":"CRISPR activation of M1AP in HEK293T cells increases MYC promoter activity and MYC mRNA/protein levels, as confirmed by doxycycline-inducible M1AP overexpression, dual-luciferase assay, and quantitative RT-PCR/western blot. This identifies M1AP as a transcriptional activator of MYC expression in this cellular context.","method":"CRISPR activation screening with MYC promoter-reporter (pMYC-promoter-Dendra2); doxycycline-inducible M1AP overexpression; dual-luciferase assay; qRT-PCR; western blot","journal":"PeerJ","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, overexpression system in non-physiological cell line (HEK293T), no mechanistic detail of how M1AP activates MYC, no loss-of-function or in vivo validation","pmids":["32411526"],"is_preprint":false},{"year":2025,"finding":"Disruption of the XPF-like domain in SHOC1 impairs recruitment of M1AP to recombination intermediates, indicating that SHOC1 (via its XPF-like domain) is required for M1AP localization to recombination sites.","method":"Immunostaining of chromosome spreads in SHOC1 XPF-domain mutant mouse/patient models; functional analysis of SHOC1 mutant effects on M1AP foci","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct cytological evidence of M1AP recruitment failure in SHOC1 mutant context, preprint, single study","pmids":["bio_10.1101_2025.05.28.656576"],"is_preprint":true}],"current_model":"M1AP (meiosis 1 associated protein) is a germ cell-specific protein that acts as a functional enhancer of meiotic recombination by interacting with the mammalian ZZS complex (SHOC1, TEX11, SPO16): it localizes to chromosomal axes in a SPO16-dependent manner, promotes recruitment of TEX11 to recombination intermediates, and facilitates class I crossover formation; loss of M1AP causes reduced crossovers and predominantly metaphase I arrest in males (but is dispensable in females), while its localization to recombination sites itself depends on the XPF-like domain of SHOC1."},"narrative":{"mechanistic_narrative":"M1AP is a germ cell-specific protein that functions as a functional enhancer of meiotic recombination, with its loss causing male meiotic arrest and infertility [PMID:23269666, PMID:36440627]. Its expression is restricted to germ cells, appearing in female germ cells upon meiotic entry and in male germ cells during spermatogenesis [PMID:16881047]. Mechanistically, M1AP physically associates with the mammalian ZZS complex (SHOC1, TEX11, SPO16) and localizes to chromosomal axes in a SPO16-dependent manner, colocalizing with TEX11; loss of M1AP does not perturb SHOC1 localization but reduces recruitment of TEX11 to recombination intermediates, decreasing recombination intermediates and class I crossovers [PMID:36440627], and its own recruitment to recombination sites depends on the XPF-like domain of SHOC1 [PMID:bio_10.1101_2025.05.28.656576]. In human males, biallelic or hemizygous loss-of-function variants cause predominant metaphase I arrest with reduced crossovers but, unlike ZZS deficiency, preserve synapsis and DSB repair, placing M1AP downstream of ZZS-mediated synapsis as a non-essential enhancer of crossover formation [PMID:40374915]; the gene is dispensable for female fertility, where M1AP localizes cytoplasmically rather than axially in fetal oocytes [PMID:36440627]. M1AP loss-of-function causes non-obstructive azoospermia and severe oligozoospermia in human patients [PMID:32673564, PMID:32017041].","teleology":[{"year":2006,"claim":"Established that M1AP is a germ cell-restricted gene of unknown family, raising the question of a meiosis-specific function despite the absence of recognizable domains.","evidence":"RT-PCR, Northern blot, in situ hybridization, and sequence analysis of germ cell expression","pmids":["16881047"],"confidence":"Medium","gaps":["No protein function or interaction partners identified","No structural or domain assignment possible from sequence"]},{"year":2013,"claim":"Defined the cellular phenotype of M1AP loss, showing it is required for male meiotic progression past metaphase I via proper synapsis and crossover formation, while being dispensable in females.","evidence":"Knockout mouse model with histology, chromosome synapsis analysis, and crossover foci immunostaining","pmids":["23269666"],"confidence":"High","gaps":["Molecular mechanism of how M1AP supports crossover formation unknown","No direct interaction partners identified","Basis of sexual dimorphism unexplained"]},{"year":2020,"claim":"Confirmed M1AP as a human male infertility gene by linking loss-of-function variants to azoospermia and oligozoospermia, translating the mouse phenotype to humans.","evidence":"Whole-exome sequencing, segregation analysis, in vitro expression of truncated protein, and patient sperm immunostaining/electron microscopy","pmids":["32673564","32017041"],"confidence":"Medium","gaps":["Mechanistic detail limited to protein truncation/absence","Mitochondrial sheath phenotype not mechanistically connected to meiotic role","Single families for some variants"]},{"year":2020,"claim":"Reported a non-meiotic transcriptional activity in which M1AP overexpression activates MYC, a context not reconciled with its germ cell meiotic role.","evidence":"CRISPR activation screen with MYC promoter reporter, inducible overexpression, luciferase, qRT-PCR, and western blot in HEK293T cells","pmids":["32411526"],"confidence":"Low","gaps":["Non-physiological overexpression in HEK293T; no loss-of-function or in vivo validation","No mechanism for how M1AP activates MYC","Relationship to meiotic function unestablished"]},{"year":2022,"claim":"Resolved the molecular mechanism by placing M1AP within the ZZS recombination machinery, showing it localizes to axes via SPO16 and promotes TEX11 recruitment to recombination intermediates.","evidence":"Co-immunoprecipitation interaction assays, immunostaining of chromosome spreads in M1AP-mutant and patient-derived models, and crossover quantification","pmids":["36440627"],"confidence":"High","gaps":["Structural basis of M1AP-ZZS interaction unknown","Direct binding stoichiometry/architecture within complex undefined","How M1AP enhances TEX11 recruitment biochemically unresolved"]},{"year":2025,"claim":"Refined M1AP's position in the pathway by showing human loss-of-function impairs class I crossovers without disrupting synapsis or DSB repair, distinguishing it as a non-essential enhancer downstream of ZZS-mediated synapsis.","evidence":"Testicular biopsy phenotyping of men with biallelic/hemizygous variants, recombination/synapsis marker immunostaining, and genetic epistasis with ZZS-deficient patients","pmids":["40374915"],"confidence":"High","gaps":["Why M1AP loss is partial rather than complete crossover block unexplained","Molecular step at which M1AP acts within crossover maturation undefined"]},{"year":2025,"claim":"Identified the upstream determinant of M1AP recruitment, showing the SHOC1 XPF-like domain is required to localize M1AP to recombination intermediates.","evidence":"Immunostaining of chromosome spreads in SHOC1 XPF-domain mutant models (preprint)","pmids":["bio_10.1101_2025.05.28.656576"],"confidence":"Medium","gaps":["Preprint, single study","Whether interaction with SHOC1 is direct not established","Structural detail of XPF-domain dependence unknown"]},{"year":null,"claim":"The biochemical mechanism by which M1AP enhances TEX11 recruitment and crossover maturation, and the structural architecture of its association with the ZZS complex, remain undefined.","evidence":"","pmids":[],"confidence":"High","gaps":["No structural model of M1AP or its ZZS interface","No reconstituted biochemical assay of M1AP activity","Reconciliation of meiotic role with reported MYC transcriptional activity outstanding"]}],"mechanism_profile":{"molecular_activity":[],"localization":[{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[2,3]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[3]}],"pathway":[{"term_id":"R-HSA-1474165","term_label":"Reproduction","supporting_discovery_ids":[0,2,4]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[0,4]}],"complexes":[],"partners":["SHOC1","TEX11","SPO16"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8TC57","full_name":"Meiosis 1 arrest protein","aliases":["Meiosis 1-arresting protein","Meiosis 1-associated protein","Spermatogenesis-associated protein 37"],"length_aa":530,"mass_kda":59.4,"function":"Required for meiosis I progression during spermatogenesis","subcellular_location":"Cytoplasm","url":"https://www.uniprot.org/uniprotkb/Q8TC57/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/M1AP","classification":"Not Classified","n_dependent_lines":6,"n_total_lines":1208,"dependency_fraction":0.004966887417218543},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/M1AP","total_profiled":1310},"omim":[{"mim_id":"619108","title":"SPERMATOGENIC FAILURE 48; SPGF48","url":"https://www.omim.org/entry/619108"},{"mim_id":"619098","title":"MEIOSIS 1-ASSOCIATED PROTEIN; M1AP","url":"https://www.omim.org/entry/619098"},{"mim_id":"258150","title":"SPERMATOGENIC FAILURE 1; SPGF1","url":"https://www.omim.org/entry/258150"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Mitochondria","reliability":"Approved"},{"location":"Nucleoli","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"testis","ntpm":6.0}],"url":"https://www.proteinatlas.org/search/M1AP"},"hgnc":{"alias_symbol":["D6Mm5e","SPATA37"],"prev_symbol":["C2orf65"]},"alphafold":{"accession":"Q8TC57","domains":[{"cath_id":"3.40.50.410","chopping":"24-183_202-227","consensus_level":"high","plddt":86.4528,"start":24,"end":227},{"cath_id":"2.40.290,2.40.290","chopping":"238-272_291-410","consensus_level":"high","plddt":80.9434,"start":238,"end":410}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TC57","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TC57-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TC57-F1-predicted_aligned_error_v6.png","plddt_mean":73.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=M1AP","jax_strain_url":"https://www.jax.org/strain/search?query=M1AP"},"sequence":{"accession":"Q8TC57","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8TC57.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8TC57/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TC57"}},"corpus_meta":[{"pmid":"32673564","id":"PMC_32673564","title":"Bi-allelic Mutations in M1AP Are a Frequent Cause of Meiotic Arrest and Severely Impaired Spermatogenesis Leading to Male Infertility.","date":"2020","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/32673564","citation_count":69,"is_preprint":false},{"pmid":"23269666","id":"PMC_23269666","title":"Meiosis I arrest abnormalities lead to severe oligozoospermia in meiosis 1 arresting protein (M1ap)-deficient mice.","date":"2013","source":"Biology of reproduction","url":"https://pubmed.ncbi.nlm.nih.gov/23269666","citation_count":28,"is_preprint":false},{"pmid":"32017041","id":"PMC_32017041","title":"An M1AP homozygous splice-site mutation associated with severe oligozoospermia in a consanguineous family.","date":"2020","source":"Clinical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/32017041","citation_count":18,"is_preprint":false},{"pmid":"36440627","id":"PMC_36440627","title":"M1AP interacts with the mammalian ZZS complex and promotes male meiotic recombination.","date":"2022","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/36440627","citation_count":17,"is_preprint":false},{"pmid":"38073178","id":"PMC_38073178","title":"Genome sequencing of Pakistani families with male infertility identifies deleterious genotypes in SPAG6, CCDC9, TKTL1, TUBA3C, and M1AP.","date":"2023","source":"Andrology","url":"https://pubmed.ncbi.nlm.nih.gov/38073178","citation_count":17,"is_preprint":false},{"pmid":"16881047","id":"PMC_16881047","title":"Expression analysis and evolutionary conservation of the mouse germ cell-specific D6Mm5e gene.","date":"2006","source":"Developmental dynamics : an official publication of the American Association of Anatomists","url":"https://pubmed.ncbi.nlm.nih.gov/16881047","citation_count":11,"is_preprint":false},{"pmid":"34604049","id":"PMC_34604049","title":"Genetic Variants of CLPP and M1AP Are Associated With Risk of Non-Small Cell Lung Cancer.","date":"2021","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/34604049","citation_count":9,"is_preprint":false},{"pmid":"32411526","id":"PMC_32411526","title":"CRISPR screening identifies M1AP as a new MYC regulator with a promoter-reporter system.","date":"2020","source":"PeerJ","url":"https://pubmed.ncbi.nlm.nih.gov/32411526","citation_count":7,"is_preprint":false},{"pmid":"35341049","id":"PMC_35341049","title":"Structural analysis of M1AP variants associated with severely impaired spermatogenesis causing male infertility.","date":"2022","source":"PeerJ","url":"https://pubmed.ncbi.nlm.nih.gov/35341049","citation_count":4,"is_preprint":false},{"pmid":"40374915","id":"PMC_40374915","title":"Genotype-specific differences in infertile men due to loss-of-function variants in M1AP or ZZS genes.","date":"2025","source":"EMBO molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40374915","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.28.656576","title":"The XPF-like domain in SHOC1 required for homologous recombination and safeguarding autosome from meiotic silencing of unsynapsed chromatin","date":"2025-06-01","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.28.656576","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":7816,"output_tokens":2351,"usd":0.029357,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9453,"output_tokens":2583,"usd":0.05592,"stage2_stop_reason":"end_turn"},"total_usd":0.085277,"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\": 2013,\n      \"finding\": \"M1AP (mouse M1ap) is expressed exclusively in germ cells from spermatogonia to secondary spermatocytes, and its knockout in male mice causes meiotic arrest predominantly at metaphase I with failure to properly align chromosomes at the metaphase plate due to abnormal chromosome synapses and failure to form crossover foci, leading to severe oligozoospermia and infertility; female knockouts have histologically normal ovaries and are fertile.\",\n      \"method\": \"Knockout mouse model (M1ap-deficient mice); histological analysis, immunostaining for meiotic markers, chromosome synapsis analysis\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean KO mouse with defined cellular phenotype, multiple orthogonal analyses (histology, chromosome synapsis, crossover foci), replicated by subsequent studies\",\n      \"pmids\": [\"23269666\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"D6Mm5e (M1AP) mRNA expression is highly restricted to germ cells: expressed in female embryonic germ cells upon entry into meiosis and in male germ cells during late spermatogenesis. Two transcripts arise from alternative splicing of exon 8 without frameshift, using the same stop codon. The protein does not belong to any known protein family and lacks known signature motifs.\",\n      \"method\": \"RT-PCR, Northern blot, in situ hybridization, sequence analysis\",\n      \"journal\": \"Developmental dynamics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal expression methods (RT-PCR, Northern blot, ISH) in single study establishing tissue-specific expression pattern\",\n      \"pmids\": [\"16881047\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"M1AP physically interacts with the mammalian ZZS complex components SHOC1, TEX11, and SPO16; M1AP localizes to chromosomal axes in a SPO16-dependent manner and colocalizes with TEX11. Ablation of M1AP does not alter SHOC1 localization but reduces recruitment of TEX11 to recombination intermediates, resulting in decreased recombination intermediates and crossovers.\",\n      \"method\": \"Co-immunoprecipitation/interaction assays identifying M1AP-ZZS binding; immunostaining of chromosome spreads in M1AP-mutant mouse models; analysis of recombination intermediate and crossover numbers in patient-derived and mouse models\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal interaction assays, genetic epistasis via mouse models, multiple orthogonal methods (co-IP, immunostaining, crossover quantification) in single study\",\n      \"pmids\": [\"36440627\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A homozygous M1AP splicing mutation (c.1074+2T>C) abolishes M1AP foci on chromosome axes and leads to metaphase I arrest with decreased crossovers in male mice; M1AP is dispensable for crossover formation and fertility in female mice, showing cytoplasmic (not axial) localization in fetal oocytes.\",\n      \"method\": \"Patient genetic analysis; mouse model of splicing mutation; immunostaining of chromosome spreads for M1AP foci and crossover markers\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — combination of human patient variant, mouse model, and cytological analysis with multiple markers in same study\",\n      \"pmids\": [\"36440627\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In human males, loss-of-function of M1AP leads to a predominant metaphase I arrest with rare haploid spermatids, caused by reduced recombination intermediates and class I crossover failure (but not incorrect synapsis or unrepaired DNA double-strand breaks, which distinguish ZZS deficiency). This establishes M1AP as an important but non-essential functional enhancer of meiotic recombination, downstream of ZZS-mediated synapsis.\",\n      \"method\": \"Testicular biopsy phenotyping of men with biallelic/hemizygous M1AP loss-of-function variants; immunostaining for recombination and synapsis markers; genetic epistasis comparison with ZZS-deficient patients\",\n      \"journal\": \"EMBO molecular medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in-depth human testicular phenotyping with multiple cytological markers, genetic epistasis with ZZS pathway, replicated across 24 individuals\",\n      \"pmids\": [\"40374915\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"A homozygous M1AP frameshift variant (c.676dup, p.Trp226LeufsTer4) results in a truncated protein as demonstrated by heterologous in vitro expression of mutant M1AP, and causes non-obstructive azoospermia in humans.\",\n      \"method\": \"In vitro heterologous expression of mutant M1AP; whole-exome sequencing; segregation analysis in consanguineous family\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro expression confirms truncation, multiple independent patients, but mechanistic detail limited to protein truncation confirmation\",\n      \"pmids\": [\"32673564\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"A homozygous splice-site mutation in M1AP (c.1435-1G>A) leads to complete absence of M1AP protein in patient spermatozoa and is associated with severe oligozoospermia and highly aberrant swollen mitochondrial sheaths with normal axonemal structures.\",\n      \"method\": \"Protein immunostaining of patient spermatozoa; electron microscopy of sperm ultrastructure; segregation analysis\",\n      \"journal\": \"Clinical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct protein absence confirmed by immunostaining in patient tissue, ultrastructural phenotype characterized, single family/patient\",\n      \"pmids\": [\"32017041\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CRISPR activation of M1AP in HEK293T cells increases MYC promoter activity and MYC mRNA/protein levels, as confirmed by doxycycline-inducible M1AP overexpression, dual-luciferase assay, and quantitative RT-PCR/western blot. This identifies M1AP as a transcriptional activator of MYC expression in this cellular context.\",\n      \"method\": \"CRISPR activation screening with MYC promoter-reporter (pMYC-promoter-Dendra2); doxycycline-inducible M1AP overexpression; dual-luciferase assay; qRT-PCR; western blot\",\n      \"journal\": \"PeerJ\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, overexpression system in non-physiological cell line (HEK293T), no mechanistic detail of how M1AP activates MYC, no loss-of-function or in vivo validation\",\n      \"pmids\": [\"32411526\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Disruption of the XPF-like domain in SHOC1 impairs recruitment of M1AP to recombination intermediates, indicating that SHOC1 (via its XPF-like domain) is required for M1AP localization to recombination sites.\",\n      \"method\": \"Immunostaining of chromosome spreads in SHOC1 XPF-domain mutant mouse/patient models; functional analysis of SHOC1 mutant effects on M1AP foci\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct cytological evidence of M1AP recruitment failure in SHOC1 mutant context, preprint, single study\",\n      \"pmids\": [\"bio_10.1101_2025.05.28.656576\"],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"M1AP (meiosis 1 associated protein) is a germ cell-specific protein that acts as a functional enhancer of meiotic recombination by interacting with the mammalian ZZS complex (SHOC1, TEX11, SPO16): it localizes to chromosomal axes in a SPO16-dependent manner, promotes recruitment of TEX11 to recombination intermediates, and facilitates class I crossover formation; loss of M1AP causes reduced crossovers and predominantly metaphase I arrest in males (but is dispensable in females), while its localization to recombination sites itself depends on the XPF-like domain of SHOC1.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"M1AP is a germ cell-specific protein that functions as a functional enhancer of meiotic recombination, with its loss causing male meiotic arrest and infertility [#0, #2]. Its expression is restricted to germ cells, appearing in female germ cells upon meiotic entry and in male germ cells during spermatogenesis [#1]. Mechanistically, M1AP physically associates with the mammalian ZZS complex (SHOC1, TEX11, SPO16) and localizes to chromosomal axes in a SPO16-dependent manner, colocalizing with TEX11; loss of M1AP does not perturb SHOC1 localization but reduces recruitment of TEX11 to recombination intermediates, decreasing recombination intermediates and class I crossovers [#2], and its own recruitment to recombination sites depends on the XPF-like domain of SHOC1 [#8]. In human males, biallelic or hemizygous loss-of-function variants cause predominant metaphase I arrest with reduced crossovers but, unlike ZZS deficiency, preserve synapsis and DSB repair, placing M1AP downstream of ZZS-mediated synapsis as a non-essential enhancer of crossover formation [#4]; the gene is dispensable for female fertility, where M1AP localizes cytoplasmically rather than axially in fetal oocytes [#3]. M1AP loss-of-function causes non-obstructive azoospermia and severe oligozoospermia in human patients [#5, #6].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established that M1AP is a germ cell-restricted gene of unknown family, raising the question of a meiosis-specific function despite the absence of recognizable domains.\",\n      \"evidence\": \"RT-PCR, Northern blot, in situ hybridization, and sequence analysis of germ cell expression\",\n      \"pmids\": [\"16881047\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No protein function or interaction partners identified\", \"No structural or domain assignment possible from sequence\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Defined the cellular phenotype of M1AP loss, showing it is required for male meiotic progression past metaphase I via proper synapsis and crossover formation, while being dispensable in females.\",\n      \"evidence\": \"Knockout mouse model with histology, chromosome synapsis analysis, and crossover foci immunostaining\",\n      \"pmids\": [\"23269666\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism of how M1AP supports crossover formation unknown\", \"No direct interaction partners identified\", \"Basis of sexual dimorphism unexplained\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Confirmed M1AP as a human male infertility gene by linking loss-of-function variants to azoospermia and oligozoospermia, translating the mouse phenotype to humans.\",\n      \"evidence\": \"Whole-exome sequencing, segregation analysis, in vitro expression of truncated protein, and patient sperm immunostaining/electron microscopy\",\n      \"pmids\": [\"32673564\", \"32017041\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanistic detail limited to protein truncation/absence\", \"Mitochondrial sheath phenotype not mechanistically connected to meiotic role\", \"Single families for some variants\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Reported a non-meiotic transcriptional activity in which M1AP overexpression activates MYC, a context not reconciled with its germ cell meiotic role.\",\n      \"evidence\": \"CRISPR activation screen with MYC promoter reporter, inducible overexpression, luciferase, qRT-PCR, and western blot in HEK293T cells\",\n      \"pmids\": [\"32411526\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Non-physiological overexpression in HEK293T; no loss-of-function or in vivo validation\", \"No mechanism for how M1AP activates MYC\", \"Relationship to meiotic function unestablished\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Resolved the molecular mechanism by placing M1AP within the ZZS recombination machinery, showing it localizes to axes via SPO16 and promotes TEX11 recruitment to recombination intermediates.\",\n      \"evidence\": \"Co-immunoprecipitation interaction assays, immunostaining of chromosome spreads in M1AP-mutant and patient-derived models, and crossover quantification\",\n      \"pmids\": [\"36440627\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of M1AP-ZZS interaction unknown\", \"Direct binding stoichiometry/architecture within complex undefined\", \"How M1AP enhances TEX11 recruitment biochemically unresolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Refined M1AP's position in the pathway by showing human loss-of-function impairs class I crossovers without disrupting synapsis or DSB repair, distinguishing it as a non-essential enhancer downstream of ZZS-mediated synapsis.\",\n      \"evidence\": \"Testicular biopsy phenotyping of men with biallelic/hemizygous variants, recombination/synapsis marker immunostaining, and genetic epistasis with ZZS-deficient patients\",\n      \"pmids\": [\"40374915\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why M1AP loss is partial rather than complete crossover block unexplained\", \"Molecular step at which M1AP acts within crossover maturation undefined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified the upstream determinant of M1AP recruitment, showing the SHOC1 XPF-like domain is required to localize M1AP to recombination intermediates.\",\n      \"evidence\": \"Immunostaining of chromosome spreads in SHOC1 XPF-domain mutant models (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.05.28.656576\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Preprint, single study\", \"Whether interaction with SHOC1 is direct not established\", \"Structural detail of XPF-domain dependence unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The biochemical mechanism by which M1AP enhances TEX11 recruitment and crossover maturation, and the structural architecture of its association with the ZZS complex, remain undefined.\",\n      \"evidence\": null,\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No structural model of M1AP or its ZZS interface\", \"No reconstituted biochemical assay of M1AP activity\", \"Reconciliation of meiotic role with reported MYC transcriptional activity outstanding\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [],\n    \"localization\": [\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1474165\", \"supporting_discovery_ids\": [0, 2, 4]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [0, 4]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"SHOC1\", \"TEX11\", \"SPO16\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":5,"faith_total":5,"faith_pct":100.0}}