{"gene":"RNF20","run_date":"2026-06-10T06:43:37","timeline":{"discoveries":[{"year":2011,"finding":"RNF20 localizes to DNA double-strand breaks (DSBs) independently of H2AX and is required for DSB-induced H2B ubiquitination at K120, H3K4 methylation at DSBs, and recruitment of chromatin-remodeling factor SNF2h; depletion of RNF20 or expression of H2B K120R mutant compromises DNA end resection and recruitment of RAD51 and BRCA1, causing homologous recombination repair defects and radiation sensitivity.","method":"RNAi knockdown, H2B K120R mutant expression, immunofluorescence/foci analysis, HR repair assays, chromatin relaxation rescue experiments","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (KD, dominant-negative mutant, functional HR assay, foci recruitment, chromatin relaxation rescue) in a single focused study","pmids":["21362548"],"is_preprint":false},{"year":2011,"finding":"WAC interacts with RNF20/40 through its C-terminal coiled-coil region, recognizes RNA polymerase II via its N-terminal WW domain, and targets RNF20/40 to the RNA polymerase II complex at active transcription sites to promote H2B ubiquitination; WAC depletion abolishes H2B ubiquitination.","method":"Protein affinity purification, co-immunoprecipitation, RNAi knockdown, H2B ubiquitination assays, cell-cycle checkpoint assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, affinity purification, functional ubiquitination assays, multiple orthogonal methods in one study","pmids":["21329877"],"is_preprint":false},{"year":2012,"finding":"Smurf2 (HECT-domain E3 ligase) targets RNF20 for proteasomal degradation, thereby regulating downstream H2B monoubiquitination and H3K4/H3K79 trimethylation; Smurf2 and RNF20 co-localize at γ-H2AX foci of DSBs.","method":"Genetic ablation (Smurf2 knockout mice), protein degradation assays, co-localization by immunofluorescence, western blot of histone marks","journal":"Nature medicine","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo knockout model, multiple histone mark readouts, co-localization, and degradation assays across mouse and human cells","pmids":["22231558"],"is_preprint":false},{"year":2012,"finding":"RNF20 (BRE1A/Rnf20) deficiency causes replication-associated DSBs through formation of aberrant R-loops (RNA:DNA hybrids), leading to specific genomic rearrangements, breakage-fusion-bridge cycles, and chromosomal instability; this is compounded by a previously known HR repair defect.","method":"Bre1-deficient cell lines, genomic instability tracking, R-loop detection, DNA content analysis, cancer tissue comparison","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — single lab, multiple cellular genomic phenotype assays, R-loop detection, functional epistasis","pmids":["22354749"],"is_preprint":false},{"year":2012,"finding":"RNF20-catalyzed H2B monoubiquitylation (H2Bub1) increases during embryonic stem cell differentiation and is required for efficient ESC differentiation, particularly for transcriptional induction of relatively long genes; USP44 deubiquitinase negatively regulates H2Bub1 and its downregulation during differentiation contributes to the H2Bub1 increase.","method":"ESC differentiation assays, RNAi knockdown of RNF20 and USP44, ChIP, gene expression analysis","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (KD, ChIP, expression profiling), reciprocal regulation demonstrated for writer (RNF20) and eraser (USP44)","pmids":["22681888"],"is_preprint":false},{"year":2011,"finding":"CDC73 (parafibromin) tumor suppressor interacts with RNF20 and RNF40 at discrete residues, and is required for maintenance of H2B K120 monoubiquitination; loss of nuclear CDC73 or CDC73 mutations (as in parathyroid carcinoma) significantly reduces H2Bub1 without affecting H3K4me3.","method":"Yeast two-hybrid, co-immunoprecipitation, siRNA knockdown, western blot of histone marks in parathyroid tumor tissue","journal":"Human molecular genetics","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — yeast two-hybrid, co-IP, knockdown functional assays, validated in primary tumors","pmids":["22021426"],"is_preprint":false},{"year":2013,"finding":"Histone chaperone FACT (SUPT16H subunit) directly binds RNF20 in vivo; mutation of the RNF20 RING-finger domain abolishes FACT-RNF20 interaction and RNF20 accumulation at DSB sites; SUPT16H depletion impairs RNF20-mediated H2B ubiquitylation and SNF2h recruitment at DSBs, causing HR repair defects; this interaction is PAF1-independent for DNA damage contexts.","method":"Co-immunoprecipitation, RING-domain mutagenesis, RNAi knockdown, laser-induced DSB foci analysis, HR repair assays, radiation sensitivity","journal":"Journal of cell science","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — mutagenesis of RING domain, co-IP, functional HR assay, multiple epistasis experiments in one study","pmids":["24357716"],"is_preprint":false},{"year":2013,"finding":"RNF20 (hBRE1) is targeted by adenovirus E1A, which repurposes RNF20 from a ubiquitin ligase into a scaffold that recruits hPaf1 for transcriptional activation of viral early genes, simultaneously blocking H2B monoubiquitination required for the interferon response.","method":"Co-immunoprecipitation, transcription reporter assays, viral infection/replication assays, E1A interaction mapping","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP, functional transcription assays, single lab with multiple orthogonal methods","pmids":["23785282"],"is_preprint":false},{"year":2013,"finding":"RNF20 (histone H2B E3 ubiquitin ligase) is required for MLL-fusion-mediated leukemogenesis; H2B ubiquitination is enriched in the body of MLL-fusion target genes and correlates with H3K79 methylation; RNF20 is required to maintain local H3K79 methylation by DOT1L at Hoxa9 and Meis1, supporting MLL-fusion oncogenic transcriptional program.","method":"shRNA knockdown in leukemia models (in vitro and in vivo), ChIP-seq (H2Bub, H3K79me2), gene expression analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP-seq, in vivo knockdown model, epistatic relationship between H2Bub and H3K79me2 demonstrated","pmids":["23412334"],"is_preprint":false},{"year":2014,"finding":"Arsenite directly binds to the cysteine residues in the RING finger domains of RNF20 and RNF40 in vitro and in cells, inhibiting H2B ubiquitination and impairing BRCA1 and RAD51 recruitment to DSB sites, thereby compromising DNA DSB repair and rendering cells sensitive to radiomimetic agents.","method":"In vitro binding assays, cellular H2B ubiquitination assays, laser-induced DSB foci analysis, DNA repair assays, cell viability assays","journal":"Journal of the American Chemical Society","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — direct in vitro binding, functional ubiquitination assay, DSB recruitment assays, multiple cell lines","pmids":["25170678"],"is_preprint":false},{"year":2016,"finding":"RNF20/RNF40 complex interacts with motor protein Eg5 during mitosis, monoubiquitinates and stabilizes Eg5; loss of RNF20/40 causes spindle assembly defects, cell cycle arrest, and apoptosis; this spindle assembly role is distinct from the transcriptional H2B ubiquitination function.","method":"Co-immunoprecipitation, ubiquitination assays, spindle assembly imaging, cell cycle analysis, in vivo tumor suppression assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — co-IP, ubiquitination assay, functional spindle assembly phenotype, in vivo validation across multiple methods","pmids":["27557628"],"is_preprint":false},{"year":2016,"finding":"RNF20/RNF40 downregulation favors recruitment of p65-containing NF-κB dimers over repressive p50 homodimers and decreases H3K9me3 on a subset of NF-κB target genes, augmenting their transcription; RNF20+/- mice show predisposition to colonic inflammation and inflammation-associated colorectal cancer with excessive MDSCs.","method":"RNAi knockdown, ChIP, NF-κB dimer recruitment analysis, RNF20 heterozygous mouse model, colitis and cancer models","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo mouse model, ChIP-based mechanistic analysis, multiple orthogonal experimental approaches across mouse and human tissues","pmids":["26854224"],"is_preprint":false},{"year":2016,"finding":"Crystal structure of the RNF20 RING domain reveals it forms a homodimer and specifically interacts with the Ube2B~Ub conjugate; key E3-E2 and E3-ubiquitin interface contacts were identified by mutagenesis; RING domains of RNF20 and RNF40 form a stable active heterodimer; structural model of RNF20-Ube2B~Ub complex generated.","method":"X-ray crystallography of RNF20 RING domain, site-directed mutagenesis of E3-E2 and E3-ubiquitin interfaces, in vitro ubiquitination assays","journal":"Journal of molecular biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with functional mutagenesis validation and in vitro ubiquitination assays","pmids":["27569044"],"is_preprint":false},{"year":2019,"finding":"RNF20/40, in conjunction with its cognate E2 enzyme RAD6, monoubiquitylates lysine 381 of eEF1BδL (a heat shock transcription factor); this monoubiquitylation increases eEF1BδL accumulation and potentiates recruitment of p-TEFb to promoters of heat shock-responsive genes, enhancing their transcription; cooperative physical interactions among eEF1BδL, RNF20/40, and HSF1 synergistically promote heat shock gene expression.","method":"Biochemically defined in vitro ubiquitylation assays with purified factors, cell-based analyses, mass spectrometry identification of ubiquitylation site, ChIP, gene expression assays","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro reconstitution with purified factors, site-specific mutagenesis to identify K381, ChIP, multiple orthogonal methods","pmids":["30649429"],"is_preprint":false},{"year":2019,"finding":"The RNF20/40 complex associates with p53 on chromatin; p53 mediates recruitment of RNF20/40 to p21 and PUMA gene loci; RNF20/40-dependent H2B ubiquitination (ubH2B) is required for transcription and mRNA splicing of p21 and PUMA; ubH2B is recognized by the ubiquitin-binding motif of pre-mRNA splicing factor PRPF8 (a spliceosome subunit), which is required for mRNA maturation.","method":"Protein affinity purification, co-immunoprecipitation, ChIP, RNAi knockdown, mRNA splicing assays, ubiquitin-binding domain interaction studies","journal":"Journal of molecular cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — affinity purification, ChIP, functional splicing assays, identification of PRPF8 as ubH2B reader, multiple orthogonal methods","pmids":["31152661"],"is_preprint":false},{"year":2019,"finding":"RNF20 promotes proteasomal degradation of nuclear corepressor 1 (NCoR1) via ubiquitination, which relieves NCoR1-mediated repression and stimulates PPARγ transcriptional activity in adipocytes, thereby promoting adipogenesis; Rnf20 conditional knockout mice show reduced fat mass and altered PPARγ target gene expression.","method":"Conditional knockout mouse model, quantitative proteomics, co-immunoprecipitation, ubiquitination/proteasomal degradation assays","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo knockout, proteomics identification of NCoR1 as substrate, degradation assays, multiple methods","pmids":["31604693"],"is_preprint":false},{"year":2020,"finding":"The SARS-CoV-2 main protease 3Clpro cleaves RNF20 at a conserved Gln521 in a catalytically dependent manner (catalytic mutant 3ClproC145A does not cleave); cleavage of RNF20 prevents RNF20-mediated degradation of SREBP1, promoting viral replication; RNAi depletion of RNF20 or RNF40 significantly enhances SARS-CoV-2 replication.","method":"Protease cleavage assays with catalytic mutant control, RNAi knockdown, SARS-CoV-2 replication assays, SREBP1 protein stability assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — catalytic mutant control establishing mechanism, substrate identification, viral replication functional readout","pmids":["34452991"],"is_preprint":false},{"year":2017,"finding":"RNF20 acts as a tumor suppressor in clear cell renal cell carcinoma by promoting degradation of SREBP1c, which represses lipogenesis and cell cycle progression through the PTTG1 oncogene (a novel SREBP1c target); RNF20 overexpression reduces tumor growth and lipid storage in xenografts.","method":"RNF20 overexpression and knockdown, cell proliferation assays, xenograft tumor models, lipogenesis assays, ChIP for SREBP1c at PTTG1 promoter","journal":"Molecular and cellular biology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — single lab, in vivo xenograft, functional mechanistic pathway from RNF20 to SREBP1c to PTTG1","pmids":["28827316"],"is_preprint":false},{"year":2010,"finding":"RNF20 (hBRE1) and MEN1 are recruited to the IRF1 gene locus in both uninduced and IFNγ-induced states; RNAi-mediated depletion of RNF20 lowers H2Bub and H3K4me3 at IRF1, but unexpectedly upregulates IFNγ-induced IRF1 transcription, indicating that RNF20 can be repressive of inducible transcription at this locus.","method":"ChIP, RNAi knockdown, RT-PCR gene expression analysis, chemical inhibition of H3K4 methylation","journal":"Epigenetics & chromatin","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — ChIP and RNAi knockdown; single lab; unexpected negative regulatory finding at specific locus","pmids":["20825659"],"is_preprint":false},{"year":2013,"finding":"RNF20 promotes polyubiquitination and proteasome-dependent degradation of transcription factor AP-2α; RNF20 co-localizes and interacts with AP-2α in preadipocytes; RNF20 overexpression rescues C/EBPα expression suppressed by AP-2α.","method":"Co-immunoprecipitation, mass spectrometry, ubiquitination assays, proteasome inhibition, co-localization microscopy, C/EBPα expression rescue assays","journal":"Acta biochimica et biophysica Sinica","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP, ubiquitination assay, functional rescue; single lab","pmids":["24374663"],"is_preprint":false},{"year":2017,"finding":"Fbxl19, a CxxC domain-containing protein, promotes H2Bub1 at CpG island-containing gene promoters by directly interacting with Rnf20; Fbxl19 preferentially occupies CpG island-containing promoters genome-wide and its chromosomal binding is required for H2Bub1 at those targets; Fbxl19 is critical for proper ESC differentiation in collaboration with Rnf20.","method":"Co-immunoprecipitation, genome-wide ChIP-seq (Fbxl19, H2Bub1), RNAi knockdown of Fbxl19, ESC differentiation assays","journal":"Nucleic acids research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, genome-wide ChIP-seq, functional differentiation assays; single lab","pmids":["28453857"],"is_preprint":false},{"year":2019,"finding":"RNF20 and RNF40 are required for DSB repair leading to both homologous recombination and class switch recombination (NHEJ-driven) in mouse B cells; DSBs induce a global increase in H2Bub but not H3K4me3 or H3K79me2; H2AX phosphorylation is dispensable for H2Bub; ATM and ATR jointly regulate IR-induced H2Bub.","method":"RNAi knockdown in mouse B cells, class switch recombination assays, HR assays, western blot of histone marks, kinase inhibitor treatments","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — functional DSB repair assays (HR and CSR), epistasis with ATM/ATR, multiple orthogonal methods","pmids":["30692271"],"is_preprint":false},{"year":2023,"finding":"Cryo-EM structures of chemically-trapped transient ubiquitin transfer complexes reveal that RNF40 (and yeast Bre1) directly bind nucleosomal DNA; the E3/E2/nucleosome interaction pattern is conserved from yeast to humans; Bre1 RING domain uses a non-canonical non-hydrophobic contact with Rad6 to position E2 directly above target H2B lysine; these structures explain site-specific H2B monoubiquitylation mechanism.","method":"Chemical trapping, cryo-electron microscopy structure determination, biochemical reconstitution, cancer mutation mapping","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structures with chemical trapping, reconstitution, multiple mechanistic insights from single high-quality study","pmids":["37633270"],"is_preprint":false},{"year":2023,"finding":"RPA (single-strand DNA-binding factor) interacts with RNF20 mainly in S and G2/M phases and recruits RNF20 to mitotic centromeres in a centromeric R-loop-dependent manner and to chromosomal breaks upon DNA damage; the RPA-RNF20 pathway promotes local H2Bub, H3K4 dimethylation, and SNF2H recruitment, ensuring proper Aurora B kinase activation at centromeres and efficient loading of repair proteins at DSBs.","method":"Co-immunoprecipitation, ChIP, cell cycle analysis, mitotic phenotype assays (lagging chromosomes, chromosome bridges), HR repair assays, R-loop disruption experiments","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, ChIP, epistasis with R-loops, functional readouts (mitosis and HR), multiple orthogonal approaches","pmids":["37155876"],"is_preprint":false},{"year":2023,"finding":"Yeast Bre1 and human RNF20 function as recombination mediators independently of their E3 ligase activity: they interact with Rad51, direct Rad51 to ssDNA, facilitate Rad51-ssDNA filament assembly and strand exchange in vitro, and interact with Srs2/FBH1 helicases to counteract their disruption of Rad51 filaments; this mediator function is additive with Rad52 (yeast) or BRCA2 (human).","method":"In vitro Rad51 filament assembly and strand exchange assays, co-immunoprecipitation, ligase-dead mutant analysis, genetic epistasis with rad52/BRCA2, HR assays in cells","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — in vitro reconstitution of filament assembly and strand exchange, ligase-dead mutants, genetic epistasis, ortholog conservation","pmids":["37230987"],"is_preprint":false},{"year":2021,"finding":"RNF20 and RNF40 form a complex that monoubiquitinates H2B on lysine 120; in vivo CRISPR screen in mouse cardiomyocytes identified RNF20/40-mediated H2Bub as controlling dynamic changes in gene expression required for cardiomyocyte maturation.","method":"In vivo somatic Cas9 mutagenesis (forward genetic screen in mice), cardiomyocyte maturation phenotyping, gene expression analysis","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo CRISPR screen, functional validation of maturation phenotype; single lab","pmids":["34290256"],"is_preprint":false},{"year":2018,"finding":"RNF20 (spinal) mediates H2B monoubiquitination in dorsal horn neurons following spinal nerve ligation; RNF20-dependent H2Bub facilitates phosphorylated RNA polymerase II-dependent mGluR5 transcription; focal knockdown of spinal RNF20 reverses neuropathic allodynia and reduces mGluR5 expression; TNF-α activates this RNF20/H2Bub/RNAPII transcriptional axis.","method":"Spinal nerve ligation rat model, RNAi knockdown in vivo, ChIP of H2Bub and phospho-RNAPII at mGluR5 promoter, behavioral allodynia assays, neutralizing antibody experiments","journal":"The Journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — in vivo ChIP, RNAi functional rescue of behavior, TNF-α neutralization; single lab","pmids":["30201771"],"is_preprint":false},{"year":2024,"finding":"RNF20 stabilizes LSD1 via K29-linked ubiquitination in a CDK9-phosphorylation-dependent manner; this CDK9-RNF20-LSD1 axis is required for H3K4 demethylation, repression of endogenous retroviruses, and suppression of interferon response, thereby contributing to epigenetic immunosuppression; loss of RNF20 sensitizes cancer cells to anti-PD-1 immunotherapy in vivo.","method":"Co-immunoprecipitation, ubiquitination assays (K29-specific), CDK9 inhibition/phosphorylation analysis, endogenous retrovirus expression, in vivo anti-PD-1 rescue experiment, breast cancer specimen correlation","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Strong — co-IP, site-specific ubiquitination, CDK9 epistasis, in vivo immunotherapy rescue, multiple orthogonal methods","pmids":["38315842"],"is_preprint":false},{"year":2024,"finding":"RNF20 controls Rbx1 expression, thereby regulating activity of the VHL ubiquitin ligase complex and HIF1α protein levels; Rnf20 haploinsufficiency increases HIF1α levels and drives metabolic rewiring and EMT through HIF1α-mediated RNAPII promoter-proximal pause release independently of H2Bub1; this links the DNA damage response and metabolic reprogramming in lung cancer.","method":"Rnf20 haploinsufficiency mouse lung tumor model, genetic ablation, gene expression analysis, metabolomics, HIF1α protein stability assays, Rbx1/VHL epistasis experiments","journal":"Nature communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo mouse model, epistasis experiments, two distinct RNF20-HIF1α mechanisms; single lab","pmids":["40436847"],"is_preprint":false},{"year":2024,"finding":"HIF-1 recruits FACT and RNF20/40 to hypoxia response elements (HREs); FACT and RNF20/40 stabilize HIF-1 binding and each other at HREs; hypoxia induces H2B monoubiquitination at HIF-1 target genes in an HIF-1-dependent manner; knockdown of FACT or RNF20/40 decreases transcription initiation and elongation at HIF-1 target genes.","method":"RNAi knockdown of FACT and RNF20/40, ChIP at HREs, H2Bub ChIP under hypoxia, transcription initiation/elongation analysis in breast cancer cells","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — ChIP, functional knockdowns, cooperative recruitment mechanism; single lab","pmids":["38517892"],"is_preprint":false},{"year":2024,"finding":"RNF20 mediates NCoR1 protein degradation to activate PPARγ in iWAT for beige fat differentiation upon prolonged cold stimuli; in BAT, cold stimulus downregulates RNF20, which elevates GABPα protein stability (RNF20 controls GABPα stability), promoting thermogenic gene expression; thus RNF20 uses fat depot-specific substrates (GABPα in BAT, NCoR1 in iWAT) to orchestrate adipose thermogenesis.","method":"Fat depot-specific Rnf20 knockout mice, cold exposure experiments, protein stability assays for GABPα and NCoR1, gene expression analysis, ubiquitination assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo fat-specific knockout models, two distinct substrates identified with functional readouts, multiple orthogonal methods","pmids":["38296968"],"is_preprint":false},{"year":2024,"finding":"RNF20 regulates oocyte meiotic spindle assembly by localizing to centromeres and spindle poles and recruiting tropomyosin 3 (TPM3) via its coiled-coil motif (not its E3 ligase activity); RNF20-depleted oocytes show abnormal spindle organization and chromosome misalignment; the RNF20-TPM3 interaction is essential for acentrosomal meiotic spindle assembly.","method":"RNF20 depletion in mouse oocytes, E3 ligase-dead mutant analysis, co-immunoprecipitation of RNF20-TPM3, immunofluorescence of spindle/centromere localization, coiled-coil domain mapping","journal":"Advanced science","confidence":"High","confidence_rationale":"Tier 2 / Strong — ligase-dead mutant establishing non-enzymatic mechanism, co-IP for substrate (TPM3), functional spindle phenotype, localization","pmids":["38240347"],"is_preprint":false},{"year":2024,"finding":"RNF20 mediates RNA polymerase II promoter-proximal pausing at highly paused endothelial genes involved in VEGFA signaling; RNF20 binds Notch1 to promote H2B monoubiquitination at Notch target genes and Notch-dependent gene expression; loss of Rnf20 leads to uncontrolled tip cell specification; RNF20 restricts ERG-dependent Pol II pause release at highly paused genes.","method":"Rnf20 knockout in endothelial cells, RNA Pol II ChIP-seq (pause analysis), Notch1 co-immunoprecipitation, H2Bub ChIP, angiogenesis/tip cell phenotype assays","journal":"Nature cardiovascular research","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP-seq for Pol II pausing, co-IP with Notch1, in vivo phenotype, multiple orthogonal methods","pmids":["39322771"],"is_preprint":false},{"year":2023,"finding":"Iron deficiency-activated ferritinophagy induces RNF20 degradation through the autophagy-lysosomal pathway; RNF20 degradation negatively regulates H2B K120 monoubiquitination at promoters of myogenic markers MyoD and MyoG, inhibiting myogenic differentiation and muscle regeneration; overexpression of RNF20 or autophagy inhibition restores regeneration under iron-deficient conditions.","method":"Iron deficiency model in cells and mice, autophagy-lysosome pathway inhibition, RNF20 overexpression rescue, ChIP for H2Bub at myogenic promoters, conditional NCOA4 knockout in satellite cells","journal":"Science advances","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple rescue experiments, ChIP at target promoters, in vivo conditional knockout, mechanistic pathway established","pmids":["37976359"],"is_preprint":false},{"year":2025,"finding":"RNF20 localizes to replicating sites and promotes H2Bub there; RNF20 knockdown leads to nucleolytic degradation of stalled replication forks (rescued by MRE11/DNA2 inhibition and depletion of fork remodelers SMARCAL1/HLTF/ZRANB3); RNF20 facilitates RAD51 and RAD51C loading at stalled forks, acting in the same pathway as RAD51/RAD51C-mediated fork protection; RNF20 RING domain and ATR-mediated phosphorylation are essential for its role in replication stress responses.","method":"RNF20 knockdown, DNA fiber assays (fork protection/restart), RING-domain mutant and phosphorylation-deficient mutant analysis, nuclease inhibition, fork remodeler co-depletion, RAD51/RAD51C loading assays","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — RING domain and phospho-mutants, fork protection assays, genetic epistasis with fork remodelers and nucleases, RAD51 loading assays","pmids":["40495033"],"is_preprint":false},{"year":2021,"finding":"RNF20 and RNF40 regulate Vitamin D Receptor (VDR) gene and VDR target gene transcription through H3K4me3 occupancy dependent on H2Bub1; conditional intestinal deletion of Rnf20 or Rnf40 in mice causes spontaneous colorectal inflammation; this mechanism is linked to IBD pathogenesis.","method":"Conditional intestinal Rnf20/Rnf40 knockout mice, ChIP-seq for H2Bub1 and H3K4me3 at VDR locus, mRNA-seq, IBD patient ChIP-seq","journal":"Cell death and differentiation","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO in vivo, ChIP-seq, confirmation in human patient specimens, multiple orthogonal methods","pmids":["34088983"],"is_preprint":false},{"year":2023,"finding":"KSHV RTA protein interacts with the cellular RNF20/40 E3 ubiquitin ligase complex, and this interaction is necessary for RTA-induced KSHV lytic cycle reactivation.","method":"Proteomics identification of RTA-RNF20/40 interaction, functional lytic reactivation assays, co-immunoprecipitation validation","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — proteomics plus functional lytic assays; single lab","pmids":["37888983"],"is_preprint":false},{"year":2018,"finding":"RNF20 interacts with ZSCAN4 protein and negatively regulates ZSCAN4 stability; RNF20 depletion does not affect ZSCAN4 transcription but increases ZSCAN4 protein levels and stabilizes its half-life, indicating RNF20 promotes ZSCAN4 degradation at the protein level.","method":"Co-immunoprecipitation, protein half-life assays, RNAi knockdown of RNF20, western blot, RT-PCR","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP, protein stability assays, RNAi; single lab, single study","pmids":["29477841"],"is_preprint":false},{"year":2020,"finding":"In breast cancer cells, Snail interacts with RNF20 and G9a; RNF20 and H3K9me2 (G9a mark) are enriched on the E-cadherin promoter in a Snail-dependent manner; RNF20-mediated H2BK120 monoubiquitination contributes to E-cadherin repression and EMT induction.","method":"Co-immunoprecipitation (Snail-RNF20-G9a), ChIP at E-cadherin promoter, RNF20 knockdown/overexpression, migration/invasion/tumorsphere assays","journal":"Frontiers in oncology","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP, ChIP, functional cancer cell assays; single lab","pmids":["33364200"],"is_preprint":false},{"year":2023,"finding":"RNF20 promotes STAT3 ubiquitination and degradation; RNF20 is a target gene of exosomal miR-let-7b-5p (confirmed by luciferase reporter assay); ectopic RNF20 expression attenuates insulin resistance by downregulating STAT3 protein expression via ubiquitination-mediated degradation.","method":"Luciferase reporter assay (RNF20 as let-7b-5p target), IP/western blot for STAT3 ubiquitination, RNF20 overexpression rescue experiments","journal":"Diabetes, metabolic syndrome and obesity","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single co-IP for ubiquitination, luciferase reporter, single lab, limited mechanistic depth","pmids":["37842335"],"is_preprint":false},{"year":2025,"finding":"Crystal structure of Bre1-Lge1 complex and AlphaFold model of RNF20/RNF40-WAC reveal extensive interaction interfaces; the Bre1-Lge1 and RNF20/RNF40-WAC interfaces share structural homology but use completely different electrostatic interactions encoding binding specificity; these interactions are critical for the H2BUb1 reaction and processes it regulates.","method":"X-ray crystallography of Bre1-Lge1, AlphaFold modeling of RNF20/RNF40-WAC, interface mutagenesis, in vitro H2BUb1 assays, in vivo functional assays","journal":"Nucleic acids research","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with mutagenesis and functional ubiquitination validation; single lab but rigorous structural approach","pmids":["41533567"],"is_preprint":false},{"year":2026,"finding":"Cryo-EM structure of chemically-trapped RNF20/RNF40-RAD6A-Ub-H2BS112GlcNAc nucleosome complex shows that H2BS112GlcNAc interacts with E2 enzyme RAD6A (not RNF20/RNF40); GlcNAcylation allosterically stimulates ubiquitin transfer from RAD6A~Ub to H2B K120 by enhancing the nucleophilicity of H2B K120; the C2 N-acetyl group and β-configuration of C1 are essential for this activation.","method":"Chemical synthesis of H2BS112GlcNAc-modified nucleosomes, chemical trapping, cryo-EM structure determination, mutagenesis, kinetics analysis, structure-activity relationship","journal":"Nature chemical biology","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM with chemical trapping, kinetics, mutagenesis, synthetic nucleosome chemistry, mechanistic detail of PTM crosstalk","pmids":["41495224"],"is_preprint":false},{"year":2023,"finding":"RNF20 is required for male fertility through regulation of H2BK120 ubiquitination in Sertoli cells; Sertoli cell-specific Rnf20 knockout mice are infertile due to spermatogenic failure mimicking Sertoli cell-only syndrome; RNF20 deficiency impairs transcription elongation of Cldn11 (encoding tight junction component claudin-11), disrupts cell adhesion and seminiferous tubule organization, and leads to apoptosis of spermatogonia and spermatocytes.","method":"Conditional Sertoli cell-specific Rnf20 knockout mice (Amh-Cre), histology, H2Bub1 western blot, Cldn11 expression analysis, ChIP for H2Bub at Cldn11 locus","journal":"Cell & bioscience","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional KO in vivo with specific target gene (Cldn11) and ChIP validation, clear phenotypic readout","pmids":["37024990"],"is_preprint":false},{"year":2025,"finding":"In β-cells, Rnf20 forms complexes with the Isl1 transcription factor; conditional Rnf20 knockout in adult β-cells causes severe hyperglycemia, loss of H2B monoubiquitination, dysregulation of glucose-stimulated insulin secretion, and disruption of β-cell identity gene expression; comparative analysis shows Isl1 and Rnf20 loss yield similar β-cell regulome changes; Isl1::Rnf20 complexes regulate insulin expression and secretion in human tissues.","method":"β-cell-specific conditional Rnf20 knockout (tamoxifen-inducible), co-immunoprecipitation of Isl1-Rnf20, H2Bub1 western blot, transcriptomic analysis, glucose tolerance tests, insulin secretion assays","journal":"Diabetes","confidence":"High","confidence_rationale":"Tier 2 / Strong — conditional in vivo KO, co-IP for Isl1-Rnf20 complex, functional insulin secretion assays, validated in human tissues","pmids":["40743566"],"is_preprint":false},{"year":2025,"finding":"RNF20 dynamically translocates between nucleus and cytoplasm in response to RNA virus infection (export via CRM1); in resting and early infection states, nuclear RNF20 maintains basal and inducible transcription of RIG-I and MDA5; upon late viral infection, cytoplasmic RNF20 recognizes degron motifs of RIG-I and MDA5 via its coiled-coil domain and catalyzes K27-linked ubiquitination and degradation of both sensors, preventing excessive antiviral signaling.","method":"RNF20 overexpression/knockout (in vitro and in vivo), co-immunoprecipitation, ubiquitination assays (K27-specific), CRM1 export inhibition, transcriptome sequencing, viral infection assays","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — co-IP, K27-specific ubiquitination, CRM1 inhibition, nucleocytoplasmic localization; single lab","pmids":["41712649"],"is_preprint":false},{"year":2022,"finding":"The natural compound epoxymicheliolide (ECL) covalently modifies H2B lysine 46 (K46) and recruits E3 ubiquitin ligase RNF20 to promote H2Bub1 at K120; ECL-mediated H2Bub1 disrupts AP-1 recruitment to proinflammatory gene promoters, inhibiting microglia-mediated neuroinflammation.","method":"Biochemical binding assays, ChIP-seq, transcriptomics, site-specific mutagenesis of H2B K46, in vitro and in vivo neuroinflammation models","journal":"Pharmacological research","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — ChIP-seq, mutagenesis, recruitment assays; indirect evidence for RNF20 recruitment via modified H2B","pmids":["35074526"],"is_preprint":false},{"year":2014,"finding":"Bre1a (RNF20 mouse ortholog) regulates neural precursor cell (NPC) differentiation and cell cycle length; knockdown of Bre1a in NPCs lengthens cell cycle through upregulation of p57kip2 and downregulation of Cdk2, and increases Hes5 expression via Fezf1 and Fezf2 to suppress NPC differentiation.","method":"RNAi knockdown in embryonic mouse NPCs, cell cycle analysis, RT-PCR for p57kip2/Cdk2/Hes5/Fezf1/Fezf2, differentiation assays","journal":"The Journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — RNAi knockdown with multiple molecular readouts; single lab, mechanistic pathway partially defined","pmids":["24553946"],"is_preprint":false},{"year":2025,"finding":"RNF20/RNF40, SET1/COMPASS, and DOT1L are dispensable for transcription restart after DNA damage repair (negative finding); levels of H2B-K120Ub and H3K4me3 do not correlate with transcription restoration following DNA damage; PAF1C promotes transcription restart independently of H2B-K120Ub deposition.","method":"RNF20/RNF40 depletion, SET1/COMPASS and DOT1L inhibition, transcription restart assays after DNA damage, ChIP for H2Bub/H3K4me3","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional transcription restart assay with multiple negative controls; preprint, single lab, orthogonal inhibitor approaches","pmids":[],"is_preprint":true}],"current_model":"RNF20, in complex with RNF40, functions as an E3 ubiquitin ligase that monoubiquitinates histone H2B at lysine 120 (H2Bub1) using the E2 enzyme RAD6/Ube2B; structural studies show RNF40 directly contacts nucleosomal DNA and RNF20 RING domain engages the Ube2B~Ub conjugate, with H2BS112 GlcNAcylation allosterically stimulating ubiquitin transfer through RAD6A; beyond transcriptional elongation and chromatin remodeling, RNF20 is recruited to DNA double-strand breaks (via RPA interaction in an R-loop-dependent manner), where H2Bub1 enables SNF2h-mediated chromatin relaxation, Rad51 filament loading, and homologous recombination repair; RNF20 also promotes Rad51 filament assembly and strand exchange independently of its ligase activity by acting as a recombination mediator and antagonizing Srs2/FBH1 helicases; at stalled replication forks, RNF20-mediated H2Bub (regulated by ATR phosphorylation) protects forks from MRE11/DNA2-dependent degradation; beyond histones, RNF20 ubiquitinates non-histone substrates including Eg5 (stabilization for spindle assembly), eEF1BδL (heat shock transcription), NCoR1 and GABPα (adipose thermogenesis), SREBP1c, LSD1 (via CDK9-dependent phosphorylation to drive epigenetic immunosuppression), AP-2α, PSMA7, STAT3, and ZSCAN4; RNF20 also recruits TPM3 to spindle poles via its coiled-coil domain independently of ligase activity for oocyte meiotic spindle assembly; the complex is regulated by Smurf2-mediated proteasomal degradation of RNF20 and by WAC (which bridges RNF20/40 to RNA Pol II), and undergoes SARS-CoV-2 Mpro cleavage at Gln521 to subvert antiviral immunity."},"narrative":{"mechanistic_narrative":"RNF20, partnered with RNF40, is the catalytic E3 ubiquitin ligase that monoubiquitinates histone H2B on lysine 120 (H2Bub1) together with the E2 enzyme RAD6/Ube2B, a chromatin mark that couples to downstream H3K4 and H3K79 methylation and drives transcriptional elongation and chromatin dynamics across diverse developmental and stress programs [PMID:21329877, PMID:27569044, PMID:34290256]. Structurally, RNF20 and RNF40 RING domains form an active heterodimer in which RNF40 contacts nucleosomal DNA and a non-canonical RING surface positions RAD6 directly above H2B K120, and this transfer is allosterically stimulated by H2BS112 GlcNAcylation acting through RAD6A [PMID:37633270, PMID:41495224]. Recruitment to chromatin is gated by accessory factors: WAC bridges the complex to RNA polymerase II at active genes, while a distinct binding interface engages the heterodimer for H2Bub1 catalysis [PMID:21329877, PMID:41533567]. Through H2Bub1, RNF20 controls transcription-dependent processes including embryonic stem cell differentiation of long genes, MLL-fusion leukemogenesis via DOT1L-dependent H3K79 methylation, cardiomyocyte and myogenic differentiation, spermatogenesis through Cldn11 elongation, beta-cell identity in complex with Isl1, and endothelial Notch- and pause-regulated angiogenic programs [PMID:22681888, PMID:23412334, PMID:34290256, PMID:37976359, PMID:37024990, PMID:40743566, PMID:39322771]. Independently of transcription, RNF20 is recruited to DNA double-strand breaks—via FACT and via an RPA/R-loop-dependent route—where H2Bub1 enables SNF2h-mediated chromatin relaxation, end resection, and RAD51/BRCA1 loading for homologous recombination [PMID:21362548, PMID:24357716, PMID:37155876]; it also acts as a non-enzymatic recombination mediator that loads RAD51 on ssDNA and antagonizes Srs2/FBH1 helicases, and protects ATR-regulated stalled forks from MRE11/DNA2 nucleolytic degradation [PMID:37230987, PMID:40495033]. Beyond histones, RNF20 ubiquitinates numerous non-histone substrates to control cell physiology, including Eg5 stabilization for spindle assembly, and degradation of NCoR1, GABPα, SREBP1c, and LSD1 to govern adipose thermogenesis, lipogenesis, and epigenetic immunosuppression [PMID:27557628, PMID:38296968, PMID:28827316, PMID:38315842]. The complex is itself regulated by Smurf2-mediated and ferritinophagy-driven degradation of RNF20, and is subverted by viral pathogens through SARS-CoV-2 Mpro cleavage at Gln521 and adenovirus E1A repurposing [PMID:22231558, PMID:37976359, PMID:34452991, PMID:23785282].","teleology":[{"year":2010,"claim":"Established that RNF20-deposited H2Bub1 and H3K4me3 are not uniformly activating, since RNF20 loss paradoxically enhanced inducible IRF1 transcription, revealing context-dependent repressive output.","evidence":"ChIP and RNAi knockdown at the IRF1 locus with H3K4 methylation inhibition","pmids":["20825659"],"confidence":"Medium","gaps":["Mechanism of locus-specific repression versus activation unresolved","Single locus, single lab"]},{"year":2011,"claim":"Defined how the RNF20/40 complex is targeted to transcription, showing WAC physically bridges the ligase to elongating RNA Pol II and is required for H2B ubiquitination.","evidence":"Affinity purification, reciprocal co-IP, RNAi, and H2B ubiquitination assays","pmids":["21329877"],"confidence":"High","gaps":["Structural basis of WAC bridging not defined here","Whether other recruiters operate at distinct gene classes unknown"]},{"year":2011,"claim":"Extended RNF20 function beyond transcription to genome maintenance, demonstrating it is recruited to DSBs independently of H2AX and that H2Bub1 drives SNF2h-dependent chromatin relaxation enabling resection and RAD51/BRCA1 loading for HR.","evidence":"RNAi, H2B K120R mutant, foci imaging, HR assays, and chromatin relaxation rescue","pmids":["21362548"],"confidence":"High","gaps":["Initial DSB-recruitment mechanism not yet identified","Did not distinguish ligase-dependent from independent contributions"]},{"year":2011,"claim":"Identified CDC73/parafibromin as a complex partner required to maintain H2Bub1, linking the ligase to a tumor-suppressor pathway.","evidence":"Yeast two-hybrid, co-IP, siRNA, and histone-mark western blots in parathyroid tumors","pmids":["22021426"],"confidence":"Medium","gaps":["How CDC73 stimulates catalysis mechanistically unclear","Selective effect on H2Bub1 but not H3K4me3 unexplained"]},{"year":2012,"claim":"Showed RNF20 abundance is set by Smurf2-mediated proteasomal degradation, establishing upstream control over global H2Bub1 and downstream methylation.","evidence":"Smurf2 knockout mice, degradation assays, and co-localization at DSB foci","pmids":["22231558"],"confidence":"High","gaps":["Signals triggering Smurf2-RNF20 turnover not defined","Quantitative coupling to repair outcomes not measured"]},{"year":2012,"claim":"Connected RNF20 loss to a source of endogenous damage, demonstrating its deficiency promotes aberrant R-loops that generate replication-associated DSBs and chromosomal instability.","evidence":"Bre1-deficient cells, R-loop detection, and genomic instability tracking","pmids":["22354749"],"confidence":"Medium","gaps":["Direct molecular link between RNF20 and R-loop resolution unclear","Single lab"]},{"year":2012,"claim":"Demonstrated a developmental requirement for RNF20-dependent H2Bub1 in inducing long genes during ESC differentiation, and identified USP44 as the opposing eraser.","evidence":"ESC differentiation, RNAi of RNF20 and USP44, ChIP, and expression profiling","pmids":["22681888"],"confidence":"High","gaps":["Why long genes are preferentially dependent not mechanistically resolved"]},{"year":2013,"claim":"Identified FACT (SUPT16H) as a direct RING-dependent partner that recruits RNF20 to DSBs in a PAF1-independent manner, separating DNA-damage recruitment from transcriptional recruitment.","evidence":"Co-IP, RING-domain mutagenesis, laser-DSB foci, and HR repair assays","pmids":["24357716"],"confidence":"High","gaps":["How FACT itself is targeted to breaks not addressed"]},{"year":2013,"claim":"Placed RNF20-mediated H2Bub1 within oncogenic transcription, showing it is required for DOT1L-dependent H3K79 methylation and the MLL-fusion leukemogenic program.","evidence":"shRNA in leukemia models in vivo, ChIP-seq for H2Bub and H3K79me2, expression analysis","pmids":["23412334"],"confidence":"High","gaps":["Direct enzymatic coupling between H2Bub1 and DOT1L at these loci not reconstituted"]},{"year":2013,"claim":"Revealed viral subversion, with adenovirus E1A converting RNF20 from a ligase into a hPaf1-recruiting scaffold that activates viral genes while blocking interferon-linked H2Bub1.","evidence":"Co-IP, transcription reporters, and viral replication assays with E1A interaction mapping","pmids":["23785282"],"confidence":"Medium","gaps":["Structural basis of scaffold conversion not defined","Single lab"]},{"year":2013,"claim":"Expanded the substrate repertoire to non-histone targets, showing RNF20 polyubiquitinates AP-2α for degradation to relieve C/EBPα repression in preadipocytes.","evidence":"Co-IP, MS, ubiquitination assays, proteasome inhibition, and rescue assays","pmids":["24374663"],"confidence":"Medium","gaps":["Ubiquitin linkage type not defined","Single lab"]},{"year":2014,"claim":"Provided chemical-biology evidence that RING-cysteine targeting inactivates the ligase, as arsenite binds RNF20/40 RING cysteines to block H2Bub1 and DSB repair.","evidence":"In vitro binding, cellular ubiquitination assays, and laser-DSB foci analysis","pmids":["25170678"],"confidence":"High","gaps":["Selectivity over other RING ligases in cells not fully mapped"]},{"year":2014,"claim":"Showed an ortholog requirement in neural development, with Bre1a regulating NPC cell-cycle length and differentiation via p57kip2, Cdk2, and Hes5/Fezf pathways.","evidence":"RNAi in mouse NPCs with cell-cycle and expression readouts","pmids":["24553946"],"confidence":"Medium","gaps":["Whether effects are H2Bub1-dependent not established","Single lab"]},{"year":2016,"claim":"Solved the RNF20 RING-domain crystal structure, defining the homodimer, the active RNF20/RNF40 heterodimer, and the E3-E2-ubiquitin interface required for catalysis.","evidence":"X-ray crystallography with interface mutagenesis and in vitro ubiquitination assays","pmids":["27569044"],"confidence":"High","gaps":["Nucleosome engagement not captured in this structure"]},{"year":2016,"claim":"Established a ligase-dependent mitotic role distinct from transcription, with RNF20/40 monoubiquitinating and stabilizing Eg5 for spindle assembly.","evidence":"Co-IP, ubiquitination assays, spindle imaging, and in vivo tumor suppression","pmids":["27557628"],"confidence":"High","gaps":["Ubiquitin linkage mediating stabilization not detailed"]},{"year":2016,"claim":"Linked RNF20 loss to inflammation-driven cancer, showing it restrains NF-κB p65 recruitment and maintains H3K9me3 at target genes, with RNF20+/- mice predisposed to colorectal cancer.","evidence":"RNAi, ChIP, NF-κB dimer analysis, and RNF20 heterozygous mouse colitis/cancer models","pmids":["26854224"],"confidence":"High","gaps":["How H2Bub1 controls H3K9me3 at these loci not mechanistically resolved"]},{"year":2017,"claim":"Defined a tumor-suppressive metabolic axis in renal carcinoma, with RNF20 degrading SREBP1c to repress lipogenesis and the PTTG1 oncogene.","evidence":"Overexpression/knockdown, xenografts, lipogenesis assays, and SREBP1c ChIP at PTTG1","pmids":["28827316"],"confidence":"Medium","gaps":["Direct SREBP1c ubiquitination biochemistry not shown","Single lab"]},{"year":2017,"claim":"Identified Fbxl19 as a CxxC-domain recruiter directing RNF20 to CpG-island promoters genome-wide for H2Bub1 and ESC differentiation.","evidence":"Co-IP, ChIP-seq of Fbxl19 and H2Bub1, RNAi, and differentiation assays","pmids":["28453857"],"confidence":"Medium","gaps":["How Fbxl19 and WAC recruitment routes are coordinated unknown","Single lab"]},{"year":2018,"claim":"Added ZSCAN4 as a degradation substrate, showing RNF20 controls ZSCAN4 protein stability without affecting its transcription.","evidence":"Co-IP, protein half-life assays, and RNAi","pmids":["29477841"],"confidence":"Medium","gaps":["Ligase-dependence and linkage type not established","Single lab"]},{"year":2018,"claim":"Demonstrated an in vivo neuronal transcriptional role, with spinal RNF20-dependent H2Bub1 driving phospho-Pol II-dependent mGluR5 transcription downstream of TNF-α in neuropathic pain.","evidence":"Nerve-ligation rat model, in vivo RNAi, ChIP at mGluR5, and behavioral assays","pmids":["30201771"],"confidence":"Medium","gaps":["Upstream signal coupling TNF-α to RNF20 not defined","Single lab"]},{"year":2019,"claim":"Showed RNF20/40-RAD6 ubiquitinates non-histone eEF1BδL at K381 to potentiate p-TEFb recruitment and heat-shock gene transcription with HSF1.","evidence":"In vitro reconstitution, MS site mapping, ChIP, and expression assays","pmids":["30649429"],"confidence":"High","gaps":["Whether eEF1BδL ubiquitination is mono- or poly-ubiquitin not fully specified"]},{"year":2019,"claim":"Connected H2Bub1 to co-transcriptional RNA processing, showing p53-recruited RNF20/40 generates ubH2B at p21/PUMA that is read by the spliceosome subunit PRPF8 for mRNA maturation.","evidence":"Affinity purification, ChIP, RNAi, splicing assays, and ubiquitin-binding domain studies","pmids":["31152661"],"confidence":"High","gaps":["Generality of PRPF8 reading of ubH2B beyond these genes not defined"]},{"year":2019,"claim":"Established RNF20 as a regulator of adipogenesis via NCoR1 degradation that activates PPARγ, validated in conditional knockout mice.","evidence":"Conditional KO mice, quantitative proteomics, co-IP, and degradation assays","pmids":["31604693"],"confidence":"High","gaps":["Direct NCoR1 ubiquitination linkage type not detailed"]},{"year":2019,"claim":"Generalized the repair role across HR and NHEJ, showing RNF20/40 are required for both HR and class-switch recombination, with H2Bub1 induced via joint ATM/ATR control and independent of H2AX.","evidence":"RNAi in mouse B cells, CSR and HR assays, histone-mark blots, and kinase inhibitors","pmids":["30692271"],"confidence":"High","gaps":["How ATM/ATR signals converge on RNF20 catalysis not mapped"]},{"year":2021,"claim":"Demonstrated an in vivo developmental transcriptional requirement, with an unbiased CRISPR screen identifying RNF20/40-mediated H2Bub1 as controlling cardiomyocyte maturation gene programs.","evidence":"In vivo somatic Cas9 screen with cardiomyocyte maturation phenotyping","pmids":["34290256"],"confidence":"Medium","gaps":["Direct target genes not individually validated","Single lab"]},{"year":2021,"claim":"Linked RNF20/40-H2Bub1 to intestinal homeostasis through H3K4me3-dependent VDR transcription, with conditional intestinal knockouts causing colitis relevant to IBD.","evidence":"Conditional intestinal KO mice, ChIP-seq at VDR, mRNA-seq, and IBD patient ChIP-seq","pmids":["34088983"],"confidence":"High","gaps":["Whether VDR is the sole driver of the colitis phenotype unresolved"]},{"year":2022,"claim":"Provided a chemical handle showing the natural compound epoxymicheliolide modifies H2B K46 to recruit RNF20 and induce H2Bub1, blocking AP-1-driven neuroinflammation.","evidence":"Binding assays, ChIP-seq, H2B K46 mutagenesis, and neuroinflammation models","pmids":["35074526"],"confidence":"Medium","gaps":["RNF20 recruitment is inferred via modified H2B rather than direct","Single lab"]},{"year":2023,"claim":"Resolved the structural basis of site-specific catalysis, with cryo-EM showing RNF40 (and yeast Bre1) bind nucleosomal DNA and a non-canonical RING contact positions RAD6 above H2B K120, conserved yeast-to-human.","evidence":"Chemical trapping and cryo-EM of E3/E2/nucleosome complexes with reconstitution","pmids":["37633270"],"confidence":"High","gaps":["Dynamics of substrate selection during transfer not captured"]},{"year":2023,"claim":"Identified RPA as a cell-cycle-regulated, R-loop-dependent recruiter of RNF20 to centromeres and breaks, coupling H2Bub1 to Aurora B activation and repair-protein loading.","evidence":"Reciprocal co-IP, ChIP, mitotic phenotyping, HR assays, and R-loop disruption","pmids":["37155876"],"confidence":"High","gaps":["How RPA and FACT recruitment routes are partitioned not defined"]},{"year":2023,"claim":"Uncovered a ligase-independent recombination-mediator activity, with RNF20/Bre1 directly loading Rad51 on ssDNA, promoting strand exchange, and antagonizing Srs2/FBH1 additively with BRCA2/Rad52.","evidence":"In vitro filament assembly and strand-exchange assays, ligase-dead mutants, and epistasis","pmids":["37230987"],"confidence":"High","gaps":["How catalytic and mediator activities are coordinated in vivo unclear"]},{"year":2023,"claim":"Showed RNF20 abundance is controlled by ferritinophagy, where iron-deficiency-driven autophagic degradation of RNF20 lowers H2Bub1 at MyoD/MyoG to block myogenic differentiation.","evidence":"Iron-deficiency models, autophagy inhibition, RNF20 overexpression rescue, ChIP, and conditional NCOA4 KO","pmids":["37976359"],"confidence":"High","gaps":["Degron/recognition basis for autophagic RNF20 turnover not defined"]},{"year":2023,"claim":"Defined a requirement in male fertility, with Sertoli-cell RNF20 driving H2Bub1-dependent Cldn11 elongation and seminiferous tubule integrity.","evidence":"Sertoli-specific conditional KO mice, histology, H2Bub1 blots, and Cldn11 ChIP","pmids":["37024990"],"confidence":"High","gaps":["Whether additional Sertoli target genes contribute not fully mapped"]},{"year":2023,"claim":"Extended viral hijacking to KSHV, where RTA engages the RNF20/40 complex to enable lytic-cycle reactivation.","evidence":"Proteomics, co-IP, and functional lytic reactivation assays","pmids":["37888983"],"confidence":"Medium","gaps":["Whether ligase activity is required for reactivation unresolved","Single lab"]},{"year":2023,"claim":"Reported RNF20-mediated STAT3 ubiquitination and degradation as an insulin-resistance modifier downstream of exosomal miR-let-7b-5p.","evidence":"Luciferase reporter, single co-IP for STAT3 ubiquitination, and overexpression rescue","pmids":["37842335"],"confidence":"Low","gaps":["Single Co-IP without reciprocal validation or linkage typing","Direct versus indirect degradation not distinguished","Single lab"]},{"year":2024,"claim":"Defined depot-specific non-histone substrate selection in thermogenesis, with RNF20 degrading NCoR1 in iWAT and controlling GABPα stability in BAT under cold stress.","evidence":"Fat-depot-specific Rnf20 KO mice, cold exposure, stability assays, and ubiquitination assays","pmids":["38296968"],"confidence":"High","gaps":["How substrate choice is restricted by depot context not mechanistically resolved"]},{"year":2024,"claim":"Established an immunosuppressive ligase axis, with CDK9-phosphorylation-dependent RNF20 K29-ubiquitination stabilizing LSD1 to repress endogenous retroviruses and the interferon response, sensitizing tumors to anti-PD-1 upon RNF20 loss.","evidence":"Co-IP, K29-specific ubiquitination, CDK9 epistasis, ERV expression, and in vivo anti-PD-1 rescue","pmids":["38315842"],"confidence":"High","gaps":["Whether K29-linkage stabilization generalizes to other substrates unknown"]},{"year":2024,"claim":"Identified an H2Bub1-independent oncogenic mechanism, with RNF20 controlling Rbx1/VHL activity and HIF1α levels to drive metabolic rewiring and EMT via Pol II pause release.","evidence":"Rnf20 haploinsufficient mouse lung tumors, metabolomics, HIF1α stability, and Rbx1/VHL epistasis","pmids":["40436847"],"confidence":"Medium","gaps":["Direct mechanism by which RNF20 sets Rbx1 levels not defined","Single lab"]},{"year":2024,"claim":"Showed cooperative recruitment under hypoxia, with HIF-1 bringing FACT and RNF20/40 to HREs to induce H2Bub1 and support transcription initiation and elongation at HIF-1 targets.","evidence":"RNAi, ChIP at HREs, and hypoxic H2Bub ChIP in breast cancer cells","pmids":["38517892"],"confidence":"Medium","gaps":["Reconciliation with the H2Bub1-independent HIF1α mechanism not addressed","Single lab"]},{"year":2024,"claim":"Revealed a non-enzymatic structural role in meiosis, with RNF20 recruiting TPM3 to spindle poles via its coiled-coil for acentrosomal oocyte spindle assembly.","evidence":"Oocyte depletion, ligase-dead mutant, RNF20-TPM3 co-IP, localization, and coiled-coil mapping","pmids":["38240347"],"confidence":"High","gaps":["How RNF20 itself targets spindle poles not defined"]},{"year":2024,"claim":"Connected RNF20 to angiogenic transcriptional control, restricting ERG-dependent Pol II pause release and promoting Notch-target H2Bub1 to constrain tip-cell specification.","evidence":"Endothelial Rnf20 KO, Pol II pausing ChIP-seq, Notch1 co-IP, H2Bub ChIP, and angiogenesis assays","pmids":["39322771"],"confidence":"High","gaps":["Direct mechanism of pause regulation versus H2Bub1 deposition not separated"]},{"year":2025,"claim":"Defined a replication-fork protective role, with ATR-phosphorylated RNF20 and its RING domain promoting H2Bub1 and RAD51/RAD51C loading to shield stalled forks from MRE11/DNA2 degradation.","evidence":"DNA fiber assays, RING and phospho-mutant analysis, nuclease inhibition, and fork-remodeler co-depletion","pmids":["40495033"],"confidence":"High","gaps":["Relative contributions of H2Bub1 versus mediator activity at forks not fully separated"]},{"year":2025,"claim":"Resolved the assembly interfaces, with crystallography and AlphaFold modeling defining the Bre1-Lge1 and RNF20/RNF40-WAC interfaces and their distinct electrostatic specificity required for H2Bub1.","evidence":"X-ray crystallography, AlphaFold modeling, interface mutagenesis, and ubiquitination assays","pmids":["41533567"],"confidence":"High","gaps":["Full RNF20/40-WAC structure experimentally unverified beyond modeling"]},{"year":2025,"claim":"Established RNF20 as a beta-cell transcriptional regulator in complex with Isl1, required for H2Bub1, beta-cell identity, and glucose-stimulated insulin secretion.","evidence":"Inducible beta-cell conditional KO, Isl1-Rnf20 co-IP, transcriptomics, and insulin secretion assays","pmids":["40743566"],"confidence":"High","gaps":["Direct Isl1-RNF20 target genes for insulin control not individually validated"]},{"year":2025,"claim":"Defined a nucleocytoplasmic antiviral switch, with CRM1-exported cytoplasmic RNF20 catalyzing K27-linked degradation of RIG-I and MDA5 via its coiled-coil while nuclear RNF20 sustains their transcription.","evidence":"Overexpression/knockout, co-IP, K27-specific ubiquitination, CRM1 inhibition, and viral infection assays","pmids":["41712649"],"confidence":"Medium","gaps":["Trigger and regulation of RNF20 export not defined","Single lab"]},{"year":2026,"claim":"Defined PTM crosstalk gating catalysis, with cryo-EM showing H2BS112 GlcNAcylation contacts RAD6A and allosterically stimulates ubiquitin transfer to H2B K120 by enhancing lysine nucleophilicity.","evidence":"Synthetic modified nucleosomes, chemical trapping, cryo-EM, kinetics, and SAR mutagenesis","pmids":["41495224"],"confidence":"High","gaps":["Physiological enzymes setting H2BS112 GlcNAc in this context not identified"]},{"year":null,"claim":"It remains unresolved how RNF20's multiple recruitment routes (WAC, FACT, RPA, Fbxl19, sequence-specific transcription factors) and its catalytic versus ligase-independent activities are coordinately partitioned across transcription, replication, repair, mitosis, and cytoplasmic immune signaling within a single cell.","evidence":"No single study integrates the competing recruitment and activity modes","pmids":[],"confidence":"Low","gaps":["No unified model of recruiter selection","Catalytic versus scaffold/mediator activity switching not mechanistically defined","A preprint reports H2Bub1 is dispensable for transcription restart after damage, complicating the repair-transcription model (37976359 not applicable; bioRxiv)"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[12,22,1,25]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[10,15,16,27,30,19,37]},{"term_id":"GO:0140110","term_label":"transcription regulator activity","supporting_discovery_ids":[1,8,32,43]},{"term_id":"GO:0003677","term_label":"DNA binding","supporting_discovery_ids":[22]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[24,31]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[1,4,14,44]},{"term_id":"GO:0000228","term_label":"nuclear chromosome","supporting_discovery_ids":[0,23,34]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[44]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[10,31]}],"pathway":[{"term_id":"R-HSA-74160","term_label":"Gene expression (Transcription)","supporting_discovery_ids":[1,4,8,32,43]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[0,6,21,23]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[1,4,22,8]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,10,15,16,27,30]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[10,23,34]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[27,44]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[15,17,28,30]}],"complexes":["RNF20/RNF40 (BRE1) H2B ubiquitin ligase complex"],"partners":["RNF40","RAD6/UBE2B","WAC","SUPT16H (FACT)","RPA","CDC73","RAD51","ISL1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q5VTR2","full_name":"E3 ubiquitin-protein ligase BRE1A","aliases":["RING finger protein 20","RING-type E3 ubiquitin transferase BRE1A"],"length_aa":975,"mass_kda":113.7,"function":"Component of the RNF20/40 E3 ubiquitin-protein ligase complex that mediates monoubiquitination of 'Lys-120' of histone H2B (H2BK120ub1). H2BK120ub1 gives a specific tag for epigenetic transcriptional activation and is also prerequisite for histone H3 'Lys-4' and 'Lys-79' methylation (H3K4me and H3K79me, respectively). It thereby plays a central role inb histone code and gene regulation. The RNF20/40 complex forms a H2B ubiquitin ligase complex in cooperation with the E2 enzyme UBE2A or UBE2B; reports about the cooperation with UBE2E1/UBCH are contradictory. Required for transcriptional activation of Hox genes. Recruited to the MDM2 promoter, probably by being recruited by p53/TP53, and thereby acts as a transcriptional coactivator. Mediates the polyubiquitination of isoform 2 of PA2G4 in cancer cells leading to its proteasome-mediated degradation (Microbial infection) Promotes the human herpesvirus 8 (KSHV) lytic cycle by inducing the expression of lytic viral genes including the latency switch gene RTA/ORF50","subcellular_location":"Nucleus","url":"https://www.uniprot.org/uniprotkb/Q5VTR2/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":true,"resolved_as":"","url":"https://depmap.org/portal/gene/RNF20","classification":"Common Essential","n_dependent_lines":1069,"n_total_lines":1208,"dependency_fraction":0.8849337748344371},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"RNF40","stoichiometry":10.0},{"gene":"PRPF19","stoichiometry":0.2},{"gene":"RBM12","stoichiometry":0.2},{"gene":"FAM117B","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/RNF20","total_profiled":1310},"omim":[{"mim_id":"617233","title":"WD REPEAT-CONTAINING PROTEIN 70; WDR70","url":"https://www.omim.org/entry/617233"},{"mim_id":"614802","title":"MSL COMPLEX SUBUNIT 2; MSL2","url":"https://www.omim.org/entry/614802"},{"mim_id":"614801","title":"MSL COMPLEX SUBUNIT 1; MSL1","url":"https://www.omim.org/entry/614801"},{"mim_id":"612860","title":"QUIESCIN Q6 SULFHYDRYL OXIDASE 2; QSOX2","url":"https://www.omim.org/entry/612860"},{"mim_id":"612116","title":"UBIQUITIN-SPECIFIC PROTEASE 22; USP22","url":"https://www.omim.org/entry/612116"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"},{"location":"Cytosol","reliability":"Additional"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/RNF20"},"hgnc":{"alias_symbol":["FLJ20382","FLJ11189","KAIA2779","BRE1A","hBRE1","BRE1"],"prev_symbol":[]},"alphafold":{"accession":"Q5VTR2","domains":[{"cath_id":"-","chopping":"63-112_153-295","consensus_level":"medium","plddt":84.8974,"start":63,"end":295},{"cath_id":"3.30.40.10","chopping":"907-970","consensus_level":"high","plddt":87.1144,"start":907,"end":970}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5VTR2","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q5VTR2-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q5VTR2-F1-predicted_aligned_error_v6.png","plddt_mean":75.38},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=RNF20","jax_strain_url":"https://www.jax.org/strain/search?query=RNF20"},"sequence":{"accession":"Q5VTR2","fasta_url":"https://rest.uniprot.org/uniprotkb/Q5VTR2.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q5VTR2/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q5VTR2"}},"corpus_meta":[{"pmid":"21362548","id":"PMC_21362548","title":"Regulation of homologous recombination by RNF20-dependent H2B ubiquitination.","date":"2011","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/21362548","citation_count":304,"is_preprint":false},{"pmid":"22681888","id":"PMC_22681888","title":"RNF20 and USP44 regulate stem cell differentiation by modulating H2B monoubiquitylation.","date":"2012","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/22681888","citation_count":177,"is_preprint":false},{"pmid":"22231558","id":"PMC_22231558","title":"A tumor suppressor function of Smurf2 associated with controlling chromatin landscape and genome stability through RNF20.","date":"2012","source":"Nature medicine","url":"https://pubmed.ncbi.nlm.nih.gov/22231558","citation_count":138,"is_preprint":false},{"pmid":"21329877","id":"PMC_21329877","title":"WAC, a functional partner of RNF20/40, regulates histone H2B ubiquitination and gene transcription.","date":"2011","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/21329877","citation_count":136,"is_preprint":false},{"pmid":"26854224","id":"PMC_26854224","title":"RNF20 Links Histone H2B Ubiquitylation with Inflammation and Inflammation-Associated Cancer.","date":"2016","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/26854224","citation_count":112,"is_preprint":false},{"pmid":"22354749","id":"PMC_22354749","title":"Deficiency in mammalian histone H2B ubiquitin ligase Bre1 (Rnf20/Rnf40) leads to replication stress and chromosomal instability.","date":"2012","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/22354749","citation_count":106,"is_preprint":false},{"pmid":"23412334","id":"PMC_23412334","title":"Histone H2B ubiquitin ligase RNF20 is required for MLL-rearranged leukemia.","date":"2013","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/23412334","citation_count":96,"is_preprint":false},{"pmid":"22021426","id":"PMC_22021426","title":"The tumor suppressor CDC73 interacts with the ring finger proteins RNF20 and RNF40 and is required for the maintenance of histone 2B monoubiquitination.","date":"2011","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/22021426","citation_count":72,"is_preprint":false},{"pmid":"27557628","id":"PMC_27557628","title":"Ubiquitin ligase RNF20/40 facilitates spindle assembly and promotes breast carcinogenesis through stabilizing motor protein Eg5.","date":"2016","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/27557628","citation_count":60,"is_preprint":false},{"pmid":"30692271","id":"PMC_30692271","title":"E3 Ubiquitin Ligases RNF20 and RNF40 Are Required for Double-Stranded Break (DSB) Repair: Evidence for Monoubiquitination of Histone H2B Lysine 120 as a Novel Axis of DSB Signaling and Repair.","date":"2019","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/30692271","citation_count":60,"is_preprint":false},{"pmid":"34290256","id":"PMC_34290256","title":"Massively parallel in vivo CRISPR screening identifies RNF20/40 as epigenetic regulators of cardiomyocyte maturation.","date":"2021","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/34290256","citation_count":54,"is_preprint":false},{"pmid":"24357716","id":"PMC_24357716","title":"Histone chaperone FACT regulates homologous recombination by chromatin remodeling through interaction with RNF20.","date":"2013","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/24357716","citation_count":53,"is_preprint":false},{"pmid":"28690313","id":"PMC_28690313","title":"The HDAC inhibitor panobinostat (LBH589) exerts in vivo anti-leukaemic activity against MLL-rearranged acute lymphoblastic leukaemia and involves the RNF20/RNF40/WAC-H2B ubiquitination axis.","date":"2017","source":"Leukemia","url":"https://pubmed.ncbi.nlm.nih.gov/28690313","citation_count":51,"is_preprint":false},{"pmid":"28157208","id":"PMC_28157208","title":"RNF20 and histone H2B ubiquitylation exert opposing effects in Basal-Like versus luminal breast cancer.","date":"2017","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/28157208","citation_count":48,"is_preprint":false},{"pmid":"25170678","id":"PMC_25170678","title":"Arsenite binds to the RING finger domains of RNF20-RNF40 histone E3 ubiquitin ligase and inhibits DNA double-strand break repair.","date":"2014","source":"Journal of the American Chemical Society","url":"https://pubmed.ncbi.nlm.nih.gov/25170678","citation_count":48,"is_preprint":false},{"pmid":"34452991","id":"PMC_34452991","title":"Protease cleavage of RNF20 facilitates coronavirus replication via stabilization of SREBP1.","date":"2021","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/34452991","citation_count":46,"is_preprint":false},{"pmid":"22155569","id":"PMC_22155569","title":"Histone H2B ubiquitin ligases RNF20 and RNF40 in androgen signaling and prostate cancer cell growth.","date":"2011","source":"Molecular and cellular endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/22155569","citation_count":46,"is_preprint":false},{"pmid":"28827316","id":"PMC_28827316","title":"RNF20 Suppresses Tumorigenesis by Inhibiting the SREBP1c-PTTG1 Axis in Kidney Cancer.","date":"2017","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/28827316","citation_count":45,"is_preprint":false},{"pmid":"29934362","id":"PMC_29934362","title":"Role of RNF20 in cancer development and progression - a comprehensive review.","date":"2018","source":"Bioscience reports","url":"https://pubmed.ncbi.nlm.nih.gov/29934362","citation_count":39,"is_preprint":false},{"pmid":"37633270","id":"PMC_37633270","title":"Mechanistic insights into nucleosomal H2B monoubiquitylation mediated by yeast Bre1-Rad6 and its human homolog RNF20/RNF40-hRAD6A.","date":"2023","source":"Molecular cell","url":"https://pubmed.ncbi.nlm.nih.gov/37633270","citation_count":32,"is_preprint":false},{"pmid":"27798111","id":"PMC_27798111","title":"The RING finger domain E3 ubiquitin ligases BRCA1 and the RNF20/RNF40 complex in global loss of the chromatin mark histone H2B monoubiquitination (H2Bub1) in cell line models and primary high-grade serous ovarian cancer.","date":"2016","source":"Human molecular genetics","url":"https://pubmed.ncbi.nlm.nih.gov/27798111","citation_count":31,"is_preprint":false},{"pmid":"27569044","id":"PMC_27569044","title":"Structure and Function of the RING Domains of RNF20 and RNF40, Dimeric E3 Ligases that Monoubiquitylate Histone H2B.","date":"2016","source":"Journal of molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/27569044","citation_count":28,"is_preprint":false},{"pmid":"20825659","id":"PMC_20825659","title":"Menin and RNF20 recruitment is associated with dynamic histone modifications that regulate signal transducer and activator of transcription 1 (STAT1)-activated transcription of the interferon regulatory factor 1 gene (IRF1).","date":"2010","source":"Epigenetics & chromatin","url":"https://pubmed.ncbi.nlm.nih.gov/20825659","citation_count":27,"is_preprint":false},{"pmid":"31604693","id":"PMC_31604693","title":"RNF20 Functions as a Transcriptional Coactivator for PPARγ by Promoting NCoR1 Degradation in Adipocytes.","date":"2019","source":"Diabetes","url":"https://pubmed.ncbi.nlm.nih.gov/31604693","citation_count":26,"is_preprint":false},{"pmid":"37155876","id":"PMC_37155876","title":"The RPA-RNF20-SNF2H cascade promotes proper chromosome segregation and homologous recombination repair.","date":"2023","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/37155876","citation_count":24,"is_preprint":false},{"pmid":"31152661","id":"PMC_31152661","title":"The RNF20/40 complex regulates p53-dependent gene transcription and mRNA splicing.","date":"2020","source":"Journal of molecular cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/31152661","citation_count":22,"is_preprint":false},{"pmid":"30649429","id":"PMC_30649429","title":"RNF20/40-mediated eEF1BδL monoubiquitylation stimulates transcription of heat shock-responsive genes.","date":"2019","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/30649429","citation_count":22,"is_preprint":false},{"pmid":"23785282","id":"PMC_23785282","title":"Viral retasking of hBre1/RNF20 to recruit hPaf1 for transcriptional activation.","date":"2013","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/23785282","citation_count":22,"is_preprint":false},{"pmid":"24553946","id":"PMC_24553946","title":"Bre1a, a histone H2B ubiquitin ligase, regulates the cell cycle and differentiation of neural precursor cells.","date":"2014","source":"The Journal of neuroscience : the official journal of the Society for Neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/24553946","citation_count":20,"is_preprint":false},{"pmid":"34088983","id":"PMC_34088983","title":"RNF20 and RNF40 regulate vitamin D receptor-dependent signaling in inflammatory bowel disease.","date":"2021","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/34088983","citation_count":19,"is_preprint":false},{"pmid":"32797353","id":"PMC_32797353","title":"Rnf20 deficiency in adipocyte impairs adipose tissue development and thermogenesis.","date":"2020","source":"Protein & cell","url":"https://pubmed.ncbi.nlm.nih.gov/32797353","citation_count":18,"is_preprint":false},{"pmid":"33364200","id":"PMC_33364200","title":"RNF20 Is Critical for Snail-Mediated E-Cadherin Repression in Human Breast Cancer.","date":"2020","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/33364200","citation_count":16,"is_preprint":false},{"pmid":"30201771","id":"PMC_30201771","title":"Spinal RNF20-Mediated Histone H2B Monoubiquitylation Regulates mGluR5 Transcription for Neuropathic Allodynia.","date":"2018","source":"The Journal of neuroscience : the official journal of the Society for Neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/30201771","citation_count":15,"is_preprint":false},{"pmid":"36103829","id":"PMC_36103829","title":"Endothelial cells regulated by RNF20 orchestrate the proliferation and differentiation of neural precursor cells during embryonic development.","date":"2022","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/36103829","citation_count":15,"is_preprint":false},{"pmid":"26184323","id":"PMC_26184323","title":"RNF20-SNF2H Pathway of Chromatin Relaxation in DNA Double-Strand Break Repair.","date":"2015","source":"Genes","url":"https://pubmed.ncbi.nlm.nih.gov/26184323","citation_count":14,"is_preprint":false},{"pmid":"24374663","id":"PMC_24374663","title":"RNF20 promotes the polyubiquitination and proteasome-dependent degradation of AP-2α protein.","date":"2013","source":"Acta biochimica et biophysica Sinica","url":"https://pubmed.ncbi.nlm.nih.gov/24374663","citation_count":14,"is_preprint":false},{"pmid":"34647359","id":"PMC_34647359","title":"RNF20 affects porcine adipocyte differentiation via regulation of mitotic clonal expansion.","date":"2021","source":"Cell proliferation","url":"https://pubmed.ncbi.nlm.nih.gov/34647359","citation_count":14,"is_preprint":false},{"pmid":"37230987","id":"PMC_37230987","title":"Bre1/RNF20 promotes Rad51-mediated strand exchange and antagonizes the Srs2/FBH1 helicases.","date":"2023","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/37230987","citation_count":13,"is_preprint":false},{"pmid":"29477841","id":"PMC_29477841","title":"ZSCAN4 is negatively regulated by the ubiquitin-proteasome system and the E3 ubiquitin ligase RNF20.","date":"2018","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/29477841","citation_count":13,"is_preprint":false},{"pmid":"37976359","id":"PMC_37976359","title":"Iron deficiency-induced ferritinophagy impairs skeletal muscle regeneration through RNF20-mediated H2Bub1 modification.","date":"2023","source":"Science advances","url":"https://pubmed.ncbi.nlm.nih.gov/37976359","citation_count":13,"is_preprint":false},{"pmid":"37842335","id":"PMC_37842335","title":"Pancreatic Cancer-Derived Exosomal miR-Let-7b-5p Stimulates Insulin Resistance in Skeletal Muscle Cells Through RNF20/STAT3/FOXO1 Axis Regulation.","date":"2023","source":"Diabetes, metabolic syndrome and obesity : targets and therapy","url":"https://pubmed.ncbi.nlm.nih.gov/37842335","citation_count":12,"is_preprint":false},{"pmid":"35074526","id":"PMC_35074526","title":"Epoxymicheliolide directly targets histone H2B to inhibit neuroinflammation via recruiting E3 ligase RNF20.","date":"2022","source":"Pharmacological research","url":"https://pubmed.ncbi.nlm.nih.gov/35074526","citation_count":12,"is_preprint":false},{"pmid":"28462496","id":"PMC_28462496","title":"Synthetic lethal targeting of RNF20 through PARP1 silencing and inhibition.","date":"2017","source":"Cellular oncology (Dordrecht, Netherlands)","url":"https://pubmed.ncbi.nlm.nih.gov/28462496","citation_count":11,"is_preprint":false},{"pmid":"38517892","id":"PMC_38517892","title":"Hypoxia-inducible factor 1 recruits FACT and RNF20/40 to mediate histone ubiquitination and transcriptional activation of target genes.","date":"2024","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/38517892","citation_count":11,"is_preprint":false},{"pmid":"40495033","id":"PMC_40495033","title":"RNF20-mediated H2B monoubiquitination protects stalled forks from degradation and promotes fork restart.","date":"2025","source":"EMBO reports","url":"https://pubmed.ncbi.nlm.nih.gov/40495033","citation_count":10,"is_preprint":false},{"pmid":"28453857","id":"PMC_28453857","title":"Fbxl19 recruitment to CpG islands is required for Rnf20-mediated H2B mono-ubiquitination.","date":"2017","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/28453857","citation_count":10,"is_preprint":false},{"pmid":"39322771","id":"PMC_39322771","title":"RNF20-mediated transcriptional pausing and VEGFA splicing orchestrate vessel growth.","date":"2024","source":"Nature cardiovascular research","url":"https://pubmed.ncbi.nlm.nih.gov/39322771","citation_count":9,"is_preprint":false},{"pmid":"37888983","id":"PMC_37888983","title":"KSHV RTA utilizes the host E3 ubiquitin ligase complex RNF20/40 to drive lytic reactivation.","date":"2023","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/37888983","citation_count":9,"is_preprint":false},{"pmid":"38278978","id":"PMC_38278978","title":"Albendazole inhibits colon cancer progression and therapy resistance by targeting ubiquitin ligase RNF20.","date":"2024","source":"British journal of cancer","url":"https://pubmed.ncbi.nlm.nih.gov/38278978","citation_count":9,"is_preprint":false},{"pmid":"36371755","id":"PMC_36371755","title":"Rnf20 inhibition enhances immunotherapy by improving regulatory T cell generation.","date":"2022","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/36371755","citation_count":8,"is_preprint":false},{"pmid":"37024990","id":"PMC_37024990","title":"RNF20 is required for male fertility through regulation of H2B ubiquitination in the Sertoli cells.","date":"2023","source":"Cell & bioscience","url":"https://pubmed.ncbi.nlm.nih.gov/37024990","citation_count":8,"is_preprint":false},{"pmid":"40436847","id":"PMC_40436847","title":"RNF20 links the DNA damage response and metabolic rewiring in lung cancer through HIF1α.","date":"2025","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/40436847","citation_count":7,"is_preprint":false},{"pmid":"38315842","id":"PMC_38315842","title":"RNF20 contributes to epigenetic immunosuppression through CDK9-dependent LSD1 stabilization.","date":"2024","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/38315842","citation_count":7,"is_preprint":false},{"pmid":"38296968","id":"PMC_38296968","title":"Ubiquitin ligase RNF20 coordinates sequential adipose thermogenesis with brown and beige fat-specific substrates.","date":"2024","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/38296968","citation_count":7,"is_preprint":false},{"pmid":"40597205","id":"PMC_40597205","title":"RNF20/RNF40 supports the aggressive behavior in cervical cancer by regulating a peroxisome-based anti-ferroptotic mechanism.","date":"2025","source":"Cell communication and signaling : CCS","url":"https://pubmed.ncbi.nlm.nih.gov/40597205","citation_count":6,"is_preprint":false},{"pmid":"38240347","id":"PMC_38240347","title":"RNF20 Regulates Oocyte Meiotic Spindle Assembly by Recruiting TPM3 to Centromeres and Spindle Poles.","date":"2024","source":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/38240347","citation_count":6,"is_preprint":false},{"pmid":"36650938","id":"PMC_36650938","title":"RNF20 deletion causes inflammation in model of sepsis through the NLRP3 activation.","date":"2023","source":"Immunopharmacology and immunotoxicology","url":"https://pubmed.ncbi.nlm.nih.gov/36650938","citation_count":5,"is_preprint":false},{"pmid":"38608146","id":"PMC_38608146","title":"RNF20 Reduces Cell Proliferation and Warburg Effect by Promoting NLRP3 Ubiquitination in Liver Cancer.","date":"2024","source":"Journal of environmental pathology, toxicology and oncology : official organ of the International Society for Environmental Toxicology and Cancer","url":"https://pubmed.ncbi.nlm.nih.gov/38608146","citation_count":4,"is_preprint":false},{"pmid":"37555936","id":"PMC_37555936","title":"Adipocyte Rnf20 ablation increases the fast-twitch fibers of skeletal muscle via lysophosphatidylcholine 16:0.","date":"2023","source":"Cellular and molecular life sciences : CMLS","url":"https://pubmed.ncbi.nlm.nih.gov/37555936","citation_count":4,"is_preprint":false},{"pmid":"22825248","id":"PMC_22825248","title":"R-loops and genomic instability in Bre1 (RNF20/40)-deficient cells.","date":"2012","source":"Cell cycle (Georgetown, Tex.)","url":"https://pubmed.ncbi.nlm.nih.gov/22825248","citation_count":4,"is_preprint":false},{"pmid":"40522008","id":"PMC_40522008","title":"Adipocyte RNF20 Knockout Leads to Hyperinsulinemia via the H2Bub-H3K4me3-Slc2a4 Axis.","date":"2025","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/40522008","citation_count":3,"is_preprint":false},{"pmid":"41145490","id":"PMC_41145490","title":"Endothelial RNF20 suppresses endothelial-to-mesenchymal transition and safeguards physiological angiocrine signaling to prevent congenital heart disease.","date":"2025","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/41145490","citation_count":2,"is_preprint":false},{"pmid":"37422774","id":"PMC_37422774","title":"RNF20/RNF40 ameliorates streptozotocin-induced type 1 diabetes by activating vitamin D receptors in vivo.","date":"2023","source":"Allergologia et immunopathologia","url":"https://pubmed.ncbi.nlm.nih.gov/37422774","citation_count":2,"is_preprint":false},{"pmid":"41533567","id":"PMC_41533567","title":"Structural insights into the Bre1-Lge1 and RNF20/RNF40-WAC interactions critical for H2B ubiquitination.","date":"2026","source":"Nucleic acids research","url":"https://pubmed.ncbi.nlm.nih.gov/41533567","citation_count":2,"is_preprint":false},{"pmid":"32443664","id":"PMC_32443664","title":"Molecular Characterization, Expression Profiling, and SNP Analysis of the Porcine RNF20 Gene.","date":"2020","source":"Animals : an open access journal from MDPI","url":"https://pubmed.ncbi.nlm.nih.gov/32443664","citation_count":2,"is_preprint":false},{"pmid":"40743566","id":"PMC_40743566","title":"The Islet-1 Interaction Partner Rnf20 Regulates Glucose Homeostasis and Pancreatic β-Cell Identity.","date":"2025","source":"Diabetes","url":"https://pubmed.ncbi.nlm.nih.gov/40743566","citation_count":1,"is_preprint":false},{"pmid":"39998124","id":"PMC_39998124","title":"RNF20 dual regulation of MDA5 signaling to maintain immune homeostasis in chickens.","date":"2025","source":"Journal of virology","url":"https://pubmed.ncbi.nlm.nih.gov/39998124","citation_count":1,"is_preprint":false},{"pmid":"41495224","id":"PMC_41495224","title":"Allosteric activation of RNF20/RNF40-RAD6A-mediated H2BK120 monoubiquitylation by H2BS112 GlcNAcylation.","date":"2026","source":"Nature chemical biology","url":"https://pubmed.ncbi.nlm.nih.gov/41495224","citation_count":1,"is_preprint":false},{"pmid":"41254650","id":"PMC_41254650","title":"Melittin inhibits NSCLC metastasis by targeting USP10 and promoting RNF20-mediated ubiquitination and degradation of PSMA7.","date":"2025","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/41254650","citation_count":0,"is_preprint":false},{"pmid":"42004469","id":"PMC_42004469","title":"Decoding RNF20: an epigenetic modifier and beyond.","date":"2026","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/42004469","citation_count":0,"is_preprint":false},{"pmid":"41712649","id":"PMC_41712649","title":"RNF20 dynamically regulates RIG-I and MDA5 transcription and degradation via nucleocytoplasmic translocation to balance antiviral signaling.","date":"2026","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/41712649","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.07.23.666359","title":"PAF1C-driven restoration of RNAPII elongation after DNA damage occurs independently of transcription-associated histone mark deposition","date":"2025-07-24","source":"bioRxiv","url":"https://doi.org/10.1101/2025.07.23.666359","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.06.02.657518","title":"A novel tool for labeling intermediary proteins between two non-interacting proteins","date":"2025-06-03","source":"bioRxiv","url":"https://doi.org/10.1101/2025.06.02.657518","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":36482,"output_tokens":13167,"usd":0.153475,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":24563,"output_tokens":11316,"usd":0.202857,"stage2_stop_reason":"end_turn"},"total_usd":0.356332,"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\": 2011,\n      \"finding\": \"RNF20 localizes to DNA double-strand breaks (DSBs) independently of H2AX and is required for DSB-induced H2B ubiquitination at K120, H3K4 methylation at DSBs, and recruitment of chromatin-remodeling factor SNF2h; depletion of RNF20 or expression of H2B K120R mutant compromises DNA end resection and recruitment of RAD51 and BRCA1, causing homologous recombination repair defects and radiation sensitivity.\",\n      \"method\": \"RNAi knockdown, H2B K120R mutant expression, immunofluorescence/foci analysis, HR repair assays, chromatin relaxation rescue experiments\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (KD, dominant-negative mutant, functional HR assay, foci recruitment, chromatin relaxation rescue) in a single focused study\",\n      \"pmids\": [\"21362548\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"WAC interacts with RNF20/40 through its C-terminal coiled-coil region, recognizes RNA polymerase II via its N-terminal WW domain, and targets RNF20/40 to the RNA polymerase II complex at active transcription sites to promote H2B ubiquitination; WAC depletion abolishes H2B ubiquitination.\",\n      \"method\": \"Protein affinity purification, co-immunoprecipitation, RNAi knockdown, H2B ubiquitination assays, cell-cycle checkpoint assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, affinity purification, functional ubiquitination assays, multiple orthogonal methods in one study\",\n      \"pmids\": [\"21329877\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Smurf2 (HECT-domain E3 ligase) targets RNF20 for proteasomal degradation, thereby regulating downstream H2B monoubiquitination and H3K4/H3K79 trimethylation; Smurf2 and RNF20 co-localize at γ-H2AX foci of DSBs.\",\n      \"method\": \"Genetic ablation (Smurf2 knockout mice), protein degradation assays, co-localization by immunofluorescence, western blot of histone marks\",\n      \"journal\": \"Nature medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo knockout model, multiple histone mark readouts, co-localization, and degradation assays across mouse and human cells\",\n      \"pmids\": [\"22231558\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"RNF20 (BRE1A/Rnf20) deficiency causes replication-associated DSBs through formation of aberrant R-loops (RNA:DNA hybrids), leading to specific genomic rearrangements, breakage-fusion-bridge cycles, and chromosomal instability; this is compounded by a previously known HR repair defect.\",\n      \"method\": \"Bre1-deficient cell lines, genomic instability tracking, R-loop detection, DNA content analysis, cancer tissue comparison\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — single lab, multiple cellular genomic phenotype assays, R-loop detection, functional epistasis\",\n      \"pmids\": [\"22354749\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"RNF20-catalyzed H2B monoubiquitylation (H2Bub1) increases during embryonic stem cell differentiation and is required for efficient ESC differentiation, particularly for transcriptional induction of relatively long genes; USP44 deubiquitinase negatively regulates H2Bub1 and its downregulation during differentiation contributes to the H2Bub1 increase.\",\n      \"method\": \"ESC differentiation assays, RNAi knockdown of RNF20 and USP44, ChIP, gene expression analysis\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (KD, ChIP, expression profiling), reciprocal regulation demonstrated for writer (RNF20) and eraser (USP44)\",\n      \"pmids\": [\"22681888\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CDC73 (parafibromin) tumor suppressor interacts with RNF20 and RNF40 at discrete residues, and is required for maintenance of H2B K120 monoubiquitination; loss of nuclear CDC73 or CDC73 mutations (as in parathyroid carcinoma) significantly reduces H2Bub1 without affecting H3K4me3.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, siRNA knockdown, western blot of histone marks in parathyroid tumor tissue\",\n      \"journal\": \"Human molecular genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — yeast two-hybrid, co-IP, knockdown functional assays, validated in primary tumors\",\n      \"pmids\": [\"22021426\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"Histone chaperone FACT (SUPT16H subunit) directly binds RNF20 in vivo; mutation of the RNF20 RING-finger domain abolishes FACT-RNF20 interaction and RNF20 accumulation at DSB sites; SUPT16H depletion impairs RNF20-mediated H2B ubiquitylation and SNF2h recruitment at DSBs, causing HR repair defects; this interaction is PAF1-independent for DNA damage contexts.\",\n      \"method\": \"Co-immunoprecipitation, RING-domain mutagenesis, RNAi knockdown, laser-induced DSB foci analysis, HR repair assays, radiation sensitivity\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — mutagenesis of RING domain, co-IP, functional HR assay, multiple epistasis experiments in one study\",\n      \"pmids\": [\"24357716\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"RNF20 (hBRE1) is targeted by adenovirus E1A, which repurposes RNF20 from a ubiquitin ligase into a scaffold that recruits hPaf1 for transcriptional activation of viral early genes, simultaneously blocking H2B monoubiquitination required for the interferon response.\",\n      \"method\": \"Co-immunoprecipitation, transcription reporter assays, viral infection/replication assays, E1A interaction mapping\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP, functional transcription assays, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"23785282\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"RNF20 (histone H2B E3 ubiquitin ligase) is required for MLL-fusion-mediated leukemogenesis; H2B ubiquitination is enriched in the body of MLL-fusion target genes and correlates with H3K79 methylation; RNF20 is required to maintain local H3K79 methylation by DOT1L at Hoxa9 and Meis1, supporting MLL-fusion oncogenic transcriptional program.\",\n      \"method\": \"shRNA knockdown in leukemia models (in vitro and in vivo), ChIP-seq (H2Bub, H3K79me2), gene expression analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP-seq, in vivo knockdown model, epistatic relationship between H2Bub and H3K79me2 demonstrated\",\n      \"pmids\": [\"23412334\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Arsenite directly binds to the cysteine residues in the RING finger domains of RNF20 and RNF40 in vitro and in cells, inhibiting H2B ubiquitination and impairing BRCA1 and RAD51 recruitment to DSB sites, thereby compromising DNA DSB repair and rendering cells sensitive to radiomimetic agents.\",\n      \"method\": \"In vitro binding assays, cellular H2B ubiquitination assays, laser-induced DSB foci analysis, DNA repair assays, cell viability assays\",\n      \"journal\": \"Journal of the American Chemical Society\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — direct in vitro binding, functional ubiquitination assay, DSB recruitment assays, multiple cell lines\",\n      \"pmids\": [\"25170678\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"RNF20/RNF40 complex interacts with motor protein Eg5 during mitosis, monoubiquitinates and stabilizes Eg5; loss of RNF20/40 causes spindle assembly defects, cell cycle arrest, and apoptosis; this spindle assembly role is distinct from the transcriptional H2B ubiquitination function.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, spindle assembly imaging, cell cycle analysis, in vivo tumor suppression assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — co-IP, ubiquitination assay, functional spindle assembly phenotype, in vivo validation across multiple methods\",\n      \"pmids\": [\"27557628\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"RNF20/RNF40 downregulation favors recruitment of p65-containing NF-κB dimers over repressive p50 homodimers and decreases H3K9me3 on a subset of NF-κB target genes, augmenting their transcription; RNF20+/- mice show predisposition to colonic inflammation and inflammation-associated colorectal cancer with excessive MDSCs.\",\n      \"method\": \"RNAi knockdown, ChIP, NF-κB dimer recruitment analysis, RNF20 heterozygous mouse model, colitis and cancer models\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo mouse model, ChIP-based mechanistic analysis, multiple orthogonal experimental approaches across mouse and human tissues\",\n      \"pmids\": [\"26854224\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Crystal structure of the RNF20 RING domain reveals it forms a homodimer and specifically interacts with the Ube2B~Ub conjugate; key E3-E2 and E3-ubiquitin interface contacts were identified by mutagenesis; RING domains of RNF20 and RNF40 form a stable active heterodimer; structural model of RNF20-Ube2B~Ub complex generated.\",\n      \"method\": \"X-ray crystallography of RNF20 RING domain, site-directed mutagenesis of E3-E2 and E3-ubiquitin interfaces, in vitro ubiquitination assays\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with functional mutagenesis validation and in vitro ubiquitination assays\",\n      \"pmids\": [\"27569044\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"RNF20/40, in conjunction with its cognate E2 enzyme RAD6, monoubiquitylates lysine 381 of eEF1BδL (a heat shock transcription factor); this monoubiquitylation increases eEF1BδL accumulation and potentiates recruitment of p-TEFb to promoters of heat shock-responsive genes, enhancing their transcription; cooperative physical interactions among eEF1BδL, RNF20/40, and HSF1 synergistically promote heat shock gene expression.\",\n      \"method\": \"Biochemically defined in vitro ubiquitylation assays with purified factors, cell-based analyses, mass spectrometry identification of ubiquitylation site, ChIP, gene expression assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro reconstitution with purified factors, site-specific mutagenesis to identify K381, ChIP, multiple orthogonal methods\",\n      \"pmids\": [\"30649429\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"The RNF20/40 complex associates with p53 on chromatin; p53 mediates recruitment of RNF20/40 to p21 and PUMA gene loci; RNF20/40-dependent H2B ubiquitination (ubH2B) is required for transcription and mRNA splicing of p21 and PUMA; ubH2B is recognized by the ubiquitin-binding motif of pre-mRNA splicing factor PRPF8 (a spliceosome subunit), which is required for mRNA maturation.\",\n      \"method\": \"Protein affinity purification, co-immunoprecipitation, ChIP, RNAi knockdown, mRNA splicing assays, ubiquitin-binding domain interaction studies\",\n      \"journal\": \"Journal of molecular cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — affinity purification, ChIP, functional splicing assays, identification of PRPF8 as ubH2B reader, multiple orthogonal methods\",\n      \"pmids\": [\"31152661\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"RNF20 promotes proteasomal degradation of nuclear corepressor 1 (NCoR1) via ubiquitination, which relieves NCoR1-mediated repression and stimulates PPARγ transcriptional activity in adipocytes, thereby promoting adipogenesis; Rnf20 conditional knockout mice show reduced fat mass and altered PPARγ target gene expression.\",\n      \"method\": \"Conditional knockout mouse model, quantitative proteomics, co-immunoprecipitation, ubiquitination/proteasomal degradation assays\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo knockout, proteomics identification of NCoR1 as substrate, degradation assays, multiple methods\",\n      \"pmids\": [\"31604693\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The SARS-CoV-2 main protease 3Clpro cleaves RNF20 at a conserved Gln521 in a catalytically dependent manner (catalytic mutant 3ClproC145A does not cleave); cleavage of RNF20 prevents RNF20-mediated degradation of SREBP1, promoting viral replication; RNAi depletion of RNF20 or RNF40 significantly enhances SARS-CoV-2 replication.\",\n      \"method\": \"Protease cleavage assays with catalytic mutant control, RNAi knockdown, SARS-CoV-2 replication assays, SREBP1 protein stability assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — catalytic mutant control establishing mechanism, substrate identification, viral replication functional readout\",\n      \"pmids\": [\"34452991\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"RNF20 acts as a tumor suppressor in clear cell renal cell carcinoma by promoting degradation of SREBP1c, which represses lipogenesis and cell cycle progression through the PTTG1 oncogene (a novel SREBP1c target); RNF20 overexpression reduces tumor growth and lipid storage in xenografts.\",\n      \"method\": \"RNF20 overexpression and knockdown, cell proliferation assays, xenograft tumor models, lipogenesis assays, ChIP for SREBP1c at PTTG1 promoter\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — single lab, in vivo xenograft, functional mechanistic pathway from RNF20 to SREBP1c to PTTG1\",\n      \"pmids\": [\"28827316\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"RNF20 (hBRE1) and MEN1 are recruited to the IRF1 gene locus in both uninduced and IFNγ-induced states; RNAi-mediated depletion of RNF20 lowers H2Bub and H3K4me3 at IRF1, but unexpectedly upregulates IFNγ-induced IRF1 transcription, indicating that RNF20 can be repressive of inducible transcription at this locus.\",\n      \"method\": \"ChIP, RNAi knockdown, RT-PCR gene expression analysis, chemical inhibition of H3K4 methylation\",\n      \"journal\": \"Epigenetics & chromatin\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — ChIP and RNAi knockdown; single lab; unexpected negative regulatory finding at specific locus\",\n      \"pmids\": [\"20825659\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"RNF20 promotes polyubiquitination and proteasome-dependent degradation of transcription factor AP-2α; RNF20 co-localizes and interacts with AP-2α in preadipocytes; RNF20 overexpression rescues C/EBPα expression suppressed by AP-2α.\",\n      \"method\": \"Co-immunoprecipitation, mass spectrometry, ubiquitination assays, proteasome inhibition, co-localization microscopy, C/EBPα expression rescue assays\",\n      \"journal\": \"Acta biochimica et biophysica Sinica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP, ubiquitination assay, functional rescue; single lab\",\n      \"pmids\": [\"24374663\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Fbxl19, a CxxC domain-containing protein, promotes H2Bub1 at CpG island-containing gene promoters by directly interacting with Rnf20; Fbxl19 preferentially occupies CpG island-containing promoters genome-wide and its chromosomal binding is required for H2Bub1 at those targets; Fbxl19 is critical for proper ESC differentiation in collaboration with Rnf20.\",\n      \"method\": \"Co-immunoprecipitation, genome-wide ChIP-seq (Fbxl19, H2Bub1), RNAi knockdown of Fbxl19, ESC differentiation assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, genome-wide ChIP-seq, functional differentiation assays; single lab\",\n      \"pmids\": [\"28453857\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"RNF20 and RNF40 are required for DSB repair leading to both homologous recombination and class switch recombination (NHEJ-driven) in mouse B cells; DSBs induce a global increase in H2Bub but not H3K4me3 or H3K79me2; H2AX phosphorylation is dispensable for H2Bub; ATM and ATR jointly regulate IR-induced H2Bub.\",\n      \"method\": \"RNAi knockdown in mouse B cells, class switch recombination assays, HR assays, western blot of histone marks, kinase inhibitor treatments\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — functional DSB repair assays (HR and CSR), epistasis with ATM/ATR, multiple orthogonal methods\",\n      \"pmids\": [\"30692271\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Cryo-EM structures of chemically-trapped transient ubiquitin transfer complexes reveal that RNF40 (and yeast Bre1) directly bind nucleosomal DNA; the E3/E2/nucleosome interaction pattern is conserved from yeast to humans; Bre1 RING domain uses a non-canonical non-hydrophobic contact with Rad6 to position E2 directly above target H2B lysine; these structures explain site-specific H2B monoubiquitylation mechanism.\",\n      \"method\": \"Chemical trapping, cryo-electron microscopy structure determination, biochemical reconstitution, cancer mutation mapping\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structures with chemical trapping, reconstitution, multiple mechanistic insights from single high-quality study\",\n      \"pmids\": [\"37633270\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RPA (single-strand DNA-binding factor) interacts with RNF20 mainly in S and G2/M phases and recruits RNF20 to mitotic centromeres in a centromeric R-loop-dependent manner and to chromosomal breaks upon DNA damage; the RPA-RNF20 pathway promotes local H2Bub, H3K4 dimethylation, and SNF2H recruitment, ensuring proper Aurora B kinase activation at centromeres and efficient loading of repair proteins at DSBs.\",\n      \"method\": \"Co-immunoprecipitation, ChIP, cell cycle analysis, mitotic phenotype assays (lagging chromosomes, chromosome bridges), HR repair assays, R-loop disruption experiments\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, ChIP, epistasis with R-loops, functional readouts (mitosis and HR), multiple orthogonal approaches\",\n      \"pmids\": [\"37155876\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Yeast Bre1 and human RNF20 function as recombination mediators independently of their E3 ligase activity: they interact with Rad51, direct Rad51 to ssDNA, facilitate Rad51-ssDNA filament assembly and strand exchange in vitro, and interact with Srs2/FBH1 helicases to counteract their disruption of Rad51 filaments; this mediator function is additive with Rad52 (yeast) or BRCA2 (human).\",\n      \"method\": \"In vitro Rad51 filament assembly and strand exchange assays, co-immunoprecipitation, ligase-dead mutant analysis, genetic epistasis with rad52/BRCA2, HR assays in cells\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — in vitro reconstitution of filament assembly and strand exchange, ligase-dead mutants, genetic epistasis, ortholog conservation\",\n      \"pmids\": [\"37230987\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"RNF20 and RNF40 form a complex that monoubiquitinates H2B on lysine 120; in vivo CRISPR screen in mouse cardiomyocytes identified RNF20/40-mediated H2Bub as controlling dynamic changes in gene expression required for cardiomyocyte maturation.\",\n      \"method\": \"In vivo somatic Cas9 mutagenesis (forward genetic screen in mice), cardiomyocyte maturation phenotyping, gene expression analysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo CRISPR screen, functional validation of maturation phenotype; single lab\",\n      \"pmids\": [\"34290256\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RNF20 (spinal) mediates H2B monoubiquitination in dorsal horn neurons following spinal nerve ligation; RNF20-dependent H2Bub facilitates phosphorylated RNA polymerase II-dependent mGluR5 transcription; focal knockdown of spinal RNF20 reverses neuropathic allodynia and reduces mGluR5 expression; TNF-α activates this RNF20/H2Bub/RNAPII transcriptional axis.\",\n      \"method\": \"Spinal nerve ligation rat model, RNAi knockdown in vivo, ChIP of H2Bub and phospho-RNAPII at mGluR5 promoter, behavioral allodynia assays, neutralizing antibody experiments\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — in vivo ChIP, RNAi functional rescue of behavior, TNF-α neutralization; single lab\",\n      \"pmids\": [\"30201771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF20 stabilizes LSD1 via K29-linked ubiquitination in a CDK9-phosphorylation-dependent manner; this CDK9-RNF20-LSD1 axis is required for H3K4 demethylation, repression of endogenous retroviruses, and suppression of interferon response, thereby contributing to epigenetic immunosuppression; loss of RNF20 sensitizes cancer cells to anti-PD-1 immunotherapy in vivo.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays (K29-specific), CDK9 inhibition/phosphorylation analysis, endogenous retrovirus expression, in vivo anti-PD-1 rescue experiment, breast cancer specimen correlation\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — co-IP, site-specific ubiquitination, CDK9 epistasis, in vivo immunotherapy rescue, multiple orthogonal methods\",\n      \"pmids\": [\"38315842\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF20 controls Rbx1 expression, thereby regulating activity of the VHL ubiquitin ligase complex and HIF1α protein levels; Rnf20 haploinsufficiency increases HIF1α levels and drives metabolic rewiring and EMT through HIF1α-mediated RNAPII promoter-proximal pause release independently of H2Bub1; this links the DNA damage response and metabolic reprogramming in lung cancer.\",\n      \"method\": \"Rnf20 haploinsufficiency mouse lung tumor model, genetic ablation, gene expression analysis, metabolomics, HIF1α protein stability assays, Rbx1/VHL epistasis experiments\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo mouse model, epistasis experiments, two distinct RNF20-HIF1α mechanisms; single lab\",\n      \"pmids\": [\"40436847\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"HIF-1 recruits FACT and RNF20/40 to hypoxia response elements (HREs); FACT and RNF20/40 stabilize HIF-1 binding and each other at HREs; hypoxia induces H2B monoubiquitination at HIF-1 target genes in an HIF-1-dependent manner; knockdown of FACT or RNF20/40 decreases transcription initiation and elongation at HIF-1 target genes.\",\n      \"method\": \"RNAi knockdown of FACT and RNF20/40, ChIP at HREs, H2Bub ChIP under hypoxia, transcription initiation/elongation analysis in breast cancer cells\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — ChIP, functional knockdowns, cooperative recruitment mechanism; single lab\",\n      \"pmids\": [\"38517892\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF20 mediates NCoR1 protein degradation to activate PPARγ in iWAT for beige fat differentiation upon prolonged cold stimuli; in BAT, cold stimulus downregulates RNF20, which elevates GABPα protein stability (RNF20 controls GABPα stability), promoting thermogenic gene expression; thus RNF20 uses fat depot-specific substrates (GABPα in BAT, NCoR1 in iWAT) to orchestrate adipose thermogenesis.\",\n      \"method\": \"Fat depot-specific Rnf20 knockout mice, cold exposure experiments, protein stability assays for GABPα and NCoR1, gene expression analysis, ubiquitination assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo fat-specific knockout models, two distinct substrates identified with functional readouts, multiple orthogonal methods\",\n      \"pmids\": [\"38296968\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF20 regulates oocyte meiotic spindle assembly by localizing to centromeres and spindle poles and recruiting tropomyosin 3 (TPM3) via its coiled-coil motif (not its E3 ligase activity); RNF20-depleted oocytes show abnormal spindle organization and chromosome misalignment; the RNF20-TPM3 interaction is essential for acentrosomal meiotic spindle assembly.\",\n      \"method\": \"RNF20 depletion in mouse oocytes, E3 ligase-dead mutant analysis, co-immunoprecipitation of RNF20-TPM3, immunofluorescence of spindle/centromere localization, coiled-coil domain mapping\",\n      \"journal\": \"Advanced science\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ligase-dead mutant establishing non-enzymatic mechanism, co-IP for substrate (TPM3), functional spindle phenotype, localization\",\n      \"pmids\": [\"38240347\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"RNF20 mediates RNA polymerase II promoter-proximal pausing at highly paused endothelial genes involved in VEGFA signaling; RNF20 binds Notch1 to promote H2B monoubiquitination at Notch target genes and Notch-dependent gene expression; loss of Rnf20 leads to uncontrolled tip cell specification; RNF20 restricts ERG-dependent Pol II pause release at highly paused genes.\",\n      \"method\": \"Rnf20 knockout in endothelial cells, RNA Pol II ChIP-seq (pause analysis), Notch1 co-immunoprecipitation, H2Bub ChIP, angiogenesis/tip cell phenotype assays\",\n      \"journal\": \"Nature cardiovascular research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP-seq for Pol II pausing, co-IP with Notch1, in vivo phenotype, multiple orthogonal methods\",\n      \"pmids\": [\"39322771\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"Iron deficiency-activated ferritinophagy induces RNF20 degradation through the autophagy-lysosomal pathway; RNF20 degradation negatively regulates H2B K120 monoubiquitination at promoters of myogenic markers MyoD and MyoG, inhibiting myogenic differentiation and muscle regeneration; overexpression of RNF20 or autophagy inhibition restores regeneration under iron-deficient conditions.\",\n      \"method\": \"Iron deficiency model in cells and mice, autophagy-lysosome pathway inhibition, RNF20 overexpression rescue, ChIP for H2Bub at myogenic promoters, conditional NCOA4 knockout in satellite cells\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple rescue experiments, ChIP at target promoters, in vivo conditional knockout, mechanistic pathway established\",\n      \"pmids\": [\"37976359\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RNF20 localizes to replicating sites and promotes H2Bub there; RNF20 knockdown leads to nucleolytic degradation of stalled replication forks (rescued by MRE11/DNA2 inhibition and depletion of fork remodelers SMARCAL1/HLTF/ZRANB3); RNF20 facilitates RAD51 and RAD51C loading at stalled forks, acting in the same pathway as RAD51/RAD51C-mediated fork protection; RNF20 RING domain and ATR-mediated phosphorylation are essential for its role in replication stress responses.\",\n      \"method\": \"RNF20 knockdown, DNA fiber assays (fork protection/restart), RING-domain mutant and phosphorylation-deficient mutant analysis, nuclease inhibition, fork remodeler co-depletion, RAD51/RAD51C loading assays\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — RING domain and phospho-mutants, fork protection assays, genetic epistasis with fork remodelers and nucleases, RAD51 loading assays\",\n      \"pmids\": [\"40495033\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"RNF20 and RNF40 regulate Vitamin D Receptor (VDR) gene and VDR target gene transcription through H3K4me3 occupancy dependent on H2Bub1; conditional intestinal deletion of Rnf20 or Rnf40 in mice causes spontaneous colorectal inflammation; this mechanism is linked to IBD pathogenesis.\",\n      \"method\": \"Conditional intestinal Rnf20/Rnf40 knockout mice, ChIP-seq for H2Bub1 and H3K4me3 at VDR locus, mRNA-seq, IBD patient ChIP-seq\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO in vivo, ChIP-seq, confirmation in human patient specimens, multiple orthogonal methods\",\n      \"pmids\": [\"34088983\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"KSHV RTA protein interacts with the cellular RNF20/40 E3 ubiquitin ligase complex, and this interaction is necessary for RTA-induced KSHV lytic cycle reactivation.\",\n      \"method\": \"Proteomics identification of RTA-RNF20/40 interaction, functional lytic reactivation assays, co-immunoprecipitation validation\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — proteomics plus functional lytic assays; single lab\",\n      \"pmids\": [\"37888983\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"RNF20 interacts with ZSCAN4 protein and negatively regulates ZSCAN4 stability; RNF20 depletion does not affect ZSCAN4 transcription but increases ZSCAN4 protein levels and stabilizes its half-life, indicating RNF20 promotes ZSCAN4 degradation at the protein level.\",\n      \"method\": \"Co-immunoprecipitation, protein half-life assays, RNAi knockdown of RNF20, western blot, RT-PCR\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP, protein stability assays, RNAi; single lab, single study\",\n      \"pmids\": [\"29477841\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"In breast cancer cells, Snail interacts with RNF20 and G9a; RNF20 and H3K9me2 (G9a mark) are enriched on the E-cadherin promoter in a Snail-dependent manner; RNF20-mediated H2BK120 monoubiquitination contributes to E-cadherin repression and EMT induction.\",\n      \"method\": \"Co-immunoprecipitation (Snail-RNF20-G9a), ChIP at E-cadherin promoter, RNF20 knockdown/overexpression, migration/invasion/tumorsphere assays\",\n      \"journal\": \"Frontiers in oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP, ChIP, functional cancer cell assays; single lab\",\n      \"pmids\": [\"33364200\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RNF20 promotes STAT3 ubiquitination and degradation; RNF20 is a target gene of exosomal miR-let-7b-5p (confirmed by luciferase reporter assay); ectopic RNF20 expression attenuates insulin resistance by downregulating STAT3 protein expression via ubiquitination-mediated degradation.\",\n      \"method\": \"Luciferase reporter assay (RNF20 as let-7b-5p target), IP/western blot for STAT3 ubiquitination, RNF20 overexpression rescue experiments\",\n      \"journal\": \"Diabetes, metabolic syndrome and obesity\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single co-IP for ubiquitination, luciferase reporter, single lab, limited mechanistic depth\",\n      \"pmids\": [\"37842335\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Crystal structure of Bre1-Lge1 complex and AlphaFold model of RNF20/RNF40-WAC reveal extensive interaction interfaces; the Bre1-Lge1 and RNF20/RNF40-WAC interfaces share structural homology but use completely different electrostatic interactions encoding binding specificity; these interactions are critical for the H2BUb1 reaction and processes it regulates.\",\n      \"method\": \"X-ray crystallography of Bre1-Lge1, AlphaFold modeling of RNF20/RNF40-WAC, interface mutagenesis, in vitro H2BUb1 assays, in vivo functional assays\",\n      \"journal\": \"Nucleic acids research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with mutagenesis and functional ubiquitination validation; single lab but rigorous structural approach\",\n      \"pmids\": [\"41533567\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Cryo-EM structure of chemically-trapped RNF20/RNF40-RAD6A-Ub-H2BS112GlcNAc nucleosome complex shows that H2BS112GlcNAc interacts with E2 enzyme RAD6A (not RNF20/RNF40); GlcNAcylation allosterically stimulates ubiquitin transfer from RAD6A~Ub to H2B K120 by enhancing the nucleophilicity of H2B K120; the C2 N-acetyl group and β-configuration of C1 are essential for this activation.\",\n      \"method\": \"Chemical synthesis of H2BS112GlcNAc-modified nucleosomes, chemical trapping, cryo-EM structure determination, mutagenesis, kinetics analysis, structure-activity relationship\",\n      \"journal\": \"Nature chemical biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM with chemical trapping, kinetics, mutagenesis, synthetic nucleosome chemistry, mechanistic detail of PTM crosstalk\",\n      \"pmids\": [\"41495224\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"RNF20 is required for male fertility through regulation of H2BK120 ubiquitination in Sertoli cells; Sertoli cell-specific Rnf20 knockout mice are infertile due to spermatogenic failure mimicking Sertoli cell-only syndrome; RNF20 deficiency impairs transcription elongation of Cldn11 (encoding tight junction component claudin-11), disrupts cell adhesion and seminiferous tubule organization, and leads to apoptosis of spermatogonia and spermatocytes.\",\n      \"method\": \"Conditional Sertoli cell-specific Rnf20 knockout mice (Amh-Cre), histology, H2Bub1 western blot, Cldn11 expression analysis, ChIP for H2Bub at Cldn11 locus\",\n      \"journal\": \"Cell & bioscience\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional KO in vivo with specific target gene (Cldn11) and ChIP validation, clear phenotypic readout\",\n      \"pmids\": [\"37024990\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"In β-cells, Rnf20 forms complexes with the Isl1 transcription factor; conditional Rnf20 knockout in adult β-cells causes severe hyperglycemia, loss of H2B monoubiquitination, dysregulation of glucose-stimulated insulin secretion, and disruption of β-cell identity gene expression; comparative analysis shows Isl1 and Rnf20 loss yield similar β-cell regulome changes; Isl1::Rnf20 complexes regulate insulin expression and secretion in human tissues.\",\n      \"method\": \"β-cell-specific conditional Rnf20 knockout (tamoxifen-inducible), co-immunoprecipitation of Isl1-Rnf20, H2Bub1 western blot, transcriptomic analysis, glucose tolerance tests, insulin secretion assays\",\n      \"journal\": \"Diabetes\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — conditional in vivo KO, co-IP for Isl1-Rnf20 complex, functional insulin secretion assays, validated in human tissues\",\n      \"pmids\": [\"40743566\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RNF20 dynamically translocates between nucleus and cytoplasm in response to RNA virus infection (export via CRM1); in resting and early infection states, nuclear RNF20 maintains basal and inducible transcription of RIG-I and MDA5; upon late viral infection, cytoplasmic RNF20 recognizes degron motifs of RIG-I and MDA5 via its coiled-coil domain and catalyzes K27-linked ubiquitination and degradation of both sensors, preventing excessive antiviral signaling.\",\n      \"method\": \"RNF20 overexpression/knockout (in vitro and in vivo), co-immunoprecipitation, ubiquitination assays (K27-specific), CRM1 export inhibition, transcriptome sequencing, viral infection assays\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — co-IP, K27-specific ubiquitination, CRM1 inhibition, nucleocytoplasmic localization; single lab\",\n      \"pmids\": [\"41712649\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The natural compound epoxymicheliolide (ECL) covalently modifies H2B lysine 46 (K46) and recruits E3 ubiquitin ligase RNF20 to promote H2Bub1 at K120; ECL-mediated H2Bub1 disrupts AP-1 recruitment to proinflammatory gene promoters, inhibiting microglia-mediated neuroinflammation.\",\n      \"method\": \"Biochemical binding assays, ChIP-seq, transcriptomics, site-specific mutagenesis of H2B K46, in vitro and in vivo neuroinflammation models\",\n      \"journal\": \"Pharmacological research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — ChIP-seq, mutagenesis, recruitment assays; indirect evidence for RNF20 recruitment via modified H2B\",\n      \"pmids\": [\"35074526\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Bre1a (RNF20 mouse ortholog) regulates neural precursor cell (NPC) differentiation and cell cycle length; knockdown of Bre1a in NPCs lengthens cell cycle through upregulation of p57kip2 and downregulation of Cdk2, and increases Hes5 expression via Fezf1 and Fezf2 to suppress NPC differentiation.\",\n      \"method\": \"RNAi knockdown in embryonic mouse NPCs, cell cycle analysis, RT-PCR for p57kip2/Cdk2/Hes5/Fezf1/Fezf2, differentiation assays\",\n      \"journal\": \"The Journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — RNAi knockdown with multiple molecular readouts; single lab, mechanistic pathway partially defined\",\n      \"pmids\": [\"24553946\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"RNF20/RNF40, SET1/COMPASS, and DOT1L are dispensable for transcription restart after DNA damage repair (negative finding); levels of H2B-K120Ub and H3K4me3 do not correlate with transcription restoration following DNA damage; PAF1C promotes transcription restart independently of H2B-K120Ub deposition.\",\n      \"method\": \"RNF20/RNF40 depletion, SET1/COMPASS and DOT1L inhibition, transcription restart assays after DNA damage, ChIP for H2Bub/H3K4me3\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional transcription restart assay with multiple negative controls; preprint, single lab, orthogonal inhibitor approaches\",\n      \"pmids\": [],\n      \"is_preprint\": true\n    }\n  ],\n  \"current_model\": \"RNF20, in complex with RNF40, functions as an E3 ubiquitin ligase that monoubiquitinates histone H2B at lysine 120 (H2Bub1) using the E2 enzyme RAD6/Ube2B; structural studies show RNF40 directly contacts nucleosomal DNA and RNF20 RING domain engages the Ube2B~Ub conjugate, with H2BS112 GlcNAcylation allosterically stimulating ubiquitin transfer through RAD6A; beyond transcriptional elongation and chromatin remodeling, RNF20 is recruited to DNA double-strand breaks (via RPA interaction in an R-loop-dependent manner), where H2Bub1 enables SNF2h-mediated chromatin relaxation, Rad51 filament loading, and homologous recombination repair; RNF20 also promotes Rad51 filament assembly and strand exchange independently of its ligase activity by acting as a recombination mediator and antagonizing Srs2/FBH1 helicases; at stalled replication forks, RNF20-mediated H2Bub (regulated by ATR phosphorylation) protects forks from MRE11/DNA2-dependent degradation; beyond histones, RNF20 ubiquitinates non-histone substrates including Eg5 (stabilization for spindle assembly), eEF1BδL (heat shock transcription), NCoR1 and GABPα (adipose thermogenesis), SREBP1c, LSD1 (via CDK9-dependent phosphorylation to drive epigenetic immunosuppression), AP-2α, PSMA7, STAT3, and ZSCAN4; RNF20 also recruits TPM3 to spindle poles via its coiled-coil domain independently of ligase activity for oocyte meiotic spindle assembly; the complex is regulated by Smurf2-mediated proteasomal degradation of RNF20 and by WAC (which bridges RNF20/40 to RNA Pol II), and undergoes SARS-CoV-2 Mpro cleavage at Gln521 to subvert antiviral immunity.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"RNF20, partnered with RNF40, is the catalytic E3 ubiquitin ligase that monoubiquitinates histone H2B on lysine 120 (H2Bub1) together with the E2 enzyme RAD6/Ube2B, a chromatin mark that couples to downstream H3K4 and H3K79 methylation and drives transcriptional elongation and chromatin dynamics across diverse developmental and stress programs [#1, #12, #25]. Structurally, RNF20 and RNF40 RING domains form an active heterodimer in which RNF40 contacts nucleosomal DNA and a non-canonical RING surface positions RAD6 directly above H2B K120, and this transfer is allosterically stimulated by H2BS112 GlcNAcylation acting through RAD6A [#22, #41]. Recruitment to chromatin is gated by accessory factors: WAC bridges the complex to RNA polymerase II at active genes, while a distinct binding interface engages the heterodimer for H2Bub1 catalysis [#1, #40]. Through H2Bub1, RNF20 controls transcription-dependent processes including embryonic stem cell differentiation of long genes, MLL-fusion leukemogenesis via DOT1L-dependent H3K79 methylation, cardiomyocyte and myogenic differentiation, spermatogenesis through Cldn11 elongation, beta-cell identity in complex with Isl1, and endothelial Notch- and pause-regulated angiogenic programs [#4, #8, #25, #33, #42, #43, #32]. Independently of transcription, RNF20 is recruited to DNA double-strand breaks—via FACT and via an RPA/R-loop-dependent route—where H2Bub1 enables SNF2h-mediated chromatin relaxation, end resection, and RAD51/BRCA1 loading for homologous recombination [#0, #6, #23]; it also acts as a non-enzymatic recombination mediator that loads RAD51 on ssDNA and antagonizes Srs2/FBH1 helicases, and protects ATR-regulated stalled forks from MRE11/DNA2 nucleolytic degradation [#24, #34]. Beyond histones, RNF20 ubiquitinates numerous non-histone substrates to control cell physiology, including Eg5 stabilization for spindle assembly, and degradation of NCoR1, GABP\\u03b1, SREBP1c, and LSD1 to govern adipose thermogenesis, lipogenesis, and epigenetic immunosuppression [#10, #30, #17, #27]. The complex is itself regulated by Smurf2-mediated and ferritinophagy-driven degradation of RNF20, and is subverted by viral pathogens through SARS-CoV-2 Mpro cleavage at Gln521 and adenovirus E1A repurposing [#2, #33, #16, #7].\",\n  \"teleology\": [\n    {\n      \"year\": 2010,\n      \"claim\": \"Established that RNF20-deposited H2Bub1 and H3K4me3 are not uniformly activating, since RNF20 loss paradoxically enhanced inducible IRF1 transcription, revealing context-dependent repressive output.\",\n      \"evidence\": \"ChIP and RNAi knockdown at the IRF1 locus with H3K4 methylation inhibition\",\n      \"pmids\": [\"20825659\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of locus-specific repression versus activation unresolved\", \"Single locus, single lab\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Defined how the RNF20/40 complex is targeted to transcription, showing WAC physically bridges the ligase to elongating RNA Pol II and is required for H2B ubiquitination.\",\n      \"evidence\": \"Affinity purification, reciprocal co-IP, RNAi, and H2B ubiquitination assays\",\n      \"pmids\": [\"21329877\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of WAC bridging not defined here\", \"Whether other recruiters operate at distinct gene classes unknown\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Extended RNF20 function beyond transcription to genome maintenance, demonstrating it is recruited to DSBs independently of H2AX and that H2Bub1 drives SNF2h-dependent chromatin relaxation enabling resection and RAD51/BRCA1 loading for HR.\",\n      \"evidence\": \"RNAi, H2B K120R mutant, foci imaging, HR assays, and chromatin relaxation rescue\",\n      \"pmids\": [\"21362548\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Initial DSB-recruitment mechanism not yet identified\", \"Did not distinguish ligase-dependent from independent contributions\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified CDC73/parafibromin as a complex partner required to maintain H2Bub1, linking the ligase to a tumor-suppressor pathway.\",\n      \"evidence\": \"Yeast two-hybrid, co-IP, siRNA, and histone-mark western blots in parathyroid tumors\",\n      \"pmids\": [\"22021426\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How CDC73 stimulates catalysis mechanistically unclear\", \"Selective effect on H2Bub1 but not H3K4me3 unexplained\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Showed RNF20 abundance is set by Smurf2-mediated proteasomal degradation, establishing upstream control over global H2Bub1 and downstream methylation.\",\n      \"evidence\": \"Smurf2 knockout mice, degradation assays, and co-localization at DSB foci\",\n      \"pmids\": [\"22231558\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signals triggering Smurf2-RNF20 turnover not defined\", \"Quantitative coupling to repair outcomes not measured\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Connected RNF20 loss to a source of endogenous damage, demonstrating its deficiency promotes aberrant R-loops that generate replication-associated DSBs and chromosomal instability.\",\n      \"evidence\": \"Bre1-deficient cells, R-loop detection, and genomic instability tracking\",\n      \"pmids\": [\"22354749\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct molecular link between RNF20 and R-loop resolution unclear\", \"Single lab\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Demonstrated a developmental requirement for RNF20-dependent H2Bub1 in inducing long genes during ESC differentiation, and identified USP44 as the opposing eraser.\",\n      \"evidence\": \"ESC differentiation, RNAi of RNF20 and USP44, ChIP, and expression profiling\",\n      \"pmids\": [\"22681888\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Why long genes are preferentially dependent not mechanistically resolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Identified FACT (SUPT16H) as a direct RING-dependent partner that recruits RNF20 to DSBs in a PAF1-independent manner, separating DNA-damage recruitment from transcriptional recruitment.\",\n      \"evidence\": \"Co-IP, RING-domain mutagenesis, laser-DSB foci, and HR repair assays\",\n      \"pmids\": [\"24357716\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How FACT itself is targeted to breaks not addressed\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Placed RNF20-mediated H2Bub1 within oncogenic transcription, showing it is required for DOT1L-dependent H3K79 methylation and the MLL-fusion leukemogenic program.\",\n      \"evidence\": \"shRNA in leukemia models in vivo, ChIP-seq for H2Bub and H3K79me2, expression analysis\",\n      \"pmids\": [\"23412334\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct enzymatic coupling between H2Bub1 and DOT1L at these loci not reconstituted\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Revealed viral subversion, with adenovirus E1A converting RNF20 from a ligase into a hPaf1-recruiting scaffold that activates viral genes while blocking interferon-linked H2Bub1.\",\n      \"evidence\": \"Co-IP, transcription reporters, and viral replication assays with E1A interaction mapping\",\n      \"pmids\": [\"23785282\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Structural basis of scaffold conversion not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Expanded the substrate repertoire to non-histone targets, showing RNF20 polyubiquitinates AP-2\\u03b1 for degradation to relieve C/EBP\\u03b1 repression in preadipocytes.\",\n      \"evidence\": \"Co-IP, MS, ubiquitination assays, proteasome inhibition, and rescue assays\",\n      \"pmids\": [\"24374663\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ubiquitin linkage type not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Provided chemical-biology evidence that RING-cysteine targeting inactivates the ligase, as arsenite binds RNF20/40 RING cysteines to block H2Bub1 and DSB repair.\",\n      \"evidence\": \"In vitro binding, cellular ubiquitination assays, and laser-DSB foci analysis\",\n      \"pmids\": [\"25170678\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Selectivity over other RING ligases in cells not fully mapped\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Showed an ortholog requirement in neural development, with Bre1a regulating NPC cell-cycle length and differentiation via p57kip2, Cdk2, and Hes5/Fezf pathways.\",\n      \"evidence\": \"RNAi in mouse NPCs with cell-cycle and expression readouts\",\n      \"pmids\": [\"24553946\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether effects are H2Bub1-dependent not established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Solved the RNF20 RING-domain crystal structure, defining the homodimer, the active RNF20/RNF40 heterodimer, and the E3-E2-ubiquitin interface required for catalysis.\",\n      \"evidence\": \"X-ray crystallography with interface mutagenesis and in vitro ubiquitination assays\",\n      \"pmids\": [\"27569044\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Nucleosome engagement not captured in this structure\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Established a ligase-dependent mitotic role distinct from transcription, with RNF20/40 monoubiquitinating and stabilizing Eg5 for spindle assembly.\",\n      \"evidence\": \"Co-IP, ubiquitination assays, spindle imaging, and in vivo tumor suppression\",\n      \"pmids\": [\"27557628\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitin linkage mediating stabilization not detailed\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Linked RNF20 loss to inflammation-driven cancer, showing it restrains NF-\\u03baB p65 recruitment and maintains H3K9me3 at target genes, with RNF20+/- mice predisposed to colorectal cancer.\",\n      \"evidence\": \"RNAi, ChIP, NF-\\u03baB dimer analysis, and RNF20 heterozygous mouse colitis/cancer models\",\n      \"pmids\": [\"26854224\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How H2Bub1 controls H3K9me3 at these loci not mechanistically resolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined a tumor-suppressive metabolic axis in renal carcinoma, with RNF20 degrading SREBP1c to repress lipogenesis and the PTTG1 oncogene.\",\n      \"evidence\": \"Overexpression/knockdown, xenografts, lipogenesis assays, and SREBP1c ChIP at PTTG1\",\n      \"pmids\": [\"28827316\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct SREBP1c ubiquitination biochemistry not shown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Identified Fbxl19 as a CxxC-domain recruiter directing RNF20 to CpG-island promoters genome-wide for H2Bub1 and ESC differentiation.\",\n      \"evidence\": \"Co-IP, ChIP-seq of Fbxl19 and H2Bub1, RNAi, and differentiation assays\",\n      \"pmids\": [\"28453857\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"How Fbxl19 and WAC recruitment routes are coordinated unknown\", \"Single lab\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Added ZSCAN4 as a degradation substrate, showing RNF20 controls ZSCAN4 protein stability without affecting its transcription.\",\n      \"evidence\": \"Co-IP, protein half-life assays, and RNAi\",\n      \"pmids\": [\"29477841\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Ligase-dependence and linkage type not established\", \"Single lab\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Demonstrated an in vivo neuronal transcriptional role, with spinal RNF20-dependent H2Bub1 driving phospho-Pol II-dependent mGluR5 transcription downstream of TNF-\\u03b1 in neuropathic pain.\",\n      \"evidence\": \"Nerve-ligation rat model, in vivo RNAi, ChIP at mGluR5, and behavioral assays\",\n      \"pmids\": [\"30201771\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Upstream signal coupling TNF-\\u03b1 to RNF20 not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Showed RNF20/40-RAD6 ubiquitinates non-histone eEF1B\\u03b4L at K381 to potentiate p-TEFb recruitment and heat-shock gene transcription with HSF1.\",\n      \"evidence\": \"In vitro reconstitution, MS site mapping, ChIP, and expression assays\",\n      \"pmids\": [\"30649429\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether eEF1B\\u03b4L ubiquitination is mono- or poly-ubiquitin not fully specified\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected H2Bub1 to co-transcriptional RNA processing, showing p53-recruited RNF20/40 generates ubH2B at p21/PUMA that is read by the spliceosome subunit PRPF8 for mRNA maturation.\",\n      \"evidence\": \"Affinity purification, ChIP, RNAi, splicing assays, and ubiquitin-binding domain studies\",\n      \"pmids\": [\"31152661\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Generality of PRPF8 reading of ubH2B beyond these genes not defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established RNF20 as a regulator of adipogenesis via NCoR1 degradation that activates PPAR\\u03b3, validated in conditional knockout mice.\",\n      \"evidence\": \"Conditional KO mice, quantitative proteomics, co-IP, and degradation assays\",\n      \"pmids\": [\"31604693\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct NCoR1 ubiquitination linkage type not detailed\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Generalized the repair role across HR and NHEJ, showing RNF20/40 are required for both HR and class-switch recombination, with H2Bub1 induced via joint ATM/ATR control and independent of H2AX.\",\n      \"evidence\": \"RNAi in mouse B cells, CSR and HR assays, histone-mark blots, and kinase inhibitors\",\n      \"pmids\": [\"30692271\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How ATM/ATR signals converge on RNF20 catalysis not mapped\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstrated an in vivo developmental transcriptional requirement, with an unbiased CRISPR screen identifying RNF20/40-mediated H2Bub1 as controlling cardiomyocyte maturation gene programs.\",\n      \"evidence\": \"In vivo somatic Cas9 screen with cardiomyocyte maturation phenotyping\",\n      \"pmids\": [\"34290256\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct target genes not individually validated\", \"Single lab\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Linked RNF20/40-H2Bub1 to intestinal homeostasis through H3K4me3-dependent VDR transcription, with conditional intestinal knockouts causing colitis relevant to IBD.\",\n      \"evidence\": \"Conditional intestinal KO mice, ChIP-seq at VDR, mRNA-seq, and IBD patient ChIP-seq\",\n      \"pmids\": [\"34088983\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether VDR is the sole driver of the colitis phenotype unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Provided a chemical handle showing the natural compound epoxymicheliolide modifies H2B K46 to recruit RNF20 and induce H2Bub1, blocking AP-1-driven neuroinflammation.\",\n      \"evidence\": \"Binding assays, ChIP-seq, H2B K46 mutagenesis, and neuroinflammation models\",\n      \"pmids\": [\"35074526\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"RNF20 recruitment is inferred via modified H2B rather than direct\", \"Single lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Resolved the structural basis of site-specific catalysis, with cryo-EM showing RNF40 (and yeast Bre1) bind nucleosomal DNA and a non-canonical RING contact positions RAD6 above H2B K120, conserved yeast-to-human.\",\n      \"evidence\": \"Chemical trapping and cryo-EM of E3/E2/nucleosome complexes with reconstitution\",\n      \"pmids\": [\"37633270\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Dynamics of substrate selection during transfer not captured\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identified RPA as a cell-cycle-regulated, R-loop-dependent recruiter of RNF20 to centromeres and breaks, coupling H2Bub1 to Aurora B activation and repair-protein loading.\",\n      \"evidence\": \"Reciprocal co-IP, ChIP, mitotic phenotyping, HR assays, and R-loop disruption\",\n      \"pmids\": [\"37155876\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How RPA and FACT recruitment routes are partitioned not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Uncovered a ligase-independent recombination-mediator activity, with RNF20/Bre1 directly loading Rad51 on ssDNA, promoting strand exchange, and antagonizing Srs2/FBH1 additively with BRCA2/Rad52.\",\n      \"evidence\": \"In vitro filament assembly and strand-exchange assays, ligase-dead mutants, and epistasis\",\n      \"pmids\": [\"37230987\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How catalytic and mediator activities are coordinated in vivo unclear\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed RNF20 abundance is controlled by ferritinophagy, where iron-deficiency-driven autophagic degradation of RNF20 lowers H2Bub1 at MyoD/MyoG to block myogenic differentiation.\",\n      \"evidence\": \"Iron-deficiency models, autophagy inhibition, RNF20 overexpression rescue, ChIP, and conditional NCOA4 KO\",\n      \"pmids\": [\"37976359\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Degron/recognition basis for autophagic RNF20 turnover not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined a requirement in male fertility, with Sertoli-cell RNF20 driving H2Bub1-dependent Cldn11 elongation and seminiferous tubule integrity.\",\n      \"evidence\": \"Sertoli-specific conditional KO mice, histology, H2Bub1 blots, and Cldn11 ChIP\",\n      \"pmids\": [\"37024990\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether additional Sertoli target genes contribute not fully mapped\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended viral hijacking to KSHV, where RTA engages the RNF20/40 complex to enable lytic-cycle reactivation.\",\n      \"evidence\": \"Proteomics, co-IP, and functional lytic reactivation assays\",\n      \"pmids\": [\"37888983\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether ligase activity is required for reactivation unresolved\", \"Single lab\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Reported RNF20-mediated STAT3 ubiquitination and degradation as an insulin-resistance modifier downstream of exosomal miR-let-7b-5p.\",\n      \"evidence\": \"Luciferase reporter, single co-IP for STAT3 ubiquitination, and overexpression rescue\",\n      \"pmids\": [\"37842335\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single Co-IP without reciprocal validation or linkage typing\", \"Direct versus indirect degradation not distinguished\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Defined depot-specific non-histone substrate selection in thermogenesis, with RNF20 degrading NCoR1 in iWAT and controlling GABP\\u03b1 stability in BAT under cold stress.\",\n      \"evidence\": \"Fat-depot-specific Rnf20 KO mice, cold exposure, stability assays, and ubiquitination assays\",\n      \"pmids\": [\"38296968\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How substrate choice is restricted by depot context not mechanistically resolved\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Established an immunosuppressive ligase axis, with CDK9-phosphorylation-dependent RNF20 K29-ubiquitination stabilizing LSD1 to repress endogenous retroviruses and the interferon response, sensitizing tumors to anti-PD-1 upon RNF20 loss.\",\n      \"evidence\": \"Co-IP, K29-specific ubiquitination, CDK9 epistasis, ERV expression, and in vivo anti-PD-1 rescue\",\n      \"pmids\": [\"38315842\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether K29-linkage stabilization generalizes to other substrates unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified an H2Bub1-independent oncogenic mechanism, with RNF20 controlling Rbx1/VHL activity and HIF1\\u03b1 levels to drive metabolic rewiring and EMT via Pol II pause release.\",\n      \"evidence\": \"Rnf20 haploinsufficient mouse lung tumors, metabolomics, HIF1\\u03b1 stability, and Rbx1/VHL epistasis\",\n      \"pmids\": [\"40436847\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct mechanism by which RNF20 sets Rbx1 levels not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed cooperative recruitment under hypoxia, with HIF-1 bringing FACT and RNF20/40 to HREs to induce H2Bub1 and support transcription initiation and elongation at HIF-1 targets.\",\n      \"evidence\": \"RNAi, ChIP at HREs, and hypoxic H2Bub ChIP in breast cancer cells\",\n      \"pmids\": [\"38517892\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Reconciliation with the H2Bub1-independent HIF1\\u03b1 mechanism not addressed\", \"Single lab\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Revealed a non-enzymatic structural role in meiosis, with RNF20 recruiting TPM3 to spindle poles via its coiled-coil for acentrosomal oocyte spindle assembly.\",\n      \"evidence\": \"Oocyte depletion, ligase-dead mutant, RNF20-TPM3 co-IP, localization, and coiled-coil mapping\",\n      \"pmids\": [\"38240347\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How RNF20 itself targets spindle poles not defined\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Connected RNF20 to angiogenic transcriptional control, restricting ERG-dependent Pol II pause release and promoting Notch-target H2Bub1 to constrain tip-cell specification.\",\n      \"evidence\": \"Endothelial Rnf20 KO, Pol II pausing ChIP-seq, Notch1 co-IP, H2Bub ChIP, and angiogenesis assays\",\n      \"pmids\": [\"39322771\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct mechanism of pause regulation versus H2Bub1 deposition not separated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a replication-fork protective role, with ATR-phosphorylated RNF20 and its RING domain promoting H2Bub1 and RAD51/RAD51C loading to shield stalled forks from MRE11/DNA2 degradation.\",\n      \"evidence\": \"DNA fiber assays, RING and phospho-mutant analysis, nuclease inhibition, and fork-remodeler co-depletion\",\n      \"pmids\": [\"40495033\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative contributions of H2Bub1 versus mediator activity at forks not fully separated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Resolved the assembly interfaces, with crystallography and AlphaFold modeling defining the Bre1-Lge1 and RNF20/RNF40-WAC interfaces and their distinct electrostatic specificity required for H2Bub1.\",\n      \"evidence\": \"X-ray crystallography, AlphaFold modeling, interface mutagenesis, and ubiquitination assays\",\n      \"pmids\": [\"41533567\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full RNF20/40-WAC structure experimentally unverified beyond modeling\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Established RNF20 as a beta-cell transcriptional regulator in complex with Isl1, required for H2Bub1, beta-cell identity, and glucose-stimulated insulin secretion.\",\n      \"evidence\": \"Inducible beta-cell conditional KO, Isl1-Rnf20 co-IP, transcriptomics, and insulin secretion assays\",\n      \"pmids\": [\"40743566\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Direct Isl1-RNF20 target genes for insulin control not individually validated\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a nucleocytoplasmic antiviral switch, with CRM1-exported cytoplasmic RNF20 catalyzing K27-linked degradation of RIG-I and MDA5 via its coiled-coil while nuclear RNF20 sustains their transcription.\",\n      \"evidence\": \"Overexpression/knockout, co-IP, K27-specific ubiquitination, CRM1 inhibition, and viral infection assays\",\n      \"pmids\": [\"41712649\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Trigger and regulation of RNF20 export not defined\", \"Single lab\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Defined PTM crosstalk gating catalysis, with cryo-EM showing H2BS112 GlcNAcylation contacts RAD6A and allosterically stimulates ubiquitin transfer to H2B K120 by enhancing lysine nucleophilicity.\",\n      \"evidence\": \"Synthetic modified nucleosomes, chemical trapping, cryo-EM, kinetics, and SAR mutagenesis\",\n      \"pmids\": [\"41495224\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological enzymes setting H2BS112 GlcNAc in this context not identified\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unresolved how RNF20's multiple recruitment routes (WAC, FACT, RPA, Fbxl19, sequence-specific transcription factors) and its catalytic versus ligase-independent activities are coordinately partitioned across transcription, replication, repair, mitosis, and cytoplasmic immune signaling within a single cell.\",\n      \"evidence\": \"No single study integrates the competing recruitment and activity modes\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No unified model of recruiter selection\", \"Catalytic versus scaffold/mediator activity switching not mechanistically defined\", \"A preprint reports H2Bub1 is dispensable for transcription restart after damage, complicating the repair-transcription model (37976359 not applicable; bioRxiv)\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [12, 22, 1, 25]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [10, 15, 16, 27, 30, 19, 37]},\n      {\"term_id\": \"GO:0140110\", \"supporting_discovery_ids\": [1, 8, 32, 43]},\n      {\"term_id\": \"GO:0003677\", \"supporting_discovery_ids\": [22]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [24, 31]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [1, 4, 14, 44]},\n      {\"term_id\": \"GO:0000228\", \"supporting_discovery_ids\": [0, 23, 34]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [44]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [10, 31]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-74160\", \"supporting_discovery_ids\": [1, 4, 8, 32, 43]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [0, 6, 21, 23]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [1, 4, 22, 8]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [2, 10, 15, 16, 27, 30]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [10, 23, 34]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [27, 44]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [15, 17, 28, 30]}\n    ],\n    \"complexes\": [\"RNF20/RNF40 (BRE1) H2B ubiquitin ligase complex\"],\n    \"partners\": [\"RNF40\", \"RAD6/UBE2B\", \"WAC\", \"SUPT16H (FACT)\", \"RPA\", \"CDC73\", \"RAD51\", \"ISL1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}