{"gene":"IFT74","run_date":"2026-06-10T01:55:22","timeline":{"discoveries":[{"year":2005,"finding":"IFT74/72 and IFT81 directly interact to form a higher-order oligomer consistent with a tetrameric complex (IFT81)2(IFT74/72)2, serving as a scaffold for IFT complex B assembly. High ionic strength dissociation revealed a 500-kDa IFT-B core containing IFT88, IFT81, IFT74/72, IFT52, IFT46, and IFT27, demonstrating IFT172, IFT80, IFT57, and IFT20 are peripheral subunits not required for core cohesion.","method":"High ionic strength fractionation of Chlamydomonas IFT-B, chemical cross-linking, yeast two-hybrid and three-hybrid analysis, vertebrate homologue interaction confirmation","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal biochemical methods (fractionation, cross-linking, Y2H, Y3H) with conservation confirmed in vertebrate homologues","pmids":["15955805"],"is_preprint":false},{"year":2015,"finding":"IFT74 is required to stabilize IFT-B complex integrity; the region aa 197–641 is sufficient for IFT-B stabilization in vivo. The N-terminus of IFT74 (aa 1–196, including coiled-coil 1) is required for IFT-A/IFT-B association at the flagellar base and for flagellar import of IFT-A, but not strictly required for tubulin entry into flagella. Loss of IFT74 in a null mutant destabilizes IFT-B and causes complete flagella assembly failure.","method":"Chlamydomonas ift74 null mutant rescue with truncated IFT74 transgenes; flagellar assembly assays, IFT protein localization, IFT injection frequency measurements","journal":"Current biology : CB","confidence":"High","confidence_rationale":"Tier 2 / Strong — domain dissection with multiple truncation alleles in vivo, reciprocal phenotypic rescue, multiple orthogonal readouts (flagellar length, IFT-A/B localization, injection frequency)","pmids":["26051893"],"is_preprint":false},{"year":2022,"finding":"The IFT25-IFT27 dimer binds the C-terminal region of the IFT74-IFT81 dimer; this binding region is deleted in BBS-causing IFT74 variants. BBS missense variants of IFT27 are impaired in IFT74-IFT81 binding and cannot rescue BBS-like phenotypes in IFT27-KO cells. BBS variants of IFT74 rescue ciliogenesis in IFT74-KO cells but produce BBS-like abnormal ciliary membrane protein export phenotypes, demonstrating that impaired IFT74-IFT81 / IFT25-IFT27 interaction specifically underlies BBSome-related ciliary defects.","method":"Co-immunoprecipitation, IFT27-KO and IFT74-KO cell rescue assays with BBS variant constructs, ciliogenesis assays, BBS-like phenotype readouts","journal":"Human molecular genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP mapping interaction domain, KO rescue with separation-of-function alleles, multiple orthogonal cellular phenotype readouts","pmids":["34888642"],"is_preprint":false},{"year":2023,"finding":"A reconstituted pentameric IFT complex containing IFT81/IFT74 acts as an unconventional GAP (GTPase-activating protein) for the small GTPase RabL2, enhancing its GTP hydrolysis rate. The GAP activity was mapped to a 70-amino-acid coiled-coil region of IFT81/74. Structural models for RabL2-containing IFT complexes were validated in vitro and in cellulo, and Chlamydomonas IFT81/74 also enhanced GTP hydrolysis of human RabL2, indicating evolutionary conservation. This GAP activity provides a molecular rationale for RabL2 dissociation from anterograde IFT trains after departure from the ciliary base.","method":"In vitro reconstitution of pentameric IFT complex with RabL2; GTP hydrolysis assays; domain mapping; structural modeling validated in vitro and in cellulo; cross-species (Chlamydomonas/human) functional comparison","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro reconstitution with biochemical activity assay, domain mapping, structural validation, evolutionary conservation tested","pmids":["37606072"],"is_preprint":false},{"year":2023,"finding":"The first 40 amino acids of IFT74 (encoded by exon 2) are dispensable for binding to other IFT subunits but are important for tubulin binding, as shown by in vitro studies with the N-terminal deletion. Loss of these 40 residues causes motile cilia defects (severely shortened motile cilia, mucociliary clearance disorder) with little effect on primary cilia structure in mice, consistent with higher tubulin transport demands in motile versus primary cilia.","method":"In vitro tubulin-binding assays with N-terminal deletion constructs; mouse knock-in allele (exon 2 deletion) phenotypic characterization; affinity purification–mass spectrometry (AP-MS) of exon 2-deleted IFT74 showing reduced IFT-B interactions","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro binding assay, mouse allele-specific phenotypes, AP-MS, and parallel human patient data across two independent papers (PMID 37315079 and 37555648)","pmids":["37315079","37555648","36865301"],"is_preprint":false},{"year":2021,"finding":"An IFT74 missense variant (c.256G>A; p.Gly86Ser) adjacent to a splice donor site affects IFT74 mRNA splicing, producing at least two distinct mutant proteins with abnormal subcellular localization along the sperm flagellum, leading to MMAF (multiple morphological abnormalities of the sperm flagellum) and male infertility without other ciliopathy features.","method":"Patient exome sequencing, RT-PCR splice analysis, immunofluorescence localization of mutant IFT74 along sperm flagellum","journal":"Human genetics","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — splice assay and protein localization in patient cells, single lab, two orthogonal methods","pmids":["33689014"],"is_preprint":false},{"year":2021,"finding":"In patient fibroblasts carrying IFT74 truncating variants, ciliogenesis is attenuated; IFT proteins and ciliary membrane proteins (ARL13B, INPP5E, GPR161) show altered ciliary distribution; and hedgehog signaling is disrupted, placing IFT74 upstream of ciliary Hedgehog pathway regulation.","method":"Patient-derived fibroblast analysis; immunofluorescence of IFT proteins and ciliary membrane markers; hedgehog signaling reporter assays; zebrafish ift74 morphant rescue with human p.Q179E variant","journal":"Genetics in medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — patient fibroblasts plus zebrafish morphant rescue with variant, multiple ciliary markers, single lab","pmids":["33531668"],"is_preprint":false},{"year":2021,"finding":"Loss of Ift74 in zebrafish leads to ciliogenesis defects in multiple organs; connecting cilia of photoreceptors are initially formed but fail to maintain, resulting in slow opsin transport and eventual photoreceptor cell death. Large amounts of maternal ift74 transcripts deposited in zebrafish eggs account for the delayed degeneration phenotype compared to other IFT-B mutants.","method":"Zebrafish ift74 mutant generation; live imaging of connecting cilia; opsin transport assays; maternal transcript quantification","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — zebrafish KO with ciliary maintenance and transport readouts, single lab, multiple phenotypic assays","pmids":["34502236"],"is_preprint":false},{"year":2010,"finding":"CMG-1 (IFT74 mammalian ortholog/alias) acts as a transcriptional regulator in mouse spermatocyte-derived GC-2 cells: its knockdown downregulates cyclin D2 at the transcriptional level via a genomic region −250 to −216 of the cyclin D2 gene, and also downregulates E-cadherin and integrin-alpha family genes (α1, α2, α10, α11), impairing collagen adhesion. siRNA-resistant CMG-1 rescue restores E-cadherin and integrin-alpha expression. FLAG-tagged CMG-1 was detected in the nuclei of transfected COS7 cells.","method":"siRNA knockdown in GC-2 cells; reporter assay with cyclin D2 promoter deletion constructs; rescue with siRNA-resistant cDNA; collagen adhesion assay; nuclear localization by FLAG immunofluorescence in COS7 cells","journal":"Genes to cells : devoted to molecular & cellular mechanisms","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, cell-line-based KD and reporter assay; nuclear transcriptional role is inconsistent with the canonical IFT74 function established by preponderance of the corpus; may reflect cell-line artifact or alias collision uncertainty","pmids":["20545763"],"is_preprint":false}],"current_model":"IFT74 is a core IFT-B subunit that heterodimerizes with IFT81 via central and C-terminal coiled-coil domains to form a (IFT74)2(IFT81)2 scaffold for IFT complex B; the IFT74-IFT81 dimer binds IFT25-IFT27 through its C-terminal region to coordinate BBSome-dependent ciliary membrane protein export; the N-terminal domain of IFT74 (including the first 40 amino acids) mediates tubulin binding and is particularly important for tubulin transport into motile cilia, as well as for IFT-A/IFT-B association at the ciliary base enabling IFT-A entry; and the IFT81/74 coiled-coil acts as an unconventional GAP for the small GTPase RabL2, triggering GTP hydrolysis and RabL2 dissociation from anterograde IFT trains at the ciliary base."},"narrative":{"mechanistic_narrative":"IFT74 is a core subunit of intraflagellar transport complex B (IFT-B) that scaffolds train assembly and coordinates the transport machinery required to build and maintain cilia [PMID:15955805, PMID:26051893]. It heterodimerizes with IFT81 to form a higher-order (IFT81)2(IFT74/72)2 tetramer that nucleates the salt-stable IFT-B core, and its central/C-terminal region (aa 197–641) is sufficient to stabilize IFT-B integrity in vivo; complete loss destabilizes the complex and abolishes ciliogenesis [PMID:15955805, PMID:26051893]. The N-terminal coiled-coil (aa 1–196) mediates IFT-A/IFT-B association at the ciliary base and is required for IFT-A import, while the extreme N-terminus (first 40 residues) binds tubulin and is especially critical for tubulin delivery into motile cilia [PMID:26051893, PMID:37315079, PMID:37555648, PMID:36865301]. The C-terminal region of the IFT74–IFT81 dimer recruits the IFT25–IFT27 dimer to coordinate BBSome-dependent export of ciliary membrane proteins, and disruption of this interface produces Bardet-Biedl-like membrane-protein export defects [PMID:34888642]. A 70-residue coiled-coil segment of IFT81/74 additionally functions as an unconventional GTPase-activating protein for RabL2, accelerating its GTP hydrolysis to drive RabL2 release from anterograde IFT trains after departure from the ciliary base [PMID:37606072]. In humans, IFT74 variants cause ciliopathy phenotypes including disrupted ciliary Hedgehog signaling and Bardet-Biedl-spectrum disease, as well as isolated sperm-flagellar defects (MMAF) [PMID:34888642, PMID:33531668, PMID:33689014].","teleology":[{"year":2005,"claim":"Established that IFT74 is not a peripheral component but a structural backbone of IFT-B, defining which subunits form the cohesive core versus accessory periphery.","evidence":"High ionic strength fractionation, cross-linking, and yeast two-/three-hybrid in Chlamydomonas with vertebrate homologue confirmation","pmids":["15955805"],"confidence":"High","gaps":["Does not resolve atomic architecture of the tetramer","Stoichiometry of additional core members within trains not defined"]},{"year":2010,"claim":"Raised the possibility of a nuclear transcriptional role for the IFT74 ortholog CMG-1, distinct from cilia.","evidence":"siRNA knockdown, cyclin D2 promoter reporter assays, and FLAG nuclear localization in mouse GC-2 and COS7 cells","pmids":["20545763"],"confidence":"Low","gaps":["Single-lab cell-line study not independently confirmed","Nuclear transcriptional function inconsistent with the cytoplasmic/ciliary IFT role and may reflect cell-line artifact or alias ambiguity","No direct DNA-binding mechanism demonstrated"]},{"year":2015,"claim":"Dissected which IFT74 domains drive distinct functions, separating IFT-B stabilization from IFT-A recruitment and tubulin entry.","evidence":"Chlamydomonas ift74 null rescue with truncated transgenes, flagellar assembly assays, and IFT injection-frequency measurements","pmids":["26051893"],"confidence":"High","gaps":["Molecular basis of how the N-terminus bridges IFT-A and IFT-B unresolved","Tubulin-binding residues not pinpointed in this study"]},{"year":2021,"claim":"Linked IFT74 loss to ciliary membrane-protein mislocalization and Hedgehog pathway disruption, and demonstrated human disease causality.","evidence":"Patient fibroblasts with truncating variants, ciliary marker immunofluorescence, Hedgehog reporters, and zebrafish morphant rescue with a human variant","pmids":["33531668"],"confidence":"Medium","gaps":["Single-lab patient cohort","Mechanistic link between IFT74 and Hedgehog effectors indirect"]},{"year":2021,"claim":"Showed IFT74 is required for ciliary maintenance and cargo (opsin) transport rather than only initial assembly, explaining tissue-specific degenerative phenotypes.","evidence":"Zebrafish ift74 mutant live imaging of connecting cilia, opsin transport assays, and maternal transcript quantification","pmids":["34502236"],"confidence":"Medium","gaps":["Maternal contribution complicates timing of null phenotype","Direct opsin-IFT74 interaction not tested"]},{"year":2021,"claim":"Demonstrated that a splice-affecting IFT74 variant can cause isolated sperm-flagellar disease without systemic ciliopathy, indicating tissue-selective requirements.","evidence":"Patient exome sequencing, RT-PCR splice analysis, and immunofluorescence of mutant IFT74 along sperm flagellum","pmids":["33689014"],"confidence":"Medium","gaps":["Single patient/lab","Functional consequence of mislocalized isoforms on transport not directly measured"]},{"year":2022,"claim":"Mapped the IFT74-IFT81 C-terminal region as the docking site for IFT25-IFT27 and showed this interface specifically underlies BBSome-dependent membrane export, with separation-of-function disease alleles.","evidence":"Reciprocal Co-IP domain mapping plus IFT27-KO and IFT74-KO rescue with BBS variant constructs and ciliary export readouts","pmids":["34888642"],"confidence":"High","gaps":["How the IFT74/81-IFT25/27 module physically couples to the BBSome not resolved","Identity of exported cargoes incompletely defined"]},{"year":2023,"claim":"Identified an enzymatic function for the IFT81/74 coiled-coil as an unconventional GAP for RabL2, explaining how RabL2 is released from anterograde trains.","evidence":"In vitro reconstitution of a pentameric IFT-RabL2 complex, GTP hydrolysis assays, domain mapping, structural modeling, and cross-species functional comparison","pmids":["37606072"],"confidence":"High","gaps":["Spatial/temporal trigger of GAP activity at the base not fully defined","Catalytic residues not pinpointed"]},{"year":2023,"claim":"Pinpointed the first 40 residues of IFT74 as a tubulin-binding determinant disproportionately required for motile cilia, rationalizing differential cilia-type phenotypes.","evidence":"In vitro tubulin-binding assays with N-terminal deletions, mouse exon 2-deletion knock-in phenotyping, and AP-MS, with parallel human patient data","pmids":["37315079","37555648","36865301"],"confidence":"High","gaps":["Structural basis of tubulin binding by the N-terminus unresolved","Why primary cilia tolerate reduced tubulin transport not mechanistically explained"]},{"year":null,"claim":"How IFT74's multiple modular activities — tubulin loading, IFT-A coupling, IFT25/27-BBSome coordination, and RabL2 GAP activity — are spatially and temporally integrated within a single train remains open.","evidence":"","pmids":[],"confidence":"High","gaps":["No integrated structural model of an assembling train carrying all activities","Regulation switching IFT74 between functional states unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0005198","term_label":"structural molecule activity","supporting_discovery_ids":[0,1]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[1,4]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[3]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[2]}],"localization":[{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[1,6,7]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,1]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[1,7]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[6]}],"complexes":["IFT complex B"],"partners":["IFT81","IFT27","IFT25","RABL2","IFT88","IFT52","IFT46"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q96LB3","full_name":"Intraflagellar transport protein 74 homolog","aliases":["Capillary morphogenesis gene 1 protein","CMG-1","Coiled-coil domain-containing protein 2"],"length_aa":600,"mass_kda":69.2,"function":"Component of the intraflagellar transport (IFT) complex B: together with IFT81, forms a tubulin-binding module that specifically mediates transport of tubulin within the cilium (PubMed:23990561). Binds beta-tubulin via its basic region (PubMed:23990561). Required for ciliogenesis (PubMed:23990561). Essential for flagellogenesis during spermatogenesis (PubMed:33689014)","subcellular_location":"Cell projection, cilium; Cytoplasmic vesicle; Cell projection, cilium, flagellum; Cytoplasmic vesicle, secretory vesicle, acrosome","url":"https://www.uniprot.org/uniprotkb/Q96LB3/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/IFT74","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"HSPB11","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/search/IFT74","total_profiled":1310},"omim":[{"mim_id":"620841","title":"INTRAFLAGELLAR TRANSPORT 25; IFT25","url":"https://www.omim.org/entry/620841"},{"mim_id":"620505","title":"INTRAFLAGELLAR TRANSPORT 22; IFT22","url":"https://www.omim.org/entry/620505"},{"mim_id":"620160","title":"IQ MOTIF-CONTAINING PROTEIN N; IQCN","url":"https://www.omim.org/entry/620160"},{"mim_id":"619585","title":"SPERMATOGENIC FAILURE 58; SPGF58","url":"https://www.omim.org/entry/619585"},{"mim_id":"619582","title":"JOUBERT SYNDROME 40; JBTS40","url":"https://www.omim.org/entry/619582"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Golgi apparatus","reliability":"Supported"},{"location":"Centriolar satellite","reliability":"Supported"},{"location":"Basal body","reliability":"Supported"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/IFT74"},"hgnc":{"alias_symbol":["CMG1","CMG-1","FLJ22621"],"prev_symbol":["CCDC2"]},"alphafold":{"accession":"Q96LB3","domains":[{"cath_id":"-","chopping":"96-224","consensus_level":"medium","plddt":91.7653,"start":96,"end":224}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96LB3","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q96LB3-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q96LB3-F1-predicted_aligned_error_v6.png","plddt_mean":80.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=IFT74","jax_strain_url":"https://www.jax.org/strain/search?query=IFT74"},"sequence":{"accession":"Q96LB3","fasta_url":"https://rest.uniprot.org/uniprotkb/Q96LB3.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q96LB3/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q96LB3"}},"corpus_meta":[{"pmid":"15955805","id":"PMC_15955805","title":"Characterization of the intraflagellar transport complex B core: direct interaction of the IFT81 and IFT74/72 subunits.","date":"2005","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/15955805","citation_count":154,"is_preprint":false},{"pmid":"17166276","id":"PMC_17166276","title":"Analysis of IFT74 as a candidate gene for chromosome 9p-linked ALS-FTD.","date":"2006","source":"BMC neurology","url":"https://pubmed.ncbi.nlm.nih.gov/17166276","citation_count":67,"is_preprint":false},{"pmid":"26051893","id":"PMC_26051893","title":"Assembly of IFT trains at the ciliary base depends on IFT74.","date":"2015","source":"Current biology : CB","url":"https://pubmed.ncbi.nlm.nih.gov/26051893","citation_count":62,"is_preprint":false},{"pmid":"33531668","id":"PMC_33531668","title":"Disrupted intraflagellar transport due to IFT74 variants causes Joubert syndrome.","date":"2021","source":"Genetics in medicine : official journal of the American College of Medical Genetics","url":"https://pubmed.ncbi.nlm.nih.gov/33531668","citation_count":34,"is_preprint":false},{"pmid":"34888642","id":"PMC_34888642","title":"Impaired cooperation between IFT74/BBS22-IFT81 and IFT25-IFT27/BBS19 causes Bardet-Biedl syndrome.","date":"2022","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/34888642","citation_count":31,"is_preprint":false},{"pmid":"33689014","id":"PMC_33689014","title":"A missense mutation in IFT74, encoding for an essential component for intraflagellar transport of Tubulin, causes asthenozoospermia and male infertility without clinical signs of Bardet-Biedl syndrome.","date":"2021","source":"Human genetics","url":"https://pubmed.ncbi.nlm.nih.gov/33689014","citation_count":25,"is_preprint":false},{"pmid":"37315079","id":"PMC_37315079","title":"IFT74 variants cause skeletal ciliopathy and motile cilia defects in mice and humans.","date":"2023","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/37315079","citation_count":20,"is_preprint":false},{"pmid":"32144365","id":"PMC_32144365","title":"Second case of Bardet-Biedl syndrome caused by biallelic variants in IFT74.","date":"2020","source":"European journal of human genetics : EJHG","url":"https://pubmed.ncbi.nlm.nih.gov/32144365","citation_count":18,"is_preprint":false},{"pmid":"33748949","id":"PMC_33748949","title":"Third case of Bardet-Biedl syndrome caused by a biallelic variant predicted to affect splicing of IFT74.","date":"2021","source":"Clinical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/33748949","citation_count":11,"is_preprint":false},{"pmid":"16705683","id":"PMC_16705683","title":"Capillary morphogenesis gene (CMG)-1 is among the genes differentially expressed in mouse male germ line stem cells and embryonic stem cells.","date":"2006","source":"Molecular reproduction and development","url":"https://pubmed.ncbi.nlm.nih.gov/16705683","citation_count":11,"is_preprint":false},{"pmid":"34502236","id":"PMC_34502236","title":"Loss of Ift74 Leads to Slow Photoreceptor Degeneration and Ciliogenesis Defects in Zebrafish.","date":"2021","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/34502236","citation_count":10,"is_preprint":false},{"pmid":"37555648","id":"PMC_37555648","title":"Defective airway intraflagellar transport underlies a combined motile and primary ciliopathy syndrome caused by IFT74 mutations.","date":"2023","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/37555648","citation_count":9,"is_preprint":false},{"pmid":"37606072","id":"PMC_37606072","title":"The IFT81-IFT74 complex acts as an unconventional RabL2 GTPase-activating protein during intraflagellar transport.","date":"2023","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/37606072","citation_count":9,"is_preprint":false},{"pmid":"34539760","id":"PMC_34539760","title":"Case Report: Second Report of Joubert Syndrome Caused by Biallelic Variants in IFT74.","date":"2021","source":"Frontiers in genetics","url":"https://pubmed.ncbi.nlm.nih.gov/34539760","citation_count":9,"is_preprint":false},{"pmid":"20545763","id":"PMC_20545763","title":"Novel role for the intraflagellar transport protein CMG-1 in regulating the transcription of cyclin-D2, E-cadherin and integrin-alpha family genes in mouse spermatocyte-derived cells.","date":"2010","source":"Genes to cells : devoted to molecular & cellular mechanisms","url":"https://pubmed.ncbi.nlm.nih.gov/20545763","citation_count":2,"is_preprint":false},{"pmid":"36865301","id":"PMC_36865301","title":"IFT74 variants cause skeletal ciliopathy and motile cilia defects in mice and humans.","date":"2023","source":"medRxiv : the preprint server for health sciences","url":"https://pubmed.ncbi.nlm.nih.gov/36865301","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":9882,"output_tokens":2991,"usd":0.037255,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":10287,"output_tokens":3309,"usd":0.06708,"stage2_stop_reason":"end_turn"},"total_usd":0.104335,"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\": 2005,\n      \"finding\": \"IFT74/72 and IFT81 directly interact to form a higher-order oligomer consistent with a tetrameric complex (IFT81)2(IFT74/72)2, serving as a scaffold for IFT complex B assembly. High ionic strength dissociation revealed a 500-kDa IFT-B core containing IFT88, IFT81, IFT74/72, IFT52, IFT46, and IFT27, demonstrating IFT172, IFT80, IFT57, and IFT20 are peripheral subunits not required for core cohesion.\",\n      \"method\": \"High ionic strength fractionation of Chlamydomonas IFT-B, chemical cross-linking, yeast two-hybrid and three-hybrid analysis, vertebrate homologue interaction confirmation\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal biochemical methods (fractionation, cross-linking, Y2H, Y3H) with conservation confirmed in vertebrate homologues\",\n      \"pmids\": [\"15955805\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"IFT74 is required to stabilize IFT-B complex integrity; the region aa 197–641 is sufficient for IFT-B stabilization in vivo. The N-terminus of IFT74 (aa 1–196, including coiled-coil 1) is required for IFT-A/IFT-B association at the flagellar base and for flagellar import of IFT-A, but not strictly required for tubulin entry into flagella. Loss of IFT74 in a null mutant destabilizes IFT-B and causes complete flagella assembly failure.\",\n      \"method\": \"Chlamydomonas ift74 null mutant rescue with truncated IFT74 transgenes; flagellar assembly assays, IFT protein localization, IFT injection frequency measurements\",\n      \"journal\": \"Current biology : CB\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — domain dissection with multiple truncation alleles in vivo, reciprocal phenotypic rescue, multiple orthogonal readouts (flagellar length, IFT-A/B localization, injection frequency)\",\n      \"pmids\": [\"26051893\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The IFT25-IFT27 dimer binds the C-terminal region of the IFT74-IFT81 dimer; this binding region is deleted in BBS-causing IFT74 variants. BBS missense variants of IFT27 are impaired in IFT74-IFT81 binding and cannot rescue BBS-like phenotypes in IFT27-KO cells. BBS variants of IFT74 rescue ciliogenesis in IFT74-KO cells but produce BBS-like abnormal ciliary membrane protein export phenotypes, demonstrating that impaired IFT74-IFT81 / IFT25-IFT27 interaction specifically underlies BBSome-related ciliary defects.\",\n      \"method\": \"Co-immunoprecipitation, IFT27-KO and IFT74-KO cell rescue assays with BBS variant constructs, ciliogenesis assays, BBS-like phenotype readouts\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP mapping interaction domain, KO rescue with separation-of-function alleles, multiple orthogonal cellular phenotype readouts\",\n      \"pmids\": [\"34888642\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A reconstituted pentameric IFT complex containing IFT81/IFT74 acts as an unconventional GAP (GTPase-activating protein) for the small GTPase RabL2, enhancing its GTP hydrolysis rate. The GAP activity was mapped to a 70-amino-acid coiled-coil region of IFT81/74. Structural models for RabL2-containing IFT complexes were validated in vitro and in cellulo, and Chlamydomonas IFT81/74 also enhanced GTP hydrolysis of human RabL2, indicating evolutionary conservation. This GAP activity provides a molecular rationale for RabL2 dissociation from anterograde IFT trains after departure from the ciliary base.\",\n      \"method\": \"In vitro reconstitution of pentameric IFT complex with RabL2; GTP hydrolysis assays; domain mapping; structural modeling validated in vitro and in cellulo; cross-species (Chlamydomonas/human) functional comparison\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro reconstitution with biochemical activity assay, domain mapping, structural validation, evolutionary conservation tested\",\n      \"pmids\": [\"37606072\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"The first 40 amino acids of IFT74 (encoded by exon 2) are dispensable for binding to other IFT subunits but are important for tubulin binding, as shown by in vitro studies with the N-terminal deletion. Loss of these 40 residues causes motile cilia defects (severely shortened motile cilia, mucociliary clearance disorder) with little effect on primary cilia structure in mice, consistent with higher tubulin transport demands in motile versus primary cilia.\",\n      \"method\": \"In vitro tubulin-binding assays with N-terminal deletion constructs; mouse knock-in allele (exon 2 deletion) phenotypic characterization; affinity purification–mass spectrometry (AP-MS) of exon 2-deleted IFT74 showing reduced IFT-B interactions\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro binding assay, mouse allele-specific phenotypes, AP-MS, and parallel human patient data across two independent papers (PMID 37315079 and 37555648)\",\n      \"pmids\": [\"37315079\", \"37555648\", \"36865301\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"An IFT74 missense variant (c.256G>A; p.Gly86Ser) adjacent to a splice donor site affects IFT74 mRNA splicing, producing at least two distinct mutant proteins with abnormal subcellular localization along the sperm flagellum, leading to MMAF (multiple morphological abnormalities of the sperm flagellum) and male infertility without other ciliopathy features.\",\n      \"method\": \"Patient exome sequencing, RT-PCR splice analysis, immunofluorescence localization of mutant IFT74 along sperm flagellum\",\n      \"journal\": \"Human genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — splice assay and protein localization in patient cells, single lab, two orthogonal methods\",\n      \"pmids\": [\"33689014\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"In patient fibroblasts carrying IFT74 truncating variants, ciliogenesis is attenuated; IFT proteins and ciliary membrane proteins (ARL13B, INPP5E, GPR161) show altered ciliary distribution; and hedgehog signaling is disrupted, placing IFT74 upstream of ciliary Hedgehog pathway regulation.\",\n      \"method\": \"Patient-derived fibroblast analysis; immunofluorescence of IFT proteins and ciliary membrane markers; hedgehog signaling reporter assays; zebrafish ift74 morphant rescue with human p.Q179E variant\",\n      \"journal\": \"Genetics in medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — patient fibroblasts plus zebrafish morphant rescue with variant, multiple ciliary markers, single lab\",\n      \"pmids\": [\"33531668\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Loss of Ift74 in zebrafish leads to ciliogenesis defects in multiple organs; connecting cilia of photoreceptors are initially formed but fail to maintain, resulting in slow opsin transport and eventual photoreceptor cell death. Large amounts of maternal ift74 transcripts deposited in zebrafish eggs account for the delayed degeneration phenotype compared to other IFT-B mutants.\",\n      \"method\": \"Zebrafish ift74 mutant generation; live imaging of connecting cilia; opsin transport assays; maternal transcript quantification\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — zebrafish KO with ciliary maintenance and transport readouts, single lab, multiple phenotypic assays\",\n      \"pmids\": [\"34502236\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CMG-1 (IFT74 mammalian ortholog/alias) acts as a transcriptional regulator in mouse spermatocyte-derived GC-2 cells: its knockdown downregulates cyclin D2 at the transcriptional level via a genomic region −250 to −216 of the cyclin D2 gene, and also downregulates E-cadherin and integrin-alpha family genes (α1, α2, α10, α11), impairing collagen adhesion. siRNA-resistant CMG-1 rescue restores E-cadherin and integrin-alpha expression. FLAG-tagged CMG-1 was detected in the nuclei of transfected COS7 cells.\",\n      \"method\": \"siRNA knockdown in GC-2 cells; reporter assay with cyclin D2 promoter deletion constructs; rescue with siRNA-resistant cDNA; collagen adhesion assay; nuclear localization by FLAG immunofluorescence in COS7 cells\",\n      \"journal\": \"Genes to cells : devoted to molecular & cellular mechanisms\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, cell-line-based KD and reporter assay; nuclear transcriptional role is inconsistent with the canonical IFT74 function established by preponderance of the corpus; may reflect cell-line artifact or alias collision uncertainty\",\n      \"pmids\": [\"20545763\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"IFT74 is a core IFT-B subunit that heterodimerizes with IFT81 via central and C-terminal coiled-coil domains to form a (IFT74)2(IFT81)2 scaffold for IFT complex B; the IFT74-IFT81 dimer binds IFT25-IFT27 through its C-terminal region to coordinate BBSome-dependent ciliary membrane protein export; the N-terminal domain of IFT74 (including the first 40 amino acids) mediates tubulin binding and is particularly important for tubulin transport into motile cilia, as well as for IFT-A/IFT-B association at the ciliary base enabling IFT-A entry; and the IFT81/74 coiled-coil acts as an unconventional GAP for the small GTPase RabL2, triggering GTP hydrolysis and RabL2 dissociation from anterograde IFT trains at the ciliary base.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"IFT74 is a core subunit of intraflagellar transport complex B (IFT-B) that scaffolds train assembly and coordinates the transport machinery required to build and maintain cilia [#0, #1]. It heterodimerizes with IFT81 to form a higher-order (IFT81)2(IFT74/72)2 tetramer that nucleates the salt-stable IFT-B core, and its central/C-terminal region (aa 197–641) is sufficient to stabilize IFT-B integrity in vivo; complete loss destabilizes the complex and abolishes ciliogenesis [#0, #1]. The N-terminal coiled-coil (aa 1–196) mediates IFT-A/IFT-B association at the ciliary base and is required for IFT-A import, while the extreme N-terminus (first 40 residues) binds tubulin and is especially critical for tubulin delivery into motile cilia [#1, #4]. The C-terminal region of the IFT74–IFT81 dimer recruits the IFT25–IFT27 dimer to coordinate BBSome-dependent export of ciliary membrane proteins, and disruption of this interface produces Bardet-Biedl-like membrane-protein export defects [#2]. A 70-residue coiled-coil segment of IFT81/74 additionally functions as an unconventional GTPase-activating protein for RabL2, accelerating its GTP hydrolysis to drive RabL2 release from anterograde IFT trains after departure from the ciliary base [#3]. In humans, IFT74 variants cause ciliopathy phenotypes including disrupted ciliary Hedgehog signaling and Bardet-Biedl-spectrum disease, as well as isolated sperm-flagellar defects (MMAF) [#2, #6, #5].\",\n  \"teleology\": [\n    {\n      \"year\": 2005,\n      \"claim\": \"Established that IFT74 is not a peripheral component but a structural backbone of IFT-B, defining which subunits form the cohesive core versus accessory periphery.\",\n      \"evidence\": \"High ionic strength fractionation, cross-linking, and yeast two-/three-hybrid in Chlamydomonas with vertebrate homologue confirmation\",\n      \"pmids\": [\"15955805\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not resolve atomic architecture of the tetramer\", \"Stoichiometry of additional core members within trains not defined\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Raised the possibility of a nuclear transcriptional role for the IFT74 ortholog CMG-1, distinct from cilia.\",\n      \"evidence\": \"siRNA knockdown, cyclin D2 promoter reporter assays, and FLAG nuclear localization in mouse GC-2 and COS7 cells\",\n      \"pmids\": [\"20545763\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single-lab cell-line study not independently confirmed\", \"Nuclear transcriptional function inconsistent with the cytoplasmic/ciliary IFT role and may reflect cell-line artifact or alias ambiguity\", \"No direct DNA-binding mechanism demonstrated\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Dissected which IFT74 domains drive distinct functions, separating IFT-B stabilization from IFT-A recruitment and tubulin entry.\",\n      \"evidence\": \"Chlamydomonas ift74 null rescue with truncated transgenes, flagellar assembly assays, and IFT injection-frequency measurements\",\n      \"pmids\": [\"26051893\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of how the N-terminus bridges IFT-A and IFT-B unresolved\", \"Tubulin-binding residues not pinpointed in this study\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked IFT74 loss to ciliary membrane-protein mislocalization and Hedgehog pathway disruption, and demonstrated human disease causality.\",\n      \"evidence\": \"Patient fibroblasts with truncating variants, ciliary marker immunofluorescence, Hedgehog reporters, and zebrafish morphant rescue with a human variant\",\n      \"pmids\": [\"33531668\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab patient cohort\", \"Mechanistic link between IFT74 and Hedgehog effectors indirect\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Showed IFT74 is required for ciliary maintenance and cargo (opsin) transport rather than only initial assembly, explaining tissue-specific degenerative phenotypes.\",\n      \"evidence\": \"Zebrafish ift74 mutant live imaging of connecting cilia, opsin transport assays, and maternal transcript quantification\",\n      \"pmids\": [\"34502236\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Maternal contribution complicates timing of null phenotype\", \"Direct opsin-IFT74 interaction not tested\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated that a splice-affecting IFT74 variant can cause isolated sperm-flagellar disease without systemic ciliopathy, indicating tissue-selective requirements.\",\n      \"evidence\": \"Patient exome sequencing, RT-PCR splice analysis, and immunofluorescence of mutant IFT74 along sperm flagellum\",\n      \"pmids\": [\"33689014\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single patient/lab\", \"Functional consequence of mislocalized isoforms on transport not directly measured\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Mapped the IFT74-IFT81 C-terminal region as the docking site for IFT25-IFT27 and showed this interface specifically underlies BBSome-dependent membrane export, with separation-of-function disease alleles.\",\n      \"evidence\": \"Reciprocal Co-IP domain mapping plus IFT27-KO and IFT74-KO rescue with BBS variant constructs and ciliary export readouts\",\n      \"pmids\": [\"34888642\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How the IFT74/81-IFT25/27 module physically couples to the BBSome not resolved\", \"Identity of exported cargoes incompletely defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identified an enzymatic function for the IFT81/74 coiled-coil as an unconventional GAP for RabL2, explaining how RabL2 is released from anterograde trains.\",\n      \"evidence\": \"In vitro reconstitution of a pentameric IFT-RabL2 complex, GTP hydrolysis assays, domain mapping, structural modeling, and cross-species functional comparison\",\n      \"pmids\": [\"37606072\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spatial/temporal trigger of GAP activity at the base not fully defined\", \"Catalytic residues not pinpointed\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Pinpointed the first 40 residues of IFT74 as a tubulin-binding determinant disproportionately required for motile cilia, rationalizing differential cilia-type phenotypes.\",\n      \"evidence\": \"In vitro tubulin-binding assays with N-terminal deletions, mouse exon 2-deletion knock-in phenotyping, and AP-MS, with parallel human patient data\",\n      \"pmids\": [\"37315079\", \"37555648\", \"36865301\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of tubulin binding by the N-terminus unresolved\", \"Why primary cilia tolerate reduced tubulin transport not mechanistically explained\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How IFT74's multiple modular activities — tubulin loading, IFT-A coupling, IFT25/27-BBSome coordination, and RabL2 GAP activity — are spatially and temporally integrated within a single train remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No integrated structural model of an assembling train carrying all activities\", \"Regulation switching IFT74 between functional states unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0005198\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [1, 4]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [1, 6, 7]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [1, 7]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [6]}\n    ],\n    \"complexes\": [\n      \"IFT complex B\"\n    ],\n    \"partners\": [\n      \"IFT81\",\n      \"IFT27\",\n      \"IFT25\",\n      \"RABL2\",\n      \"IFT88\",\n      \"IFT52\",\n      \"IFT46\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}