{"gene":"C9ORF78","run_date":"2026-06-09T22:02:45","timeline":{"discoveries":[{"year":2014,"finding":"The fission yeast ortholog of C9ORF78, Tls1, associates with the spliceosome subunit Brr2 (as shown by protein purification and mass spectrometry). Tls1 regulates splicing of a subset of mRNAs including the shelterin components rap1+ and poz1+; missplicing of these components causes telomeric heterochromatin defects and altered telomere length in tls1Δ cells. Epistasis experiments (replacing rap1+ and poz1+ with cDNAs) confirmed that missplicing of shelterin mRNAs is the mechanistic cause of heterochromatin defects. The human homologue C9ORF78 was also shown to associate with the spliceosome.","method":"Protein purification and mass spectrometry (Tls1 interactors), RNA sequencing (splicing analysis), genetic epistasis (cDNA rescue experiments), deletion library screen","journal":"Nucleic Acids Research","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (purification/MS, RNA-seq, genetic epistasis with cDNA rescue) in a single rigorous study; human C9ORF78 spliceosome association also directly shown","pmids":["25245948"],"is_preprint":false},{"year":2022,"finding":"C9ORF78 tightly interacts with BRR2 (a spliceosome remodeling helicase) in vitro. Cryo-EM structures reveal that C9ORF78 and the spliceosomal B complex protein FBP21 wrap around the C-terminal helicase cassette of BRR2 in a mutually exclusive manner, defining a multi-factor trafficking site. Affinity purification/mass spectrometry and RNA UV-crosslinking identified additional C9ORF78 interactors in spliceosomes, including the 3'-splice site regulating helicase PRPF22. Knockdown of C9ORF78 leads to alternative NAGNAG 3'-splice site usage and exon skipping, with the exon skipping dependent on BRR2. C9ORF78 is established as a late-stage splicing regulatory protein.","method":"In vitro binding assay, cryo-EM structure determination, affinity purification/mass spectrometry, RNA UV-crosslinking, siRNA knockdown with RNA-seq splicing analysis","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with functional validation, in vitro binding, AP-MS, UV-crosslinking, and KD phenotype with BRR2-dependency test; multiple orthogonal methods in single rigorous study","pmids":["35241646"],"is_preprint":false},{"year":2022,"finding":"GFP-tagged and endogenous C9ORF78 localizes to nuclei in interphase but, unexpectedly, a subpopulation co-localizes with kinetochore/centromere markers ACA, Mad1, and Ndc80 in mitotic cells. C9ORF78 levels at centromere/kinetochore increase upon mitotic checkpoint activation. Knockdown of C9ORF78 caused mitotic defects (chromosome segregation errors). Protein abundance is dramatically reduced in confluent/serum-deprived cells and re-expressed upon serum stimulation; overexpression of E2F1 or N-Myc elevated C9ORF78 expression, suggesting these transcription factors mediate serum-dependent upregulation.","method":"GFP tagging and immunofluorescence, co-localization with centromere/kinetochore markers (ACA, Mad1, Ndc80), siRNA knockdown with mitotic defect readout, overexpression of E2F1/N-Myc with Western blot","journal":"Experimental Cell Research","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — immunofluorescence co-localization and KD phenotype from a single lab; multiple markers used but no reconstitution or structural confirmation of centromere interaction","pmids":["35167828"],"is_preprint":false},{"year":2025,"finding":"C9ORF78 forms a complex with FAM50A specifically at the S121 residue of C9ORF78, and this complex enhances ASNS (asparagine synthetase) transcription and accelerates asparagine biosynthesis, facilitating breast cancer brain metastasis.","method":"Co-immunoprecipitation (FAM50A-C9ORF78 complex), site-specific interaction mapping (S121 residue), ASNS transcription assay, asparagine synthesis assay, genetic suppression and pharmacological inhibition with metastasis readout","journal":"Science Advances","confidence":"Medium","confidence_rationale":"Tier 2–3 / Weak — Co-IP interaction with site mapping and functional transcription/metabolic assays reported, but single lab and abstract-level detail limits full method assessment","pmids":["40531994"],"is_preprint":false}],"current_model":"C9ORF78 is an intrinsically unstructured late-stage splicing regulatory protein that binds the spliceosome remodeling helicase BRR2 at a multi-factor trafficking site (competing with FBP21), contacts additional spliceosomal factors including PRPF22, and regulates alternative NAGNAG 3'-splice site selection and BRR2-dependent exon skipping; it also partially localizes to centromeres/kinetochores during mitosis and its loss causes chromosome segregation defects, and it can interact with FAM50A to enhance ASNS transcription and asparagine biosynthesis."},"narrative":{"mechanistic_narrative":"C9ORF78 is a late-stage splicing regulatory protein that engages the spliceosome to control alternative splice-site selection [PMID:25245948, PMID:35241646]. Its fission yeast ortholog Tls1 associates with the spliceosome remodeling helicase Brr2 and governs splicing of a defined subset of mRNAs, including shelterin components whose missplicing drives telomeric heterochromatin and telomere-length defects; human C9ORF78 likewise associates with the spliceosome [PMID:25245948]. Mechanistically, C9ORF78 binds tightly to the C-terminal helicase cassette of BRR2, wrapping around it in a manner mutually exclusive with the B-complex protein FBP21, thereby defining a multi-factor trafficking site on the helicase; it additionally contacts other spliceosomal factors including the 3'-splice-site-regulating helicase PRPF22, and its depletion shifts NAGNAG 3'-splice-site usage and causes BRR2-dependent exon skipping [PMID:35241646]. Beyond splicing, a subpopulation of C9ORF78 localizes to centromeres/kinetochores during mitosis where it co-localizes with ACA, Mad1, and Ndc80, accumulates upon mitotic checkpoint activation, and is required for accurate chromosome segregation [PMID:35167828]. C9ORF78 also forms a complex with FAM50A at its S121 residue to enhance ASNS transcription and asparagine biosynthesis, promoting breast cancer brain metastasis [PMID:40531994].","teleology":[{"year":2014,"claim":"Established that C9ORF78's conserved function lies in spliceosome-associated regulation of a specific mRNA subset, by showing the fission yeast ortholog Tls1 binds Brr2 and controls shelterin splicing with downstream chromatin consequences.","evidence":"Protein purification/MS, RNA-seq, and genetic cDNA-rescue epistasis in fission yeast, with human C9ORF78 spliceosome association also shown","pmids":["25245948"],"confidence":"High","gaps":["Direct biochemical mechanism of human C9ORF78 within the spliceosome not resolved here","Whether human C9ORF78 regulates an orthologous mRNA set is untested"]},{"year":2022,"claim":"Defined the molecular mechanism of C9ORF78 as a late-stage splicing factor that occupies a multi-factor trafficking site on BRR2 to control 3'-splice-site choice and exon inclusion.","evidence":"In vitro binding, cryo-EM of the C9ORF78/FBP21-BRR2 helicase cassette, AP-MS, RNA UV-crosslinking, and siRNA knockdown with BRR2-dependency test","pmids":["35241646"],"confidence":"High","gaps":["Functional consequence of competition with FBP21 in vivo not defined","Role of PRPF22 contact in splice-site selection unresolved"]},{"year":2022,"claim":"Revealed an unexpected mitotic role by showing a C9ORF78 subpopulation localizes to centromeres/kinetochores and is required for faithful chromosome segregation.","evidence":"GFP-tagging/immunofluorescence with centromere/kinetochore markers, siRNA knockdown mitotic-defect readout, and E2F1/N-Myc overexpression Western blot","pmids":["35167828"],"confidence":"Medium","gaps":["No reconstitution or structural confirmation of a centromere interaction","Mechanism linking splicing role to segregation function unknown","Direct binding partner at the kinetochore not identified"]},{"year":2025,"claim":"Connected C9ORF78 to a metabolic-transcriptional axis by showing it complexes with FAM50A to drive ASNS transcription and asparagine biosynthesis in cancer.","evidence":"Co-IP with S121 site mapping, ASNS transcription and asparagine synthesis assays, and genetic/pharmacological perturbation with metastasis readout","pmids":["40531994"],"confidence":"Medium","gaps":["Single-lab finding without reciprocal structural validation","Mechanism by which the complex enhances ASNS transcription undefined","Relationship between this transcriptional role and the spliceosomal function unclear"]},{"year":null,"claim":"How C9ORF78's distinct activities — late-stage splicing regulation, mitotic centromere localization, and FAM50A-dependent transcriptional control — are integrated into one protein's biology remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unifying model connecting splicing, segregation, and metabolic roles","Tissue- and context-specific functions not delineated"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[1]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[2]},{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[2]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,1]}],"complexes":["spliceosome"],"partners":["BRR2","FBP21","PRPF22","FAM50A"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NZ63","full_name":"Splicing factor C9orf78","aliases":["Hepatocellular carcinoma-associated antigen 59"],"length_aa":289,"mass_kda":33.7,"function":"Plays a role in pre-mRNA splicing by promoting usage of the upstream 3'-splice site at alternative NAGNAG splice sites; these are sites featuring alternative acceptor motifs separated by only a few nucleotides (PubMed:35241646). May also modulate exon inclusion events (PubMed:35241646). Plays a role in spliceosomal remodeling by displacing WBP4 from SNRNP200 and may act to inhibit SNRNP200 helicase activity (PubMed:35241646). Binds U5 snRNA (PubMed:35241646). Required for proper chromosome segregation (PubMed:35167828). Not required for splicing of shelterin components (PubMed:35167828)","subcellular_location":"Nucleus; Chromosome, centromere","url":"https://www.uniprot.org/uniprotkb/Q9NZ63/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/C9ORF78"},"opencell":{"profiled":true,"resolved_as":"","ensg_id":"ENSG00000136819","cell_line_id":"CID000961","localizations":[{"compartment":"nucleoplasm","grade":3}],"interactors":[{"gene":"CD2BP2","stoichiometry":0.2},{"gene":"FAM50B","stoichiometry":0.2},{"gene":"FAM50A","stoichiometry":0.2},{"gene":"EFTUD2","stoichiometry":0.2},{"gene":"PRPF8","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/target/CID000961","total_profiled":1310},"omim":[{"mim_id":"619569","title":"CHROMOSOME 9 OPEN READING FRAME 78; C9ORF78","url":"https://www.omim.org/entry/619569"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/C9ORF78"},"hgnc":{"alias_symbol":["HSPC220","HCA59","CSU2"],"prev_symbol":[]},"alphafold":{"accession":"Q9NZ63","domains":[{"cath_id":"-","chopping":"22-64","consensus_level":"medium","plddt":80.7702,"start":22,"end":64}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NZ63","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NZ63-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NZ63-F1-predicted_aligned_error_v6.png","plddt_mean":68.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=C9ORF78","jax_strain_url":"https://www.jax.org/strain/search?query=C9ORF78"},"sequence":{"accession":"Q9NZ63","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NZ63.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NZ63/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NZ63"}},"corpus_meta":[{"pmid":"12097419","id":"PMC_12097419","title":"Large scale identification of human hepatocellular carcinoma-associated antigens by autoantibodies.","date":"2002","source":"Journal of immunology (Baltimore, Md. : 1950)","url":"https://pubmed.ncbi.nlm.nih.gov/12097419","citation_count":160,"is_preprint":false},{"pmid":"20065082","id":"PMC_20065082","title":"Combined interphase fluorescence in situ hybridization elucidates the genetic heterogeneity of T-cell acute lymphoblastic leukemia in adults.","date":"2010","source":"Haematologica","url":"https://pubmed.ncbi.nlm.nih.gov/20065082","citation_count":35,"is_preprint":false},{"pmid":"30871600","id":"PMC_30871600","title":"Hepatocellular carcinoma-associated antigen 59 of Haemonchus contortus modulates the functions of PBMCs and the differentiation and maturation of monocyte-derived dendritic cells of goats in vitro.","date":"2019","source":"Parasites & vectors","url":"https://pubmed.ncbi.nlm.nih.gov/30871600","citation_count":20,"is_preprint":false},{"pmid":"25245948","id":"PMC_25245948","title":"Tls1 regulates splicing of shelterin components to control telomeric heterochromatin assembly and telomere length.","date":"2014","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/25245948","citation_count":16,"is_preprint":false},{"pmid":"31412573","id":"PMC_31412573","title":"Combined Use of Indirect ELISA and Western Blotting with Recombinant Hepatocellular Carcinoma-Associated Antigen 59 Is a Potential Immunodiagnostic Tool for the Detection of Prepatent Haemonchus contortus Infection in Goat.","date":"2019","source":"Animals : an open access journal from MDPI","url":"https://pubmed.ncbi.nlm.nih.gov/31412573","citation_count":14,"is_preprint":false},{"pmid":"33981142","id":"PMC_33981142","title":"Nanoparticles of Chitosan/Poly(D,L-Lactide-Co-Glycolide) Enhanced the Immune Responses of Haemonchus contortus HCA59 Antigen in Model Mice.","date":"2021","source":"International journal of nanomedicine","url":"https://pubmed.ncbi.nlm.nih.gov/33981142","citation_count":10,"is_preprint":false},{"pmid":"35241646","id":"PMC_35241646","title":"A multi-factor trafficking site on the spliceosome remodeling enzyme BRR2 recruits C9ORF78 to regulate alternative splicing.","date":"2022","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/35241646","citation_count":10,"is_preprint":false},{"pmid":"33677347","id":"PMC_33677347","title":"Haemonchus contortus hepatocellular carcinoma-associated antigen 59 with poly (lactic-co-glycolic acid): A promising nanovaccine candidate against Haemonchus contortus infection.","date":"2021","source":"Veterinary parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/33677347","citation_count":10,"is_preprint":false},{"pmid":"38215077","id":"PMC_38215077","title":"AGO2-RIP-Seq reveals miR-34/miR-449 cluster targetome in sinonasal cancers.","date":"2024","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/38215077","citation_count":5,"is_preprint":false},{"pmid":"40531994","id":"PMC_40531994","title":"FAM50A drives breast cancer brain metastasis through interaction with C9ORF78 to enhance ʟ-asparagine production.","date":"2025","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/40531994","citation_count":4,"is_preprint":false},{"pmid":"35167828","id":"PMC_35167828","title":"C9ORF78 partially localizes to centromeres and plays a role in chromosome segregation.","date":"2022","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/35167828","citation_count":2,"is_preprint":false},{"pmid":"42235246","id":"PMC_42235246","title":"Transcriptome and functional analyses reveal key regulators of flowering time in Camellia sinensis.","date":"2026","source":"Plant physiology and biochemistry : PPB","url":"https://pubmed.ncbi.nlm.nih.gov/42235246","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8675,"output_tokens":1944,"usd":0.027592,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":8218,"output_tokens":1998,"usd":0.04552,"stage2_stop_reason":"end_turn"},"total_usd":0.073112,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"Looking at the papers, I need to identify which ones are about the canonical human/mammalian C9ORF78 protein (aliases: HSPC220, HCA59, CSU2):\n\n- Paper [1]: About human HCA59 as a tumor antigen in HCC - identified but no mechanistic experiments on the protein's function. Skip for discoveries.\n- Paper [2]: Mentions C9ORF78 downregulation in leukemia but no mechanistic experiments. Skip.\n- Paper [3]: About *Haemonchus contortus* HCA59 (a parasite protein) - symbol collision, fundamentally different organism/context. Skip.\n- Paper [4]: About fission yeast Tls1, the homologue of C9ORF78 - ortholog paper with mechanistic findings. Include.\n- Paper [5]: Parasite HCA59 - collision. Skip.\n- Paper [6]: Parasite HCA59 - collision. Skip.\n- Paper [7]: Direct mechanistic study of human C9ORF78 in splicing. Include.\n- Paper [8]: Parasite HCA59 - collision. Skip.\n- Paper [9]: miRNA targeting study, C9ORF78 as target gene - expression/biomarker, no mechanistic experiments on C9ORF78. Skip.\n- Paper [10]: FAM50A-C9ORF78 interaction in cancer. Include.\n- Paper [11]: Direct mechanistic study of C9ORF78 localization and mitotic function. Include.\n- Paper [12]: CsCSU2 is a tea plant gene - symbol collision (CSU2 in Camellia sinensis is a flowering regulator, completely different context). Skip.\n\n```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2014,\n      \"finding\": \"The fission yeast ortholog of C9ORF78, Tls1, associates with the spliceosome subunit Brr2 (as shown by protein purification and mass spectrometry). Tls1 regulates splicing of a subset of mRNAs including the shelterin components rap1+ and poz1+; missplicing of these components causes telomeric heterochromatin defects and altered telomere length in tls1Δ cells. Epistasis experiments (replacing rap1+ and poz1+ with cDNAs) confirmed that missplicing of shelterin mRNAs is the mechanistic cause of heterochromatin defects. The human homologue C9ORF78 was also shown to associate with the spliceosome.\",\n      \"method\": \"Protein purification and mass spectrometry (Tls1 interactors), RNA sequencing (splicing analysis), genetic epistasis (cDNA rescue experiments), deletion library screen\",\n      \"journal\": \"Nucleic Acids Research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (purification/MS, RNA-seq, genetic epistasis with cDNA rescue) in a single rigorous study; human C9ORF78 spliceosome association also directly shown\",\n      \"pmids\": [\"25245948\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"C9ORF78 tightly interacts with BRR2 (a spliceosome remodeling helicase) in vitro. Cryo-EM structures reveal that C9ORF78 and the spliceosomal B complex protein FBP21 wrap around the C-terminal helicase cassette of BRR2 in a mutually exclusive manner, defining a multi-factor trafficking site. Affinity purification/mass spectrometry and RNA UV-crosslinking identified additional C9ORF78 interactors in spliceosomes, including the 3'-splice site regulating helicase PRPF22. Knockdown of C9ORF78 leads to alternative NAGNAG 3'-splice site usage and exon skipping, with the exon skipping dependent on BRR2. C9ORF78 is established as a late-stage splicing regulatory protein.\",\n      \"method\": \"In vitro binding assay, cryo-EM structure determination, affinity purification/mass spectrometry, RNA UV-crosslinking, siRNA knockdown with RNA-seq splicing analysis\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with functional validation, in vitro binding, AP-MS, UV-crosslinking, and KD phenotype with BRR2-dependency test; multiple orthogonal methods in single rigorous study\",\n      \"pmids\": [\"35241646\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"GFP-tagged and endogenous C9ORF78 localizes to nuclei in interphase but, unexpectedly, a subpopulation co-localizes with kinetochore/centromere markers ACA, Mad1, and Ndc80 in mitotic cells. C9ORF78 levels at centromere/kinetochore increase upon mitotic checkpoint activation. Knockdown of C9ORF78 caused mitotic defects (chromosome segregation errors). Protein abundance is dramatically reduced in confluent/serum-deprived cells and re-expressed upon serum stimulation; overexpression of E2F1 or N-Myc elevated C9ORF78 expression, suggesting these transcription factors mediate serum-dependent upregulation.\",\n      \"method\": \"GFP tagging and immunofluorescence, co-localization with centromere/kinetochore markers (ACA, Mad1, Ndc80), siRNA knockdown with mitotic defect readout, overexpression of E2F1/N-Myc with Western blot\",\n      \"journal\": \"Experimental Cell Research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — immunofluorescence co-localization and KD phenotype from a single lab; multiple markers used but no reconstitution or structural confirmation of centromere interaction\",\n      \"pmids\": [\"35167828\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"C9ORF78 forms a complex with FAM50A specifically at the S121 residue of C9ORF78, and this complex enhances ASNS (asparagine synthetase) transcription and accelerates asparagine biosynthesis, facilitating breast cancer brain metastasis.\",\n      \"method\": \"Co-immunoprecipitation (FAM50A-C9ORF78 complex), site-specific interaction mapping (S121 residue), ASNS transcription assay, asparagine synthesis assay, genetic suppression and pharmacological inhibition with metastasis readout\",\n      \"journal\": \"Science Advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Weak — Co-IP interaction with site mapping and functional transcription/metabolic assays reported, but single lab and abstract-level detail limits full method assessment\",\n      \"pmids\": [\"40531994\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"C9ORF78 is an intrinsically unstructured late-stage splicing regulatory protein that binds the spliceosome remodeling helicase BRR2 at a multi-factor trafficking site (competing with FBP21), contacts additional spliceosomal factors including PRPF22, and regulates alternative NAGNAG 3'-splice site selection and BRR2-dependent exon skipping; it also partially localizes to centromeres/kinetochores during mitosis and its loss causes chromosome segregation defects, and it can interact with FAM50A to enhance ASNS transcription and asparagine biosynthesis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"C9ORF78 is a late-stage splicing regulatory protein that engages the spliceosome to control alternative splice-site selection [#0, #1]. Its fission yeast ortholog Tls1 associates with the spliceosome remodeling helicase Brr2 and governs splicing of a defined subset of mRNAs, including shelterin components whose missplicing drives telomeric heterochromatin and telomere-length defects; human C9ORF78 likewise associates with the spliceosome [#0]. Mechanistically, C9ORF78 binds tightly to the C-terminal helicase cassette of BRR2, wrapping around it in a manner mutually exclusive with the B-complex protein FBP21, thereby defining a multi-factor trafficking site on the helicase; it additionally contacts other spliceosomal factors including the 3'-splice-site-regulating helicase PRPF22, and its depletion shifts NAGNAG 3'-splice-site usage and causes BRR2-dependent exon skipping [#1]. Beyond splicing, a subpopulation of C9ORF78 localizes to centromeres/kinetochores during mitosis where it co-localizes with ACA, Mad1, and Ndc80, accumulates upon mitotic checkpoint activation, and is required for accurate chromosome segregation [#2]. C9ORF78 also forms a complex with FAM50A at its S121 residue to enhance ASNS transcription and asparagine biosynthesis, promoting breast cancer brain metastasis [#3].\",\n  \"teleology\": [\n    {\n      \"year\": 2014,\n      \"claim\": \"Established that C9ORF78's conserved function lies in spliceosome-associated regulation of a specific mRNA subset, by showing the fission yeast ortholog Tls1 binds Brr2 and controls shelterin splicing with downstream chromatin consequences.\",\n      \"evidence\": \"Protein purification/MS, RNA-seq, and genetic cDNA-rescue epistasis in fission yeast, with human C9ORF78 spliceosome association also shown\",\n      \"pmids\": [\"25245948\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct biochemical mechanism of human C9ORF78 within the spliceosome not resolved here\", \"Whether human C9ORF78 regulates an orthologous mRNA set is untested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Defined the molecular mechanism of C9ORF78 as a late-stage splicing factor that occupies a multi-factor trafficking site on BRR2 to control 3'-splice-site choice and exon inclusion.\",\n      \"evidence\": \"In vitro binding, cryo-EM of the C9ORF78/FBP21-BRR2 helicase cassette, AP-MS, RNA UV-crosslinking, and siRNA knockdown with BRR2-dependency test\",\n      \"pmids\": [\"35241646\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Functional consequence of competition with FBP21 in vivo not defined\", \"Role of PRPF22 contact in splice-site selection unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Revealed an unexpected mitotic role by showing a C9ORF78 subpopulation localizes to centromeres/kinetochores and is required for faithful chromosome segregation.\",\n      \"evidence\": \"GFP-tagging/immunofluorescence with centromere/kinetochore markers, siRNA knockdown mitotic-defect readout, and E2F1/N-Myc overexpression Western blot\",\n      \"pmids\": [\"35167828\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No reconstitution or structural confirmation of a centromere interaction\", \"Mechanism linking splicing role to segregation function unknown\", \"Direct binding partner at the kinetochore not identified\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Connected C9ORF78 to a metabolic-transcriptional axis by showing it complexes with FAM50A to drive ASNS transcription and asparagine biosynthesis in cancer.\",\n      \"evidence\": \"Co-IP with S121 site mapping, ASNS transcription and asparagine synthesis assays, and genetic/pharmacological perturbation with metastasis readout\",\n      \"pmids\": [\"40531994\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab finding without reciprocal structural validation\", \"Mechanism by which the complex enhances ASNS transcription undefined\", \"Relationship between this transcriptional role and the spliceosomal function unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How C9ORF78's distinct activities — late-stage splicing regulation, mitotic centromere localization, and FAM50A-dependent transcriptional control — are integrated into one protein's biology remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unifying model connecting splicing, segregation, and metabolic roles\", \"Tissue- and context-specific functions not delineated\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"complexes\": [\"spliceosome\"],\n    \"partners\": [\"BRR2\", \"FBP21\", \"PRPF22\", \"FAM50A\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":5,"faith_pct":100.0}}