{"gene":"IFT56","run_date":"2026-06-10T01:55:22","timeline":{"discoveries":[{"year":2014,"finding":"TTC26/DYF13 is a component of IFT complex B in both mammalian cells and Chlamydomonas reinhardtii, as demonstrated by biochemical fractionation and live imaging. In dyf13 mutant flagella, IFT particle assembly and speed are normal (unlike other IFT complex B mutants), but a specific subset of motility-related proteins is depleted, indicating TTC26 is selectively required for transport of motility cargo rather than general IFT.","method":"Live imaging, biochemical fractionation, proteomic analysis of dyf13 mutant flagella in Chlamydomonas; morpholino knockdown in zebrafish","journal":"eLife","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (live imaging, biochemistry, proteomics) across two model organisms, replicated in mammalian cells","pmids":["24596149"],"is_preprint":false},{"year":2014,"finding":"Wild-type Ttc26 binds directly to the Ift46 subunit of IFT complex B, as shown by protein-protein interaction assays. Loss of Ttc26 impairs Hedgehog signaling by blocking Gli dissociation from Sufu after Gli accumulates at the ciliary tip, rather than preventing Gli accumulation itself.","method":"Protein-protein interaction assay (co-IP/pulldown), analysis of Gli localization and Sufu interaction in embryonic fibroblasts from hop (Ttc26 mutant) mice","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct binding assay plus functional cellular epistasis with Gli/Sufu dissociation step, single lab but two orthogonal methods","pmids":["25340710"],"is_preprint":false},{"year":2017,"finding":"KIF17 (a homodimeric kinesin-2) interacts with the IFT46-IFT56 dimer within the IFT-B complex through its C-terminal sequence immediately upstream of its nuclear localization signal (NLS), as shown by visible immunoprecipitation assay. This interaction is required for KIF17 entry into cilia across the ciliary permeability barrier, but not for intraciliary trafficking once inside.","method":"Visible immunoprecipitation (VIP) assay, domain mapping with C-terminal truncations/mutations, ciliary entry rescue experiments in mammalian cells","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal/direct interaction assay with domain mapping and functional rescue, single lab, two orthogonal methods","pmids":["28077622"],"is_preprint":false},{"year":2017,"finding":"IFT56 is specifically required for the integrity of the IFT-B complex within cilia: in Ift56 mouse mutants, core IFT-B proteins (IFT88, IFT81, IFT27) fail to accumulate normally inside cilia, leading to abnormal ciliary microtubule doublet positioning/number and failure to accumulate Gli proteins, causing defective Shh signaling. IFT-A components and IFT-A-dependent proteins are unaffected.","method":"Mouse genetic mutant analysis, immunofluorescence of IFT-B/IFT-A components in cilia, Gli accumulation assays, limb and neural tube patterning readouts","journal":"Development (Cambridge, England)","confidence":"High","confidence_rationale":"Tier 2 / Moderate — clean KO mouse with defined molecular phenotype (IFT-B accumulation) and specific pathway readout (Gli/Shh), single lab, multiple orthogonal cellular assays","pmids":["28264835"],"is_preprint":false},{"year":2012,"finding":"Ttc26 localizes to the transition zone of photoreceptor sensory cilia and primary cilia in mIMCD3 renal cells. Knockdown of Ttc26 in mIMCD3 cells produces shortened and defective primary cilia, and morpholino knockdown in zebrafish causes ciliary defects in the pronephric kidney and shortened/absent photoreceptor outer segments, establishing a direct requirement for Ttc26 in ciliogenesis.","method":"Immunofluorescence localization, scanning electron microscopy, morpholino knockdown in zebrafish, siRNA knockdown in mIMCD3 cells","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization experiment with functional loss-of-function phenotype in two model systems, single lab","pmids":["22718903"],"is_preprint":false},{"year":2010,"finding":"In Trypanosoma brucei, the DYF-13 orthologue PIFTC3 participates in a ~660 kDa macromolecular complex containing multiple IFT complex B components and putative IFT factors (DYF-1, DYF-3, DYF-11/Elipsa, IFTA-2), as identified by affinity purification and mass spectrometry. Co-immunoprecipitation also detected an interaction between PIFTC3 and IFT122 (IFT complex A), suggesting a bridging role between IFT-A and IFT-B.","method":"Affinity purification and mass spectrometry, co-immunoprecipitation in T. brucei","journal":"Molecular microbiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS-based interactome with co-IP validation in a divergent eukaryote (relevant as ortholog study), single lab","pmids":["20923419"],"is_preprint":false},{"year":2022,"finding":"A CRISPR-replicated point mutation in Ttc26 recapitulates the hop mouse morphological phenotype (absent ventral midline, fused lumbar spinal cord, misplaced notochord, reduced ventral progenitor domains), directly demonstrating that TTC26 is required for normal ventral spinal cord patterning and CPG circuit assembly, likely through ciliary Hedgehog signaling in the floor plate.","method":"CRISPR knock-in of point mutation in Ttc26, histological analysis of spinal cord morphology, fictive locomotion electrophysiology","journal":"eNeuro","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR-replicated mutation with defined morphological and functional phenotype, single lab, genetic confirmation of causality","pmids":["35210288"],"is_preprint":false},{"year":2026,"finding":"TTC26 acts as a scaffold protein that recruits the deubiquitinase MINDY3 to promote K48-linked deubiquitination of RACK1, thereby stabilizing RACK1 protein and activating NFATc1 transcription factor to drive osteoclast differentiation. Six specific TTC26 residues (N533, T534, E537, R541, K545, K548) are essential for this scaffold activity, identified by structural analysis.","method":"Co-immunoprecipitation, Western blotting, RNA sequencing, in vitro osteoclast differentiation assays, protein structural analysis, osteoclast-specific Ttc26 conditional knockout and shRNA knockdown in OVX mice","journal":"Journal of orthopaedic translation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus functional differentiation assays and structural residue mapping, single lab, multiple orthogonal methods","pmids":["42006908"],"is_preprint":false},{"year":2026,"finding":"TTC26 is required for the localization of protein arginine methyltransferase 7 (PRMT7) to the primary cilium, enabling methylation of the transcription factor GLI2. This ciliary methylation of GLI2 is required for its subsequent nuclear import (mediated by FLNB binding to methylated GLI2), thereby activating Shh-GLI2 signaling and maintaining intervertebral disc matrix homeostasis.","method":"Mouse double heterozygous (Flnb/Ttc26) genetic model, transcriptomic analysis, cellular localization of PRMT7 and GLI2, Shh signaling rescue in vivo","journal":"Genome biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis in double-mutant mice with cellular mechanistic follow-up (PRMT7 localization, GLI2 methylation), single lab, multiple methods","pmids":["42178579"],"is_preprint":false},{"year":2025,"finding":"A modifier locus for the Ift56hop phenotype maps to Chromosome 4 in mice, as shown by SNP-based mapping comparing Balb/cByJ (viable, hopping adult) and C57BL/6J (perinatal lethal with multiple organ defects including tracheoesophageal fistulas) backgrounds carrying the same Ift56hop mutation. This demonstrates that genetic background dramatically modifies IFT56 loss-of-function phenotypic severity.","method":"SNP mapping in congenic mouse crosses, phenotypic characterization of Ift56hop across two genetic backgrounds","journal":"Developmental biology","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — genetic mapping with defined chromosomal locus, single lab, modifier gene not yet identified","pmids":["41352382"],"is_preprint":false}],"current_model":"IFT56/TTC26 is an IFT complex B protein that forms a direct dimer with IFT46, functions as a scaffold for cargo-specific intraflagellar transport (particularly motility-related proteins), is required for IFT-B complex integrity within cilia (enabling accumulation of IFT88, IFT81, IFT27 and Gli proteins), regulates ciliary localization of PRMT7 to enable GLI2 methylation and Hedgehog signaling, and outside the cilium acts as a scaffold recruiting the deubiquitinase MINDY3 to stabilize RACK1 and activate NFATc1-driven osteoclastogenesis."},"narrative":{"mechanistic_narrative":"IFT56 (TTC26/DYF13/PIFTC3) is a component of intraflagellar transport (IFT) complex B that is selectively required for the transport of motility-related cargo and for the structural integrity of the IFT-B complex within cilia [PMID:24596149, PMID:28264835]. It binds directly to the IFT46 subunit of IFT-B, and the resulting IFT46–IFT56 dimer serves as a docking site within the complex [PMID:25340710, PMID:28077622]. In mouse mutants, IFT56 is specifically needed for normal ciliary accumulation of core IFT-B proteins (IFT88, IFT81, IFT27) while leaving IFT-A components unaffected, defining its role as an IFT-B-intrinsic scaffold [PMID:28264835]. Through this function IFT56 supports ciliogenesis: its loss produces shortened and defective primary cilia and abnormal photoreceptor outer segments [PMID:22718903]. IFT56 is required for Hedgehog signal transduction at multiple steps — enabling Gli protein accumulation at the ciliary tip, the dissociation of Gli from Sufu, and the ciliary localization of PRMT7 that methylates GLI2 to permit its nuclear import and activation of Shh-GLI2 targets [PMID:25340710, PMID:28264835, PMID:42178579]. These ciliary Hedgehog roles underlie its requirement in ventral spinal cord patterning and intervertebral disc matrix homeostasis [PMID:35210288, PMID:42178579]. The IFT46–IFT56 dimer also recruits the homodimeric kinesin-2 motor KIF17 into cilia across the ciliary permeability barrier via KIF17's C-terminal sequence upstream of its nuclear localization signal [PMID:28077622]. Beyond the cilium, IFT56 acts as a scaffold that recruits the deubiquitinase MINDY3 to deubiquitinate and stabilize RACK1, activating NFATc1-driven osteoclast differentiation [PMID:42006908].","teleology":[{"year":2010,"claim":"Established that the DYF-13 orthologue is embedded in an IFT-B macromolecular complex, defining its biochemical context before its specific function was known.","evidence":"Affinity purification, mass spectrometry, and co-IP of PIFTC3 in Trypanosoma brucei","pmids":["20923419"],"confidence":"Medium","gaps":["Bridging interaction with IFT122 (IFT-A) not validated in other systems","No functional consequence of complex membership defined","Divergent organism may not reflect mammalian complex composition"]},{"year":2012,"claim":"Showed that TTC26 is directly required for ciliogenesis, localizing to the transition zone and being needed for cilium formation and length.","evidence":"Immunofluorescence, SEM, siRNA in mIMCD3 cells, and morpholino knockdown in zebrafish","pmids":["22718903"],"confidence":"Medium","gaps":["Molecular basis of the ciliary defect not resolved","Transition zone localization mechanism unknown","Morpholino phenotypes not confirmed by genetic mutants"]},{"year":2014,"claim":"Distinguished TTC26 from general IFT-B subunits by showing it is selectively required for transport of motility cargo rather than for IFT particle assembly or speed, and that it binds IFT46 directly while controlling the Gli–Sufu dissociation step in Hedgehog signaling.","evidence":"Live imaging, biochemistry and proteomics in Chlamydomonas dyf13 mutants plus zebrafish knockdown; protein interaction assays and Gli/Sufu analysis in hop mouse fibroblasts","pmids":["24596149","25340710"],"confidence":"High","gaps":["Structural basis of the IFT46–TTC26 interface not resolved","How cargo selectivity is achieved mechanistically unclear","Mechanism linking IFT56 to Gli–Sufu dissociation not defined"]},{"year":2017,"claim":"Defined IFT56 as required for in-cilium IFT-B integrity and as the docking site that imports the kinesin-2 motor KIF17 into cilia.","evidence":"Mouse Ift56 mutant immunofluorescence of IFT-B/IFT-A components and Gli assays; visible immunoprecipitation with KIF17 domain mapping and ciliary entry rescue in mammalian cells","pmids":["28264835","28077622"],"confidence":"High","gaps":["How the IFT46–IFT56 dimer enables KIF17 barrier crossing mechanistically unknown","Whether IFT-B disassembly is cause or consequence of microtubule defects unresolved","Other cargos selected by the dimer not catalogued"]},{"year":2022,"claim":"Confirmed by a clean genetic replication that TTC26 is causally required for ventral spinal cord patterning and locomotor circuit assembly through ciliary Hedgehog signaling.","evidence":"CRISPR knock-in point mutation in Ttc26, histological spinal cord analysis, and fictive locomotion electrophysiology","pmids":["35210288"],"confidence":"Medium","gaps":["Direct demonstration that floor-plate Hedgehog defects drive the circuit phenotype not shown","Cell-type-specific contributions not dissected"]},{"year":2025,"claim":"Demonstrated that genetic background strongly modifies the severity of IFT56 loss-of-function, mapping a modifier to Chromosome 4.","evidence":"SNP-based mapping in congenic mouse crosses comparing two backgrounds carrying the same Ift56hop allele","pmids":["41352382"],"confidence":"Medium","gaps":["Modifier gene not identified","Molecular mechanism of phenotypic modification unknown"]},{"year":2026,"claim":"Extended IFT56 function into two new mechanistic arms: ciliary recruitment of PRMT7 to methylate GLI2 for nuclear import, and a non-ciliary scaffold role recruiting MINDY3 to stabilize RACK1 and drive osteoclastogenesis.","evidence":"Flnb/Ttc26 double-heterozygous mice with PRMT7/GLI2 localization and Shh rescue; co-IP, RNA-seq, osteoclast differentiation assays, conditional KO and structural residue mapping","pmids":["42178579","42006908"],"confidence":"Medium","gaps":["Direct enzymatic readout of GLI2 methylation by PRMT7 in cilia not shown","Structural model of the MINDY3/RACK1 scaffold residues not experimentally solved","Relationship between ciliary and non-ciliary scaffold functions unclear"]},{"year":null,"claim":"How the IFT46–IFT56 dimer achieves cargo selectivity and what governs the choice between ciliary IFT-B and non-ciliary scaffold functions remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No high-resolution structure of the IFT46–IFT56 dimer with cargo","Full repertoire of motility cargo dependent on IFT56 not defined","Regulatory switch between cilium-resident and cytoplasmic roles unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[1,2,7]},{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,3]}],"localization":[{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[0,3,4,8]},{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[4]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,3]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,3,8]},{"term_id":"R-HSA-1266738","term_label":"Developmental Biology","supporting_discovery_ids":[6,8]}],"complexes":["IFT complex B"],"partners":["IFT46","KIF17","MINDY3","RACK1","PRMT7"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"A0AVF1","full_name":"Intraflagellar transport protein 56","aliases":["Tetratricopeptide repeat protein 26","TPR repeat protein 26"],"length_aa":554,"mass_kda":64.2,"function":"Component of the intraflagellar transport (IFT) complex B required for transport of proteins in the motile cilium. Required for transport of specific ciliary cargo proteins related to motility, while it is neither required for IFT complex B assembly or motion nor for cilium assembly. Required for efficient coupling between the accumulation of GLI2 and GLI3 at the ciliary tips and their dissociation from the negative regulator SUFU. Plays a key role in maintaining the integrity of the IFT complex B and the proper ciliary localization of the IFT complex B components. Not required for IFT complex A ciliary localization or function. Essential for maintaining proper microtubule organization within the ciliary axoneme","subcellular_location":"Cell projection, cilium","url":"https://www.uniprot.org/uniprotkb/A0AVF1/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/IFT56","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":[{"gene":"HSPB11","stoichiometry":10.0},{"gene":"FKBP5","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/IFT56","total_profiled":1310},"omim":[{"mim_id":"617453","title":"TETRATRICOPEPTIDE REPEAT DOMAIN-CONTAINING PROTEIN 26; TTC26","url":"https://www.omim.org/entry/617453"}],"hpa":{"profiled":true,"resolved_as":"TTC26","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Primary cilium","reliability":"Additional"},{"location":"Mitochondria","reliability":"Additional"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/TTC26"},"hgnc":{"alias_symbol":["FLJ12571","dyf-13","DYF13"],"prev_symbol":["TTC26"]},"alphafold":{"accession":"A0AVF1","domains":[{"cath_id":"1.25.40.10","chopping":"149-270","consensus_level":"medium","plddt":91.6932,"start":149,"end":270},{"cath_id":"1.25.40,1.25.40","chopping":"276-386","consensus_level":"medium","plddt":95.6531,"start":276,"end":386},{"cath_id":"1.25.40,1.25.40","chopping":"390-475","consensus_level":"medium","plddt":96.7972,"start":390,"end":475},{"cath_id":"-","chopping":"476-554","consensus_level":"medium","plddt":93.4666,"start":476,"end":554}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/A0AVF1","model_url":"https://alphafold.ebi.ac.uk/files/AF-A0AVF1-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-A0AVF1-F1-predicted_aligned_error_v6.png","plddt_mean":91.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=IFT56","jax_strain_url":"https://www.jax.org/strain/search?query=IFT56"},"sequence":{"accession":"A0AVF1","fasta_url":"https://rest.uniprot.org/uniprotkb/A0AVF1.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/A0AVF1/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/A0AVF1"}},"corpus_meta":[{"pmid":"24596149","id":"PMC_24596149","title":"TTC26/DYF13 is an intraflagellar transport protein required for transport of motility-related proteins into flagella.","date":"2014","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/24596149","citation_count":62,"is_preprint":false},{"pmid":"28077622","id":"PMC_28077622","title":"Ciliary entry of KIF17 is dependent on its binding to the IFT-B complex via IFT46-IFT56 as well as on its nuclear localization signal.","date":"2017","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/28077622","citation_count":51,"is_preprint":false},{"pmid":"28264835","id":"PMC_28264835","title":"IFT56 regulates vertebrate developmental patterning by maintaining IFTB complex integrity and ciliary microtubule architecture.","date":"2017","source":"Development (Cambridge, England)","url":"https://pubmed.ncbi.nlm.nih.gov/28264835","citation_count":33,"is_preprint":false},{"pmid":"22718903","id":"PMC_22718903","title":"Knockdown of ttc26 disrupts ciliogenesis of the photoreceptor cells and the pronephros in zebrafish.","date":"2012","source":"Molecular biology of the cell","url":"https://pubmed.ncbi.nlm.nih.gov/22718903","citation_count":25,"is_preprint":false},{"pmid":"25340710","id":"PMC_25340710","title":"A mutation in the mouse ttc26 gene leads to impaired hedgehog signaling.","date":"2014","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/25340710","citation_count":24,"is_preprint":false},{"pmid":"20923419","id":"PMC_20923419","title":"Biochemical analysis of PIFTC3, the Trypanosoma brucei orthologue of nematode DYF-13, reveals interactions with established and putative intraflagellar transport components.","date":"2010","source":"Molecular microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/20923419","citation_count":19,"is_preprint":false},{"pmid":"32617964","id":"PMC_32617964","title":"Pituitary stalk interruption syndrome broadens the clinical spectrum of the TTC26 ciliopathy.","date":"2020","source":"Clinical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/32617964","citation_count":12,"is_preprint":false},{"pmid":"34177428","id":"PMC_34177428","title":"Identification of the TTC26 Splice Variant in a Novel Complex Ciliopathy Syndrome with Biliary, Renal, Neurological, and Skeletal Manifestations.","date":"2021","source":"Molecular syndromology","url":"https://pubmed.ncbi.nlm.nih.gov/34177428","citation_count":10,"is_preprint":false},{"pmid":"35210288","id":"PMC_35210288","title":"Hop Mice Display Synchronous Hindlimb Locomotion and a Ventrally Fused Lumbar Spinal Cord Caused by a Point Mutation in Ttc26.","date":"2022","source":"eNeuro","url":"https://pubmed.ncbi.nlm.nih.gov/35210288","citation_count":3,"is_preprint":false},{"pmid":"38135897","id":"PMC_38135897","title":"A novel TTC26 variant in a patient with hexadactyly, pituitary stalk interruption, hepatopathy, nephropathy, and bilateral lip-palate cleft: A case report and expansion of the phenotype.","date":"2023","source":"American journal of medical genetics. Part A","url":"https://pubmed.ncbi.nlm.nih.gov/38135897","citation_count":3,"is_preprint":false},{"pmid":"41352382","id":"PMC_41352382","title":"Genetic background influences the extent and severity of cilia-related congenital anomalies in Ift56/Ttc26 mutant mice.","date":"2025","source":"Developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/41352382","citation_count":0,"is_preprint":false},{"pmid":"42006908","id":"PMC_42006908","title":"TTC26 scaffolds MINDY3-mediated RACK1 deubiquitination to drive osteoclastogenesis and pathological bone resorption.","date":"2026","source":"Journal of orthopaedic translation","url":"https://pubmed.ncbi.nlm.nih.gov/42006908","citation_count":0,"is_preprint":false},{"pmid":"42178579","id":"PMC_42178579","title":"FLNB and TTC26 regulate ciliary Hedgehog signaling to maintain intervertebral disc matrix homeostasis in adolescent idiopathic scoliosis.","date":"2026","source":"Genome biology","url":"https://pubmed.ncbi.nlm.nih.gov/42178579","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8477,"output_tokens":2799,"usd":0.033708,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10073,"output_tokens":3081,"usd":0.063695,"stage2_stop_reason":"end_turn"},"total_usd":0.097403,"stage1_batch_id":"msgbatch_01DUPACZi7AAWwuZiUjseQF4","stage2_batch_id":"msgbatch_018yt7chE3f48PEFtDwtj6AE","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"TTC26/DYF13 is a component of IFT complex B in both mammalian cells and Chlamydomonas reinhardtii, as demonstrated by biochemical fractionation and live imaging. In dyf13 mutant flagella, IFT particle assembly and speed are normal (unlike other IFT complex B mutants), but a specific subset of motility-related proteins is depleted, indicating TTC26 is selectively required for transport of motility cargo rather than general IFT.\",\n      \"method\": \"Live imaging, biochemical fractionation, proteomic analysis of dyf13 mutant flagella in Chlamydomonas; morpholino knockdown in zebrafish\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (live imaging, biochemistry, proteomics) across two model organisms, replicated in mammalian cells\",\n      \"pmids\": [\"24596149\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Wild-type Ttc26 binds directly to the Ift46 subunit of IFT complex B, as shown by protein-protein interaction assays. Loss of Ttc26 impairs Hedgehog signaling by blocking Gli dissociation from Sufu after Gli accumulates at the ciliary tip, rather than preventing Gli accumulation itself.\",\n      \"method\": \"Protein-protein interaction assay (co-IP/pulldown), analysis of Gli localization and Sufu interaction in embryonic fibroblasts from hop (Ttc26 mutant) mice\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct binding assay plus functional cellular epistasis with Gli/Sufu dissociation step, single lab but two orthogonal methods\",\n      \"pmids\": [\"25340710\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"KIF17 (a homodimeric kinesin-2) interacts with the IFT46-IFT56 dimer within the IFT-B complex through its C-terminal sequence immediately upstream of its nuclear localization signal (NLS), as shown by visible immunoprecipitation assay. This interaction is required for KIF17 entry into cilia across the ciliary permeability barrier, but not for intraciliary trafficking once inside.\",\n      \"method\": \"Visible immunoprecipitation (VIP) assay, domain mapping with C-terminal truncations/mutations, ciliary entry rescue experiments in mammalian cells\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal/direct interaction assay with domain mapping and functional rescue, single lab, two orthogonal methods\",\n      \"pmids\": [\"28077622\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"IFT56 is specifically required for the integrity of the IFT-B complex within cilia: in Ift56 mouse mutants, core IFT-B proteins (IFT88, IFT81, IFT27) fail to accumulate normally inside cilia, leading to abnormal ciliary microtubule doublet positioning/number and failure to accumulate Gli proteins, causing defective Shh signaling. IFT-A components and IFT-A-dependent proteins are unaffected.\",\n      \"method\": \"Mouse genetic mutant analysis, immunofluorescence of IFT-B/IFT-A components in cilia, Gli accumulation assays, limb and neural tube patterning readouts\",\n      \"journal\": \"Development (Cambridge, England)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO mouse with defined molecular phenotype (IFT-B accumulation) and specific pathway readout (Gli/Shh), single lab, multiple orthogonal cellular assays\",\n      \"pmids\": [\"28264835\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Ttc26 localizes to the transition zone of photoreceptor sensory cilia and primary cilia in mIMCD3 renal cells. Knockdown of Ttc26 in mIMCD3 cells produces shortened and defective primary cilia, and morpholino knockdown in zebrafish causes ciliary defects in the pronephric kidney and shortened/absent photoreceptor outer segments, establishing a direct requirement for Ttc26 in ciliogenesis.\",\n      \"method\": \"Immunofluorescence localization, scanning electron microscopy, morpholino knockdown in zebrafish, siRNA knockdown in mIMCD3 cells\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment with functional loss-of-function phenotype in two model systems, single lab\",\n      \"pmids\": [\"22718903\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"In Trypanosoma brucei, the DYF-13 orthologue PIFTC3 participates in a ~660 kDa macromolecular complex containing multiple IFT complex B components and putative IFT factors (DYF-1, DYF-3, DYF-11/Elipsa, IFTA-2), as identified by affinity purification and mass spectrometry. Co-immunoprecipitation also detected an interaction between PIFTC3 and IFT122 (IFT complex A), suggesting a bridging role between IFT-A and IFT-B.\",\n      \"method\": \"Affinity purification and mass spectrometry, co-immunoprecipitation in T. brucei\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based interactome with co-IP validation in a divergent eukaryote (relevant as ortholog study), single lab\",\n      \"pmids\": [\"20923419\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A CRISPR-replicated point mutation in Ttc26 recapitulates the hop mouse morphological phenotype (absent ventral midline, fused lumbar spinal cord, misplaced notochord, reduced ventral progenitor domains), directly demonstrating that TTC26 is required for normal ventral spinal cord patterning and CPG circuit assembly, likely through ciliary Hedgehog signaling in the floor plate.\",\n      \"method\": \"CRISPR knock-in of point mutation in Ttc26, histological analysis of spinal cord morphology, fictive locomotion electrophysiology\",\n      \"journal\": \"eNeuro\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR-replicated mutation with defined morphological and functional phenotype, single lab, genetic confirmation of causality\",\n      \"pmids\": [\"35210288\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"TTC26 acts as a scaffold protein that recruits the deubiquitinase MINDY3 to promote K48-linked deubiquitination of RACK1, thereby stabilizing RACK1 protein and activating NFATc1 transcription factor to drive osteoclast differentiation. Six specific TTC26 residues (N533, T534, E537, R541, K545, K548) are essential for this scaffold activity, identified by structural analysis.\",\n      \"method\": \"Co-immunoprecipitation, Western blotting, RNA sequencing, in vitro osteoclast differentiation assays, protein structural analysis, osteoclast-specific Ttc26 conditional knockout and shRNA knockdown in OVX mice\",\n      \"journal\": \"Journal of orthopaedic translation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus functional differentiation assays and structural residue mapping, single lab, multiple orthogonal methods\",\n      \"pmids\": [\"42006908\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"TTC26 is required for the localization of protein arginine methyltransferase 7 (PRMT7) to the primary cilium, enabling methylation of the transcription factor GLI2. This ciliary methylation of GLI2 is required for its subsequent nuclear import (mediated by FLNB binding to methylated GLI2), thereby activating Shh-GLI2 signaling and maintaining intervertebral disc matrix homeostasis.\",\n      \"method\": \"Mouse double heterozygous (Flnb/Ttc26) genetic model, transcriptomic analysis, cellular localization of PRMT7 and GLI2, Shh signaling rescue in vivo\",\n      \"journal\": \"Genome biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis in double-mutant mice with cellular mechanistic follow-up (PRMT7 localization, GLI2 methylation), single lab, multiple methods\",\n      \"pmids\": [\"42178579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A modifier locus for the Ift56hop phenotype maps to Chromosome 4 in mice, as shown by SNP-based mapping comparing Balb/cByJ (viable, hopping adult) and C57BL/6J (perinatal lethal with multiple organ defects including tracheoesophageal fistulas) backgrounds carrying the same Ift56hop mutation. This demonstrates that genetic background dramatically modifies IFT56 loss-of-function phenotypic severity.\",\n      \"method\": \"SNP mapping in congenic mouse crosses, phenotypic characterization of Ift56hop across two genetic backgrounds\",\n      \"journal\": \"Developmental biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — genetic mapping with defined chromosomal locus, single lab, modifier gene not yet identified\",\n      \"pmids\": [\"41352382\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"IFT56/TTC26 is an IFT complex B protein that forms a direct dimer with IFT46, functions as a scaffold for cargo-specific intraflagellar transport (particularly motility-related proteins), is required for IFT-B complex integrity within cilia (enabling accumulation of IFT88, IFT81, IFT27 and Gli proteins), regulates ciliary localization of PRMT7 to enable GLI2 methylation and Hedgehog signaling, and outside the cilium acts as a scaffold recruiting the deubiquitinase MINDY3 to stabilize RACK1 and activate NFATc1-driven osteoclastogenesis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"IFT56 (TTC26/DYF13/PIFTC3) is a component of intraflagellar transport (IFT) complex B that is selectively required for the transport of motility-related cargo and for the structural integrity of the IFT-B complex within cilia [#0, #3]. It binds directly to the IFT46 subunit of IFT-B, and the resulting IFT46–IFT56 dimer serves as a docking site within the complex [#1, #2]. In mouse mutants, IFT56 is specifically needed for normal ciliary accumulation of core IFT-B proteins (IFT88, IFT81, IFT27) while leaving IFT-A components unaffected, defining its role as an IFT-B-intrinsic scaffold [#3]. Through this function IFT56 supports ciliogenesis: its loss produces shortened and defective primary cilia and abnormal photoreceptor outer segments [#4]. IFT56 is required for Hedgehog signal transduction at multiple steps — enabling Gli protein accumulation at the ciliary tip, the dissociation of Gli from Sufu, and the ciliary localization of PRMT7 that methylates GLI2 to permit its nuclear import and activation of Shh-GLI2 targets [#1, #3, #8]. These ciliary Hedgehog roles underlie its requirement in ventral spinal cord patterning and intervertebral disc matrix homeostasis [#6, #8]. The IFT46–IFT56 dimer also recruits the homodimeric kinesin-2 motor KIF17 into cilia across the ciliary permeability barrier via KIF17's C-terminal sequence upstream of its nuclear localization signal [#2]. Beyond the cilium, IFT56 acts as a scaffold that recruits the deubiquitinase MINDY3 to deubiquitinate and stabilize RACK1, activating NFATc1-driven osteoclast differentiation [#7].\",\n  \"teleology\": [\n    {\n      \"year\": 2010,\n      \"claim\": \"Established that the DYF-13 orthologue is embedded in an IFT-B macromolecular complex, defining its biochemical context before its specific function was known.\",\n      \"evidence\": \"Affinity purification, mass spectrometry, and co-IP of PIFTC3 in Trypanosoma brucei\",\n      \"pmids\": [\"20923419\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Bridging interaction with IFT122 (IFT-A) not validated in other systems\", \"No functional consequence of complex membership defined\", \"Divergent organism may not reflect mammalian complex composition\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed that TTC26 is directly required for ciliogenesis, localizing to the transition zone and being needed for cilium formation and length.\",\n      \"evidence\": \"Immunofluorescence, SEM, siRNA in mIMCD3 cells, and morpholino knockdown in zebrafish\",\n      \"pmids\": [\"22718903\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of the ciliary defect not resolved\", \"Transition zone localization mechanism unknown\", \"Morpholino phenotypes not confirmed by genetic mutants\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Distinguished TTC26 from general IFT-B subunits by showing it is selectively required for transport of motility cargo rather than for IFT particle assembly or speed, and that it binds IFT46 directly while controlling the Gli–Sufu dissociation step in Hedgehog signaling.\",\n      \"evidence\": \"Live imaging, biochemistry and proteomics in Chlamydomonas dyf13 mutants plus zebrafish knockdown; protein interaction assays and Gli/Sufu analysis in hop mouse fibroblasts\",\n      \"pmids\": [\"24596149\", \"25340710\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the IFT46–TTC26 interface not resolved\", \"How cargo selectivity is achieved mechanistically unclear\", \"Mechanism linking IFT56 to Gli–Sufu dissociation not defined\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined IFT56 as required for in-cilium IFT-B integrity and as the docking site that imports the kinesin-2 motor KIF17 into cilia.\",\n      \"evidence\": \"Mouse Ift56 mutant immunofluorescence of IFT-B/IFT-A components and Gli assays; visible immunoprecipitation with KIF17 domain mapping and ciliary entry rescue in mammalian cells\",\n      \"pmids\": [\"28264835\", \"28077622\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the IFT46–IFT56 dimer enables KIF17 barrier crossing mechanistically unknown\", \"Whether IFT-B disassembly is cause or consequence of microtubule defects unresolved\", \"Other cargos selected by the dimer not catalogued\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Confirmed by a clean genetic replication that TTC26 is causally required for ventral spinal cord patterning and locomotor circuit assembly through ciliary Hedgehog signaling.\",\n      \"evidence\": \"CRISPR knock-in point mutation in Ttc26, histological spinal cord analysis, and fictive locomotion electrophysiology\",\n      \"pmids\": [\"35210288\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct demonstration that floor-plate Hedgehog defects drive the circuit phenotype not shown\", \"Cell-type-specific contributions not dissected\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated that genetic background strongly modifies the severity of IFT56 loss-of-function, mapping a modifier to Chromosome 4.\",\n      \"evidence\": \"SNP-based mapping in congenic mouse crosses comparing two backgrounds carrying the same Ift56hop allele\",\n      \"pmids\": [\"41352382\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Modifier gene not identified\", \"Molecular mechanism of phenotypic modification unknown\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Extended IFT56 function into two new mechanistic arms: ciliary recruitment of PRMT7 to methylate GLI2 for nuclear import, and a non-ciliary scaffold role recruiting MINDY3 to stabilize RACK1 and drive osteoclastogenesis.\",\n      \"evidence\": \"Flnb/Ttc26 double-heterozygous mice with PRMT7/GLI2 localization and Shh rescue; co-IP, RNA-seq, osteoclast differentiation assays, conditional KO and structural residue mapping\",\n      \"pmids\": [\"42178579\", \"42006908\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct enzymatic readout of GLI2 methylation by PRMT7 in cilia not shown\", \"Structural model of the MINDY3/RACK1 scaffold residues not experimentally solved\", \"Relationship between ciliary and non-ciliary scaffold functions unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How the IFT46–IFT56 dimer achieves cargo selectivity and what governs the choice between ciliary IFT-B and non-ciliary scaffold functions remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No high-resolution structure of the IFT46–IFT56 dimer with cargo\", \"Full repertoire of motility cargo dependent on IFT56 not defined\", \"Regulatory switch between cilium-resident and cytoplasmic roles unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [1, 2, 7]},\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 3]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [0, 3, 4, 8]},\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 3]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 3, 8]},\n      {\"term_id\": \"R-HSA-1266738\", \"supporting_discovery_ids\": [6, 8]}\n    ],\n    \"complexes\": [\n      \"IFT complex B\"\n    ],\n    \"partners\": [\n      \"IFT46\",\n      \"KIF17\",\n      \"MINDY3\",\n      \"RACK1\",\n      \"PRMT7\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}