{"gene":"RCCD1","run_date":"2026-06-10T06:43:36","timeline":{"discoveries":[{"year":2018,"finding":"JMJD5 catalyses stereoselective C-3 hydroxylation of arginine residues in sequences from human RCCD1 (and ribosomal protein RPS6), identifying RCCD1 as a substrate of the arginyl hydroxylase JMJD5. High-resolution crystallographic analyses of JMJD5 support its assignment as an arginine hydroxylase rather than a histone demethylase.","method":"In vitro peptide-based screening assay with JMJD5, high-resolution X-ray crystallography of JMJD5 active site with substrate peptides","journal":"Nature Communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro enzymatic reconstitution with peptide substrates plus high-resolution crystal structures; multiple orthogonal methods in a single rigorous study","pmids":["29563586"],"is_preprint":false},{"year":2022,"finding":"Crystallographic analyses of human JMJD5 complexed with 2-oxoglutarate analogues further support JMJD5 as a protein hydroxylase (arginine C-3 hydroxylase) acting on RCCD1-derived sequences, revealing an unusually compact 2OG-binding pocket.","method":"X-ray crystallography of JMJD5 with 2OG analogues; functional context from prior RCCD1 peptide hydroxylation data","journal":"Scientific Reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — structural study replicating and extending the JMJD5:RCCD1 hydroxylation finding with multiple ligand-bound crystal structures","pmids":["36450832"],"is_preprint":false},{"year":2017,"finding":"RCCD1 depletion (siRNA) in NSCLC cells increases acetylated α-tubulin levels and stabilizes cytoskeletal microtubules, inhibiting cell migration and attenuating TGF-β-induced EMT (altered Occludin, Vimentin, Snail expression). RCCD1 modulates microtubule stability through interaction with JMJD5.","method":"siRNA knockdown, Western blot for acetylated α-tubulin and EMT markers, cell migration assay, co-immunoprecipitation (RCCD1–JMJD5 interaction)","journal":"Cancer Letters","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — clean KD with defined cellular phenotype plus Co-IP for JMJD5 interaction; single lab, multiple orthogonal readouts","pmids":["28455245"],"is_preprint":false},{"year":2023,"finding":"RCCD1 localizes to the mitochondrial matrix (in addition to the nucleus), where it interacts with the mitochondrial contact site/cristae organizing system (MICOS) and mitochondrial DNA (mtDNA) to regulate mtDNA transcription, oxidative phosphorylation, and reactive oxygen species production. Under hypoxia, RCCD1 is upregulated, decreases ROS generation, and alleviates apoptosis. RCCD1 promotes breast cancer cell proliferation in vitro and tumor growth in vivo.","method":"Subcellular fractionation/mitochondrial isolation, co-immunoprecipitation with MICOS components, mtDNA transcription assays, ROS measurement, apoptosis assays, in vitro proliferation and in vivo xenograft experiments","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by fractionation, Co-IP with MICOS, functional mtDNA/ROS assays, and in vivo tumor growth; single lab with multiple orthogonal methods","pmids":["37903896"],"is_preprint":false},{"year":2024,"finding":"E2F1 binds to the upstream promoter of RCCD1 to transcriptionally regulate RCCD1 expression, which in turn affects EMT-related marker expression and promotes proliferation, migration, and invasion of colorectal cancer cells.","method":"DNA pull-down assay, dual luciferase reporter assay (promoter activity), Western blot / qPCR for EMT markers, EdU proliferation assay, wound-healing and Transwell invasion assays","journal":"Pathology, Research and Practice","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — direct promoter binding confirmed by two methods (pull-down + luciferase), functional rescue/knockdown; single lab","pmids":["39024731"],"is_preprint":false},{"year":2025,"finding":"JMJD5 and RCCD1 form a protein hydroxylase complex; cancer-associated missense mutations in JMJD5 disrupt the JMJD5:RCCD1 interaction and impair JMJD5 hydroxylase activity. The JMJD5:RCCD1 complex is required for suppressing replication stress and genome instability in tumour cells, and the complex interacts with RAD51 paralogues to enable normal replication fork restart.","method":"Structural analysis of cancer missense mutations, in vitro hydroxylase activity assays, co-immunoprecipitation of JMJD5:RCCD1 and RAD51 paralogues, replication fork restart assays (DNA fiber assay), genome instability readouts","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — structural + enzymatic + Co-IP + functional replication assays; preprint, not yet peer-reviewed, single study","pmids":["bio_10.1101_2025.11.22.689938"],"is_preprint":true},{"year":2026,"finding":"In cancer-associated fibroblasts (CAFs), RCCD1 activates AMPK/mTOR/ULK1 signaling to enhance autophagy, driving WNT5A secretion. Secreted WNT5A activates the Wnt/CaMKII/ERK pathway in colon tumor cells, promoting EMT, proliferation, and invasion. Effects were reversed by autophagy inhibition or WNT5A neutralization.","method":"Coculture assays (CAFs/HCT116 cells), single-cell RNA-seq, Western blot for AMPK/mTOR/ULK1 pathway and EMT markers, autophagy inhibition, WNT5A neutralization, clinical sample validation","journal":"Journal of the Royal Society of New Zealand","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — functional coculture with multiple pathway readouts and rescue experiments; single study with several orthogonal approaches","pmids":["41798776"],"is_preprint":false}],"current_model":"RCCD1 is a substrate of the arginyl hydroxylase JMJD5 (which hydroxylates an arginine residue in RCCD1), and the two proteins form an essential complex that suppresses replication stress and enables replication fork restart, likely through interaction with RAD51 paralogues; independently, RCCD1 localizes to the mitochondrial matrix where it associates with MICOS and mtDNA to regulate oxidative phosphorylation and ROS under hypoxia, and in the cytoplasm it modulates microtubule stability (via JMJD5 interaction) to control TGF-β-induced EMT and cell migration, while its transcription is driven by E2F1 and its expression promotes cancer cell proliferation, metastasis, and CAF-mediated tumor progression through autophagy-dependent WNT5A secretion."},"narrative":{"mechanistic_narrative":"RCCD1 is a multifunctional protein that operates as a substrate and stable partner of the 2-oxoglutarate-dependent oxygenase JMJD5, contributing to genome maintenance, mitochondrial physiology, and cancer cell behavior [PMID:29563586, PMID:bio_10.1101_2025.11.22.689938]. JMJD5 catalyzes stereoselective C-3 hydroxylation of arginine residues within RCCD1-derived sequences, an assignment supported by in vitro reconstitution and high-resolution crystal structures of the enzyme active site [PMID:29563586, PMID:36450832]. The two proteins form a hydroxylase complex required to suppress replication stress and genome instability; this complex engages RAD51 paralogues to enable normal replication fork restart, and cancer-associated JMJD5 missense mutations disrupt the JMJD5:RCCD1 interaction while impairing hydroxylase activity [PMID:bio_10.1101_2025.11.22.689938]. Beyond the nucleus, RCCD1 localizes to the mitochondrial matrix, where it associates with the MICOS complex and mtDNA to regulate mtDNA transcription, oxidative phosphorylation, and reactive oxygen species, and is upregulated under hypoxia to limit ROS and apoptosis [PMID:37903896]. In the cytoplasm, RCCD1 destabilizes microtubules through its interaction with JMJD5, reducing acetylated alpha-tubulin and promoting TGF-beta-induced EMT and cell migration [PMID:28455245]. RCCD1 transcription is driven by E2F1 binding to its promoter, and its expression promotes proliferation, migration, and invasion across breast, colorectal, and colon cancer models, including a cancer-associated-fibroblast program in which RCCD1 activates AMPK/mTOR/ULK1 autophagy to drive WNT5A secretion [PMID:37903896, PMID:39024731, PMID:41798776].","teleology":[{"year":2017,"claim":"Established the first cellular function for RCCD1, showing it controls microtubule dynamics and TGF-beta-driven EMT through a physical link to JMJD5.","evidence":"siRNA knockdown in NSCLC cells with Western blot for acetylated alpha-tubulin and EMT markers, migration assays, and RCCD1-JMJD5 co-immunoprecipitation","pmids":["28455245"],"confidence":"Medium","gaps":["Does not establish whether the JMJD5-RCCD1 interaction acts via hydroxylation in this context","Direct molecular link between RCCD1 and tubulin acetylation enzymes not defined","Single lab, single cell-type context"]},{"year":2018,"claim":"Defined RCCD1 biochemically as a substrate of JMJD5, reassigning JMJD5 as an arginine C-3 hydroxylase rather than a histone demethylase.","evidence":"In vitro peptide hydroxylation assays with JMJD5 and high-resolution crystallography of the JMJD5 active site with substrate peptides","pmids":["29563586"],"confidence":"High","gaps":["Hydroxylation demonstrated on peptides, not full-length RCCD1 in cells","Functional consequence of RCCD1 arginine hydroxylation not determined","Identity of the modified arginine residue's downstream role unknown"]},{"year":2022,"claim":"Reinforced the protein-hydroxylase model by detailing the JMJD5 cofactor-binding pocket acting on RCCD1-derived sequences.","evidence":"X-ray crystallography of JMJD5 with 2-oxoglutarate analogues","pmids":["36450832"],"confidence":"High","gaps":["Structural work centers on JMJD5; no structure of the RCCD1 complex","Does not address cellular regulation of the hydroxylation reaction"]},{"year":2023,"claim":"Revealed an unexpected mitochondrial role for RCCD1, placing it in the matrix in association with MICOS and mtDNA to regulate respiration and ROS under hypoxia.","evidence":"Subcellular fractionation, co-IP with MICOS components, mtDNA transcription and ROS/apoptosis assays, and breast cancer proliferation/xenograft models","pmids":["37903896"],"confidence":"Medium","gaps":["Mechanism by which RCCD1 enters and acts in the matrix is undefined","Whether JMJD5 hydroxylation governs mitochondrial RCCD1 function unknown","Single lab"]},{"year":2024,"claim":"Identified the upstream transcriptional control of RCCD1, showing E2F1 directly drives its expression to promote EMT and tumor cell aggressiveness.","evidence":"DNA pull-down and dual-luciferase promoter assays, plus proliferation, wound-healing, and invasion assays in colorectal cancer cells","pmids":["39024731"],"confidence":"Medium","gaps":["Does not connect E2F1-driven expression to any specific RCCD1 molecular activity","Single lab"]},{"year":2025,"claim":"Demonstrated that the JMJD5:RCCD1 complex safeguards genome stability by enabling replication fork restart via RAD51 paralogues, and that cancer mutations in JMJD5 break this complex.","evidence":"Structural analysis of cancer missense mutations, in vitro hydroxylase assays, co-IP of JMJD5:RCCD1 and RAD51 paralogues, and DNA fiber replication fork restart assays (preprint)","pmids":["bio_10.1101_2025.11.22.689938"],"confidence":"Medium","gaps":["Preprint, not yet peer-reviewed","Whether RAD51 paralogue interaction is direct or hydroxylation-dependent unresolved","Mechanistic role of RCCD1 hydroxylation in fork restart not isolated"]},{"year":2026,"claim":"Extended RCCD1's tumor-promoting role to the stroma, showing it drives autophagy-dependent WNT5A secretion from cancer-associated fibroblasts to activate Wnt signaling in tumor cells.","evidence":"CAF/tumor coculture, single-cell RNA-seq, AMPK/mTOR/ULK1 pathway Western blots, autophagy inhibition and WNT5A neutralization rescue, and clinical sample validation","pmids":["41798776"],"confidence":"Medium","gaps":["How RCCD1 engages AMPK/mTOR/ULK1 mechanistically is undefined","Relationship to RCCD1's nuclear and mitochondrial functions unclear","Single study"]},{"year":null,"claim":"It remains unknown how RCCD1's distinct nuclear (replication/genome stability), mitochondrial (OXPHOS/ROS), and cytoplasmic (microtubule/EMT) activities are coordinated, and whether JMJD5-mediated arginine hydroxylation regulates each of these roles.","evidence":"No integrating study in the available corpus","pmids":[],"confidence":"Low","gaps":["No demonstrated functional consequence of RCCD1 arginine hydroxylation in cells","No unified model linking the three subcellular localizations","No structural model of the full-length JMJD5:RCCD1 complex"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[2]}],"localization":[{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[3]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[3]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[2]}],"pathway":[{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[5]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[5]}],"complexes":["JMJD5:RCCD1 hydroxylase complex","MICOS"],"partners":["JMJD5","RAD51 PARALOGUES","MICOS"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"A6NED2","full_name":"RCC1 domain-containing protein 1","aliases":[],"length_aa":376,"mass_kda":40.1,"function":"Plays a role in transcriptional repression of satellite repeats, possibly by regulating H3K36 methylation levels in centromeric regions together with KDM8 (PubMed:24981860). Possibly together with KDM8, is involved in proper mitotic spindle organization and chromosome segregation (PubMed:24981860). Plays a role in regulating alpha-tubulin deacetylation and cytoskeletal microtubule stability, thereby promoting cell migration and TGF-beta-induced epithelial to mesenchymal transition (EMT), potentially through the inhibition of KDM8 (PubMed:28455245)","subcellular_location":"Chromosome","url":"https://www.uniprot.org/uniprotkb/A6NED2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/RCCD1","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":[],"url":"https://opencell.sf.czbiohub.org/search/RCCD1","total_profiled":1310},"omim":[{"mim_id":"617997","title":"RCC1 DOMAIN-CONTAINING PROTEIN 1; RCCD1","url":"https://www.omim.org/entry/617997"},{"mim_id":"611917","title":"LYSINE DEMETHYLASE 8; KDM8","url":"https://www.omim.org/entry/611917"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Plasma membrane","reliability":"Enhanced"},{"location":"Cytosol","reliability":"Enhanced"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in many","driving_tissues":[],"url":"https://www.proteinatlas.org/search/RCCD1"},"hgnc":{"alias_symbol":["MGC14386"],"prev_symbol":[]},"alphafold":{"accession":"A6NED2","domains":[{"cath_id":"-","chopping":"37-118_131-153","consensus_level":"medium","plddt":89.602,"start":37,"end":153},{"cath_id":"2.130.10.30","chopping":"158-257_283-376","consensus_level":"medium","plddt":94.0846,"start":158,"end":376}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/A6NED2","model_url":"https://alphafold.ebi.ac.uk/files/AF-A6NED2-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-A6NED2-F1-predicted_aligned_error_v6.png","plddt_mean":87.12},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=RCCD1","jax_strain_url":"https://www.jax.org/strain/search?query=RCCD1"},"sequence":{"accession":"A6NED2","fasta_url":"https://rest.uniprot.org/uniprotkb/A6NED2.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/A6NED2/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/A6NED2"}},"corpus_meta":[{"pmid":"27432226","id":"PMC_27432226","title":"Genome-Wide 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by stabilizing cytoskeletal microtubules in NSCLC cells.","date":"2017","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/28455245","citation_count":42,"is_preprint":false},{"pmid":"32907841","id":"PMC_32907841","title":"A Transcriptome-Wide Association Study Identifies Candidate Susceptibility Genes for Pancreatic Cancer Risk.","date":"2020","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/32907841","citation_count":36,"is_preprint":false},{"pmid":"18094031","id":"PMC_18094031","title":"Role of AQP2 in activation of calcium entry by hypotonicity: implications in cell volume regulation.","date":"2007","source":"American journal of physiology. 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Role in the long term regulation of ion transport.","date":"2002","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/12000747","citation_count":30,"is_preprint":false},{"pmid":"11155210","id":"PMC_11155210","title":"Vasopressin regulates water flow in a rat cortical collecting duct cell line not containing known aquaporins.","date":"2001","source":"The Journal of membrane biology","url":"https://pubmed.ncbi.nlm.nih.gov/11155210","citation_count":24,"is_preprint":false},{"pmid":"31582214","id":"PMC_31582214","title":"LINC01419 promotes cell proliferation and metastasis in lung adenocarcinoma via sponging miR-519b-3p to up-regulate RCCD1.","date":"2019","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/31582214","citation_count":20,"is_preprint":false},{"pmid":"22786728","id":"PMC_22786728","title":"Aquaporin 2-increased renal cell proliferation is associated with cell volume regulation.","date":"2012","source":"Journal of cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/22786728","citation_count":20,"is_preprint":false},{"pmid":"37130759","id":"PMC_37130759","title":"Polymorphic variants involved in methylation regulation: a strategy to discover risk loci for pancreatic ductal adenocarcinoma.","date":"2023","source":"Journal of medical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/37130759","citation_count":19,"is_preprint":false},{"pmid":"38590415","id":"PMC_38590415","title":"Multi-omics Mendelian randomization integrating GWAS, eQTL, and mQTL data identified genes associated with breast cancer.","date":"2024","source":"American journal of cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/38590415","citation_count":17,"is_preprint":false},{"pmid":"12474076","id":"PMC_12474076","title":"Differential role of Na+/H+ exchange isoforms NHE-1 and NHE-2 in a rat cortical collecting duct cell line.","date":"2002","source":"The Journal of membrane biology","url":"https://pubmed.ncbi.nlm.nih.gov/12474076","citation_count":17,"is_preprint":false},{"pmid":"11354997","id":"PMC_11354997","title":"Coordinate control of Na,K-atpase mRNA expression by aldosterone, vasopressin and cell sodium delivery in the cortical collecting duct.","date":"2001","source":"Cellular and molecular biology (Noisy-le-Grand, France)","url":"https://pubmed.ncbi.nlm.nih.gov/11354997","citation_count":16,"is_preprint":false},{"pmid":"10556530","id":"PMC_10556530","title":"Vasopressin stimulates long-term net chloride secretion in cortical collecting duct cells.","date":"1999","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/10556530","citation_count":12,"is_preprint":false},{"pmid":"29243846","id":"PMC_29243846","title":"AQP2 can modulate the pattern of Ca2+ transients induced by store-operated Ca2+ entry under TRPV4 activation.","date":"2018","source":"Journal of cellular 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biologicas","url":"https://pubmed.ncbi.nlm.nih.gov/15666000","citation_count":6,"is_preprint":false},{"pmid":"37903896","id":"PMC_37903896","title":"RCCD1 promotes breast carcinogenesis through regulating hypoxia-associated mitochondrial homeostasis.","date":"2023","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/37903896","citation_count":5,"is_preprint":false},{"pmid":"36450832","id":"PMC_36450832","title":"Structural analysis of the 2-oxoglutarate binding site of the circadian rhythm linked oxygenase JMJD5.","date":"2022","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/36450832","citation_count":5,"is_preprint":false},{"pmid":"16301827","id":"PMC_16301827","title":"Functional and molecular adaptation of Cl/HCO3- exchanger to chronic alkaline media in renal cells.","date":"2005","source":"Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and 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neurodevelopmental disorders","date":"2025-11-25","source":"bioRxiv","url":"https://doi.org/10.1101/2025.11.22.689938","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":21284,"output_tokens":2096,"usd":0.047646,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":9056,"output_tokens":3085,"usd":0.061202,"stage2_stop_reason":"end_turn"},"total_usd":0.108848,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2018,\n      \"finding\": \"JMJD5 catalyses stereoselective C-3 hydroxylation of arginine residues in sequences from human RCCD1 (and ribosomal protein RPS6), identifying RCCD1 as a substrate of the arginyl hydroxylase JMJD5. High-resolution crystallographic analyses of JMJD5 support its assignment as an arginine hydroxylase rather than a histone demethylase.\",\n      \"method\": \"In vitro peptide-based screening assay with JMJD5, high-resolution X-ray crystallography of JMJD5 active site with substrate peptides\",\n      \"journal\": \"Nature Communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro enzymatic reconstitution with peptide substrates plus high-resolution crystal structures; multiple orthogonal methods in a single rigorous study\",\n      \"pmids\": [\"29563586\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"Crystallographic analyses of human JMJD5 complexed with 2-oxoglutarate analogues further support JMJD5 as a protein hydroxylase (arginine C-3 hydroxylase) acting on RCCD1-derived sequences, revealing an unusually compact 2OG-binding pocket.\",\n      \"method\": \"X-ray crystallography of JMJD5 with 2OG analogues; functional context from prior RCCD1 peptide hydroxylation data\",\n      \"journal\": \"Scientific Reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — structural study replicating and extending the JMJD5:RCCD1 hydroxylation finding with multiple ligand-bound crystal structures\",\n      \"pmids\": [\"36450832\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"RCCD1 depletion (siRNA) in NSCLC cells increases acetylated α-tubulin levels and stabilizes cytoskeletal microtubules, inhibiting cell migration and attenuating TGF-β-induced EMT (altered Occludin, Vimentin, Snail expression). RCCD1 modulates microtubule stability through interaction with JMJD5.\",\n      \"method\": \"siRNA knockdown, Western blot for acetylated α-tubulin and EMT markers, cell migration assay, co-immunoprecipitation (RCCD1–JMJD5 interaction)\",\n      \"journal\": \"Cancer Letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — clean KD with defined cellular phenotype plus Co-IP for JMJD5 interaction; single lab, multiple orthogonal readouts\",\n      \"pmids\": [\"28455245\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RCCD1 localizes to the mitochondrial matrix (in addition to the nucleus), where it interacts with the mitochondrial contact site/cristae organizing system (MICOS) and mitochondrial DNA (mtDNA) to regulate mtDNA transcription, oxidative phosphorylation, and reactive oxygen species production. Under hypoxia, RCCD1 is upregulated, decreases ROS generation, and alleviates apoptosis. RCCD1 promotes breast cancer cell proliferation in vitro and tumor growth in vivo.\",\n      \"method\": \"Subcellular fractionation/mitochondrial isolation, co-immunoprecipitation with MICOS components, mtDNA transcription assays, ROS measurement, apoptosis assays, in vitro proliferation and in vivo xenograft experiments\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by fractionation, Co-IP with MICOS, functional mtDNA/ROS assays, and in vivo tumor growth; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"37903896\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"E2F1 binds to the upstream promoter of RCCD1 to transcriptionally regulate RCCD1 expression, which in turn affects EMT-related marker expression and promotes proliferation, migration, and invasion of colorectal cancer cells.\",\n      \"method\": \"DNA pull-down assay, dual luciferase reporter assay (promoter activity), Western blot / qPCR for EMT markers, EdU proliferation assay, wound-healing and Transwell invasion assays\",\n      \"journal\": \"Pathology, Research and Practice\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — direct promoter binding confirmed by two methods (pull-down + luciferase), functional rescue/knockdown; single lab\",\n      \"pmids\": [\"39024731\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"JMJD5 and RCCD1 form a protein hydroxylase complex; cancer-associated missense mutations in JMJD5 disrupt the JMJD5:RCCD1 interaction and impair JMJD5 hydroxylase activity. The JMJD5:RCCD1 complex is required for suppressing replication stress and genome instability in tumour cells, and the complex interacts with RAD51 paralogues to enable normal replication fork restart.\",\n      \"method\": \"Structural analysis of cancer missense mutations, in vitro hydroxylase activity assays, co-immunoprecipitation of JMJD5:RCCD1 and RAD51 paralogues, replication fork restart assays (DNA fiber assay), genome instability readouts\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — structural + enzymatic + Co-IP + functional replication assays; preprint, not yet peer-reviewed, single study\",\n      \"pmids\": [\"bio_10.1101_2025.11.22.689938\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In cancer-associated fibroblasts (CAFs), RCCD1 activates AMPK/mTOR/ULK1 signaling to enhance autophagy, driving WNT5A secretion. Secreted WNT5A activates the Wnt/CaMKII/ERK pathway in colon tumor cells, promoting EMT, proliferation, and invasion. Effects were reversed by autophagy inhibition or WNT5A neutralization.\",\n      \"method\": \"Coculture assays (CAFs/HCT116 cells), single-cell RNA-seq, Western blot for AMPK/mTOR/ULK1 pathway and EMT markers, autophagy inhibition, WNT5A neutralization, clinical sample validation\",\n      \"journal\": \"Journal of the Royal Society of New Zealand\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — functional coculture with multiple pathway readouts and rescue experiments; single study with several orthogonal approaches\",\n      \"pmids\": [\"41798776\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"RCCD1 is a substrate of the arginyl hydroxylase JMJD5 (which hydroxylates an arginine residue in RCCD1), and the two proteins form an essential complex that suppresses replication stress and enables replication fork restart, likely through interaction with RAD51 paralogues; independently, RCCD1 localizes to the mitochondrial matrix where it associates with MICOS and mtDNA to regulate oxidative phosphorylation and ROS under hypoxia, and in the cytoplasm it modulates microtubule stability (via JMJD5 interaction) to control TGF-β-induced EMT and cell migration, while its transcription is driven by E2F1 and its expression promotes cancer cell proliferation, metastasis, and CAF-mediated tumor progression through autophagy-dependent WNT5A secretion.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RCCD1 is a multifunctional protein that operates as a substrate and stable partner of the 2-oxoglutarate-dependent oxygenase JMJD5, contributing to genome maintenance, mitochondrial physiology, and cancer cell behavior [#0, #5]. JMJD5 catalyzes stereoselective C-3 hydroxylation of arginine residues within RCCD1-derived sequences, an assignment supported by in vitro reconstitution and high-resolution crystal structures of the enzyme active site [#0, #1]. The two proteins form a hydroxylase complex required to suppress replication stress and genome instability; this complex engages RAD51 paralogues to enable normal replication fork restart, and cancer-associated JMJD5 missense mutations disrupt the JMJD5:RCCD1 interaction while impairing hydroxylase activity [#5]. Beyond the nucleus, RCCD1 localizes to the mitochondrial matrix, where it associates with the MICOS complex and mtDNA to regulate mtDNA transcription, oxidative phosphorylation, and reactive oxygen species, and is upregulated under hypoxia to limit ROS and apoptosis [#3]. In the cytoplasm, RCCD1 destabilizes microtubules through its interaction with JMJD5, reducing acetylated alpha-tubulin and promoting TGF-beta-induced EMT and cell migration [#2]. RCCD1 transcription is driven by E2F1 binding to its promoter, and its expression promotes proliferation, migration, and invasion across breast, colorectal, and colon cancer models, including a cancer-associated-fibroblast program in which RCCD1 activates AMPK/mTOR/ULK1 autophagy to drive WNT5A secretion [#3, #4, #6].\",\n  \"teleology\": [\n    {\n      \"year\": 2017,\n      \"claim\": \"Established the first cellular function for RCCD1, showing it controls microtubule dynamics and TGF-beta-driven EMT through a physical link to JMJD5.\",\n      \"evidence\": \"siRNA knockdown in NSCLC cells with Western blot for acetylated alpha-tubulin and EMT markers, migration assays, and RCCD1-JMJD5 co-immunoprecipitation\",\n      \"pmids\": [\"28455245\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Does not establish whether the JMJD5-RCCD1 interaction acts via hydroxylation in this context\",\n        \"Direct molecular link between RCCD1 and tubulin acetylation enzymes not defined\",\n        \"Single lab, single cell-type context\"\n      ]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Defined RCCD1 biochemically as a substrate of JMJD5, reassigning JMJD5 as an arginine C-3 hydroxylase rather than a histone demethylase.\",\n      \"evidence\": \"In vitro peptide hydroxylation assays with JMJD5 and high-resolution crystallography of the JMJD5 active site with substrate peptides\",\n      \"pmids\": [\"29563586\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Hydroxylation demonstrated on peptides, not full-length RCCD1 in cells\",\n        \"Functional consequence of RCCD1 arginine hydroxylation not determined\",\n        \"Identity of the modified arginine residue's downstream role unknown\"\n      ]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Reinforced the protein-hydroxylase model by detailing the JMJD5 cofactor-binding pocket acting on RCCD1-derived sequences.\",\n      \"evidence\": \"X-ray crystallography of JMJD5 with 2-oxoglutarate analogues\",\n      \"pmids\": [\"36450832\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"Structural work centers on JMJD5; no structure of the RCCD1 complex\",\n        \"Does not address cellular regulation of the hydroxylation reaction\"\n      ]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Revealed an unexpected mitochondrial role for RCCD1, placing it in the matrix in association with MICOS and mtDNA to regulate respiration and ROS under hypoxia.\",\n      \"evidence\": \"Subcellular fractionation, co-IP with MICOS components, mtDNA transcription and ROS/apoptosis assays, and breast cancer proliferation/xenograft models\",\n      \"pmids\": [\"37903896\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Mechanism by which RCCD1 enters and acts in the matrix is undefined\",\n        \"Whether JMJD5 hydroxylation governs mitochondrial RCCD1 function unknown\",\n        \"Single lab\"\n      ]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified the upstream transcriptional control of RCCD1, showing E2F1 directly drives its expression to promote EMT and tumor cell aggressiveness.\",\n      \"evidence\": \"DNA pull-down and dual-luciferase promoter assays, plus proliferation, wound-healing, and invasion assays in colorectal cancer cells\",\n      \"pmids\": [\"39024731\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Does not connect E2F1-driven expression to any specific RCCD1 molecular activity\",\n        \"Single lab\"\n      ]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated that the JMJD5:RCCD1 complex safeguards genome stability by enabling replication fork restart via RAD51 paralogues, and that cancer mutations in JMJD5 break this complex.\",\n      \"evidence\": \"Structural analysis of cancer missense mutations, in vitro hydroxylase assays, co-IP of JMJD5:RCCD1 and RAD51 paralogues, and DNA fiber replication fork restart assays (preprint)\",\n      \"pmids\": [\"bio_10.1101_2025.11.22.689938\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Preprint, not yet peer-reviewed\",\n        \"Whether RAD51 paralogue interaction is direct or hydroxylation-dependent unresolved\",\n        \"Mechanistic role of RCCD1 hydroxylation in fork restart not isolated\"\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Extended RCCD1's tumor-promoting role to the stroma, showing it drives autophagy-dependent WNT5A secretion from cancer-associated fibroblasts to activate Wnt signaling in tumor cells.\",\n      \"evidence\": \"CAF/tumor coculture, single-cell RNA-seq, AMPK/mTOR/ULK1 pathway Western blots, autophagy inhibition and WNT5A neutralization rescue, and clinical sample validation\",\n      \"pmids\": [\"41798776\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"How RCCD1 engages AMPK/mTOR/ULK1 mechanistically is undefined\",\n        \"Relationship to RCCD1's nuclear and mitochondrial functions unclear\",\n        \"Single study\"\n      ]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown how RCCD1's distinct nuclear (replication/genome stability), mitochondrial (OXPHOS/ROS), and cytoplasmic (microtubule/EMT) activities are coordinated, and whether JMJD5-mediated arginine hydroxylation regulates each of these roles.\",\n      \"evidence\": \"No integrating study in the available corpus\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\n        \"No demonstrated functional consequence of RCCD1 arginine hydroxylation in cells\",\n        \"No unified model linking the three subcellular localizations\",\n        \"No structural model of the full-length JMJD5:RCCD1 complex\"\n      ]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [5]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"complexes\": [\n      \"JMJD5:RCCD1 hydroxylase complex\",\n      \"MICOS\"\n    ],\n    \"partners\": [\n      \"JMJD5\",\n      \"RAD51 paralogues\",\n      \"MICOS\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"tie","faith_supported":5,"faith_total":5,"faith_pct":100.0}}