{"gene":"DNAH3","run_date":"2026-04-28T17:46:02","timeline":{"discoveries":[{"year":2024,"finding":"Bi-allelic loss-of-function variants in DNAH3 cause asthenoteratozoospermia in humans and mice. DNAH3 localizes to sperm flagella and its deficiency results in loss of the central pair of microtubules, dislocated mitochondrial sheath and fibrous sheath, and partial absence of inner dynein arms (IDAs). Dnah3 knockout mice recapitulate the human phenotype with severe reduction in sperm movement.","method":"Whole-exome sequencing, Dnah3 knockout mouse generation, H&E staining, transmission electron microscopy, immunostaining, CASA","journal":"Human reproduction open","confidence":"High","confidence_rationale":"Tier 2 — KO mouse phenotype with multiple orthogonal methods (TEM, immunostaining, CASA), replicated in two independent KO lines and patient samples","pmids":["38312775"],"is_preprint":false},{"year":2024,"finding":"DNAH3 is a component of the inner dynein arm (IDA) in sperm flagella. Biallelic DNAH3 variants in infertile men cause deficient DNAH3 expression in sperm flagella, severe disruption of flagellar ultrastructure, and decreased expression of other IDA-associated proteins DNAH1, DNAH6, and DNALI1, indicating DNAH3 is required for IDA assembly.","method":"Whole-exome sequencing, immunofluorescence staining, transmission electron microscopy, Dnah3 knockout mice, Western blot","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 — reciprocal loss-of-function in patients and KO mice with TEM ultrastructure and co-localization evidence for IDA complex members","pmids":["39503742"],"is_preprint":false},{"year":2025,"finding":"DNAH3 deficiency in Dnah3-knockout mice causes asthenozoospermia through disturbance of sperm energy metabolism, specifically reducing L-palmitoylcarnitine and glycocholic acid levels, rather than through gross morphological abnormalities of the sperm flagellum.","method":"Dnah3 knockout mouse generation, CASA, H&E staining, transmission electron microscopy, sperm metabolomics","journal":"Biology of reproduction","confidence":"Medium","confidence_rationale":"Tier 2 — KO mouse with metabolomics, but single lab and metabolic mechanism not yet fully characterized","pmids":["39774634"],"is_preprint":false},{"year":2026,"finding":"DNAH3 physically interacts with DNALI1, forming a complex required for inner dynein arm assembly and sperm flagellum function. Dnah3 knockout mice lack IDAs and are infertile, and scRNA-seq showed that Dnah3 deficiency in elongating spermatids alters expression of sperm motility genes.","method":"Co-immunoprecipitation, mass spectrometry, comparative proteomics, CRISPR/Cas9 knockout mice, electron microscopy, immunostaining, scRNA-seq","journal":"Reproductive biology and endocrinology : RB&E","confidence":"High","confidence_rationale":"Tier 1–2 — co-IP with mass spectrometry identifying DNALI1 as binding partner, supported by KO mouse phenotype with multiple orthogonal methods","pmids":["41731497"],"is_preprint":false},{"year":2024,"finding":"Novel biallelic DNAH3 variants (including premature stop codons) in infertile men cause oligoasthenoteratozoospermia with fibrous sheath dysplasia and multiple tail malformations, associated with significant reduction in DNAH3 protein expression and altered expression of related proteins.","method":"Whole-exome sequencing, Papanicolaou staining, scanning electron microscopy, transmission electron microscopy, RT-qPCR, Western blot, protein structure prediction","journal":"Frontiers in endocrinology","confidence":"Medium","confidence_rationale":"Tier 2–3 — patient variants with protein expression validation and ultrastructural phenotyping, single lab","pmids":["39588341"],"is_preprint":false},{"year":2008,"finding":"Retroviral insertional mutagenesis in mouse colon epithelial cells identified Dnah3 as a common integration site associated with transformation. Upregulation or truncation of Dnah3 by viral integration was demonstrated, and Dnah3-overexpressing IMCE cells showed impairment of microtubule function, suggesting a role in cytoskeletal regulation relevant to Apc-related tumor development.","method":"Retroviral insertional mutagenesis, identification of common integration sites, Dnah3 overexpression in cell line, microtubule function assay","journal":"Cancer science","confidence":"Medium","confidence_rationale":"Tier 3 — functional cell-line assay with overexpression showing microtubule impairment, single lab","pmids":["18294281"],"is_preprint":false},{"year":2016,"finding":"DNAH3 protein was detected in human spermatozoa by mass spectrometry, providing proteomics evidence that DNAH3 is expressed as a protein in sperm.","method":"Mining of PRIDE mass spectrometry database, targeted proteomics (SRM/MRM assays)","journal":"Journal of proteome research","confidence":"Medium","confidence_rationale":"Tier 2 — validated by targeted proteomics (SRM/MRM), single study","pmids":["27581094"],"is_preprint":false},{"year":2025,"finding":"DNAH3 expression in the spermaduct was identified as a key regulator of sperm motility in drakes; it co-expressed with other dynein components (DNAH1, DNAH7, DNAH10, DNAH12, DNAI1, DNAI2, DNALI1) in protein-protein interaction network analysis, consistent with a conserved role in flagellar dynein arm function.","method":"Comparative transcriptome RNA-seq, co-expression network and protein-protein interaction network analysis","journal":"Poultry science","confidence":"Low","confidence_rationale":"Tier 4 — computational/network analysis only, no direct functional experiment on DNAH3","pmids":["36881978"],"is_preprint":false}],"current_model":"DNAH3 is an axonemal dynein heavy chain that localizes to the inner dynein arm (IDA) of sperm flagella, where it physically interacts with DNALI1 and supports the assembly of the IDA complex (including DNAH1, DNAH6, and DNALI1); loss of DNAH3 function—through biallelic human variants or Dnah3 knockout in mice—causes IDA deficiency, flagellar ultrastructural defects, and asthenoteratozoospermia, with evidence also suggesting a role in sperm energy metabolism."},"narrative":{"teleology":[{"year":2008,"claim":"Before any connection to flagellar biology, retroviral insertional mutagenesis identified Dnah3 as a gene whose overexpression or truncation impairs microtubule function in colon epithelial cells, providing the first hint that DNAH3 influences cytoskeletal dynamics.","evidence":"Retroviral integration mapping and Dnah3 overexpression in mouse IMCE colon epithelial cells with microtubule function assays","pmids":["18294281"],"confidence":"Medium","gaps":["Overexpression artefact in a non-ciliated cell type; relevance to endogenous DNAH3 function unclear","No loss-of-function experiment performed","Connection to dynein arm biology not established"]},{"year":2016,"claim":"Detection of DNAH3 protein in human spermatozoa by targeted proteomics confirmed that this dynein heavy chain is expressed in the relevant cell type for motility function.","evidence":"Mining of PRIDE mass spectrometry data with SRM/MRM validation in human sperm","pmids":["27581094"],"confidence":"Medium","gaps":["Sub-flagellar localization not determined","No functional data linking DNAH3 protein to sperm motility"]},{"year":2024,"claim":"Genetic and ultrastructural studies in patients and knockout mice established that biallelic DNAH3 loss causes asthenoteratozoospermia by disrupting IDA assembly, central pair microtubules, and flagellar accessory structures, resolving the question of whether DNAH3 is required for sperm motility.","evidence":"WES in infertile men, two independent Dnah3-KO mouse lines, TEM, immunostaining, CASA, and Western blot across multiple studies","pmids":["38312775","39503742","39588341"],"confidence":"High","gaps":["Whether DNAH3 is a structural subunit versus an assembly factor for IDA was not resolved","Mechanism by which DNAH3 loss disrupts the central pair of microtubules remains unknown","No structural or biochemical reconstitution of the DNAH3-containing IDA complex"]},{"year":2025,"claim":"Metabolomic profiling of Dnah3-KO mouse sperm revealed that DNAH3 deficiency alters energy metabolism — reducing L-palmitoylcarnitine and glycocholic acid — suggesting that impaired motility is not solely due to structural defects but also involves metabolic dysregulation.","evidence":"Sperm metabolomics in Dnah3-KO mice with CASA and histology","pmids":["39774634"],"confidence":"Medium","gaps":["Causal relationship between metabolite changes and motility loss not established","Whether metabolic effects are direct or secondary to structural disorganization is unknown","Single-lab finding not independently replicated"]},{"year":2026,"claim":"Co-immunoprecipitation and mass spectrometry identified DNALI1 as a direct physical interactor of DNAH3, establishing that DNAH3 forms a complex with a known IDA light intermediate chain and that this interaction is required for IDA integrity and sperm motility gene expression programs.","evidence":"Co-IP/MS in testis, comparative proteomics, scRNA-seq of elongating spermatids, and Dnah3-KO mice with EM and immunostaining","pmids":["41731497"],"confidence":"High","gaps":["Full stoichiometry and additional subunits of the DNAH3-containing IDA complex not determined","Whether DNAH3 interacts directly with DNAH1 or DNAH6 was not tested by reciprocal co-IP","scRNA-seq changes in motility gene expression lack causal mechanistic follow-up"]},{"year":null,"claim":"Key unresolved questions include the precise molecular architecture of the DNAH3-containing IDA complex, whether DNAH3 has ATPase/motor activity distinct from other dynein heavy chains, how its loss leads to central pair defects, and whether its metabolic role is direct or secondary.","evidence":"","pmids":[],"confidence":"Low","gaps":["No in vitro reconstitution or cryo-EM structure of DNAH3-containing IDA","ATPase activity of DNAH3 not directly measured","Mechanism linking IDA deficiency to central pair and accessory structure loss is unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003774","term_label":"cytoskeletal motor activity","supporting_discovery_ids":[0,1,3]}],"localization":[{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[0,1,3,4]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,1,3]}],"pathway":[{"term_id":"R-HSA-1474165","term_label":"Reproduction","supporting_discovery_ids":[0,1,3,4]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[0,1,3]}],"complexes":["Inner dynein arm (IDA) complex"],"partners":["DNALI1","DNAH1","DNAH6"],"other_free_text":[]},"mechanistic_narrative":"DNAH3 is an axonemal dynein heavy chain essential for inner dynein arm (IDA) assembly and motility of sperm flagella. It localizes to the sperm flagellum, physically interacts with DNALI1, and is required for stable expression of other IDA components including DNAH1, DNAH6, and DNALI1; loss of DNAH3 causes IDA deficiency, central pair microtubule loss, mitochondrial and fibrous sheath dislocation, and severely impaired sperm motility [PMID:38312775, PMID:39503742, PMID:41731497]. Biallelic loss-of-function variants in DNAH3 cause asthenoteratozoospermia and male infertility in humans, a phenotype faithfully recapitulated by Dnah3-knockout mice [PMID:38312775, PMID:39503742, PMID:39588341]. DNAH3 deficiency also disturbs sperm energy metabolism, reducing levels of L-palmitoylcarnitine and glycocholic acid, suggesting an additional role in metabolic support of flagellar function [PMID:39774634]."},"prefetch_data":{"uniprot":{"accession":"Q8TD57","full_name":"Dynein axonemal heavy chain 3","aliases":["Axonemal beta dynein heavy chain 3","HsADHC3","Ciliary dynein heavy chain 3","Dnahc3-b"],"length_aa":4116,"mass_kda":470.8,"function":"Force generating protein of respiratory cilia. Produces force towards the minus ends of microtubules. Dynein has ATPase activity; the force-producing power stroke is thought to occur on release of ADP. Involved in sperm motility; implicated in sperm flagellar assembly (By similarity). Essential for the assembly and structural organization of the sperm flagellum's axoneme, including its inner dynein arms and accessory components, which are critical for maintaining flagellar integrity and motility (PubMed:39503742, PubMed:38312775)","subcellular_location":"Cytoplasm, cytoskeleton, cilium axoneme; Cytoplasm; Cell projection, cilium, flagellum","url":"https://www.uniprot.org/uniprotkb/Q8TD57/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/DNAH3","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/DNAH3","total_profiled":1310},"omim":[{"mim_id":"603334","title":"DYNEIN, AXONEMAL, HEAVY CHAIN 3; DNAH3","url":"https://www.omim.org/entry/603334"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Mid piece","reliability":"Approved"},{"location":"Principal piece","reliability":"Approved"},{"location":"Equatorial segment","reliability":"Additional"},{"location":"Flagellar centriole","reliability":"Additional"}],"tissue_specificity":"Group enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"fallopian tube","ntpm":5.1},{"tissue":"testis","ntpm":3.2}],"url":"https://www.proteinatlas.org/search/DNAH3"},"hgnc":{"alias_symbol":["Dnahc3b","DLP3","Hsadhc3","DKFZp434N074"],"prev_symbol":[]},"alphafold":{"accession":"Q8TD57","domains":[],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TD57","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TD57-3-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TD57-3-F1-predicted_aligned_error_v6.png","plddt_mean":76.25},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=DNAH3","jax_strain_url":"https://www.jax.org/strain/search?query=DNAH3"},"sequence":{"accession":"Q8TD57","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8TD57.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8TD57/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TD57"}},"corpus_meta":[{"pmid":"10839998","id":"PMC_10839998","title":"Defensin-like peptide-2 from platypus venom: member of a class of peptides with a distinct structural fold.","date":"2000","source":"The Biochemical journal","url":"https://pubmed.ncbi.nlm.nih.gov/10839998","citation_count":51,"is_preprint":false},{"pmid":"3656451","id":"PMC_3656451","title":"Spectroscopic studies on histone-DNA interactions. 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DNAH3 localizes to sperm flagella and its deficiency results in loss of the central pair of microtubules, dislocated mitochondrial sheath and fibrous sheath, and partial absence of inner dynein arms (IDAs). Dnah3 knockout mice recapitulate the human phenotype with severe reduction in sperm movement.\",\n      \"method\": \"Whole-exome sequencing, Dnah3 knockout mouse generation, H&E staining, transmission electron microscopy, immunostaining, CASA\",\n      \"journal\": \"Human reproduction open\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — KO mouse phenotype with multiple orthogonal methods (TEM, immunostaining, CASA), replicated in two independent KO lines and patient samples\",\n      \"pmids\": [\"38312775\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"DNAH3 is a component of the inner dynein arm (IDA) in sperm flagella. Biallelic DNAH3 variants in infertile men cause deficient DNAH3 expression in sperm flagella, severe disruption of flagellar ultrastructure, and decreased expression of other IDA-associated proteins DNAH1, DNAH6, and DNALI1, indicating DNAH3 is required for IDA assembly.\",\n      \"method\": \"Whole-exome sequencing, immunofluorescence staining, transmission electron microscopy, Dnah3 knockout mice, Western blot\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal loss-of-function in patients and KO mice with TEM ultrastructure and co-localization evidence for IDA complex members\",\n      \"pmids\": [\"39503742\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DNAH3 deficiency in Dnah3-knockout mice causes asthenozoospermia through disturbance of sperm energy metabolism, specifically reducing L-palmitoylcarnitine and glycocholic acid levels, rather than through gross morphological abnormalities of the sperm flagellum.\",\n      \"method\": \"Dnah3 knockout mouse generation, CASA, H&E staining, transmission electron microscopy, sperm metabolomics\",\n      \"journal\": \"Biology of reproduction\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — KO mouse with metabolomics, but single lab and metabolic mechanism not yet fully characterized\",\n      \"pmids\": [\"39774634\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"DNAH3 physically interacts with DNALI1, forming a complex required for inner dynein arm assembly and sperm flagellum function. Dnah3 knockout mice lack IDAs and are infertile, and scRNA-seq showed that Dnah3 deficiency in elongating spermatids alters expression of sperm motility genes.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, comparative proteomics, CRISPR/Cas9 knockout mice, electron microscopy, immunostaining, scRNA-seq\",\n      \"journal\": \"Reproductive biology and endocrinology : RB&E\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 — co-IP with mass spectrometry identifying DNALI1 as binding partner, supported by KO mouse phenotype with multiple orthogonal methods\",\n      \"pmids\": [\"41731497\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Novel biallelic DNAH3 variants (including premature stop codons) in infertile men cause oligoasthenoteratozoospermia with fibrous sheath dysplasia and multiple tail malformations, associated with significant reduction in DNAH3 protein expression and altered expression of related proteins.\",\n      \"method\": \"Whole-exome sequencing, Papanicolaou staining, scanning electron microscopy, transmission electron microscopy, RT-qPCR, Western blot, protein structure prediction\",\n      \"journal\": \"Frontiers in endocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 — patient variants with protein expression validation and ultrastructural phenotyping, single lab\",\n      \"pmids\": [\"39588341\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Retroviral insertional mutagenesis in mouse colon epithelial cells identified Dnah3 as a common integration site associated with transformation. Upregulation or truncation of Dnah3 by viral integration was demonstrated, and Dnah3-overexpressing IMCE cells showed impairment of microtubule function, suggesting a role in cytoskeletal regulation relevant to Apc-related tumor development.\",\n      \"method\": \"Retroviral insertional mutagenesis, identification of common integration sites, Dnah3 overexpression in cell line, microtubule function assay\",\n      \"journal\": \"Cancer science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — functional cell-line assay with overexpression showing microtubule impairment, single lab\",\n      \"pmids\": [\"18294281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"DNAH3 protein was detected in human spermatozoa by mass spectrometry, providing proteomics evidence that DNAH3 is expressed as a protein in sperm.\",\n      \"method\": \"Mining of PRIDE mass spectrometry database, targeted proteomics (SRM/MRM assays)\",\n      \"journal\": \"Journal of proteome research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — validated by targeted proteomics (SRM/MRM), single study\",\n      \"pmids\": [\"27581094\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"DNAH3 expression in the spermaduct was identified as a key regulator of sperm motility in drakes; it co-expressed with other dynein components (DNAH1, DNAH7, DNAH10, DNAH12, DNAI1, DNAI2, DNALI1) in protein-protein interaction network analysis, consistent with a conserved role in flagellar dynein arm function.\",\n      \"method\": \"Comparative transcriptome RNA-seq, co-expression network and protein-protein interaction network analysis\",\n      \"journal\": \"Poultry science\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 — computational/network analysis only, no direct functional experiment on DNAH3\",\n      \"pmids\": [\"36881978\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"DNAH3 is an axonemal dynein heavy chain that localizes to the inner dynein arm (IDA) of sperm flagella, where it physically interacts with DNALI1 and supports the assembly of the IDA complex (including DNAH1, DNAH6, and DNALI1); loss of DNAH3 function—through biallelic human variants or Dnah3 knockout in mice—causes IDA deficiency, flagellar ultrastructural defects, and asthenoteratozoospermia, with evidence also suggesting a role in sperm energy metabolism.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"DNAH3 is an axonemal dynein heavy chain essential for inner dynein arm (IDA) assembly and motility of sperm flagella. It localizes to the sperm flagellum, physically interacts with DNALI1, and is required for stable expression of other IDA components including DNAH1, DNAH6, and DNALI1; loss of DNAH3 causes IDA deficiency, central pair microtubule loss, mitochondrial and fibrous sheath dislocation, and severely impaired sperm motility [PMID:38312775, PMID:39503742, PMID:41731497]. Biallelic loss-of-function variants in DNAH3 cause asthenoteratozoospermia and male infertility in humans, a phenotype faithfully recapitulated by Dnah3-knockout mice [PMID:38312775, PMID:39503742, PMID:39588341]. DNAH3 deficiency also disturbs sperm energy metabolism, reducing levels of L-palmitoylcarnitine and glycocholic acid, suggesting an additional role in metabolic support of flagellar function [PMID:39774634].\",\n  \"teleology\": [\n    {\n      \"year\": 2008,\n      \"claim\": \"Before any connection to flagellar biology, retroviral insertional mutagenesis identified Dnah3 as a gene whose overexpression or truncation impairs microtubule function in colon epithelial cells, providing the first hint that DNAH3 influences cytoskeletal dynamics.\",\n      \"evidence\": \"Retroviral integration mapping and Dnah3 overexpression in mouse IMCE colon epithelial cells with microtubule function assays\",\n      \"pmids\": [\"18294281\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Overexpression artefact in a non-ciliated cell type; relevance to endogenous DNAH3 function unclear\",\n        \"No loss-of-function experiment performed\",\n        \"Connection to dynein arm biology not established\"\n      ]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Detection of DNAH3 protein in human spermatozoa by targeted proteomics confirmed that this dynein heavy chain is expressed in the relevant cell type for motility function.\",\n      \"evidence\": \"Mining of PRIDE mass spectrometry data with SRM/MRM validation in human sperm\",\n      \"pmids\": [\"27581094\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Sub-flagellar localization not determined\",\n        \"No functional data linking DNAH3 protein to sperm motility\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Genetic and ultrastructural studies in patients and knockout mice established that biallelic DNAH3 loss causes asthenoteratozoospermia by disrupting IDA assembly, central pair microtubules, and flagellar accessory structures, resolving the question of whether DNAH3 is required for sperm motility.\",\n      \"evidence\": \"WES in infertile men, two independent Dnah3-KO mouse lines, TEM, immunostaining, CASA, and Western blot across multiple studies\",\n      \"pmids\": [\"38312775\", \"39503742\", \"39588341\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Whether DNAH3 is a structural subunit versus an assembly factor for IDA was not resolved\",\n        \"Mechanism by which DNAH3 loss disrupts the central pair of microtubules remains unknown\",\n        \"No structural or biochemical reconstitution of the DNAH3-containing IDA complex\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Metabolomic profiling of Dnah3-KO mouse sperm revealed that DNAH3 deficiency alters energy metabolism — reducing L-palmitoylcarnitine and glycocholic acid — suggesting that impaired motility is not solely due to structural defects but also involves metabolic dysregulation.\",\n      \"evidence\": \"Sperm metabolomics in Dnah3-KO mice with CASA and histology\",\n      \"pmids\": [\"39774634\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Causal relationship between metabolite changes and motility loss not established\",\n        \"Whether metabolic effects are direct or secondary to structural disorganization is unknown\",\n        \"Single-lab finding not independently replicated\"\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Co-immunoprecipitation and mass spectrometry identified DNALI1 as a direct physical interactor of DNAH3, establishing that DNAH3 forms a complex with a known IDA light intermediate chain and that this interaction is required for IDA integrity and sperm motility gene expression programs.\",\n      \"evidence\": \"Co-IP/MS in testis, comparative proteomics, scRNA-seq of elongating spermatids, and Dnah3-KO mice with EM and immunostaining\",\n      \"pmids\": [\"41731497\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Full stoichiometry and additional subunits of the DNAH3-containing IDA complex not determined\",\n        \"Whether DNAH3 interacts directly with DNAH1 or DNAH6 was not tested by reciprocal co-IP\",\n        \"scRNA-seq changes in motility gene expression lack causal mechanistic follow-up\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key unresolved questions include the precise molecular architecture of the DNAH3-containing IDA complex, whether DNAH3 has ATPase/motor activity distinct from other dynein heavy chains, how its loss leads to central pair defects, and whether its metabolic role is direct or secondary.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"No in vitro reconstitution or cryo-EM structure of DNAH3-containing IDA\",\n        \"ATPase activity of DNAH3 not directly measured\",\n        \"Mechanism linking IDA deficiency to central pair and accessory structure loss is unknown\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\n        \"term_id\": \"GO:0003774\",\n        \"supporting_discovery_ids\": [0, 1, 3]\n      }\n    ],\n    \"localization\": [\n      {\n        \"term_id\": \"GO:0005929\",\n        \"supporting_discovery_ids\": [0, 1, 3, 4]\n      },\n      {\n        \"term_id\": \"GO:0005856\",\n        \"supporting_discovery_ids\": [0, 1, 3]\n      }\n    ],\n    \"pathway\": [\n      {\n        \"term_id\": \"R-HSA-1474165\",\n        \"supporting_discovery_ids\": [0, 1, 3, 4]\n      },\n      {\n        \"term_id\": \"R-HSA-1852241\",\n        \"supporting_discovery_ids\": [0, 1, 3]\n      }\n    ],\n    \"complexes\": [\n      \"Inner dynein arm (IDA) complex\"\n    ],\n    \"partners\": [\n      \"DNALI1\",\n      \"DNAH1\",\n      \"DNAH6\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}