{"gene":"CHTOP","run_date":"2026-06-09T22:57:18","timeline":{"discoveries":[{"year":2013,"finding":"Chtop is a novel component of the TREX mRNA export complex. Chtop activates the ATPase and RNA helicase activities of Uap56 (similar to Alyref). Uap56 recruits both Alyref and Chtop onto mRNA. Chtop binds to the NTF2-like domain of Nxf1 in a manner requiring arginine methylation of Chtop, and this interaction is mutually exclusive with Thoc5 binding. Chtop binds Uap56 in a mutually exclusive manner with Alyref. Co-knockdown of Alyref and Chtop results in a potent mRNA export block. Together, these interactions indicate TREX and Nxf1 undergo dynamic remodelling driven by Uap56 ATPase activity and Chtop post-translational modifications.","method":"Co-immunoprecipitation, RNAi knockdown, ATPase/helicase activity assays, mRNA export assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, functional biochemical assays, RNAi with defined cellular phenotype, multiple orthogonal methods in a single rigorous study","pmids":["23299939"],"is_preprint":false},{"year":2012,"finding":"Chtop interacts with a nuclear complex called Five Friends of Methylated Chtop (5FMC), which can only be recruited when Chtop is arginine-methylated by Prmt1. 5FMC consists of Pelp1 (core scaffold), Senp3 (SUMO-specific protease), Wdr18, Tex10, and Las1L. Recruitment of 5FMC to the zinc-finger transcription factor Zbp-89 affects its sumoylation status and transactivation potential, providing a mechanistic link between arginine methylation and desumoylation in transcriptional control.","method":"Biotinylation-proteomics (BioID-like approach), mass spectrometry, co-immunoprecipitation, sumoylation/transactivation assays","journal":"Molecular & cellular proteomics : MCP","confidence":"High","confidence_rationale":"Tier 2 / Moderate — MS-based interactome with functional follow-up (sumoylation and transactivation assays), multiple orthogonal methods, single lab","pmids":["22872859"],"is_preprint":false},{"year":2014,"finding":"CHTOP binds to 5-hydroxymethylcytosine (5hmC) on chromatin and is associated with the arginine methyltransferase methylosome complex. This CHTOP-methylosome complex promotes PRMT1-mediated methylation of arginine 3 of histone H4 (H4R3) at genes involved in glioblastomagenesis (EGFR, AKT3, CDK6, CCND2, BRAF), thereby activating their transcription. CHTOP and PRMT1 are both required for expression of these genes, and CHTOP is required for glioblastoma cell tumorigenicity.","method":"Chromatin immunoprecipitation, co-immunoprecipitation, RNAi knockdown, in vitro methylation assay, gene expression analysis","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Moderate — ChIP, Co-IP, RNAi with defined transcriptional and tumorigenic phenotype, multiple orthogonal methods in a single lab","pmids":["25284789"],"is_preprint":false},{"year":2009,"finding":"SRAG (CHTOP) localizes to the nucleus and nucleolus; nucleolar localization is regulated by the N-terminal domain. CHTOP can interact with RNA. Full-length CHTOP protein levels are highest in resting cells and reduced in proliferating cells, with reduction mapping to the G2/M phase. Overexpression of CHTOP reduces the percentage of cells in G2/M and increases cell death.","method":"Subcellular fractionation, immunofluorescence microscopy, N-terminal truncation mutants, cell cycle analysis (flow cytometry), overexpression studies","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 / Weak — direct localization experiments with truncation mutants, overexpression phenotype, single lab, no in vitro reconstitution","pmids":["19254951"],"is_preprint":false},{"year":2022,"finding":"Arginine methylation of CHTOP by Type I PRMTs regulates post-transcriptional intron detention. Mutagenesis of all methylarginine sites in CHTOP recapitulated the retained-intron splicing changes seen with Type I PRMT inhibition. CHTOP occupancy on chromatin-associated polyadenylated RNA is altered upon Type I PRMT inhibition. Retained introns are enriched in nucleoplasm and chromatin fractions, and CHTOP arginine methylation is required for their post-transcriptional processing.","method":"PRMT inhibition (pharmacological), targeted mutagenesis of methylarginine sites, SKaTER-seq (co-transcriptional splicing kinetics), actinomycin D transcription block, proteomic analysis of chromatin fractions, subcellular fractionation","journal":"eLife","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — targeted mutagenesis recapitulating pharmacological phenotype combined with multiple sequencing and proteomic methods, single lab but multiple orthogonal approaches","pmids":["34984976"],"is_preprint":false},{"year":2018,"finding":"Chtop binds competitively to both PRMT1 and PRMT5, thereby promoting either asymmetric or symmetric methylation of arginine residues depending on which enzyme is bound. Cellular Chtop levels are autoregulated by a mechanism involving intron retention and nonsense-mediated mRNA decay (NMD).","method":"Competitive binding assays (Co-IP), intron retention and NMD analysis (RNA analysis with NMD inhibitors)","journal":"RNA biology","confidence":"Medium","confidence_rationale":"Tier 3 / Weak — review/primary study combination; competitive binding reported but abstract compresses methods; NMD-based autoregulation described as a recent finding but single lab, limited methodological detail in abstract","pmids":["29683372"],"is_preprint":false},{"year":2016,"finding":"KSHV ORF57 protein interacts with CHTOP (and CIP29) as components of the human TREX complex. Depletion of CHTOP affects ORF57-mediated viral mRNA processing, indicating CHTOP is recruited into an ORF57-mediated viral ribonucleoprotein particle (vRNP) and is functionally required for viral mRNA export.","method":"Co-immunoprecipitation, siRNA depletion, viral mRNA processing assays","journal":"The Journal of general virology","confidence":"Medium","confidence_rationale":"Tier 3 / Weak — single Co-IP and depletion study, single lab, limited mechanistic detail in abstract","pmids":["27189710"],"is_preprint":false}],"current_model":"CHTOP is a vertebrate-specific, arginine-rich nuclear/nucleolar protein that functions as a dynamic component of the TREX mRNA export complex (activating Uap56 ATPase/helicase activity and bridging mRNA to the export receptor Nxf1 in an arginine-methylation-dependent manner), while also acting as an epigenetic co-activator by binding 5-hydroxymethylcytosine on chromatin and recruiting the PRMT1-containing methylosome to drive H4R3 methylation and gene activation; its arginine methylation by PRMT1 is additionally required for assembly of the 5FMC desumoylation complex (containing Pelp1, Senp3, and others) that modulates transcription factor activity, and for post-transcriptional regulation of nuclear retained introns, with cellular CHTOP levels themselves autoregulated through intron retention and nonsense-mediated mRNA decay."},"narrative":{"mechanistic_narrative":"CHTOP is a vertebrate arginine-rich nuclear and nucleolar protein that couples mRNA export to arginine-methylation-dependent control of gene expression [PMID:23299939, PMID:19254951]. As a dynamic component of the TREX complex, CHTOP activates the ATPase and RNA helicase activities of Uap56 and is loaded with Alyref onto mRNA, then binds the NTF2-like domain of Nxf1 in a manner requiring its own arginine methylation; its Nxf1 and Uap56 contacts are mutually exclusive with Thoc5 and Alyref respectively, so that Uap56 ATPase activity and CHTOP modification drive remodelling of TREX during export, and co-depletion of CHTOP and Alyref blocks bulk mRNA export [PMID:23299939]. CHTOP binds competitively to PRMT1 and PRMT5, directing either asymmetric or symmetric arginine methylation, and this methylation state governs its functional partnerships [PMID:29683372]. On chromatin, CHTOP binds 5-hydroxymethylcytosine and recruits the PRMT1-containing methylosome to promote H4R3 methylation and transcriptional activation of growth-promoting genes, a role required for glioblastoma cell tumorigenicity [PMID:25284789]. PRMT1-dependent methylation of CHTOP is also the trigger for assembly of the 5FMC complex (Pelp1, Senp3, Wdr18, Tex10, Las1L), which alters the sumoylation and transactivation of the transcription factor Zbp-89 [PMID:22872859], and is required for the post-transcriptional processing of nucleoplasm- and chromatin-retained detained introns [PMID:34984976]. CHTOP abundance is autoregulated through intron retention coupled to nonsense-mediated mRNA decay [PMID:29683372], and the protein is appropriated by KSHV ORF57 to support viral mRNA export [PMID:27189710].","teleology":[{"year":2009,"claim":"Established the basic cell biology of CHTOP: where it resides, that it contacts RNA, and that its level tracks proliferative state, framing it as a regulated nuclear/nucleolar RNA-associated factor.","evidence":"Subcellular fractionation, immunofluorescence, N-terminal truncation mutants, and cell-cycle flow cytometry in cultured cells","pmids":["19254951"],"confidence":"Medium","gaps":["No molecular partners or biochemical activity identified","Mechanism linking CHTOP level to G2/M and cell death not defined","RNA-binding specificity unresolved"]},{"year":2012,"claim":"Showed that arginine methylation of CHTOP is the switch that nucleates a defined nuclear complex, connecting a post-translational mark to transcriptional control via desumoylation.","evidence":"Biotinylation-proteomics, mass spectrometry, Co-IP, and sumoylation/transactivation assays defining the 5FMC complex and its effect on Zbp-89","pmids":["22872859"],"confidence":"High","gaps":["Generality of 5FMC targets beyond Zbp-89 unknown","Structural basis of methyl-CHTOP recognition by Pelp1 not resolved","Direct desumoylation of Zbp-89 by Senp3 within the complex not reconstituted"]},{"year":2013,"claim":"Identified CHTOP as a core, dynamic TREX subunit and defined the biochemical logic of export-complex remodelling, explaining how mRNA is handed to the Nxf1 export receptor.","evidence":"Reciprocal Co-IP, Uap56 ATPase/helicase assays, RNAi co-knockdown with mRNA export readout in human cells","pmids":["23299939"],"confidence":"High","gaps":["Identity of the methyltransferase acting on CHTOP within TREX not pinned in this study","Stoichiometry and order of Alyref/CHTOP exchange on individual transcripts unknown","Structure of the CHTOP-Nxf1 interface not solved"]},{"year":2014,"claim":"Connected CHTOP to chromatin directly by showing it reads 5hmC and writes an activating histone mark through the methylosome, establishing a co-activator role with oncogenic consequences.","evidence":"ChIP, Co-IP, in vitro methylation assays, and RNAi with transcriptional and tumorigenicity readouts in glioblastoma cells","pmids":["25284789"],"confidence":"High","gaps":["Domain mediating 5hmC recognition not mapped","Whether export and chromatin functions are physically separable not addressed","Breadth of CHTOP-methylosome target genes beyond the named oncogenes unknown"]},{"year":2018,"claim":"Revealed that CHTOP partitions between two antagonistic PRMTs and that its own abundance is feedback-controlled, positioning it as a tunable hub for methylation-dependent outputs.","evidence":"Competitive Co-IP binding assays and intron-retention/NMD analysis with NMD inhibitors","pmids":["29683372"],"confidence":"Medium","gaps":["Cellular signals that bias PRMT1 versus PRMT5 binding not identified","Quantitative contribution of NMD autoregulation to steady-state CHTOP unknown","Methods compressed in abstract; competitive binding not independently confirmed"]},{"year":2022,"claim":"Demonstrated that CHTOP arginine methylation directly governs post-transcriptional processing of detained introns, linking its modification state to splicing outcomes downstream of transcription.","evidence":"Type I PRMT inhibition, methylarginine-site mutagenesis, SKaTER-seq, actinomycin D chase, and chromatin-fraction proteomics in human cells","pmids":["34984976"],"confidence":"High","gaps":["Mechanism by which methyl-CHTOP promotes intron removal not defined","Splicing factors recruited by CHTOP for detained introns unidentified","Overlap with TREX export function on the same transcripts unresolved"]},{"year":2016,"claim":"Showed a viral pathogen co-opts CHTOP, establishing that its TREX-based export activity is required for processing and export of viral mRNAs.","evidence":"Co-IP and siRNA depletion with viral mRNA processing assays for KSHV ORF57","pmids":["27189710"],"confidence":"Medium","gaps":["Direct versus TREX-mediated CHTOP-ORF57 contact not distinguished","Single Co-IP/depletion study","Selectivity for viral over host transcripts not quantified"]},{"year":null,"claim":"How CHTOP's separate functions — TREX-mediated export, 5hmC/histone co-activation, 5FMC desumoylation, and intron-detention processing — are coordinated, and what determines its choice of PRMT and downstream complex on a given transcript or locus, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model of CHTOP or its methyl-dependent interfaces","No genome-wide integration of its chromatin versus RNA-export roles","Signals controlling PRMT1/PRMT5 partitioning unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[0,3]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[2,1]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3]},{"term_id":"GO:0005730","term_label":"nucleolus","supporting_discovery_ids":[3]},{"term_id":"GO:0005654","term_label":"nucleoplasm","supporting_discovery_ids":[4]},{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[2,4]}],"pathway":[{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[0,4]},{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[2,1]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[2]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[2,6]}],"complexes":["TREX complex","5FMC complex","methylosome"],"partners":["UAP56","NXF1","ALYREF","PRMT1","PRMT5","PELP1","SENP3","ZNF148"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9Y3Y2","full_name":"Chromatin target of PRMT1 protein","aliases":["Friend of PRMT1 protein","Small arginine- and glycine-rich protein","SRAG"],"length_aa":248,"mass_kda":26.4,"function":"Plays an important role in the ligand-dependent activation of estrogen receptor target genes (PubMed:19858291). May play a role in the silencing of fetal globin genes (PubMed:20688955). Recruits the 5FMC complex to ZNF148, leading to desumoylation of ZNF148 and subsequent transactivation of ZNF148 target genes (By similarity). Plays an important role in the tumorigenicity of glioblastoma cells. Binds to 5-hydroxymethylcytosine (5hmC) and associates with the methylosome complex containing PRMT1, PRMT5, MEP50 and ERH. The CHTOP-methylosome complex associated with 5hmC is recruited to selective sites on the chromosome, where it methylates H4R3 and activates the transcription of genes involved in glioblastomagenesis (PubMed:25284789) Required for effective mRNA nuclear export and is a component of the TREX complex which is thought to couple mRNA transcription, processing and nuclear export, and specifically associates with spliced mRNA and not with unspliced pre-mRNA. TREX is recruited to spliced mRNAs by a transcription-independent mechanism, binds to mRNA upstream of the exon-junction complex (EJC) and is recruited in a splicing- and cap-dependent manner to a region near the 5' end of the mRNA where it functions in mRNA export to the cytoplasm via the TAP/NFX1 pathway. The TREX complex is essential for the export of Kaposi's sarcoma-associated herpesvirus (KSHV) intronless mRNAs and infectious virus production. Stimulates DDX39B ATPase and helicase activities. In cooperation with ALYREF/THOC4 enhances NXF1 RNA binding activity (PubMed:23299939)","subcellular_location":"Nucleus; Nucleus, nucleolus; Nucleus, nucleoplasm; Nucleus speckle","url":"https://www.uniprot.org/uniprotkb/Q9Y3Y2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CHTOP","classification":"Not Classified","n_dependent_lines":314,"n_total_lines":1208,"dependency_fraction":0.2599337748344371},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"PRPF4B","stoichiometry":10.0},{"gene":"SSRP1","stoichiometry":4.0},{"gene":"TOP1","stoichiometry":4.0},{"gene":"CPSF6","stoichiometry":0.2},{"gene":"DDX21","stoichiometry":0.2},{"gene":"RBM33","stoichiometry":0.2},{"gene":"RBM39","stoichiometry":0.2},{"gene":"RBM8A","stoichiometry":0.2},{"gene":"SF3A1","stoichiometry":0.2},{"gene":"SNRPA","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/CHTOP","total_profiled":1310},"omim":[{"mim_id":"614206","title":"CHROMATIN TARGET OF PRMT1; CHTOP","url":"https://www.omim.org/entry/614206"},{"mim_id":"191164","title":"EPHRIN A1; EFNA1","url":"https://www.omim.org/entry/191164"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nuclear speckles","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/CHTOP"},"hgnc":{"alias_symbol":["DKFZP547E1010","SRAG","FOP"],"prev_symbol":["C1orf77"]},"alphafold":{"accession":"Q9Y3Y2","domains":[{"cath_id":"-","chopping":"19-62","consensus_level":"medium","plddt":79.8677,"start":19,"end":62}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y3Y2","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y3Y2-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9Y3Y2-F1-predicted_aligned_error_v6.png","plddt_mean":61.22},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CHTOP","jax_strain_url":"https://www.jax.org/strain/search?query=CHTOP"},"sequence":{"accession":"Q9Y3Y2","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9Y3Y2.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9Y3Y2/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9Y3Y2"}},"corpus_meta":[{"pmid":"25284789","id":"PMC_25284789","title":"5-Hydroxymethylcytosine plays a critical role in glioblastomagenesis by recruiting the CHTOP-methylosome complex.","date":"2014","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/25284789","citation_count":109,"is_preprint":false},{"pmid":"23299939","id":"PMC_23299939","title":"Chtop is a component of the dynamic TREX mRNA export complex.","date":"2013","source":"The EMBO journal","url":"https://pubmed.ncbi.nlm.nih.gov/23299939","citation_count":98,"is_preprint":false},{"pmid":"22872859","id":"PMC_22872859","title":"Five friends of methylated chromatin target of protein-arginine-methyltransferase[prmt]-1 (chtop), a complex linking arginine methylation to desumoylation.","date":"2012","source":"Molecular & cellular proteomics : MCP","url":"https://pubmed.ncbi.nlm.nih.gov/22872859","citation_count":49,"is_preprint":false},{"pmid":"34984976","id":"PMC_34984976","title":"Type I and II PRMTs inversely regulate post-transcriptional intron detention through Sm and CHTOP methylation.","date":"2022","source":"eLife","url":"https://pubmed.ncbi.nlm.nih.gov/34984976","citation_count":33,"is_preprint":false},{"pmid":"34012271","id":"PMC_34012271","title":"CircHIPK2 Contributes to DDP Resistance and Malignant Behaviors of DDP-Resistant Ovarian Cancer Cells Both in vitro and in vivo Through circHIPK2/miR-338-3p/CHTOP ceRNA Pathway.","date":"2021","source":"OncoTargets and therapy","url":"https://pubmed.ncbi.nlm.nih.gov/34012271","citation_count":22,"is_preprint":false},{"pmid":"32736653","id":"PMC_32736653","title":"Circulating unmethylated CHTOP and INS DNA fragments provide evidence of possible islet cell death in youth with obesity and diabetes.","date":"2020","source":"Clinical epigenetics","url":"https://pubmed.ncbi.nlm.nih.gov/32736653","citation_count":21,"is_preprint":false},{"pmid":"19254951","id":"PMC_19254951","title":"Identification of the small protein rich in arginine and glycine (SRAG): a newly identified nucleolar protein that can regulate cell proliferation.","date":"2009","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/19254951","citation_count":16,"is_preprint":false},{"pmid":"29683372","id":"PMC_29683372","title":"Modulating the expression of Chtop, a versatile regulator of gene-specific transcription and mRNA export.","date":"2018","source":"RNA biology","url":"https://pubmed.ncbi.nlm.nih.gov/29683372","citation_count":14,"is_preprint":false},{"pmid":"27495318","id":"PMC_27495318","title":"The small RNA SraG participates in PNPase homeostasis.","date":"2016","source":"RNA (New York, N.Y.)","url":"https://pubmed.ncbi.nlm.nih.gov/27495318","citation_count":13,"is_preprint":false},{"pmid":"22428705","id":"PMC_22428705","title":"Small non-coding RNA SraG regulates the operon YPK_1206-1205 in Yersinia pseudotuberculosis.","date":"2012","source":"FEMS microbiology letters","url":"https://pubmed.ncbi.nlm.nih.gov/22428705","citation_count":13,"is_preprint":false},{"pmid":"32722765","id":"PMC_32722765","title":"Srag Regulates Autophagy via Integrating into a Preexisting Autophagy Pathway in Testis.","date":"2021","source":"Molecular biology and evolution","url":"https://pubmed.ncbi.nlm.nih.gov/32722765","citation_count":10,"is_preprint":false},{"pmid":"31380263","id":"PMC_31380263","title":"CHTOP in Chemoresistant Epithelial Ovarian Cancer: A Novel and Potential Therapeutic Target.","date":"2019","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/31380263","citation_count":10,"is_preprint":false},{"pmid":"27189710","id":"PMC_27189710","title":"Interactions between KSHV ORF57 and the novel human TREX proteins, CHTOP and CIP29.","date":"2016","source":"The Journal of general virology","url":"https://pubmed.ncbi.nlm.nih.gov/27189710","citation_count":8,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":8093,"output_tokens":2245,"usd":0.028977,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9209,"output_tokens":3346,"usd":0.064848,"stage2_stop_reason":"end_turn"},"total_usd":0.093825,"stage1_batch_id":"msgbatch_01ACAxJQabDsht39JXeWG22d","stage2_batch_id":"msgbatch_01QEU22A9U5sbPTgsMzTmvSf","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"Chtop is a novel component of the TREX mRNA export complex. Chtop activates the ATPase and RNA helicase activities of Uap56 (similar to Alyref). Uap56 recruits both Alyref and Chtop onto mRNA. Chtop binds to the NTF2-like domain of Nxf1 in a manner requiring arginine methylation of Chtop, and this interaction is mutually exclusive with Thoc5 binding. Chtop binds Uap56 in a mutually exclusive manner with Alyref. Co-knockdown of Alyref and Chtop results in a potent mRNA export block. Together, these interactions indicate TREX and Nxf1 undergo dynamic remodelling driven by Uap56 ATPase activity and Chtop post-translational modifications.\",\n      \"method\": \"Co-immunoprecipitation, RNAi knockdown, ATPase/helicase activity assays, mRNA export assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, functional biochemical assays, RNAi with defined cellular phenotype, multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"23299939\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Chtop interacts with a nuclear complex called Five Friends of Methylated Chtop (5FMC), which can only be recruited when Chtop is arginine-methylated by Prmt1. 5FMC consists of Pelp1 (core scaffold), Senp3 (SUMO-specific protease), Wdr18, Tex10, and Las1L. Recruitment of 5FMC to the zinc-finger transcription factor Zbp-89 affects its sumoylation status and transactivation potential, providing a mechanistic link between arginine methylation and desumoylation in transcriptional control.\",\n      \"method\": \"Biotinylation-proteomics (BioID-like approach), mass spectrometry, co-immunoprecipitation, sumoylation/transactivation assays\",\n      \"journal\": \"Molecular & cellular proteomics : MCP\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS-based interactome with functional follow-up (sumoylation and transactivation assays), multiple orthogonal methods, single lab\",\n      \"pmids\": [\"22872859\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CHTOP binds to 5-hydroxymethylcytosine (5hmC) on chromatin and is associated with the arginine methyltransferase methylosome complex. This CHTOP-methylosome complex promotes PRMT1-mediated methylation of arginine 3 of histone H4 (H4R3) at genes involved in glioblastomagenesis (EGFR, AKT3, CDK6, CCND2, BRAF), thereby activating their transcription. CHTOP and PRMT1 are both required for expression of these genes, and CHTOP is required for glioblastoma cell tumorigenicity.\",\n      \"method\": \"Chromatin immunoprecipitation, co-immunoprecipitation, RNAi knockdown, in vitro methylation assay, gene expression analysis\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP, Co-IP, RNAi with defined transcriptional and tumorigenic phenotype, multiple orthogonal methods in a single lab\",\n      \"pmids\": [\"25284789\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"SRAG (CHTOP) localizes to the nucleus and nucleolus; nucleolar localization is regulated by the N-terminal domain. CHTOP can interact with RNA. Full-length CHTOP protein levels are highest in resting cells and reduced in proliferating cells, with reduction mapping to the G2/M phase. Overexpression of CHTOP reduces the percentage of cells in G2/M and increases cell death.\",\n      \"method\": \"Subcellular fractionation, immunofluorescence microscopy, N-terminal truncation mutants, cell cycle analysis (flow cytometry), overexpression studies\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Weak — direct localization experiments with truncation mutants, overexpression phenotype, single lab, no in vitro reconstitution\",\n      \"pmids\": [\"19254951\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Arginine methylation of CHTOP by Type I PRMTs regulates post-transcriptional intron detention. Mutagenesis of all methylarginine sites in CHTOP recapitulated the retained-intron splicing changes seen with Type I PRMT inhibition. CHTOP occupancy on chromatin-associated polyadenylated RNA is altered upon Type I PRMT inhibition. Retained introns are enriched in nucleoplasm and chromatin fractions, and CHTOP arginine methylation is required for their post-transcriptional processing.\",\n      \"method\": \"PRMT inhibition (pharmacological), targeted mutagenesis of methylarginine sites, SKaTER-seq (co-transcriptional splicing kinetics), actinomycin D transcription block, proteomic analysis of chromatin fractions, subcellular fractionation\",\n      \"journal\": \"eLife\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — targeted mutagenesis recapitulating pharmacological phenotype combined with multiple sequencing and proteomic methods, single lab but multiple orthogonal approaches\",\n      \"pmids\": [\"34984976\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Chtop binds competitively to both PRMT1 and PRMT5, thereby promoting either asymmetric or symmetric methylation of arginine residues depending on which enzyme is bound. Cellular Chtop levels are autoregulated by a mechanism involving intron retention and nonsense-mediated mRNA decay (NMD).\",\n      \"method\": \"Competitive binding assays (Co-IP), intron retention and NMD analysis (RNA analysis with NMD inhibitors)\",\n      \"journal\": \"RNA biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Weak — review/primary study combination; competitive binding reported but abstract compresses methods; NMD-based autoregulation described as a recent finding but single lab, limited methodological detail in abstract\",\n      \"pmids\": [\"29683372\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"KSHV ORF57 protein interacts with CHTOP (and CIP29) as components of the human TREX complex. Depletion of CHTOP affects ORF57-mediated viral mRNA processing, indicating CHTOP is recruited into an ORF57-mediated viral ribonucleoprotein particle (vRNP) and is functionally required for viral mRNA export.\",\n      \"method\": \"Co-immunoprecipitation, siRNA depletion, viral mRNA processing assays\",\n      \"journal\": \"The Journal of general virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP and depletion study, single lab, limited mechanistic detail in abstract\",\n      \"pmids\": [\"27189710\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CHTOP is a vertebrate-specific, arginine-rich nuclear/nucleolar protein that functions as a dynamic component of the TREX mRNA export complex (activating Uap56 ATPase/helicase activity and bridging mRNA to the export receptor Nxf1 in an arginine-methylation-dependent manner), while also acting as an epigenetic co-activator by binding 5-hydroxymethylcytosine on chromatin and recruiting the PRMT1-containing methylosome to drive H4R3 methylation and gene activation; its arginine methylation by PRMT1 is additionally required for assembly of the 5FMC desumoylation complex (containing Pelp1, Senp3, and others) that modulates transcription factor activity, and for post-transcriptional regulation of nuclear retained introns, with cellular CHTOP levels themselves autoregulated through intron retention and nonsense-mediated mRNA decay.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CHTOP is a vertebrate arginine-rich nuclear and nucleolar protein that couples mRNA export to arginine-methylation-dependent control of gene expression [#0, #3]. As a dynamic component of the TREX complex, CHTOP activates the ATPase and RNA helicase activities of Uap56 and is loaded with Alyref onto mRNA, then binds the NTF2-like domain of Nxf1 in a manner requiring its own arginine methylation; its Nxf1 and Uap56 contacts are mutually exclusive with Thoc5 and Alyref respectively, so that Uap56 ATPase activity and CHTOP modification drive remodelling of TREX during export, and co-depletion of CHTOP and Alyref blocks bulk mRNA export [#0]. CHTOP binds competitively to PRMT1 and PRMT5, directing either asymmetric or symmetric arginine methylation, and this methylation state governs its functional partnerships [#5]. On chromatin, CHTOP binds 5-hydroxymethylcytosine and recruits the PRMT1-containing methylosome to promote H4R3 methylation and transcriptional activation of growth-promoting genes, a role required for glioblastoma cell tumorigenicity [#2]. PRMT1-dependent methylation of CHTOP is also the trigger for assembly of the 5FMC complex (Pelp1, Senp3, Wdr18, Tex10, Las1L), which alters the sumoylation and transactivation of the transcription factor Zbp-89 [#1], and is required for the post-transcriptional processing of nucleoplasm- and chromatin-retained detained introns [#4]. CHTOP abundance is autoregulated through intron retention coupled to nonsense-mediated mRNA decay [#5], and the protein is appropriated by KSHV ORF57 to support viral mRNA export [#6].\",\n  \"teleology\": [\n    {\n      \"year\": 2009,\n      \"claim\": \"Established the basic cell biology of CHTOP: where it resides, that it contacts RNA, and that its level tracks proliferative state, framing it as a regulated nuclear/nucleolar RNA-associated factor.\",\n      \"evidence\": \"Subcellular fractionation, immunofluorescence, N-terminal truncation mutants, and cell-cycle flow cytometry in cultured cells\",\n      \"pmids\": [\"19254951\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No molecular partners or biochemical activity identified\", \"Mechanism linking CHTOP level to G2/M and cell death not defined\", \"RNA-binding specificity unresolved\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed that arginine methylation of CHTOP is the switch that nucleates a defined nuclear complex, connecting a post-translational mark to transcriptional control via desumoylation.\",\n      \"evidence\": \"Biotinylation-proteomics, mass spectrometry, Co-IP, and sumoylation/transactivation assays defining the 5FMC complex and its effect on Zbp-89\",\n      \"pmids\": [\"22872859\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Generality of 5FMC targets beyond Zbp-89 unknown\", \"Structural basis of methyl-CHTOP recognition by Pelp1 not resolved\", \"Direct desumoylation of Zbp-89 by Senp3 within the complex not reconstituted\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identified CHTOP as a core, dynamic TREX subunit and defined the biochemical logic of export-complex remodelling, explaining how mRNA is handed to the Nxf1 export receptor.\",\n      \"evidence\": \"Reciprocal Co-IP, Uap56 ATPase/helicase assays, RNAi co-knockdown with mRNA export readout in human cells\",\n      \"pmids\": [\"23299939\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Identity of the methyltransferase acting on CHTOP within TREX not pinned in this study\", \"Stoichiometry and order of Alyref/CHTOP exchange on individual transcripts unknown\", \"Structure of the CHTOP-Nxf1 interface not solved\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Connected CHTOP to chromatin directly by showing it reads 5hmC and writes an activating histone mark through the methylosome, establishing a co-activator role with oncogenic consequences.\",\n      \"evidence\": \"ChIP, Co-IP, in vitro methylation assays, and RNAi with transcriptional and tumorigenicity readouts in glioblastoma cells\",\n      \"pmids\": [\"25284789\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Domain mediating 5hmC recognition not mapped\", \"Whether export and chromatin functions are physically separable not addressed\", \"Breadth of CHTOP-methylosome target genes beyond the named oncogenes unknown\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Revealed that CHTOP partitions between two antagonistic PRMTs and that its own abundance is feedback-controlled, positioning it as a tunable hub for methylation-dependent outputs.\",\n      \"evidence\": \"Competitive Co-IP binding assays and intron-retention/NMD analysis with NMD inhibitors\",\n      \"pmids\": [\"29683372\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Cellular signals that bias PRMT1 versus PRMT5 binding not identified\", \"Quantitative contribution of NMD autoregulation to steady-state CHTOP unknown\", \"Methods compressed in abstract; competitive binding not independently confirmed\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Demonstrated that CHTOP arginine methylation directly governs post-transcriptional processing of detained introns, linking its modification state to splicing outcomes downstream of transcription.\",\n      \"evidence\": \"Type I PRMT inhibition, methylarginine-site mutagenesis, SKaTER-seq, actinomycin D chase, and chromatin-fraction proteomics in human cells\",\n      \"pmids\": [\"34984976\"],\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Mechanism by which methyl-CHTOP promotes intron removal not defined\", \"Splicing factors recruited by CHTOP for detained introns unidentified\", \"Overlap with TREX export function on the same transcripts unresolved\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Showed a viral pathogen co-opts CHTOP, establishing that its TREX-based export activity is required for processing and export of viral mRNAs.\",\n      \"evidence\": \"Co-IP and siRNA depletion with viral mRNA processing assays for KSHV ORF57\",\n      \"pmids\": [\"27189710\"],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"Direct versus TREX-mediated CHTOP-ORF57 contact not distinguished\", \"Single Co-IP/depletion study\", \"Selectivity for viral over host transcripts not quantified\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CHTOP's separate functions — TREX-mediated export, 5hmC/histone co-activation, 5FMC desumoylation, and intron-detention processing — are coordinated, and what determines its choice of PRMT and downstream complex on a given transcript or locus, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"\",\n      \"gaps\": [\"No structural model of CHTOP or its methyl-dependent interfaces\", \"No genome-wide integration of its chromatin versus RNA-export roles\", \"Signals controlling PRMT1/PRMT5 partitioning unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [0, 3]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [2, 1]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005730\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005654\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [2, 4]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [2, 1]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [2]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [2, 6]}\n    ],\n    \"complexes\": [\"TREX complex\", \"5FMC complex\", \"methylosome\"],\n    \"partners\": [\"UAP56\", \"NXF1\", \"ALYREF\", \"PRMT1\", \"PRMT5\", \"PELP1\", \"SENP3\", \"ZNF148\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":5,"faith_total":6,"faith_pct":83.33333333333333}}