{"gene":"ANKEF1","run_date":"2026-06-09T22:02:43","timeline":{"discoveries":[{"year":2025,"finding":"ANKEF1 (ANKRD5) is a component of the sperm axoneme that physically interacts with DRC5/TCTE1 and DRC4/GAS8, two key components of the nexin-dynein regulatory complex (N-DRC), and these interactions occur independently of calcium regulation. Loss of ANKEF1 in mice causes impaired sperm motility and male infertility. Cryo-electron tomography of Ankef1-/- sperm revealed a structurally intact '9+2' axoneme with intact doublet microtubules (DMTs) but with pronounced morphological variability and increased structural heterogeneity of DMTs, suggesting ANKEF1 attenuates N-DRC mechanical buffering between adjacent DMTs under high mechanical stress during flagellar beating. ANKEF1 deficiency did not alter ATP levels, reactive oxygen species levels, or mitochondrial membrane potential.","method":"Co-immunoprecipitation (protein-protein interaction with N-DRC components), knockout mouse model (Ankef1-/-), cryo-electron tomography (axoneme ultrastructure), ATP/ROS/mitochondrial membrane potential assays","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (Co-IP for interactions, KO mouse with defined fertility phenotype, cryo-ET for structural analysis) in a single rigorous peer-reviewed study, replicated by companion preprint","pmids":["41460250"],"is_preprint":false},{"year":2024,"finding":"ANKEF1 (ANKRD5) interacts with DRC5/TCTE1 and DRC4/GAS8 of the nexin-dynein regulatory complex (N-DRC) in the sperm axoneme, with interactions independent of calcium. Ankrd5-/- male mice are infertile with impaired sperm motility; cryo-electron tomography shows intact '9+2' structure but increased DMT morphological variability and structural heterogeneity without changes in ATP, ROS, or mitochondrial membrane potential.","method":"Co-immunoprecipitation, knockout mouse, cryo-electron tomography, metabolic assays (ATP, ROS, mitochondrial membrane potential)","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — multiple orthogonal methods, but preprint not yet peer-reviewed; largely mirrors the published eLife study","pmids":["bio_10.1101_2024.12.03.626701"],"is_preprint":true},{"year":2023,"finding":"The zebrafish ortholog Ankef1a localizes to the kinocilia of hair cells in sensory organs, exhibiting a distinct regionalized localization pattern along the proximal-distal axis of the kinocilium and within the cell body, as demonstrated by transgenic fluorescently tagged protein expression.","method":"Transgenic zebrafish expressing fluorescently tagged Ankef1a; live imaging of kinocilial localization","journal":"Journal of developmental biology","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — direct localization by transgenic fluorescent tagging in zebrafish; no functional loss-of-function phenotype reported for Ankef1a specifically","pmids":["37367482"],"is_preprint":false}],"current_model":"ANKEF1 (ANKRD5) is an axonemal protein that associates with the nexin-dynein regulatory complex (N-DRC) via direct interactions with DRC5/TCTE1 and DRC4/GAS8 independently of calcium, and is required for maintaining doublet microtubule structural stability and normal sperm motility in mice; its zebrafish ortholog additionally localizes in a regionalized pattern along the kinocilium of sensory hair cells."},"narrative":{"mechanistic_narrative":"ANKEF1 (ANKRD5) is an axonemal protein required for sperm motility that maintains the structural integrity of flagellar doublet microtubules [PMID:41460250]. It is a component of the sperm axoneme that physically associates with the nexin-dynein regulatory complex (N-DRC) through calcium-independent interactions with its DRC5/TCTE1 and DRC4/GAS8 subunits [PMID:41460250]. Loss of ANKEF1 in mice causes male infertility and impaired sperm motility; cryo-electron tomography of knockout sperm shows a structurally intact 9+2 axoneme but with pronounced morphological variability and increased structural heterogeneity of doublet microtubules, while ATP, reactive oxygen species, and mitochondrial membrane potential are unaffected — placing ANKEF1 in N-DRC mechanical buffering between adjacent doublets rather than in energy metabolism [PMID:41460250]. In zebrafish, the ortholog Ankef1a localizes in a regionalized pattern along the proximal-distal axis of sensory hair-cell kinocilia, consistent with a broader role at motile/sensory ciliary axonemes [PMID:37367482]. Beyond these findings, no additional mechanistic detail has been characterized in the available corpus.","teleology":[{"year":2023,"claim":"Before functional studies, it was unknown where ANKEF1 acts; localization of the zebrafish ortholog to hair-cell kinocilia established a ciliary/axonemal context for the protein.","evidence":"Transgenic zebrafish expressing fluorescently tagged Ankef1a with live imaging of kinocilial localization","pmids":["37367482"],"confidence":"Medium","gaps":["No loss-of-function phenotype reported for Ankef1a in zebrafish","Functional significance of the regionalized kinocilial localization unresolved","No molecular partners identified in this system"]},{"year":2024,"claim":"To define ANKEF1's molecular role, its interactions and the consequences of its loss were probed, identifying it as an N-DRC-associated axonemal protein required for sperm motility.","evidence":"Co-IP for N-DRC interactions, Ankrd5-/- knockout mouse fertility phenotype, cryo-electron tomography, and metabolic assays (preprint)","pmids":["bio_10.1101_2024.12.03.626701"],"confidence":"Medium","gaps":["Preprint not yet peer-reviewed at this stage","Direct versus indirect nature of N-DRC interactions not fully resolved","Mechanism of doublet stabilization not directly demonstrated"]},{"year":2025,"claim":"Peer-reviewed work confirmed ANKEF1 binds DRC5/TCTE1 and DRC4/GAS8 calcium-independently and is required to maintain doublet microtubule structural homogeneity, establishing a mechanical-buffering role distinct from axonemal energetics.","evidence":"Co-IP, Ankef1-/- knockout mouse with infertility/motility phenotype, cryo-electron tomography of axoneme ultrastructure, and ATP/ROS/mitochondrial membrane potential assays","pmids":["41460250"],"confidence":"High","gaps":["Stoichiometry and structural position of ANKEF1 within the N-DRC not resolved","Whether ANKEF1 contributes to motile cilia outside sperm not addressed","Molecular mechanism linking N-DRC association to doublet structural homogeneity not directly demonstrated"]},{"year":null,"claim":"How ANKEF1 mechanistically stabilizes doublet microtubules and whether it functions in sensory/motile cilia beyond the sperm flagellum remains open.","evidence":"","pmids":[],"confidence":"High","gaps":["No structural model of ANKEF1 within the N-DRC","Human disease link not established in the corpus","Functional role at hair-cell kinocilia not tested by loss-of-function"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[0,2]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0]}],"pathway":[],"complexes":["nexin-dynein regulatory complex (N-DRC)"],"partners":["TCTE1","GAS8"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NU02","full_name":"Ankyrin repeat and EF-hand domain-containing protein 1","aliases":["Ankyrin repeat domain-containing protein 5"],"length_aa":776,"mass_kda":86.7,"function":"","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q9NU02/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ANKEF1","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/ANKEF1","total_profiled":1310},"omim":[],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoli fibrillar center","reliability":"Approved"},{"location":"Plasma membrane","reliability":"Approved"},{"location":"Cytosol","reliability":"Approved"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in many","driving_tissues":[{"tissue":"pancreas","ntpm":13.2}],"url":"https://www.proteinatlas.org/search/ANKEF1"},"hgnc":{"alias_symbol":["FLJ21669","dJ839B4.6"],"prev_symbol":["ANKRD5"]},"alphafold":{"accession":"Q9NU02","domains":[{"cath_id":"1.25.40.20","chopping":"2-108","consensus_level":"medium","plddt":90.9215,"start":2,"end":108},{"cath_id":"1.25.40.20","chopping":"219-319","consensus_level":"medium","plddt":91.9477,"start":219,"end":319},{"cath_id":"1.10.238,1.10.225","chopping":"325-411","consensus_level":"high","plddt":85.0748,"start":325,"end":411},{"cath_id":"1.25.40","chopping":"435-463_492-535","consensus_level":"medium","plddt":81.599,"start":435,"end":535},{"cath_id":"1.25.40.20","chopping":"552-650_737-776","consensus_level":"medium","plddt":84.3754,"start":552,"end":776}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NU02","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NU02-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NU02-F1-predicted_aligned_error_v6.png","plddt_mean":81.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ANKEF1","jax_strain_url":"https://www.jax.org/strain/search?query=ANKEF1"},"sequence":{"accession":"Q9NU02","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NU02.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NU02/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NU02"}},"corpus_meta":[{"pmid":"28673325","id":"PMC_28673325","title":"Maternal intake of high n-6 polyunsaturated fatty acid diet during pregnancy causes transgenerational increase in mammary cancer risk in mice.","date":"2017","source":"Breast cancer research : BCR","url":"https://pubmed.ncbi.nlm.nih.gov/28673325","citation_count":30,"is_preprint":false},{"pmid":"17142250","id":"PMC_17142250","title":"Sex-specific linkage to total serum immunoglobulin E in families of children with asthma in Costa Rica.","date":"2006","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/17142250","citation_count":20,"is_preprint":false},{"pmid":"34382870","id":"PMC_34382870","title":"Genome-Wide Scan for Parent-of-Origin Effects in a sub-Saharan African Cohort With Nonsyndromic Cleft Lip and/or Cleft Palate (CL/P).","date":"2021","source":"The Cleft palate-craniofacial journal : official publication of the American Cleft Palate-Craniofacial Association","url":"https://pubmed.ncbi.nlm.nih.gov/34382870","citation_count":8,"is_preprint":false},{"pmid":"37367482","id":"PMC_37367482","title":"Regionalized Protein Localization Domains in the Zebrafish Hair Cell Kinocilium.","date":"2023","source":"Journal of developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/37367482","citation_count":7,"is_preprint":false},{"pmid":"41460250","id":"PMC_41460250","title":"ANKEF1 is a key axonemal component essential for murine sperm motility and male fertility.","date":"2025","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/41460250","citation_count":1,"is_preprint":false},{"pmid":"39613498","id":"PMC_39613498","title":"MicroRNAs bta-novel-miR-117, bta-novel-miR-234 and bta-novel-miR-417 have adverse effects on blastocyst formation.","date":"2024","source":"Theriogenology","url":"https://pubmed.ncbi.nlm.nih.gov/39613498","citation_count":1,"is_preprint":false},{"pmid":"31149201","id":"PMC_31149201","title":"NOVEL GENETIC FINDINGS IN A CHINESE FAMILY WITH EARLY-ONSET FEMALE-RELATED TYPE 2 DIABETES.","date":"2017","source":"Acta endocrinologica (Bucharest, Romania : 2005)","url":"https://pubmed.ncbi.nlm.nih.gov/31149201","citation_count":1,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2024.12.03.626701","title":"ANKRD5: a key component of the axoneme required for sperm motility and male fertility","date":"2024-12-07","source":"bioRxiv","url":"https://doi.org/10.1101/2024.12.03.626701","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":6659,"output_tokens":1025,"usd":0.017676,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7602,"output_tokens":1720,"usd":0.040505,"stage2_stop_reason":"end_turn"},"total_usd":0.058181,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2025,\n      \"finding\": \"ANKEF1 (ANKRD5) is a component of the sperm axoneme that physically interacts with DRC5/TCTE1 and DRC4/GAS8, two key components of the nexin-dynein regulatory complex (N-DRC), and these interactions occur independently of calcium regulation. Loss of ANKEF1 in mice causes impaired sperm motility and male infertility. Cryo-electron tomography of Ankef1-/- sperm revealed a structurally intact '9+2' axoneme with intact doublet microtubules (DMTs) but with pronounced morphological variability and increased structural heterogeneity of DMTs, suggesting ANKEF1 attenuates N-DRC mechanical buffering between adjacent DMTs under high mechanical stress during flagellar beating. ANKEF1 deficiency did not alter ATP levels, reactive oxygen species levels, or mitochondrial membrane potential.\",\n      \"method\": \"Co-immunoprecipitation (protein-protein interaction with N-DRC components), knockout mouse model (Ankef1-/-), cryo-electron tomography (axoneme ultrastructure), ATP/ROS/mitochondrial membrane potential assays\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (Co-IP for interactions, KO mouse with defined fertility phenotype, cryo-ET for structural analysis) in a single rigorous peer-reviewed study, replicated by companion preprint\",\n      \"pmids\": [\"41460250\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ANKEF1 (ANKRD5) interacts with DRC5/TCTE1 and DRC4/GAS8 of the nexin-dynein regulatory complex (N-DRC) in the sperm axoneme, with interactions independent of calcium. Ankrd5-/- male mice are infertile with impaired sperm motility; cryo-electron tomography shows intact '9+2' structure but increased DMT morphological variability and structural heterogeneity without changes in ATP, ROS, or mitochondrial membrane potential.\",\n      \"method\": \"Co-immunoprecipitation, knockout mouse, cryo-electron tomography, metabolic assays (ATP, ROS, mitochondrial membrane potential)\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — multiple orthogonal methods, but preprint not yet peer-reviewed; largely mirrors the published eLife study\",\n      \"pmids\": [\"bio_10.1101_2024.12.03.626701\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The zebrafish ortholog Ankef1a localizes to the kinocilia of hair cells in sensory organs, exhibiting a distinct regionalized localization pattern along the proximal-distal axis of the kinocilium and within the cell body, as demonstrated by transgenic fluorescently tagged protein expression.\",\n      \"method\": \"Transgenic zebrafish expressing fluorescently tagged Ankef1a; live imaging of kinocilial localization\",\n      \"journal\": \"Journal of developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — direct localization by transgenic fluorescent tagging in zebrafish; no functional loss-of-function phenotype reported for Ankef1a specifically\",\n      \"pmids\": [\"37367482\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ANKEF1 (ANKRD5) is an axonemal protein that associates with the nexin-dynein regulatory complex (N-DRC) via direct interactions with DRC5/TCTE1 and DRC4/GAS8 independently of calcium, and is required for maintaining doublet microtubule structural stability and normal sperm motility in mice; its zebrafish ortholog additionally localizes in a regionalized pattern along the kinocilium of sensory hair cells.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ANKEF1 (ANKRD5) is an axonemal protein required for sperm motility that maintains the structural integrity of flagellar doublet microtubules [#0]. It is a component of the sperm axoneme that physically associates with the nexin-dynein regulatory complex (N-DRC) through calcium-independent interactions with its DRC5/TCTE1 and DRC4/GAS8 subunits [#0]. Loss of ANKEF1 in mice causes male infertility and impaired sperm motility; cryo-electron tomography of knockout sperm shows a structurally intact 9+2 axoneme but with pronounced morphological variability and increased structural heterogeneity of doublet microtubules, while ATP, reactive oxygen species, and mitochondrial membrane potential are unaffected — placing ANKEF1 in N-DRC mechanical buffering between adjacent doublets rather than in energy metabolism [#0]. In zebrafish, the ortholog Ankef1a localizes in a regionalized pattern along the proximal-distal axis of sensory hair-cell kinocilia, consistent with a broader role at motile/sensory ciliary axonemes [#2]. Beyond these findings, no additional mechanistic detail has been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2023,\n      \"claim\": \"Before functional studies, it was unknown where ANKEF1 acts; localization of the zebrafish ortholog to hair-cell kinocilia established a ciliary/axonemal context for the protein.\",\n      \"evidence\": \"Transgenic zebrafish expressing fluorescently tagged Ankef1a with live imaging of kinocilial localization\",\n      \"pmids\": [\"37367482\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No loss-of-function phenotype reported for Ankef1a in zebrafish\",\n        \"Functional significance of the regionalized kinocilial localization unresolved\",\n        \"No molecular partners identified in this system\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"To define ANKEF1's molecular role, its interactions and the consequences of its loss were probed, identifying it as an N-DRC-associated axonemal protein required for sperm motility.\",\n      \"evidence\": \"Co-IP for N-DRC interactions, Ankrd5-/- knockout mouse fertility phenotype, cryo-electron tomography, and metabolic assays (preprint)\",\n      \"pmids\": [\"bio_10.1101_2024.12.03.626701\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Preprint not yet peer-reviewed at this stage\",\n        \"Direct versus indirect nature of N-DRC interactions not fully resolved\",\n        \"Mechanism of doublet stabilization not directly demonstrated\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Peer-reviewed work confirmed ANKEF1 binds DRC5/TCTE1 and DRC4/GAS8 calcium-independently and is required to maintain doublet microtubule structural homogeneity, establishing a mechanical-buffering role distinct from axonemal energetics.\",\n      \"evidence\": \"Co-IP, Ankef1-/- knockout mouse with infertility/motility phenotype, cryo-electron tomography of axoneme ultrastructure, and ATP/ROS/mitochondrial membrane potential assays\",\n      \"pmids\": [\"41460250\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Stoichiometry and structural position of ANKEF1 within the N-DRC not resolved\",\n        \"Whether ANKEF1 contributes to motile cilia outside sperm not addressed\",\n        \"Molecular mechanism linking N-DRC association to doublet structural homogeneity not directly demonstrated\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How ANKEF1 mechanistically stabilizes doublet microtubules and whether it functions in sensory/motile cilia beyond the sperm flagellum remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No structural model of ANKEF1 within the N-DRC\",\n        \"Human disease link not established in the corpus\",\n        \"Functional role at hair-cell kinocilia not tested by loss-of-function\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [0, 2]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [],\n    \"complexes\": [\"nexin-dynein regulatory complex (N-DRC)\"],\n    \"partners\": [\"TCTE1\", \"GAS8\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"faith_supported":4,"faith_total":4,"faith_pct":100.0}}