{"gene":"CKAP2L","run_date":"2026-06-09T22:57:18","timeline":{"discoveries":[{"year":2013,"finding":"Radmis/CKAP2L is a microtubule-associated protein (MAP) enriched in neural stem/progenitor cells (NSPCs) that localizes to bipolar mitotic spindles and radial fibers during NSPC division. Overexpression induces hyper-stabilization of microtubules and severe defects in mitotic spindle formation leading to mitotic arrest. Loss-of-function via shRNA induces multipolar mitotic spindle structures and chromosome segregation defects including long chromosome bridges. Radmis is a putative substrate for the APC/C E3-ubiquitin ligase and is degraded via the KEN box.","method":"Differential subtractive screening, shRNA knockdown, overexpression in vitro and in vivo (in utero electroporation), immunofluorescence localization","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (KD, OE, in vivo, localization) in a single foundational study, replicated by subsequent papers","pmids":["24260314"],"is_preprint":false},{"year":2014,"finding":"Loss-of-function mutations in CKAP2L cause Filippi syndrome. In dividing lymphoblastoid cell lines from homozygous frameshift mutation carriers, CKAP2L is absent from spindle poles, and cells exhibit multipolar spindle configurations and chromosome segregation defects, establishing CKAP2L's role at the spindle pole as essential for faithful mitosis in human cells.","method":"Homozygosity mapping, whole-exome sequencing, Sanger sequencing, immunofluorescence of patient-derived lymphoblastoid cell lines","journal":"American journal of human genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — patient-derived cell lines with defined mutations, direct localization assay, phenotypic readout, replicated across multiple families","pmids":["25439729"],"is_preprint":false},{"year":2021,"finding":"CKAP2L directly interacts with RNA Polymerase II and regulates transcription elongation of genes involved in spindle assembly checkpoint, chromosome segregation, cell cycle, and E2F signaling. Depletion of CKAP2L increased sensitivity of NSCLC cells to alvocidib (pan-CDK inhibitor).","method":"Co-immunoprecipitation (CKAP2L–RNA Pol II interaction), RNAi screen, in vitro and in vivo proliferation/growth assays","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — reciprocal Co-IP reported in single study, single lab, novel mechanism not yet independently replicated","pmids":["33472893"],"is_preprint":false},{"year":2022,"finding":"FOXP3 transcriptionally represses CKAP2L by binding to its promoter, and CKAP2L in turn activates the AKT/mTOR signaling pathway in breast cancer cells, as evidenced by increased phosphorylation of AKT, mTOR, and p70S6K upon CKAP2L overexpression.","method":"Chromatin immunoprecipitation (ChIP), promoter-binding assay, Western blot for pathway phosphorylation, CKAP2L overexpression and silencing in BC cell lines","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — ChIP confirms FOXP3 promoter binding; pathway activation by single lab with two orthogonal methods but not independently replicated","pmids":["35065924"],"is_preprint":false},{"year":2024,"finding":"Human CKAP2L protein levels are cell cycle phase-dependent, peaking at G2/M. During interphase, CKAP2L localizes predominantly to the nucleus, while during mitosis it localizes to the mitotic spindle apparatus. Ectopic overexpression causes microtubule bundling and prolonged mitosis.","method":"Cell cycle synchronization, immunofluorescence, ectopic overexpression in human cancer cell lines","journal":"FEBS open bio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization experiments with functional consequence (prolonged mitosis), two orthogonal methods (cell fractionation/IF + overexpression phenotype), single lab","pmids":["39073037"],"is_preprint":false},{"year":2025,"finding":"CKAP2L is a bona fide microtubule-associated protein that localizes to the centrosome, mitotic spindle, and ciliary basal body. Depletion of CKAP2L leads to shortened mitotic spindles and cytokinesis failure resulting in multinucleation. CKAP2L acts as a negative regulator of primary cilium length; its loss markedly increases ciliary length in both human and mouse cells. CKAP2L knockout mice exhibit reduced male fertility with decreased sperm count, impaired motility, and abnormally elongated flagella.","method":"Ckap2l knockout mouse generation, immunofluorescence localization to microtubule-based organelles, ciliary length measurements in human and mouse cells, sperm analysis","journal":"Journal of molecular cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo knockout model combined with multiple orthogonal cellular assays (spindle length, cytokinesis, ciliogenesis) across human and mouse systems in a single rigorous study","pmids":["41370039"],"is_preprint":false},{"year":2025,"finding":"FOXM1 transcriptionally activates CKAP2L by binding to its promoter, as confirmed by ChIP-qPCR. Cigarette smoke extract upregulates FOXM1 and CKAP2L expression, and CKAP2L knockdown arrests the cell cycle at S and G2/M phases in bladder cancer cells.","method":"Chromatin immunoprecipitation (ChIP)-qPCR, CKAP2L knockdown, cell cycle flow cytometry","journal":"Journal of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — ChIP-qPCR confirms FOXM1 promoter binding; single lab, single study, not independently replicated","pmids":["40646610"],"is_preprint":false},{"year":2025,"finding":"RFX5 acts as an upstream transcription factor that directly activates CKAP2L transcription, as confirmed by dual-luciferase assay and chromatin immunoprecipitation. CKAP2L overexpression rescues the antiproliferative and anti-migratory effects of RFX5 knockdown in colorectal cancer cells, and CKAP2L inactivates the AKT/mTOR pathway downstream.","method":"Dual-luciferase reporter assay, ChIP, siRNA knockdown, CKAP2L overexpression rescue experiments, Western blot","journal":"Discovery medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — two orthogonal methods (luciferase + ChIP) confirm transcriptional regulation; single lab, not independently replicated","pmids":["39851233"],"is_preprint":false},{"year":2026,"finding":"CKAP2L stabilizes phosphorylated AKT by inhibiting its ubiquitin-proteasome degradation. Co-immunoprecipitation and ubiquitination assays showed that CKAP2L interacts with AKT and overexpression of CKAP2L decreases AKT ubiquitination, sustaining AKT phosphorylation and activating PI3K/AKT signaling in endometrial cancer cells.","method":"Co-immunoprecipitation (Co-IP), ubiquitination assay, cycloheximide chase, MG132 proteasome inhibitor assay, Western blot","journal":"Frontiers in oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP plus functional ubiquitination/degradation assays, multiple orthogonal methods, single lab, not independently replicated","pmids":["42245708"],"is_preprint":false},{"year":2026,"finding":"CKAP2L promotes AREG (amphiregulin) expression in colorectal cancer cells by increasing STAT3 phosphorylation; activated STAT3 then binds the AREG promoter (confirmed by ChIP) to transcriptionally activate AREG, thereby engaging the AREG/EGFR pathway to drive cancer progression.","method":"RNA sequencing, CKAP2L knockdown, ChIP assay (STAT3 at AREG promoter), Western blot for pSTAT3, rescue experiments with AREG suppression","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — ChIP confirms STAT3-AREG promoter binding; RNA-seq + rescue experiments; single lab, single study","pmids":["41823522"],"is_preprint":false},{"year":2025,"finding":"Proximity-labeling proteomics of radial glial cell primary cilia in the embryonic brain identified CKAP2L as a ciliary protein component in the developing brain, and its ciliary localization was experimentally validated.","method":"Proximity labeling (BioID), quantitative proteomics, immunofluorescence validation of ciliary localization in embryonic brain","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 3 / Weak — preprint, proximity labeling with validation, single study, no functional follow-up on the ciliary localization itself","pmids":[],"is_preprint":true}],"current_model":"CKAP2L (Radmis) is a microtubule-associated protein that localizes to the centrosome, mitotic spindle, and ciliary basal body; it is degraded by APC/C via its KEN box after mitosis, and its cell cycle-regulated expression (peaking at G2/M) is transcriptionally controlled by factors including FOXP3 (repressor), FOXM1 and RFX5 (activators); during mitosis it stabilizes microtubules and is required for bipolar spindle formation, correct chromosome segregation, and cytokinesis, while its loss leads to multipolar spindles, multinucleation, and elongated primary cilia; beyond its structural spindle role, CKAP2L also interacts with RNA Pol II to regulate transcription elongation and with AKT to suppress its ubiquitin-mediated degradation, thereby activating PI3K/AKT/mTOR signaling."},"narrative":{"mechanistic_narrative":"CKAP2L (Radmis) is a cell cycle-regulated microtubule-associated protein that governs the fidelity of mitotic spindle assembly and the homeostasis of microtubule-based organelles [PMID:24260314, PMID:41370039]. Its protein levels peak at G2/M, and it relocalizes from the interphase nucleus to the centrosome, mitotic spindle, and ciliary basal body during division [PMID:39073037, PMID:41370039]; its destruction after mitosis depends on APC/C-mediated degradation through a KEN box [PMID:24260314]. At the spindle, CKAP2L stabilizes and bundles microtubules and is required for bipolar spindle formation, correct chromosome segregation, and cytokinesis, such that its loss produces multipolar spindles, chromosome bridges, shortened spindles, and multinucleation [PMID:24260314, PMID:41370039]. Loss-of-function mutations in CKAP2L cause Filippi syndrome, with patient cells showing absence of CKAP2L from spindle poles together with multipolar spindles and segregation defects [PMID:25439729]. Beyond mitosis, CKAP2L acts as a negative regulator of primary cilium length, and its knockout in mice produces abnormally elongated flagella and reduced male fertility [PMID:41370039]. Its transcription is repressed by FOXP3 and activated by FOXM1 and RFX5 [PMID:35065924, PMID:40646610, PMID:39851233]. A separate body of work places CKAP2L in transcriptional and oncogenic signaling roles: it interacts with RNA Polymerase II to regulate transcription elongation of cell-cycle and spindle genes [PMID:33472893], and modulates PI3K/AKT/mTOR signaling, in part by binding AKT and reducing its ubiquitin-mediated degradation [PMID:42245708].","teleology":[{"year":2013,"claim":"Established CKAP2L/Radmis as a microtubule-associated protein required for normal mitotic spindle architecture in neural progenitors, defining its core cellular role.","evidence":"Subtractive screening, shRNA knockdown, overexpression in vitro and in vivo, immunofluorescence in NSPCs","pmids":["24260314"],"confidence":"High","gaps":["Molecular basis of microtubule stabilization not defined","KEN-box/APC/C degradation inferred but direct ubiquitination not shown"]},{"year":2014,"claim":"Connected CKAP2L loss-of-function to a human Mendelian disease, demonstrating its spindle-pole function is essential for faithful mitosis in patient cells.","evidence":"Homozygosity mapping, whole-exome sequencing, immunofluorescence of patient lymphoblastoid lines","pmids":["25439729"],"confidence":"High","gaps":["Does not resolve how spindle defects translate to the developmental phenotypes of Filippi syndrome"]},{"year":2021,"claim":"Proposed a non-spindle function in which CKAP2L binds RNA Pol II to regulate transcription elongation of cell-cycle and checkpoint genes.","evidence":"Reciprocal Co-IP, RNAi screen, proliferation assays in NSCLC cells","pmids":["33472893"],"confidence":"Medium","gaps":["Single-lab Co-IP not independently replicated","Direct vs indirect effect on elongation machinery unresolved"]},{"year":2022,"claim":"Identified FOXP3 as a transcriptional repressor of CKAP2L and linked CKAP2L to AKT/mTOR pathway activation in cancer cells.","evidence":"ChIP/promoter binding, Western blot of pathway phosphorylation, overexpression/silencing in breast cancer lines","pmids":["35065924"],"confidence":"Medium","gaps":["Mechanism connecting a spindle MAP to AKT/mTOR activation not defined","Not independently replicated"]},{"year":2024,"claim":"Defined the cell-cycle and subcellular dynamics of human CKAP2L, showing G2/M-peaking levels and an interphase-nuclear to mitotic-spindle relocalization with a microtubule-bundling phenotype.","evidence":"Cell cycle synchronization, immunofluorescence, ectopic overexpression in cancer cells","pmids":["39073037"],"confidence":"Medium","gaps":["Function of interphase nuclear pool unclear","Mechanism of relocalization unknown"]},{"year":2025,"claim":"Extended CKAP2L function beyond the spindle to microtubule-based organelles, establishing it as a negative regulator of cilium/flagellum length with an in vivo fertility phenotype.","evidence":"Ckap2l knockout mouse, immunofluorescence to centrosome/spindle/basal body, ciliary length measurements, sperm analysis","pmids":["41370039"],"confidence":"High","gaps":["Molecular mechanism limiting cilium length unknown","Relationship between ciliary and spindle roles not resolved"]},{"year":2025,"claim":"Identified FOXM1 and RFX5 as transcriptional activators of CKAP2L, embedding it in proliferative transcriptional programs in cancer.","evidence":"ChIP-qPCR, dual-luciferase reporter, knockdown, rescue experiments in bladder and colorectal cancer cells","pmids":["40646610","39851233"],"confidence":"Medium","gaps":["Conflicting reports on whether CKAP2L activates or inactivates AKT/mTOR downstream","Single-lab studies not independently replicated"]},{"year":2026,"claim":"Proposed a direct biochemical mechanism for oncogenic signaling, in which CKAP2L binds AKT and suppresses its ubiquitin-proteasome degradation to sustain PI3K/AKT signaling, and engages a STAT3/AREG/EGFR axis.","evidence":"Co-IP, ubiquitination assays, cycloheximide chase, MG132 treatment, RNA-seq, ChIP in endometrial and colorectal cancer cells","pmids":["42245708","41823522"],"confidence":"Medium","gaps":["No structural basis for CKAP2L-AKT interaction","How a microtubule MAP physically protects AKT from ubiquitination unexplained","Single-lab studies"]},{"year":null,"claim":"How CKAP2L's spindle/ciliary structural role mechanistically integrates with its reported transcriptional (RNA Pol II) and signaling (AKT/mTOR, STAT3) functions remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying model linking microtubule and signaling functions","Direction of AKT/mTOR regulation conflicts across studies","Signaling mechanisms rely on single-lab evidence"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[0,4,5]}],"localization":[{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[5]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,4,5]},{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[5,10]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[4]}],"pathway":[{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[0,4,5]}],"complexes":[],"partners":["AKT1","POLR2A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8IYA6","full_name":"Cytoskeleton-associated protein 2-like","aliases":["Radial fiber and mitotic spindle protein","Radmis"],"length_aa":745,"mass_kda":83.6,"function":"Microtubule-associated protein required for mitotic spindle formation and cell-cycle progression in neural progenitor cells","subcellular_location":"Cytoplasm, cytoskeleton, spindle pole","url":"https://www.uniprot.org/uniprotkb/Q8IYA6/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CKAP2L","classification":"Not Classified","n_dependent_lines":84,"n_total_lines":1208,"dependency_fraction":0.0695364238410596},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000169607","cell_line_id":"CID001898","localizations":[{"compartment":"centrosome","grade":3},{"compartment":"cytoskeleton","grade":3},{"compartment":"cytoplasmic","grade":2},{"compartment":"nucleoplasm","grade":2},{"compartment":"nuclear_membrane","grade":1}],"interactors":[{"gene":"SIRT5","stoichiometry":10.0},{"gene":"MAP4","stoichiometry":0.2},{"gene":"STX6","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID001898","total_profiled":1310},"omim":[{"mim_id":"616174","title":"CYTOSKELETON-ASSOCIATED PROTEIN 2-LIKE; CKAP2L","url":"https://www.omim.org/entry/616174"},{"mim_id":"272440","title":"FILIPPI SYNDROME; FLPIS","url":"https://www.omim.org/entry/272440"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Microtubules","reliability":"Supported"},{"location":"Mitotic spindle","reliability":"Additional"},{"location":"Primary cilium","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"bone marrow","ntpm":12.4},{"tissue":"lymphoid tissue","ntpm":11.2}],"url":"https://www.proteinatlas.org/search/CKAP2L"},"hgnc":{"alias_symbol":["FLJ40629","radmis"],"prev_symbol":[]},"alphafold":{"accession":"Q8IYA6","domains":[{"cath_id":"-","chopping":"492-605","consensus_level":"high","plddt":86.2993,"start":492,"end":605},{"cath_id":"-","chopping":"663-729","consensus_level":"high","plddt":81.0101,"start":663,"end":729}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8IYA6","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8IYA6-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8IYA6-F1-predicted_aligned_error_v6.png","plddt_mean":54.28},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CKAP2L","jax_strain_url":"https://www.jax.org/strain/search?query=CKAP2L"},"sequence":{"accession":"Q8IYA6","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8IYA6.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8IYA6/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8IYA6"}},"corpus_meta":[{"pmid":"25439729","id":"PMC_25439729","title":"Mutations in CKAP2L, the human homolog of the mouse Radmis gene, cause Filippi syndrome.","date":"2014","source":"American journal of human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25439729","citation_count":41,"is_preprint":false},{"pmid":"24260314","id":"PMC_24260314","title":"Radmis, a novel mitotic spindle protein that functions in cell division of neural progenitors.","date":"2013","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/24260314","citation_count":38,"is_preprint":false},{"pmid":"33472893","id":"PMC_33472893","title":"CKAP2L Promotes Non-Small Cell Lung Cancer Progression through Regulation of Transcription Elongation.","date":"2021","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/33472893","citation_count":21,"is_preprint":false},{"pmid":"33375517","id":"PMC_33375517","title":"CKAP2L Knockdown Exerts Antitumor Effects by Increasing miR-4496 in Glioblastoma Cell Lines.","date":"2020","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/33375517","citation_count":19,"is_preprint":false},{"pmid":"35065924","id":"PMC_35065924","title":"CKAP2L, transcriptionally inhibited by FOXP3, promotes breast carcinogenesis through the AKT/mTOR pathway.","date":"2022","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/35065924","citation_count":14,"is_preprint":false},{"pmid":"36090903","id":"PMC_36090903","title":"CKAP2L Promotes Esophageal Squamous Cell Carcinoma Progression and Drug-Resistance by Modulating Cell Cycle.","date":"2022","source":"Journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/36090903","citation_count":7,"is_preprint":false},{"pmid":"35715760","id":"PMC_35715760","title":"CKAP2L, a crucial target of miR-326, promotes prostate cancer progression.","date":"2022","source":"BMC cancer","url":"https://pubmed.ncbi.nlm.nih.gov/35715760","citation_count":6,"is_preprint":false},{"pmid":"40646610","id":"PMC_40646610","title":"Smoking promotes the progression of bladder cancer through FOXM1/CKAP2L axis.","date":"2025","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40646610","citation_count":3,"is_preprint":false},{"pmid":"40229458","id":"PMC_40229458","title":"Exosomal circ_0001583 Drives Glioblastoma Cell Advancement Through the miR-647/CKAP2L Pathway.","date":"2025","source":"Molecular neurobiology","url":"https://pubmed.ncbi.nlm.nih.gov/40229458","citation_count":3,"is_preprint":false},{"pmid":"39073037","id":"PMC_39073037","title":"Human CKAP2L shows a cell cycle-dependent expression pattern and exhibits microtubule-stabilizing properties.","date":"2024","source":"FEBS open bio","url":"https://pubmed.ncbi.nlm.nih.gov/39073037","citation_count":2,"is_preprint":false},{"pmid":"39851233","id":"PMC_39851233","title":"CKAP2L Plays a Pivotal Role in Colorectal Cancer Progression via the Dual Regulation of Cell Cycle and Epithelial-Mesenchymal Transition.","date":"2025","source":"Discovery medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39851233","citation_count":2,"is_preprint":false},{"pmid":"41370039","id":"PMC_41370039","title":"Filippi syndrome-associated CKAP2L modulates microtubule dynamics essential for mitosis and ciliary length regulation.","date":"2025","source":"Journal of molecular cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/41370039","citation_count":0,"is_preprint":false},{"pmid":"42245708","id":"PMC_42245708","title":"CKAP2L promotes endometrial cancer progression by suppressing AKT ubiquitination and activating the PI3K/AKT signaling pathway.","date":"2026","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/42245708","citation_count":0,"is_preprint":false},{"pmid":"41823522","id":"PMC_41823522","title":"Smoking promotes colorectal cancer via the CKAP2L/AREG axis.","date":"2026","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/41823522","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.03.652041","title":"Proteomic profiling of primary cilia in the developing brain uncovers new regulators of cortical development","date":"2025-05-04","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.03.652041","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":10122,"output_tokens":2953,"usd":0.037331,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10265,"output_tokens":2855,"usd":0.06135,"stage2_stop_reason":"end_turn"},"total_usd":0.098681,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"Radmis/CKAP2L is a microtubule-associated protein (MAP) enriched in neural stem/progenitor cells (NSPCs) that localizes to bipolar mitotic spindles and radial fibers during NSPC division. Overexpression induces hyper-stabilization of microtubules and severe defects in mitotic spindle formation leading to mitotic arrest. Loss-of-function via shRNA induces multipolar mitotic spindle structures and chromosome segregation defects including long chromosome bridges. Radmis is a putative substrate for the APC/C E3-ubiquitin ligase and is degraded via the KEN box.\",\n      \"method\": \"Differential subtractive screening, shRNA knockdown, overexpression in vitro and in vivo (in utero electroporation), immunofluorescence localization\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (KD, OE, in vivo, localization) in a single foundational study, replicated by subsequent papers\",\n      \"pmids\": [\"24260314\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Loss-of-function mutations in CKAP2L cause Filippi syndrome. In dividing lymphoblastoid cell lines from homozygous frameshift mutation carriers, CKAP2L is absent from spindle poles, and cells exhibit multipolar spindle configurations and chromosome segregation defects, establishing CKAP2L's role at the spindle pole as essential for faithful mitosis in human cells.\",\n      \"method\": \"Homozygosity mapping, whole-exome sequencing, Sanger sequencing, immunofluorescence of patient-derived lymphoblastoid cell lines\",\n      \"journal\": \"American journal of human genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — patient-derived cell lines with defined mutations, direct localization assay, phenotypic readout, replicated across multiple families\",\n      \"pmids\": [\"25439729\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CKAP2L directly interacts with RNA Polymerase II and regulates transcription elongation of genes involved in spindle assembly checkpoint, chromosome segregation, cell cycle, and E2F signaling. Depletion of CKAP2L increased sensitivity of NSCLC cells to alvocidib (pan-CDK inhibitor).\",\n      \"method\": \"Co-immunoprecipitation (CKAP2L–RNA Pol II interaction), RNAi screen, in vitro and in vivo proliferation/growth assays\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — reciprocal Co-IP reported in single study, single lab, novel mechanism not yet independently replicated\",\n      \"pmids\": [\"33472893\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"FOXP3 transcriptionally represses CKAP2L by binding to its promoter, and CKAP2L in turn activates the AKT/mTOR signaling pathway in breast cancer cells, as evidenced by increased phosphorylation of AKT, mTOR, and p70S6K upon CKAP2L overexpression.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP), promoter-binding assay, Western blot for pathway phosphorylation, CKAP2L overexpression and silencing in BC cell lines\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — ChIP confirms FOXP3 promoter binding; pathway activation by single lab with two orthogonal methods but not independently replicated\",\n      \"pmids\": [\"35065924\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"Human CKAP2L protein levels are cell cycle phase-dependent, peaking at G2/M. During interphase, CKAP2L localizes predominantly to the nucleus, while during mitosis it localizes to the mitotic spindle apparatus. Ectopic overexpression causes microtubule bundling and prolonged mitosis.\",\n      \"method\": \"Cell cycle synchronization, immunofluorescence, ectopic overexpression in human cancer cell lines\",\n      \"journal\": \"FEBS open bio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiments with functional consequence (prolonged mitosis), two orthogonal methods (cell fractionation/IF + overexpression phenotype), single lab\",\n      \"pmids\": [\"39073037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"CKAP2L is a bona fide microtubule-associated protein that localizes to the centrosome, mitotic spindle, and ciliary basal body. Depletion of CKAP2L leads to shortened mitotic spindles and cytokinesis failure resulting in multinucleation. CKAP2L acts as a negative regulator of primary cilium length; its loss markedly increases ciliary length in both human and mouse cells. CKAP2L knockout mice exhibit reduced male fertility with decreased sperm count, impaired motility, and abnormally elongated flagella.\",\n      \"method\": \"Ckap2l knockout mouse generation, immunofluorescence localization to microtubule-based organelles, ciliary length measurements in human and mouse cells, sperm analysis\",\n      \"journal\": \"Journal of molecular cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo knockout model combined with multiple orthogonal cellular assays (spindle length, cytokinesis, ciliogenesis) across human and mouse systems in a single rigorous study\",\n      \"pmids\": [\"41370039\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"FOXM1 transcriptionally activates CKAP2L by binding to its promoter, as confirmed by ChIP-qPCR. Cigarette smoke extract upregulates FOXM1 and CKAP2L expression, and CKAP2L knockdown arrests the cell cycle at S and G2/M phases in bladder cancer cells.\",\n      \"method\": \"Chromatin immunoprecipitation (ChIP)-qPCR, CKAP2L knockdown, cell cycle flow cytometry\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — ChIP-qPCR confirms FOXM1 promoter binding; single lab, single study, not independently replicated\",\n      \"pmids\": [\"40646610\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RFX5 acts as an upstream transcription factor that directly activates CKAP2L transcription, as confirmed by dual-luciferase assay and chromatin immunoprecipitation. CKAP2L overexpression rescues the antiproliferative and anti-migratory effects of RFX5 knockdown in colorectal cancer cells, and CKAP2L inactivates the AKT/mTOR pathway downstream.\",\n      \"method\": \"Dual-luciferase reporter assay, ChIP, siRNA knockdown, CKAP2L overexpression rescue experiments, Western blot\",\n      \"journal\": \"Discovery medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — two orthogonal methods (luciferase + ChIP) confirm transcriptional regulation; single lab, not independently replicated\",\n      \"pmids\": [\"39851233\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"CKAP2L stabilizes phosphorylated AKT by inhibiting its ubiquitin-proteasome degradation. Co-immunoprecipitation and ubiquitination assays showed that CKAP2L interacts with AKT and overexpression of CKAP2L decreases AKT ubiquitination, sustaining AKT phosphorylation and activating PI3K/AKT signaling in endometrial cancer cells.\",\n      \"method\": \"Co-immunoprecipitation (Co-IP), ubiquitination assay, cycloheximide chase, MG132 proteasome inhibitor assay, Western blot\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP plus functional ubiquitination/degradation assays, multiple orthogonal methods, single lab, not independently replicated\",\n      \"pmids\": [\"42245708\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"CKAP2L promotes AREG (amphiregulin) expression in colorectal cancer cells by increasing STAT3 phosphorylation; activated STAT3 then binds the AREG promoter (confirmed by ChIP) to transcriptionally activate AREG, thereby engaging the AREG/EGFR pathway to drive cancer progression.\",\n      \"method\": \"RNA sequencing, CKAP2L knockdown, ChIP assay (STAT3 at AREG promoter), Western blot for pSTAT3, rescue experiments with AREG suppression\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — ChIP confirms STAT3-AREG promoter binding; RNA-seq + rescue experiments; single lab, single study\",\n      \"pmids\": [\"41823522\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Proximity-labeling proteomics of radial glial cell primary cilia in the embryonic brain identified CKAP2L as a ciliary protein component in the developing brain, and its ciliary localization was experimentally validated.\",\n      \"method\": \"Proximity labeling (BioID), quantitative proteomics, immunofluorescence validation of ciliary localization in embryonic brain\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — preprint, proximity labeling with validation, single study, no functional follow-up on the ciliary localization itself\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"CKAP2L (Radmis) is a microtubule-associated protein that localizes to the centrosome, mitotic spindle, and ciliary basal body; it is degraded by APC/C via its KEN box after mitosis, and its cell cycle-regulated expression (peaking at G2/M) is transcriptionally controlled by factors including FOXP3 (repressor), FOXM1 and RFX5 (activators); during mitosis it stabilizes microtubules and is required for bipolar spindle formation, correct chromosome segregation, and cytokinesis, while its loss leads to multipolar spindles, multinucleation, and elongated primary cilia; beyond its structural spindle role, CKAP2L also interacts with RNA Pol II to regulate transcription elongation and with AKT to suppress its ubiquitin-mediated degradation, thereby activating PI3K/AKT/mTOR signaling.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CKAP2L (Radmis) is a cell cycle-regulated microtubule-associated protein that governs the fidelity of mitotic spindle assembly and the homeostasis of microtubule-based organelles [#0, #5]. Its protein levels peak at G2/M, and it relocalizes from the interphase nucleus to the centrosome, mitotic spindle, and ciliary basal body during division [#4, #5]; its destruction after mitosis depends on APC/C-mediated degradation through a KEN box [#0]. At the spindle, CKAP2L stabilizes and bundles microtubules and is required for bipolar spindle formation, correct chromosome segregation, and cytokinesis, such that its loss produces multipolar spindles, chromosome bridges, shortened spindles, and multinucleation [#0, #5]. Loss-of-function mutations in CKAP2L cause Filippi syndrome, with patient cells showing absence of CKAP2L from spindle poles together with multipolar spindles and segregation defects [#1]. Beyond mitosis, CKAP2L acts as a negative regulator of primary cilium length, and its knockout in mice produces abnormally elongated flagella and reduced male fertility [#5]. Its transcription is repressed by FOXP3 and activated by FOXM1 and RFX5 [#3, #6, #7]. A separate body of work places CKAP2L in transcriptional and oncogenic signaling roles: it interacts with RNA Polymerase II to regulate transcription elongation of cell-cycle and spindle genes [#2], and modulates PI3K/AKT/mTOR signaling, in part by binding AKT and reducing its ubiquitin-mediated degradation [#8].\",\n  \"teleology\": [\n    {\n      \"year\": 2013,\n      \"claim\": \"Established CKAP2L/Radmis as a microtubule-associated protein required for normal mitotic spindle architecture in neural progenitors, defining its core cellular role.\",\n      \"evidence\": \"Subtractive screening, shRNA knockdown, overexpression in vitro and in vivo, immunofluorescence in NSPCs\",\n      \"pmids\": [\"24260314\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of microtubule stabilization not defined\", \"KEN-box/APC/C degradation inferred but direct ubiquitination not shown\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected CKAP2L loss-of-function to a human Mendelian disease, demonstrating its spindle-pole function is essential for faithful mitosis in patient cells.\",\n      \"evidence\": \"Homozygosity mapping, whole-exome sequencing, immunofluorescence of patient lymphoblastoid lines\",\n      \"pmids\": [\"25439729\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve how spindle defects translate to the developmental phenotypes of Filippi syndrome\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Proposed a non-spindle function in which CKAP2L binds RNA Pol II to regulate transcription elongation of cell-cycle and checkpoint genes.\",\n      \"evidence\": \"Reciprocal Co-IP, RNAi screen, proliferation assays in NSCLC cells\",\n      \"pmids\": [\"33472893\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab Co-IP not independently replicated\", \"Direct vs indirect effect on elongation machinery unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified FOXP3 as a transcriptional repressor of CKAP2L and linked CKAP2L to AKT/mTOR pathway activation in cancer cells.\",\n      \"evidence\": \"ChIP/promoter binding, Western blot of pathway phosphorylation, overexpression/silencing in breast cancer lines\",\n      \"pmids\": [\"35065924\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism connecting a spindle MAP to AKT/mTOR activation not defined\", \"Not independently replicated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined the cell-cycle and subcellular dynamics of human CKAP2L, showing G2/M-peaking levels and an interphase-nuclear to mitotic-spindle relocalization with a microtubule-bundling phenotype.\",\n      \"evidence\": \"Cell cycle synchronization, immunofluorescence, ectopic overexpression in cancer cells\",\n      \"pmids\": [\"39073037\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Function of interphase nuclear pool unclear\", \"Mechanism of relocalization unknown\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Extended CKAP2L function beyond the spindle to microtubule-based organelles, establishing it as a negative regulator of cilium/flagellum length with an in vivo fertility phenotype.\",\n      \"evidence\": \"Ckap2l knockout mouse, immunofluorescence to centrosome/spindle/basal body, ciliary length measurements, sperm analysis\",\n      \"pmids\": [\"41370039\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism limiting cilium length unknown\", \"Relationship between ciliary and spindle roles not resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Identified FOXM1 and RFX5 as transcriptional activators of CKAP2L, embedding it in proliferative transcriptional programs in cancer.\",\n      \"evidence\": \"ChIP-qPCR, dual-luciferase reporter, knockdown, rescue experiments in bladder and colorectal cancer cells\",\n      \"pmids\": [\"40646610\", \"39851233\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Conflicting reports on whether CKAP2L activates or inactivates AKT/mTOR downstream\", \"Single-lab studies not independently replicated\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Proposed a direct biochemical mechanism for oncogenic signaling, in which CKAP2L binds AKT and suppresses its ubiquitin-proteasome degradation to sustain PI3K/AKT signaling, and engages a STAT3/AREG/EGFR axis.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, cycloheximide chase, MG132 treatment, RNA-seq, ChIP in endometrial and colorectal cancer cells\",\n      \"pmids\": [\"42245708\", \"41823522\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural basis for CKAP2L-AKT interaction\", \"How a microtubule MAP physically protects AKT from ubiquitination unexplained\", \"Single-lab studies\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CKAP2L's spindle/ciliary structural role mechanistically integrates with its reported transcriptional (RNA Pol II) and signaling (AKT/mTOR, STAT3) functions remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying model linking microtubule and signaling functions\", \"Direction of AKT/mTOR regulation conflicts across studies\", \"Signaling mechanisms rely on single-lab evidence\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [0, 4, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [5]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 4, 5]},\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [5, 10]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [0, 4, 5]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"AKT1\", \"POLR2A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}