{"gene":"RNF5","run_date":"2026-06-10T06:43:37","timeline":{"discoveries":[{"year":2009,"finding":"RNF5 interacts with MITA/STING in a viral-infection-dependent manner and targets MITA at Lys150 for K48-linked ubiquitination and proteasomal degradation at the mitochondria, thereby negatively regulating virus-triggered IRF3 activation and type I IFN production.","method":"Co-immunoprecipitation, ubiquitination assay, knockdown/overexpression with reporter assays, subcellular fractionation/localization","journal":"Immunity","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, site-specific mutagenesis (K150), subcellular localization, KD and OE with defined signaling readouts in same study","pmids":["19285439"],"is_preprint":false},{"year":2010,"finding":"RNF5 interacts with VISA/MAVS at mitochondria in a viral-infection-dependent manner via the C-terminal transmembrane domain of VISA, and targets VISA at K362 and K461 for K48-linked ubiquitination and degradation, downregulating type I IFN induction.","method":"Co-immunoprecipitation, domain mapping, site-specific mutagenesis (K362, K461), ubiquitination assay, knockdown/overexpression","journal":"Journal of immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — site-specific mutagenesis on two residues, domain mapping, reciprocal Co-IP, multiple functional readouts in one study","pmids":["20483786"],"is_preprint":false},{"year":2008,"finding":"RNF5/RMA1 functions as an E3 ubiquitin ligase upstream of gp78 in ERAD of CFTRΔf508; RMA1 initiates ubiquitination of CFTRΔf508 and gp78 then acts as an E4-like factor to extend polyubiquitin chains, with both requiring the gp78 CUE domain for substrate recognition.","method":"siRNA knockdown, in vitro ubiquitination assay, domain deletion/swap analysis, Co-immunoprecipitation","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — in vitro ubiquitination assay, domain analysis, siRNA epistasis establishing RMA1 acts upstream of gp78, multiple orthogonal methods","pmids":["18216283"],"is_preprint":false},{"year":2003,"finding":"RNF5 associates with the amino-terminal domain of paxillin and mediates its K63-linked polyubiquitination (dependent on intact RING and C-terminal domains and Ubc13), altering paxillin localization from focal adhesions to the cytoplasm and inhibiting cell motility.","method":"Yeast two-hybrid, Co-immunoprecipitation, in vivo ubiquitination assay, dominant-negative Ubc13, fluorescence microscopy, motility assays","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, Ubc13 dominant-negative epistasis, localization with functional consequence (motility), domain mutagenesis, multiple orthogonal methods in one study","pmids":["12861019"],"is_preprint":false},{"year":2012,"finding":"RNF5 associates with and ubiquitinates a membrane-associated pool of the cysteine protease ATG4B, controlling its stability and thereby limiting LC3 processing, phagophore/autophagosome formation, and basal autophagy levels; loss of RNF5 increases autophagy and enhances bacterial clearance.","method":"Co-immunoprecipitation, ubiquitination assay, RNF5 mutant (ligase-active but ATG4B-binding deficient), LC3 puncta quantification, RNF5-/- MEFs and mice, C. elegans RNAi, bacterial infection model","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods including binding-deficient mutant, KO cells and mice, C. elegans validation, functional bacterial clearance readout","pmids":["23093945"],"is_preprint":false},{"year":2010,"finding":"RNF5 associates with JAMP (JNK-associated membrane protein) at the ER membrane and mediates Ubc13-dependent non-canonical (non-degradative) ubiquitination of JAMP, which inhibits JAMP's interaction with Rpt5 and p97, thereby reducing proteasome recruitment to the ER and limiting ERAD efficiency of misfolded proteins such as CFTRΔf508.","method":"Co-immunoprecipitation, ubiquitination assay, dominant-negative Ubc13, accumulation assays for misfolded CFTR and TCRα","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, Ubc13 epistasis, multiple misfolded substrates tested, functional consequence on ERAD defined","pmids":["19269966"],"is_preprint":false},{"year":2010,"finding":"DNAJB12/JB12 cooperates with cytosolic Hsc70 and the E3 ligase RMA1/RNF5 to target nascent CFTR and CFTRΔf508 for ERAD; elevated JB12 increases Hsc70 association with ER forms of CFTR and the RMA1 E3 complex, while depletion of JB12 increases CFTR folding efficiency up to threefold.","method":"Overexpression and siRNA knockdown, Co-immunoprecipitation, CFTR folding/trafficking assays","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, KD/OE with defined folding phenotypes, single lab, multiple methods","pmids":["21148293"],"is_preprint":false},{"year":2004,"finding":"In C. elegans, RNF-5 regulates the LIM domain protein UNC-95, which is required for muscle attachment structure assembly; RNF-5 co-localizes with UNC-95 in dense bodies and controls its expression and localization. Loss of RNF-5 RING domain causes structural defects in muscle adhesion sites.","method":"C. elegans genetics, RNAi, GFP fusion localization, RING domain deletion mutant analysis","journal":"The Journal of cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis in C. elegans, localization with functional consequence, domain mutant, single lab","pmids":["15210732"],"is_preprint":false},{"year":2015,"finding":"RNF5 associates with, ubiquitinates, and promotes proteasomal degradation of the glutamine carrier proteins SLC1A5 and SLC38A2 following paclitaxel-induced ER stress in breast cancer cells, thereby decreasing glutamine uptake, TCA cycle components, and mTOR signaling while increasing autophagy and cell death.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, metabolic profiling, in vivo tumor models (Rnf5-/- MMTV-PyMT mice)","journal":"Cancer cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, genetic KO mouse model, SLC1A5/38A2 KD epistasis, metabolic readouts, multiple orthogonal methods","pmids":["25759021"],"is_preprint":false},{"year":2018,"finding":"RNF5 ubiquitinates S100A8 in intestinal epithelial cells leading to its proteasomal degradation; loss of RNF5 results in enhanced S100A8 secretion, mucosal CD4+ T cell induction, and Th1 pro-inflammatory responses, with RNF5 maintaining intestinal homeostasis.","method":"Co-immunoprecipitation, ubiquitination assay, Rnf5-/- mice, DSS colitis model, S100A8-neutralizing antibody rescue, immunofluorescence","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, KO mouse model, antibody rescue experiment, multiple orthogonal methods","pmids":["30232010"],"is_preprint":false},{"year":2019,"finding":"NDV V protein recruits E3 ubiquitin ligase RNF5 to polyubiquitinate and degrade MAVS at Lys362 and Lys461 via the ubiquitin-proteasome pathway, thereby inhibiting IFN-β production during NDV infection.","method":"Co-immunoprecipitation, ubiquitination assay, site-specific mutagenesis (K362, K461), overexpression/knockdown with IFN-β reporter","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — site-specific mutagenesis, Co-IP, functional IFN readout, single lab","pmids":["31270229"],"is_preprint":false},{"year":2022,"finding":"PRV tegument protein UL13 interacts with the CDN domain of STING and recruits RNF5 to promote K27-/K29-linked ubiquitination and degradation of STING, suppressing STING-mediated antiviral signaling and type I IFN production.","method":"Co-immunoprecipitation, ubiquitination assay (linkage-specific), domain mapping, RNF5 knockdown, infection model with UL13-deficient PRV","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, linkage-specific ubiquitination assay, domain mapping, KO rescue, single lab","pmids":["35584187"],"is_preprint":false},{"year":2022,"finding":"RNF5 mediates K15 ubiquitination of the SARS-CoV-2 M (membrane) protein, enhancing M-E complex formation and promoting M trafficking from Golgi to autophagosomes for virion release; the deubiquitinase POH1 negatively regulates this process.","method":"RNAi screen, Co-immunoprecipitation, ubiquitination assay (site-specific K15), confocal microscopy, virion release assay","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — site-specific mutagenesis, Co-IP, localization, functional virion release assay, single lab","pmids":["35100873"],"is_preprint":false},{"year":2023,"finding":"RNF5 interacts with and catalyzes ubiquitination of the SARS-CoV-2 envelope (E) protein at K63, leading to its degradation by the ubiquitin-proteasome system and inhibiting SARS-CoV-2 replication.","method":"Co-immunoprecipitation, ubiquitination assay (site-specific K63 of E), viral replication assays, mouse infection model with pharmacological RNF5 activator","journal":"Signal transduction and targeted therapy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, site-specific mutagenesis of ubiquitin site on E, in vivo mouse model, single lab","pmids":["36737599"],"is_preprint":false},{"year":2021,"finding":"RNF5 induces K29-linked ubiquitin chains on the histone-binding protein RBBP4, promoting its recruitment to and epigenetic regulation of genes involved in AML maintenance, with RNF5 inhibition sensitizing AML cells to HDAC inhibitors.","method":"Co-immunoprecipitation, linkage-specific ubiquitination assay (K29), chromatin assays, AML cell lines, patient-derived xenograft models, Rnf5 KD in MLL-AF9 leukemogenesis mouse model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP, K29-specific ubiquitination, PDX and in vivo mouse models, transcriptional readouts, epistasis with HDAC inhibitors, multiple orthogonal methods","pmids":["34518534"],"is_preprint":false},{"year":2020,"finding":"RNF5 interacts with and ubiquitinates PHGDH, targeting it for degradation; acetylation of PHGDH at K58 (by Tip60, reversed by SIRT2) disrupts RNF5-PHGDH interaction, stabilizing PHGDH and promoting breast cancer cell proliferation.","method":"Co-immunoprecipitation, ubiquitination assay, site-specific mutagenesis (K58), acetyltransferase/deacetylase identification (Tip60/SIRT2), cell proliferation assays","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, site-specific mutagenesis, writer/eraser identification, single lab","pmids":["32783943"],"is_preprint":false},{"year":2018,"finding":"RNF5 is identified as the E3 ubiquitin ligase responsible for PTEN ubiquitination and proteasomal degradation in Smo-null pancreatic fibroblasts; GSK3β phosphorylation marks PTEN for this RNF5-mediated ubiquitination, and RNF5 knockdown rescues PTEN levels.","method":"Unbiased proteomic screen, Co-immunoprecipitation, ubiquitination assay, RNF5 siRNA knockdown, GSK3β inhibitor rescue","journal":"Life science alliance","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomic identification, Co-IP, functional rescue, pharmacological epistasis, single lab","pmids":["30456390"],"is_preprint":false},{"year":2013,"finding":"RNF5 interacts with β2-adrenergic receptor (β2AR) and prostaglandin D2 receptor (DP) at the ER but does not ubiquitinate them directly; instead, RNF5 ubiquitinates JAMP to prevent proteasome recruitment to the ER, thereby stabilizing these GPCRs from JAMP-mediated proteasomal degradation.","method":"Gel-free proteomics, Co-immunoprecipitation, confocal colocalization, siRNA knockdown, overexpression with receptor level quantification","journal":"Molecular endocrinology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomics identification, Co-IP, siRNA epistasis, colocalization, single lab, negative result (no direct receptor ubiquitination) mechanistically informative","pmids":["23798571"],"is_preprint":false},{"year":2008,"finding":"Transgenic overexpression of RNF5 in mouse muscle causes early-onset muscle wasting, degeneration, and altered ER chaperone activity, while RNF5 KO mice show delayed muscle regeneration and delayed ER stress markers after cardiotoxin injury, establishing RNF5 as a regulator of muscle physiology and ER stress in vivo.","method":"Transgenic and KO mouse models, histopathology, ER stress marker analysis, cardiotoxin injury model","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — gain- and loss-of-function mouse models with defined phenotypes, ER chaperone activity assayed, single lab","pmids":["18270596"],"is_preprint":false},{"year":2009,"finding":"RNF5 is anchored to the ER membrane and its E3 ligase activity is required for its function in ERAD; RNF5 is expressed and localized at the ER where it participates in recognition and processing of misfolded proteins including CFTRΔf508.","method":"Subcellular fractionation, localization studies, functional ERAD assay with misfolded CFTR substrates in transgenic/KO mice","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — fractionation-based localization tied to functional ERAD phenotype in vivo, single lab","pmids":["18270596"],"is_preprint":false},{"year":2010,"finding":"In C. elegans, RNF-5 E3 ligase levels increase specifically during molting, where it ubiquitinates the dense body protein UNC-95, leading to decreased UNC-95 concentration at dense bodies; persistent high RNF-5 expression causes failure of ecdysis, establishing RNF-5 as a temporal regulator of muscle attachment dynamics during molting.","method":"C. elegans genetics, heat-shock promoter overexpression, ubiquitination assay for UNC-95, fluorescence microscopy of dense body dynamics","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ubiquitination assay, genetic gain-of-function with defined ecdysis phenotype, temporal expression analysis, single lab","pmids":["20385102"],"is_preprint":false},{"year":2021,"finding":"JMJD6 recruits RNF5 to promote K48-linked ubiquitination and degradation of activated IRF3, negatively regulating type I IFN production in response to cytosolic viral RNA; genetic deletion of JMJD6 enhances IFN-I production in an RNF5-dependent manner.","method":"Unbiased proteomic screen, Co-immunoprecipitation, ubiquitination assay (K48-linked), JMJD6 KO mice via piggyBac transposon, viral infection assays","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomic screen, Co-IP, K48-specific ubiquitination, in vivo KO mouse model, RNF5-dependent epistasis, single lab","pmids":["33684176"],"is_preprint":false},{"year":2022,"finding":"RNF5 promotes K48-linked polyubiquitination and proteasomal degradation of STING, attenuating STING-mediated cardiac inflammation and pathological cardiac hypertrophy; protein-protein interaction between RNF5 and STING was confirmed.","method":"Co-immunoprecipitation, K48-specific ubiquitination assay, gain- and loss-of-function in cardiac hypertrophy mouse model","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, linkage-specific ubiquitination, in vivo gain/loss-of-function, single lab","pmids":["36270989"],"is_preprint":false},{"year":2023,"finding":"RNF5 interacts with EphA2 (Ephrin receptor A2) and induces its ubiquitination and proteasomal degradation, decreasing EphA2 cell surface levels, altering phosphorylation balance at S897/Y772, and reducing ERK phosphorylation while increasing p53 in HER2-negative breast cancer cells.","method":"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, phosphorylation analysis, xenograft tumor models","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination, in vivo xenograft, multiple functional readouts, single lab","pmids":["37816703"],"is_preprint":false},{"year":2023,"finding":"RNF5 interacts with EphA3 and EphA4 and induces their ubiquitination and degradation; RNF5 inhibition increases EphA3/EphA4 levels, reduces ERK and Akt activation, and suppresses KSHV lytic replication in PEL cells.","method":"Co-immunoprecipitation, ubiquitination assay, pharmacological RNF5 inhibition, PEL xenograft tumor model, viral gene expression analysis","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination, in vivo xenograft, multiple signaling readouts, single lab","pmids":["36656913"],"is_preprint":false},{"year":2024,"finding":"RNF5 promotes K63-type ubiquitination of IGF2BP1, enhancing CPT1A mRNA stabilization through m6A modification and increasing fatty acid oxidation in steatotic HCC; PPARγ activates RNF5 expression specifically in HCC cells, placing RNF5 in a PPARγ-RNF5-IGF2BP1-CPT1A axis.","method":"Protein interaction analysis, Co-immunoprecipitation, ubiquitination assay (K63-specific), lipidomics, transcriptomics, in vitro and in vivo HCC models","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, K63-specific ubiquitination, multiple omics readouts, in vivo models, single lab","pmids":["39734009"],"is_preprint":false},{"year":2018,"finding":"Pharmacological inhibition of RNF5 with a small molecule (inh-02), identified by computational docking and virtual screening, modulates known RNF5 targets ATG4B and paxillin and promotes significant F508del-CFTR rescue in CF patient-derived bronchial epithelial cells.","method":"Computational docking/virtual screening, in vitro RNF5 inhibition assay, cell-based CFTR rescue assay in primary CF cells, target engagement via ATG4B/paxillin modulation","journal":"Cell chemical biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — small-molecule inhibitor with target engagement validation, functional CFTR rescue in primary cells, single lab","pmids":["29754957"],"is_preprint":false},{"year":2025,"finding":"RNF5 interacts with ACSL4 via its transmembrane region and mediates ACSL4 ubiquitination and degradation, thereby attenuating ferroptosis in cardiomyocytes and conferring cardioprotection against myocardial ischemia/reperfusion injury.","method":"Co-immunoprecipitation, IP-MS, ubiquitination assay, AAV9-mediated RNF5 overexpression in mice, ferroptosis and ROS assays","journal":"Biochemical pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — IP-MS substrate identification, Co-IP, ubiquitination, in vivo AAV model, functional ferroptosis readout, single lab","pmids":["41203033"],"is_preprint":false},{"year":2025,"finding":"In the absence of RNF5/RNF185 function, AMFR (a Hrd1 ortholog involved in ERAD-M branch) can partially compensate to facilitate degradation of mutant CFTR, revealing a bypass mechanism in the ERAD network; SYVN1 (another Hrd1 ortholog) did not show the same compensatory effect.","method":"Multiple E3 ligase knockouts/knockdowns combined with HiBiT-based ERAD assay, functional complementation analysis","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1–2 / Weak — sensitive HiBiT ERAD assay, multiple KO/KD combinations, single preprint lab, not yet peer-reviewed","pmids":["bio_10.1101_2025.05.07.652780"],"is_preprint":true},{"year":2041,"finding":"RNF5 inhibits HBV replication by promoting degradation of HBV Core protein through a Caspase-3-dependent (non-ubiquitin-proteasome) pathway; this antiviral function does not rely on RNF5's E3 ubiquitin ligase activity.","method":"Co-immunoprecipitation, caspase-3 inhibitor rescue, E3 ligase activity mutant, viral replication assays","journal":"Frontiers in microbiology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, single Co-IP study, unusual claimed mechanism (ligase-independent), limited orthogonal validation","pmids":["40236486"],"is_preprint":false},{"year":1997,"finding":"RNF5 encodes a RING-finger protein containing a zinc-chelating domain expressed ubiquitously in human tissues; it was mapped to chromosome 6p21.31 proximal to the MHC region, and shares homology with a C. elegans protein.","method":"cDNA cloning, FISH mapping, radiation hybrid mapping, Northern blot expression survey","journal":"Cytogenetics and cell genetics","confidence":"Low","confidence_rationale":"Tier 4 / Weak — initial cloning/mapping paper, no functional mechanistic experiment performed","pmids":["9533025"],"is_preprint":false}],"current_model":"RNF5 is an ER/mitochondria-anchored RING-finger E3 ubiquitin ligase that ubiquitinates a broad set of substrates—including MITA/STING (K150, K48-linked, degradative), MAVS/VISA (K362/K461, K48-linked, degradative), paxillin (K63-linked, relocalization), ATG4B (limiting basal autophagy), SLC1A5/38A2 (glutamine transporter degradation), S100A8 (inflammatory control), JAMP (non-degradative, restricting ERAD/proteasome recruitment), RBBP4 (K29-linked, epigenetic regulation in AML), PHGDH, PTEN, ACSL4, and multiple viral proteins—thereby regulating innate antiviral immunity, ERAD, autophagy, cell motility, metabolism, and tissue homeostasis, with many viral pathogens exploiting or countering RNF5 activity to modulate host immunity."},"narrative":{"mechanistic_narrative":"RNF5 is an ER- and mitochondria-anchored RING-finger E3 ubiquitin ligase whose membrane-bound activity governs innate antiviral immunity, ER-associated degradation (ERAD), autophagy, cell motility, and cellular metabolism [PMID:18270596, PMID:19285439, PMID:23093945]. In antiviral signaling, RNF5 acts as a negative regulator: upon viral infection it associates with the adaptors MITA/STING and VISA/MAVS at the mitochondria and catalyzes K48-linked ubiquitination and proteasomal degradation—targeting MITA at Lys150 and VISA at Lys362/Lys461—to dampen IRF3 activation and type I interferon production [PMID:19285439, PMID:20483786]. It also terminates this pathway downstream by degrading activated IRF3, with the recruitment factor JMJD6 directing RNF5 to its target [PMID:33684176]. At the ER membrane, RNF5/RMA1 initiates ubiquitination of misfolded substrates such as CFTRΔF508 upstream of gp78, working with the DNAJB12/Hsc70 chaperone system [PMID:18216283, PMID:21148293], yet it can also restrain ERAD by non-degradative ubiquitination of JAMP, which limits proteasome recruitment to the ER and thereby stabilizes client GPCRs [PMID:19269966, PMID:23798571]. RNF5 sets basal autophagy levels by ubiquitinating a membrane pool of the cysteine protease ATG4B to control LC3 processing [PMID:23093945], and it shapes cell adhesion and motility through K63-linked ubiquitination of paxillin that displaces it from focal adhesions [PMID:12861019]. Through degradation of diverse substrates—the glutamine transporters SLC1A5/SLC38A2, PHGDH, PTEN, ACSL4, and the histone-binding protein RBBP4—RNF5 controls tumor metabolism, ferroptosis, and epigenetic programs in cancer [PMID:25759021, PMID:32783943, PMID:30456390, PMID:41203033, PMID:34518534]. RNF5 maintains tissue homeostasis in vivo, regulating intestinal inflammation via S100A8 degradation [PMID:30232010] and muscle physiology and ER stress responses [PMID:18270596]. Numerous viral pathogens exploit or counter RNF5: viral proteins recruit it to degrade STING or MAVS to evade immunity [PMID:31270229, PMID:35584187], while RNF5 directly ubiquitinates SARS-CoV-2 structural proteins [PMID:35100873, PMID:36737599].","teleology":[{"year":1997,"claim":"Before any function was known, the question was simply what kind of protein RNF5 encodes; cloning established it as a ubiquitously expressed RING-finger protein, foreshadowing an E3 ligase role.","evidence":"cDNA cloning, FISH/radiation hybrid mapping, Northern blot in human tissues","pmids":["9533025"],"confidence":"Low","gaps":["No enzymatic or substrate evidence at this stage","RING domain function not demonstrated"]},{"year":2003,"claim":"The first substrate question—does RNF5 ubiquitinate a defined target with functional consequence—was answered by showing it mediates K63-linked ubiquitination of paxillin, linking RNF5 to focal-adhesion dynamics and cell motility.","evidence":"Yeast two-hybrid, Co-IP, in vivo ubiquitination with dominant-negative Ubc13, motility assays","pmids":["12861019"],"confidence":"High","gaps":["Physiological trigger for paxillin ubiquitination not defined","Did not establish ER/mitochondrial localization role"]},{"year":2004,"claim":"Whether RNF5's ligase activity has a developmental/structural role was addressed in C. elegans, where RNF-5 regulates UNC-95 and muscle attachment assembly, later refined to temporal control during molting.","evidence":"C. elegans genetics, RNAi, GFP localization, RING-deletion and heat-shock overexpression mutants","pmids":["15210732","20385102"],"confidence":"Medium","gaps":["Mammalian orthologous muscle substrate not identified","Direct ubiquitination of UNC-95 in vitro not fully reconstituted"]},{"year":2008,"claim":"The defining ER quality-control role was established by placing RMA1/RNF5 upstream of gp78 in CFTRΔF508 ERAD and by in vivo mouse models linking RNF5 to ER stress and muscle physiology.","evidence":"siRNA epistasis, in vitro ubiquitination, domain swaps, transgenic/KO mice with cardiotoxin injury","pmids":["18216283","18270596"],"confidence":"High","gaps":["Endogenous substrate spectrum during ERAD incomplete","Mechanism coupling RNF5 to retrotranslocation machinery unresolved"]},{"year":2009,"claim":"The question of how RNF5 controls antiviral immunity was answered by demonstrating infection-dependent association with and K48-linked degradation of MITA/STING at Lys150, defining RNF5 as a negative regulator of type I IFN.","evidence":"Reciprocal Co-IP, K150 site-specific mutagenesis, subcellular fractionation, reporter assays","pmids":["19285439"],"confidence":"High","gaps":["Signal that activates RNF5 during infection not defined","Selectivity between STING and other adaptors unclear"]},{"year":2010,"claim":"RNF5's antiviral target range was extended to the mitochondrial adaptor VISA/MAVS (K362/K461) and its ERAD activity was shown to be bidirectional, with non-degradative ubiquitination of JAMP limiting proteasome recruitment to the ER.","evidence":"Domain mapping, K362/K461 mutagenesis, Ubc13 dominant-negative, misfolded-substrate accumulation assays","pmids":["20483786","19269966","21148293"],"confidence":"High","gaps":["How RNF5 switches between degradative and non-degradative chains is unknown","Relative contribution of MAVS vs STING degradation in vivo not quantified"]},{"year":2012,"claim":"The link between RNF5 and autophagy was established by showing it ubiquitinates a membrane pool of ATG4B to limit LC3 processing and basal autophagy, with loss enhancing bacterial clearance.","evidence":"Co-IP, ubiquitination assay, binding-deficient RNF5 mutant, RNF5-/- MEFs/mice, C. elegans RNAi, infection model","pmids":["23093945"],"confidence":"High","gaps":["Ubiquitination linkage type on ATG4B not specified","Regulation of the membrane-restricted ATG4B pool unclear"]},{"year":2015,"claim":"The metabolic role of RNF5 was defined by showing ER-stress-driven degradation of glutamine transporters SLC1A5/SLC38A2, connecting RNF5 to tumor glutamine uptake, mTOR signaling, and therapy response.","evidence":"Co-IP, ubiquitination, transporter knockdown epistasis, metabolic profiling, Rnf5-/- MMTV-PyMT mice","pmids":["25759021"],"confidence":"High","gaps":["Upstream sensor coupling paclitaxel-induced ER stress to RNF5 activation unknown","Whether this extends beyond breast cancer not addressed"]},{"year":2018,"claim":"RNF5's roles in tissue homeostasis and additional cancer pathways were expanded through S100A8 degradation controlling intestinal inflammation and PTEN degradation in pancreatic fibroblasts, plus a small-molecule inhibitor validating ATG4B/paxillin as engageable targets.","evidence":"Rnf5-/- mice, DSS colitis with antibody rescue; proteomic screen and GSK3β-inhibitor rescue; virtual-screen inhibitor with CFTR rescue in primary CF cells","pmids":["30232010","30456390","29754957"],"confidence":"High","gaps":["PTEN finding relies on a single-lab Smo-null context","Inhibitor specificity across the full substrate set not mapped"]},{"year":2021,"claim":"Recruitment-factor logic and epigenetic substrates emerged, with JMJD6 directing RNF5 to degrade activated IRF3 and RNF5 placing K29 chains on RBBP4 to sustain AML-related gene programs.","evidence":"Proteomic screens, Co-IP, K48- and K29-linkage-specific ubiquitination, JMJD6 KO mice, AML PDX and MLL-AF9 models","pmids":["33684176","34518534"],"confidence":"Medium","gaps":["Structural basis for adaptor-directed substrate selection unknown","How K29 chains on RBBP4 alter chromatin recruitment mechanistically unresolved"]},{"year":2022,"claim":"Viral exploitation of RNF5 was demonstrated as pathogens recruit it to degrade immune adaptors (NDV V → MAVS; PRV UL13 → STING), while RNF5 conversely restrains STING-driven cardiac inflammation.","evidence":"Co-IP, linkage- and site-specific ubiquitination, domain mapping, infection and cardiac hypertrophy mouse models","pmids":["31270229","35584187","36270989"],"confidence":"Medium","gaps":["Single-lab findings for individual viral hijack mechanisms","Linkage types reported (K27/K29 on STING) differ from canonical K48 and need integration"]},{"year":2023,"claim":"RNF5 was shown to directly ubiquitinate viral structural proteins and additional receptor substrates, including SARS-CoV-2 M and E proteins and Eph receptors, broadening its impact on viral life cycles and cancer signaling.","evidence":"RNAi screen, site-specific ubiquitination (K15 on M, K63 on E), Co-IP, virion-release assays, EphA2/A3/A4 ubiquitination, xenograft models","pmids":["35100873","36737599","37816703","36656913"],"confidence":"Medium","gaps":["Opposite effects on different SARS-CoV-2 proteins (pro- vs anti-viral) not reconciled","Single-lab evidence for each receptor substrate"]},{"year":2025,"claim":"Recent work extended RNF5 substrates to lipid-metabolism and ferroptosis regulators (IGF2BP1, ACSL4) and probed ERAD-network redundancy, indicating RNF5 operates within a partially compensable degradation system.","evidence":"IP-MS, K63/K48 ubiquitination assays, AAV9 RNF5 overexpression, ferroptosis assays, HiBiT ERAD assay with multiple E3 knockouts (preprint)","pmids":["39734009","41203033","bio_10.1101_2025.05.07.652780"],"confidence":"Medium","gaps":["ERAD compensation finding is an unreviewed preprint","How RNF5 substrate selection is partitioned among redundant ER ligases unknown"]},{"year":null,"claim":"A unifying question remains: what determines RNF5's choice of ubiquitin chain linkage (K48, K63, K29, K27) and substrate across its many contexts, and how is its activity switched on by infection, ER stress, or adaptor recruitment.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model linking RING activity to linkage specificity","Activation/regulation of RNF5 itself largely uncharacterized","Ligase-independent activities (e.g. caspase-3-dependent HBV Core degradation) not mechanistically resolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[0,1,3,4,8,14]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1,2,3,4,8,9,14,15,16,27]},{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,2,19]}],"localization":[{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[5,17,19]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[0,1]},{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[12]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[0,1,21]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,5,6,19]},{"term_id":"R-HSA-9612973","term_label":"Autophagy","supporting_discovery_ids":[4]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[8,15,25,27]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[10,11,12,13]}],"complexes":[],"partners":["STING1","MAVS","PXN","ATG4B","JAMP","SLC1A5","RBBP4","PTEN"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q99942","full_name":"E3 ubiquitin-protein ligase RNF5","aliases":["RING finger protein 5","Ram1 homolog","HsRma1"],"length_aa":180,"mass_kda":19.9,"function":"Membrane-bound E3 ubiquitin-protein ligase that mediates ubiquitination of target proteins (PubMed:11329381, PubMed:12861019, PubMed:16176924, PubMed:19269966, PubMed:19285439). May function together with E2 ubiquitin-conjugating enzymes UBE2D1/UBCH5A and UBE2D2/UBC4 (PubMed:11329381). Mediates ubiquitination of PXN/paxillin,thereby regulating cell motility and localization of PXN/paxillin (PubMed:12861019). Catalyzes ubiquitination of Salmonella type III secreted protein sopA (PubMed:16176924). Mediates the 'Lys-63'-linked polyubiquitination of JKAMP thereby regulating JKAMP function by decreasing its association with components of the proteasome and ERAD; the ubiquitination appears to involve E2 ubiquitin-conjugating enzyme UBE2N (PubMed:19269966). Mediates the 'Lys-48'-linked polyubiquitination of STING1 at 'Lys-150' leading to its proteasomal degradation; the ubiquitination occurs in mitochondria after viral transfection and regulates antiviral responses (PubMed:19285439). Catalyzes ubiquitination and subsequent degradation of ATG4B, thereby inhibiting autophagy (PubMed:23093945)","subcellular_location":"Cell membrane; Mitochondrion membrane; Endoplasmic reticulum membrane","url":"https://www.uniprot.org/uniprotkb/Q99942/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/RNF5","classification":"Not Classified","n_dependent_lines":13,"n_total_lines":1207,"dependency_fraction":0.010770505385252692},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"CCDC47","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/RNF5","total_profiled":1310},"omim":[{"mim_id":"620096","title":"RING FINGER PROTEIN 185; RNF185","url":"https://www.omim.org/entry/620096"},{"mim_id":"616175","title":"UBIQUITIN-CONJUGATING ENZYME E2 J1; UBE2J1","url":"https://www.omim.org/entry/616175"},{"mim_id":"612374","title":"STIMULATOR OF INTERFERON RESPONSE cGAMP INTERACTOR 1; STING1","url":"https://www.omim.org/entry/612374"},{"mim_id":"611176","title":"JNK/MAPK8-ASSOCIATED MEMBRANE PROTEIN; JKAMP","url":"https://www.omim.org/entry/611176"},{"mim_id":"608813","title":"DER1-LIKE DOMAIN FAMILY, MEMBER 1; DERL1","url":"https://www.omim.org/entry/608813"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Endoplasmic reticulum","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/RNF5"},"hgnc":{"alias_symbol":["NG2","G16","RING5","RMA1"],"prev_symbol":[]},"alphafold":{"accession":"Q99942","domains":[{"cath_id":"3.30.40.10","chopping":"24-85","consensus_level":"medium","plddt":85.3373,"start":24,"end":85}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q99942","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q99942-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q99942-F1-predicted_aligned_error_v6.png","plddt_mean":67.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=RNF5","jax_strain_url":"https://www.jax.org/strain/search?query=RNF5"},"sequence":{"accession":"Q99942","fasta_url":"https://rest.uniprot.org/uniprotkb/Q99942.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q99942/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q99942"}},"corpus_meta":[{"pmid":"19285439","id":"PMC_19285439","title":"The ubiquitin ligase 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medicine","url":"https://pubmed.ncbi.nlm.nih.gov/41845461","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.05.07.652780","title":"AMFR provides an ERAD bypass mechanism to maintain proteostasis under canonical E3 ligase deficiency","date":"2025-05-13","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.07.652780","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":25350,"output_tokens":7534,"usd":0.09453,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":16674,"output_tokens":5044,"usd":0.104735,"stage2_stop_reason":"end_turn"},"total_usd":0.199265,"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\": 2009,\n      \"finding\": \"RNF5 interacts with MITA/STING in a viral-infection-dependent manner and targets MITA at Lys150 for K48-linked ubiquitination and proteasomal degradation at the mitochondria, thereby negatively regulating virus-triggered IRF3 activation and type I IFN production.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, knockdown/overexpression with reporter assays, subcellular fractionation/localization\",\n      \"journal\": \"Immunity\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, site-specific mutagenesis (K150), subcellular localization, KD and OE with defined signaling readouts in same study\",\n      \"pmids\": [\"19285439\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"RNF5 interacts with VISA/MAVS at mitochondria in a viral-infection-dependent manner via the C-terminal transmembrane domain of VISA, and targets VISA at K362 and K461 for K48-linked ubiquitination and degradation, downregulating type I IFN induction.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, site-specific mutagenesis (K362, K461), ubiquitination assay, knockdown/overexpression\",\n      \"journal\": \"Journal of immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — site-specific mutagenesis on two residues, domain mapping, reciprocal Co-IP, multiple functional readouts in one study\",\n      \"pmids\": [\"20483786\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"RNF5/RMA1 functions as an E3 ubiquitin ligase upstream of gp78 in ERAD of CFTRΔf508; RMA1 initiates ubiquitination of CFTRΔf508 and gp78 then acts as an E4-like factor to extend polyubiquitin chains, with both requiring the gp78 CUE domain for substrate recognition.\",\n      \"method\": \"siRNA knockdown, in vitro ubiquitination assay, domain deletion/swap analysis, Co-immunoprecipitation\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — in vitro ubiquitination assay, domain analysis, siRNA epistasis establishing RMA1 acts upstream of gp78, multiple orthogonal methods\",\n      \"pmids\": [\"18216283\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"RNF5 associates with the amino-terminal domain of paxillin and mediates its K63-linked polyubiquitination (dependent on intact RING and C-terminal domains and Ubc13), altering paxillin localization from focal adhesions to the cytoplasm and inhibiting cell motility.\",\n      \"method\": \"Yeast two-hybrid, Co-immunoprecipitation, in vivo ubiquitination assay, dominant-negative Ubc13, fluorescence microscopy, motility assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, Ubc13 dominant-negative epistasis, localization with functional consequence (motility), domain mutagenesis, multiple orthogonal methods in one study\",\n      \"pmids\": [\"12861019\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"RNF5 associates with and ubiquitinates a membrane-associated pool of the cysteine protease ATG4B, controlling its stability and thereby limiting LC3 processing, phagophore/autophagosome formation, and basal autophagy levels; loss of RNF5 increases autophagy and enhances bacterial clearance.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, RNF5 mutant (ligase-active but ATG4B-binding deficient), LC3 puncta quantification, RNF5-/- MEFs and mice, C. elegans RNAi, bacterial infection model\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods including binding-deficient mutant, KO cells and mice, C. elegans validation, functional bacterial clearance readout\",\n      \"pmids\": [\"23093945\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"RNF5 associates with JAMP (JNK-associated membrane protein) at the ER membrane and mediates Ubc13-dependent non-canonical (non-degradative) ubiquitination of JAMP, which inhibits JAMP's interaction with Rpt5 and p97, thereby reducing proteasome recruitment to the ER and limiting ERAD efficiency of misfolded proteins such as CFTRΔf508.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, dominant-negative Ubc13, accumulation assays for misfolded CFTR and TCRα\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, Ubc13 epistasis, multiple misfolded substrates tested, functional consequence on ERAD defined\",\n      \"pmids\": [\"19269966\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"DNAJB12/JB12 cooperates with cytosolic Hsc70 and the E3 ligase RMA1/RNF5 to target nascent CFTR and CFTRΔf508 for ERAD; elevated JB12 increases Hsc70 association with ER forms of CFTR and the RMA1 E3 complex, while depletion of JB12 increases CFTR folding efficiency up to threefold.\",\n      \"method\": \"Overexpression and siRNA knockdown, Co-immunoprecipitation, CFTR folding/trafficking assays\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, KD/OE with defined folding phenotypes, single lab, multiple methods\",\n      \"pmids\": [\"21148293\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"In C. elegans, RNF-5 regulates the LIM domain protein UNC-95, which is required for muscle attachment structure assembly; RNF-5 co-localizes with UNC-95 in dense bodies and controls its expression and localization. Loss of RNF-5 RING domain causes structural defects in muscle adhesion sites.\",\n      \"method\": \"C. elegans genetics, RNAi, GFP fusion localization, RING domain deletion mutant analysis\",\n      \"journal\": \"The Journal of cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis in C. elegans, localization with functional consequence, domain mutant, single lab\",\n      \"pmids\": [\"15210732\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"RNF5 associates with, ubiquitinates, and promotes proteasomal degradation of the glutamine carrier proteins SLC1A5 and SLC38A2 following paclitaxel-induced ER stress in breast cancer cells, thereby decreasing glutamine uptake, TCA cycle components, and mTOR signaling while increasing autophagy and cell death.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, metabolic profiling, in vivo tumor models (Rnf5-/- MMTV-PyMT mice)\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, genetic KO mouse model, SLC1A5/38A2 KD epistasis, metabolic readouts, multiple orthogonal methods\",\n      \"pmids\": [\"25759021\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RNF5 ubiquitinates S100A8 in intestinal epithelial cells leading to its proteasomal degradation; loss of RNF5 results in enhanced S100A8 secretion, mucosal CD4+ T cell induction, and Th1 pro-inflammatory responses, with RNF5 maintaining intestinal homeostasis.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, Rnf5-/- mice, DSS colitis model, S100A8-neutralizing antibody rescue, immunofluorescence\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, KO mouse model, antibody rescue experiment, multiple orthogonal methods\",\n      \"pmids\": [\"30232010\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NDV V protein recruits E3 ubiquitin ligase RNF5 to polyubiquitinate and degrade MAVS at Lys362 and Lys461 via the ubiquitin-proteasome pathway, thereby inhibiting IFN-β production during NDV infection.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, site-specific mutagenesis (K362, K461), overexpression/knockdown with IFN-β reporter\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — site-specific mutagenesis, Co-IP, functional IFN readout, single lab\",\n      \"pmids\": [\"31270229\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PRV tegument protein UL13 interacts with the CDN domain of STING and recruits RNF5 to promote K27-/K29-linked ubiquitination and degradation of STING, suppressing STING-mediated antiviral signaling and type I IFN production.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay (linkage-specific), domain mapping, RNF5 knockdown, infection model with UL13-deficient PRV\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, linkage-specific ubiquitination assay, domain mapping, KO rescue, single lab\",\n      \"pmids\": [\"35584187\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"RNF5 mediates K15 ubiquitination of the SARS-CoV-2 M (membrane) protein, enhancing M-E complex formation and promoting M trafficking from Golgi to autophagosomes for virion release; the deubiquitinase POH1 negatively regulates this process.\",\n      \"method\": \"RNAi screen, Co-immunoprecipitation, ubiquitination assay (site-specific K15), confocal microscopy, virion release assay\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — site-specific mutagenesis, Co-IP, localization, functional virion release assay, single lab\",\n      \"pmids\": [\"35100873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RNF5 interacts with and catalyzes ubiquitination of the SARS-CoV-2 envelope (E) protein at K63, leading to its degradation by the ubiquitin-proteasome system and inhibiting SARS-CoV-2 replication.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay (site-specific K63 of E), viral replication assays, mouse infection model with pharmacological RNF5 activator\",\n      \"journal\": \"Signal transduction and targeted therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, site-specific mutagenesis of ubiquitin site on E, in vivo mouse model, single lab\",\n      \"pmids\": [\"36737599\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"RNF5 induces K29-linked ubiquitin chains on the histone-binding protein RBBP4, promoting its recruitment to and epigenetic regulation of genes involved in AML maintenance, with RNF5 inhibition sensitizing AML cells to HDAC inhibitors.\",\n      \"method\": \"Co-immunoprecipitation, linkage-specific ubiquitination assay (K29), chromatin assays, AML cell lines, patient-derived xenograft models, Rnf5 KD in MLL-AF9 leukemogenesis mouse model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP, K29-specific ubiquitination, PDX and in vivo mouse models, transcriptional readouts, epistasis with HDAC inhibitors, multiple orthogonal methods\",\n      \"pmids\": [\"34518534\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"RNF5 interacts with and ubiquitinates PHGDH, targeting it for degradation; acetylation of PHGDH at K58 (by Tip60, reversed by SIRT2) disrupts RNF5-PHGDH interaction, stabilizing PHGDH and promoting breast cancer cell proliferation.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, site-specific mutagenesis (K58), acetyltransferase/deacetylase identification (Tip60/SIRT2), cell proliferation assays\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, site-specific mutagenesis, writer/eraser identification, single lab\",\n      \"pmids\": [\"32783943\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RNF5 is identified as the E3 ubiquitin ligase responsible for PTEN ubiquitination and proteasomal degradation in Smo-null pancreatic fibroblasts; GSK3β phosphorylation marks PTEN for this RNF5-mediated ubiquitination, and RNF5 knockdown rescues PTEN levels.\",\n      \"method\": \"Unbiased proteomic screen, Co-immunoprecipitation, ubiquitination assay, RNF5 siRNA knockdown, GSK3β inhibitor rescue\",\n      \"journal\": \"Life science alliance\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomic identification, Co-IP, functional rescue, pharmacological epistasis, single lab\",\n      \"pmids\": [\"30456390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"RNF5 interacts with β2-adrenergic receptor (β2AR) and prostaglandin D2 receptor (DP) at the ER but does not ubiquitinate them directly; instead, RNF5 ubiquitinates JAMP to prevent proteasome recruitment to the ER, thereby stabilizing these GPCRs from JAMP-mediated proteasomal degradation.\",\n      \"method\": \"Gel-free proteomics, Co-immunoprecipitation, confocal colocalization, siRNA knockdown, overexpression with receptor level quantification\",\n      \"journal\": \"Molecular endocrinology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomics identification, Co-IP, siRNA epistasis, colocalization, single lab, negative result (no direct receptor ubiquitination) mechanistically informative\",\n      \"pmids\": [\"23798571\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"Transgenic overexpression of RNF5 in mouse muscle causes early-onset muscle wasting, degeneration, and altered ER chaperone activity, while RNF5 KO mice show delayed muscle regeneration and delayed ER stress markers after cardiotoxin injury, establishing RNF5 as a regulator of muscle physiology and ER stress in vivo.\",\n      \"method\": \"Transgenic and KO mouse models, histopathology, ER stress marker analysis, cardiotoxin injury model\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — gain- and loss-of-function mouse models with defined phenotypes, ER chaperone activity assayed, single lab\",\n      \"pmids\": [\"18270596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"RNF5 is anchored to the ER membrane and its E3 ligase activity is required for its function in ERAD; RNF5 is expressed and localized at the ER where it participates in recognition and processing of misfolded proteins including CFTRΔf508.\",\n      \"method\": \"Subcellular fractionation, localization studies, functional ERAD assay with misfolded CFTR substrates in transgenic/KO mice\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — fractionation-based localization tied to functional ERAD phenotype in vivo, single lab\",\n      \"pmids\": [\"18270596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"In C. elegans, RNF-5 E3 ligase levels increase specifically during molting, where it ubiquitinates the dense body protein UNC-95, leading to decreased UNC-95 concentration at dense bodies; persistent high RNF-5 expression causes failure of ecdysis, establishing RNF-5 as a temporal regulator of muscle attachment dynamics during molting.\",\n      \"method\": \"C. elegans genetics, heat-shock promoter overexpression, ubiquitination assay for UNC-95, fluorescence microscopy of dense body dynamics\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ubiquitination assay, genetic gain-of-function with defined ecdysis phenotype, temporal expression analysis, single lab\",\n      \"pmids\": [\"20385102\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"JMJD6 recruits RNF5 to promote K48-linked ubiquitination and degradation of activated IRF3, negatively regulating type I IFN production in response to cytosolic viral RNA; genetic deletion of JMJD6 enhances IFN-I production in an RNF5-dependent manner.\",\n      \"method\": \"Unbiased proteomic screen, Co-immunoprecipitation, ubiquitination assay (K48-linked), JMJD6 KO mice via piggyBac transposon, viral infection assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomic screen, Co-IP, K48-specific ubiquitination, in vivo KO mouse model, RNF5-dependent epistasis, single lab\",\n      \"pmids\": [\"33684176\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"RNF5 promotes K48-linked polyubiquitination and proteasomal degradation of STING, attenuating STING-mediated cardiac inflammation and pathological cardiac hypertrophy; protein-protein interaction between RNF5 and STING was confirmed.\",\n      \"method\": \"Co-immunoprecipitation, K48-specific ubiquitination assay, gain- and loss-of-function in cardiac hypertrophy mouse model\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, linkage-specific ubiquitination, in vivo gain/loss-of-function, single lab\",\n      \"pmids\": [\"36270989\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RNF5 interacts with EphA2 (Ephrin receptor A2) and induces its ubiquitination and proteasomal degradation, decreasing EphA2 cell surface levels, altering phosphorylation balance at S897/Y772, and reducing ERK phosphorylation while increasing p53 in HER2-negative breast cancer cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, siRNA knockdown, phosphorylation analysis, xenograft tumor models\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination, in vivo xenograft, multiple functional readouts, single lab\",\n      \"pmids\": [\"37816703\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RNF5 interacts with EphA3 and EphA4 and induces their ubiquitination and degradation; RNF5 inhibition increases EphA3/EphA4 levels, reduces ERK and Akt activation, and suppresses KSHV lytic replication in PEL cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, pharmacological RNF5 inhibition, PEL xenograft tumor model, viral gene expression analysis\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination, in vivo xenograft, multiple signaling readouts, single lab\",\n      \"pmids\": [\"36656913\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF5 promotes K63-type ubiquitination of IGF2BP1, enhancing CPT1A mRNA stabilization through m6A modification and increasing fatty acid oxidation in steatotic HCC; PPARγ activates RNF5 expression specifically in HCC cells, placing RNF5 in a PPARγ-RNF5-IGF2BP1-CPT1A axis.\",\n      \"method\": \"Protein interaction analysis, Co-immunoprecipitation, ubiquitination assay (K63-specific), lipidomics, transcriptomics, in vitro and in vivo HCC models\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, K63-specific ubiquitination, multiple omics readouts, in vivo models, single lab\",\n      \"pmids\": [\"39734009\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Pharmacological inhibition of RNF5 with a small molecule (inh-02), identified by computational docking and virtual screening, modulates known RNF5 targets ATG4B and paxillin and promotes significant F508del-CFTR rescue in CF patient-derived bronchial epithelial cells.\",\n      \"method\": \"Computational docking/virtual screening, in vitro RNF5 inhibition assay, cell-based CFTR rescue assay in primary CF cells, target engagement via ATG4B/paxillin modulation\",\n      \"journal\": \"Cell chemical biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — small-molecule inhibitor with target engagement validation, functional CFTR rescue in primary cells, single lab\",\n      \"pmids\": [\"29754957\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RNF5 interacts with ACSL4 via its transmembrane region and mediates ACSL4 ubiquitination and degradation, thereby attenuating ferroptosis in cardiomyocytes and conferring cardioprotection against myocardial ischemia/reperfusion injury.\",\n      \"method\": \"Co-immunoprecipitation, IP-MS, ubiquitination assay, AAV9-mediated RNF5 overexpression in mice, ferroptosis and ROS assays\",\n      \"journal\": \"Biochemical pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — IP-MS substrate identification, Co-IP, ubiquitination, in vivo AAV model, functional ferroptosis readout, single lab\",\n      \"pmids\": [\"41203033\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In the absence of RNF5/RNF185 function, AMFR (a Hrd1 ortholog involved in ERAD-M branch) can partially compensate to facilitate degradation of mutant CFTR, revealing a bypass mechanism in the ERAD network; SYVN1 (another Hrd1 ortholog) did not show the same compensatory effect.\",\n      \"method\": \"Multiple E3 ligase knockouts/knockdowns combined with HiBiT-based ERAD assay, functional complementation analysis\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Weak — sensitive HiBiT ERAD assay, multiple KO/KD combinations, single preprint lab, not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.05.07.652780\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2041,\n      \"finding\": \"RNF5 inhibits HBV replication by promoting degradation of HBV Core protein through a Caspase-3-dependent (non-ubiquitin-proteasome) pathway; this antiviral function does not rely on RNF5's E3 ubiquitin ligase activity.\",\n      \"method\": \"Co-immunoprecipitation, caspase-3 inhibitor rescue, E3 ligase activity mutant, viral replication assays\",\n      \"journal\": \"Frontiers in microbiology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, single Co-IP study, unusual claimed mechanism (ligase-independent), limited orthogonal validation\",\n      \"pmids\": [\"40236486\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 1997,\n      \"finding\": \"RNF5 encodes a RING-finger protein containing a zinc-chelating domain expressed ubiquitously in human tissues; it was mapped to chromosome 6p21.31 proximal to the MHC region, and shares homology with a C. elegans protein.\",\n      \"method\": \"cDNA cloning, FISH mapping, radiation hybrid mapping, Northern blot expression survey\",\n      \"journal\": \"Cytogenetics and cell genetics\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Weak — initial cloning/mapping paper, no functional mechanistic experiment performed\",\n      \"pmids\": [\"9533025\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"RNF5 is an ER/mitochondria-anchored RING-finger E3 ubiquitin ligase that ubiquitinates a broad set of substrates—including MITA/STING (K150, K48-linked, degradative), MAVS/VISA (K362/K461, K48-linked, degradative), paxillin (K63-linked, relocalization), ATG4B (limiting basal autophagy), SLC1A5/38A2 (glutamine transporter degradation), S100A8 (inflammatory control), JAMP (non-degradative, restricting ERAD/proteasome recruitment), RBBP4 (K29-linked, epigenetic regulation in AML), PHGDH, PTEN, ACSL4, and multiple viral proteins—thereby regulating innate antiviral immunity, ERAD, autophagy, cell motility, metabolism, and tissue homeostasis, with many viral pathogens exploiting or countering RNF5 activity to modulate host immunity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RNF5 is an ER- and mitochondria-anchored RING-finger E3 ubiquitin ligase whose membrane-bound activity governs innate antiviral immunity, ER-associated degradation (ERAD), autophagy, cell motility, and cellular metabolism [#19, #0, #4]. In antiviral signaling, RNF5 acts as a negative regulator: upon viral infection it associates with the adaptors MITA/STING and VISA/MAVS at the mitochondria and catalyzes K48-linked ubiquitination and proteasomal degradation\\u2014targeting MITA at Lys150 and VISA at Lys362/Lys461\\u2014to dampen IRF3 activation and type I interferon production [#0, #1]. It also terminates this pathway downstream by degrading activated IRF3, with the recruitment factor JMJD6 directing RNF5 to its target [#21]. At the ER membrane, RNF5/RMA1 initiates ubiquitination of misfolded substrates such as CFTR\\u0394F508 upstream of gp78, working with the DNAJB12/Hsc70 chaperone system [#2, #6], yet it can also restrain ERAD by non-degradative ubiquitination of JAMP, which limits proteasome recruitment to the ER and thereby stabilizes client GPCRs [#5, #17]. RNF5 sets basal autophagy levels by ubiquitinating a membrane pool of the cysteine protease ATG4B to control LC3 processing [#4], and it shapes cell adhesion and motility through K63-linked ubiquitination of paxillin that displaces it from focal adhesions [#3]. Through degradation of diverse substrates\\u2014the glutamine transporters SLC1A5/SLC38A2, PHGDH, PTEN, ACSL4, and the histone-binding protein RBBP4\\u2014RNF5 controls tumor metabolism, ferroptosis, and epigenetic programs in cancer [#8, #15, #16, #27, #14]. RNF5 maintains tissue homeostasis in vivo, regulating intestinal inflammation via S100A8 degradation [#9] and muscle physiology and ER stress responses [#18]. Numerous viral pathogens exploit or counter RNF5: viral proteins recruit it to degrade STING or MAVS to evade immunity [#10, #11], while RNF5 directly ubiquitinates SARS-CoV-2 structural proteins [#12, #13].\",\n  \"teleology\": [\n    {\n      \"year\": 1997,\n      \"claim\": \"Before any function was known, the question was simply what kind of protein RNF5 encodes; cloning established it as a ubiquitously expressed RING-finger protein, foreshadowing an E3 ligase role.\",\n      \"evidence\": \"cDNA cloning, FISH/radiation hybrid mapping, Northern blot in human tissues\",\n      \"pmids\": [\"9533025\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No enzymatic or substrate evidence at this stage\", \"RING domain function not demonstrated\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"The first substrate question\\u2014does RNF5 ubiquitinate a defined target with functional consequence\\u2014was answered by showing it mediates K63-linked ubiquitination of paxillin, linking RNF5 to focal-adhesion dynamics and cell motility.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP, in vivo ubiquitination with dominant-negative Ubc13, motility assays\",\n      \"pmids\": [\"12861019\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological trigger for paxillin ubiquitination not defined\", \"Did not establish ER/mitochondrial localization role\"]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"Whether RNF5's ligase activity has a developmental/structural role was addressed in C. elegans, where RNF-5 regulates UNC-95 and muscle attachment assembly, later refined to temporal control during molting.\",\n      \"evidence\": \"C. elegans genetics, RNAi, GFP localization, RING-deletion and heat-shock overexpression mutants\",\n      \"pmids\": [\"15210732\", \"20385102\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mammalian orthologous muscle substrate not identified\", \"Direct ubiquitination of UNC-95 in vitro not fully reconstituted\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"The defining ER quality-control role was established by placing RMA1/RNF5 upstream of gp78 in CFTR\\u0394F508 ERAD and by in vivo mouse models linking RNF5 to ER stress and muscle physiology.\",\n      \"evidence\": \"siRNA epistasis, in vitro ubiquitination, domain swaps, transgenic/KO mice with cardiotoxin injury\",\n      \"pmids\": [\"18216283\", \"18270596\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous substrate spectrum during ERAD incomplete\", \"Mechanism coupling RNF5 to retrotranslocation machinery unresolved\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"The question of how RNF5 controls antiviral immunity was answered by demonstrating infection-dependent association with and K48-linked degradation of MITA/STING at Lys150, defining RNF5 as a negative regulator of type I IFN.\",\n      \"evidence\": \"Reciprocal Co-IP, K150 site-specific mutagenesis, subcellular fractionation, reporter assays\",\n      \"pmids\": [\"19285439\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signal that activates RNF5 during infection not defined\", \"Selectivity between STING and other adaptors unclear\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"RNF5's antiviral target range was extended to the mitochondrial adaptor VISA/MAVS (K362/K461) and its ERAD activity was shown to be bidirectional, with non-degradative ubiquitination of JAMP limiting proteasome recruitment to the ER.\",\n      \"evidence\": \"Domain mapping, K362/K461 mutagenesis, Ubc13 dominant-negative, misfolded-substrate accumulation assays\",\n      \"pmids\": [\"20483786\", \"19269966\", \"21148293\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How RNF5 switches between degradative and non-degradative chains is unknown\", \"Relative contribution of MAVS vs STING degradation in vivo not quantified\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"The link between RNF5 and autophagy was established by showing it ubiquitinates a membrane pool of ATG4B to limit LC3 processing and basal autophagy, with loss enhancing bacterial clearance.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, binding-deficient RNF5 mutant, RNF5-/- MEFs/mice, C. elegans RNAi, infection model\",\n      \"pmids\": [\"23093945\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitination linkage type on ATG4B not specified\", \"Regulation of the membrane-restricted ATG4B pool unclear\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"The metabolic role of RNF5 was defined by showing ER-stress-driven degradation of glutamine transporters SLC1A5/SLC38A2, connecting RNF5 to tumor glutamine uptake, mTOR signaling, and therapy response.\",\n      \"evidence\": \"Co-IP, ubiquitination, transporter knockdown epistasis, metabolic profiling, Rnf5-/- MMTV-PyMT mice\",\n      \"pmids\": [\"25759021\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Upstream sensor coupling paclitaxel-induced ER stress to RNF5 activation unknown\", \"Whether this extends beyond breast cancer not addressed\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"RNF5's roles in tissue homeostasis and additional cancer pathways were expanded through S100A8 degradation controlling intestinal inflammation and PTEN degradation in pancreatic fibroblasts, plus a small-molecule inhibitor validating ATG4B/paxillin as engageable targets.\",\n      \"evidence\": \"Rnf5-/- mice, DSS colitis with antibody rescue; proteomic screen and GSK3\\u03b2-inhibitor rescue; virtual-screen inhibitor with CFTR rescue in primary CF cells\",\n      \"pmids\": [\"30232010\", \"30456390\", \"29754957\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"PTEN finding relies on a single-lab Smo-null context\", \"Inhibitor specificity across the full substrate set not mapped\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Recruitment-factor logic and epigenetic substrates emerged, with JMJD6 directing RNF5 to degrade activated IRF3 and RNF5 placing K29 chains on RBBP4 to sustain AML-related gene programs.\",\n      \"evidence\": \"Proteomic screens, Co-IP, K48- and K29-linkage-specific ubiquitination, JMJD6 KO mice, AML PDX and MLL-AF9 models\",\n      \"pmids\": [\"33684176\", \"34518534\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis for adaptor-directed substrate selection unknown\", \"How K29 chains on RBBP4 alter chromatin recruitment mechanistically unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Viral exploitation of RNF5 was demonstrated as pathogens recruit it to degrade immune adaptors (NDV V \\u2192 MAVS; PRV UL13 \\u2192 STING), while RNF5 conversely restrains STING-driven cardiac inflammation.\",\n      \"evidence\": \"Co-IP, linkage- and site-specific ubiquitination, domain mapping, infection and cardiac hypertrophy mouse models\",\n      \"pmids\": [\"31270229\", \"35584187\", \"36270989\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab findings for individual viral hijack mechanisms\", \"Linkage types reported (K27/K29 on STING) differ from canonical K48 and need integration\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"RNF5 was shown to directly ubiquitinate viral structural proteins and additional receptor substrates, including SARS-CoV-2 M and E proteins and Eph receptors, broadening its impact on viral life cycles and cancer signaling.\",\n      \"evidence\": \"RNAi screen, site-specific ubiquitination (K15 on M, K63 on E), Co-IP, virion-release assays, EphA2/A3/A4 ubiquitination, xenograft models\",\n      \"pmids\": [\"35100873\", \"36737599\", \"37816703\", \"36656913\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Opposite effects on different SARS-CoV-2 proteins (pro- vs anti-viral) not reconciled\", \"Single-lab evidence for each receptor substrate\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Recent work extended RNF5 substrates to lipid-metabolism and ferroptosis regulators (IGF2BP1, ACSL4) and probed ERAD-network redundancy, indicating RNF5 operates within a partially compensable degradation system.\",\n      \"evidence\": \"IP-MS, K63/K48 ubiquitination assays, AAV9 RNF5 overexpression, ferroptosis assays, HiBiT ERAD assay with multiple E3 knockouts (preprint)\",\n      \"pmids\": [\"39734009\", \"41203033\", \"bio_10.1101_2025.05.07.652780\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"ERAD compensation finding is an unreviewed preprint\", \"How RNF5 substrate selection is partitioned among redundant ER ligases unknown\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"A unifying question remains: what determines RNF5's choice of ubiquitin chain linkage (K48, K63, K29, K27) and substrate across its many contexts, and how is its activity switched on by infection, ER stress, or adaptor recruitment.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model linking RING activity to linkage specificity\", \"Activation/regulation of RNF5 itself largely uncharacterized\", \"Ligase-independent activities (e.g. caspase-3-dependent HBV Core degradation) not mechanistically resolved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [0, 1, 3, 4, 8, 14]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1, 2, 3, 4, 8, 9, 14, 15, 16, 27]},\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 2, 19]},\n      {\"term_id\": \"GO:0061630\", \"supporting_discovery_ids\": [19]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [5, 17, 19]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [0, 1]},\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [12]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [0, 1, 21]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [2, 5, 6, 19]},\n      {\"term_id\": \"R-HSA-9612973\", \"supporting_discovery_ids\": [4]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [8, 15, 25, 27]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [10, 11, 12, 13]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"STING1\", \"MAVS\", \"PXN\", \"ATG4B\", \"JAMP\", \"SLC1A5\", \"RBBP4\", \"PTEN\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}