{"gene":"ADRM1","run_date":"2026-06-09T22:02:42","timeline":{"discoveries":[{"year":2008,"finding":"Rpn13/ADRM1 was identified as a novel proteasomal ubiquitin receptor that binds ubiquitin through a conserved N-terminal pleckstrin-like receptor for ubiquitin (Pru) domain, binding K48-linked diubiquitin with ~90 nM affinity. Like Rpn10/S5a, Rpn13 also binds UBL domains of UBL-UBA shuttle proteins. In yeast, combining specific ubiquitin-binding mutations in both Rpn10 and Rpn13 produces a synthetic phenotype, indicating functional linkage. Rpn13 also serves as the proteasomal receptor for deubiquitinating enzyme Uch37, suggesting coupling of chain recognition and disassembly.","method":"Biochemical binding assays, NMR, mutational analysis, yeast genetic epistasis (synthetic phenotype screen)","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (NMR, affinity measurements, mutagenesis, genetic epistasis), independently foundational, replicated by subsequent studies","pmids":["18497817"],"is_preprint":false},{"year":2006,"finding":"Uch37 (UCH-L5) is recruited to mammalian PA700 (19S) proteasomes via direct binding to Adrm1/hRpn13, which is itself bound to the S1/Rpn2 subunit of the 19S complex. The C-terminal tail of Uch37 (distinct from its UCH catalytic domain) mediates binding to Adrm1. Adrm1 binding relieves Uch37 autoinhibition and accelerates hydrolysis of ubiquitin-AMC. Full incorporation into the 19S complex (not Uch37-Adrm1 or Uch37-Adrm1-S1 alone) is required for efficient polyubiquitin chain processing. PA700 isopeptidase disassembles K48-linked polyubiquitin from the distal end of the chain.","method":"Co-immunoprecipitation, in vitro biochemical reconstitution, ubiquitin-AMC fluorogenic assay, di-ubiquitin hydrolysis assay","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution with multiple biochemical assays, replicated by independent group (PMID:17139257)","pmids":["16906146"],"is_preprint":false},{"year":2006,"finding":"ADRM1/hRpn13 is a novel 46-kDa subunit of the 19S regulatory complex whose N-terminal half is homologous to yeast Rpn13 and incorporates into the 26S proteasome, while its C-terminal half binds directly to UCH37 and enhances its isopeptidase activity. Knockdown of hRpn13 in 293T cells increases cellular ubiquitin conjugates and decreases degradation of short-lived proteins. Overproduction of hRpn13 or transfection of the C-terminal half (dominant-negative) also reduces proteolysis and induces cell death.","method":"Affinity purification, co-immunoprecipitation, siRNA knockdown, ubiquitin conjugate accumulation assay, protein degradation assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods, replicated independently (PMID:16906146)","pmids":["17139257"],"is_preprint":false},{"year":2006,"finding":"Adrm1 is a component of the regulatory ATPase complex of the 26S proteasome, present almost exclusively in soluble 26S proteasomes at equimolar amounts with core subunit S6a (Rpn1). A small fraction is membrane-associated. No free pool of Adrm1 exists in cells, but recombinant Adrm1 can bind pre-existing 26S proteasomes in cell extracts. Knockdown of Adrm1 in HeLa cells had no effect on proteasome amount, bulk protein degradation, or polyubiquitinated protein accumulation, indicating a specialized rather than general role in proteasome function.","method":"Co-immunoprecipitation, gel-filtration chromatography, non-denaturing PAGE, siRNA knockdown, protein degradation assay","journal":"Journal of molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple biochemical methods, single lab, negative functional result also reported","pmids":["16815440"],"is_preprint":false},{"year":2013,"finding":"The bis-benzylidine piperidone RA190 covalently binds to cysteine 88 (C88) of ubiquitin receptor RPN13 in the 19S regulatory particle, inhibiting proteasome function and triggering rapid accumulation of polyubiquitinated proteins, ER stress-related apoptosis. CRISPR/Cas9 Rpn13-knockout abrogates RA190-induced substrate accumulation at proteasomes.","method":"Covalent drug binding assay, cell-based proteasome activity assay, CRISPR/Cas9 knockout, western blotting for polyubiquitinated proteins","journal":"Cancer cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — covalent binding site identified, CRISPR KO validation, single lab; note: a subsequent paper (PMID:32857985) challenged Rpn13 as the physiologically relevant target of RA190","pmids":["24332045"],"is_preprint":false},{"year":2014,"finding":"During proteotoxic stress or partial proteasome inhibition, the proteasome-associated ubiquitin ligase Ube3c/Hul5 selectively poly-ubiquitinates Rpn13 on the 26S proteasome. This autoubiquitination of Rpn13 strongly decreases the proteasome's ability to bind and degrade ubiquitin-conjugated proteins without affecting activity against peptide substrates, constituting an autoinhibitory mechanism that prevents ubiquitin conjugate binding to stalled or defective proteasomes.","method":"Mass spectrometry identification of ubiquitinated sites, in vitro ubiquitination assay with purified 26S proteasomes, cell-based stress treatments, proteasome activity assays","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro reconstitution with purified proteasomes, MS identification of sites, cell-based validation with multiple stress conditions","pmids":["24811749"],"is_preprint":false},{"year":2015,"finding":"Crystal structures of UCH-L5/Uch37 in complex with the DEUBAD domain of RPN13 reveal that RPN13 activates Uch37 by repositioning its C-terminal ULD domain and crossover loop to promote substrate binding and catalysis. The related DEUBAD domain of INO80G inhibits UCH-L5 by exploiting the same structural elements but driving a radically different conformation and employing molecular mimicry to block ubiquitin docking.","method":"Crystal structure determination, mutagenesis, enzymatic activity assays","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structures with mutagenesis and functional validation, provides atomic-level mechanism","pmids":["25702870"],"is_preprint":false},{"year":2017,"finding":"The structure of hRpn13 bound to a segment of hRpn2 (its proteasome docking site) was solved; a proline-rich C-terminal hRpn2 extension stretches across a narrow canyon of the ubiquitin-binding hRpn13 Pru domain, blocking an RA190-binding surface. hRpn13 binds preferentially to hRpn2/proteasomes over RA190. RA190 does not affect the hRpn13-Uch37 interaction but directly binds and inactivates Uch37. hRpn13 deletion from HCT116 cells abrogates RA190-induced substrate accumulation at proteasomes.","method":"NMR structure determination, biophysical binding assays (SPR, fluorescence polarization), CRISPR/Cas9 cell-based assays","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — NMR structure, multiple biophysical methods, cell-based genetic validation","pmids":["28598414"],"is_preprint":false},{"year":2017,"finding":"Crystal structures of the RPN13 PRU domain in complex with peptides corresponding to the RPN2 C terminus and ubiquitin were determined. The RPN13-binding epitope maps to the C-terminal 14 residues of RPN2. RPN2, ubiquitin, and UCH37 each bind RPN13 with independent energetics. Mutational analysis validated the RPN2-binding interface by SPR and fluorescence polarization.","method":"Crystal structure determination, SPR, fluorescence polarization, mutational analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structures with quantitative binding validation using multiple orthogonal biophysical methods","pmids":["28442575"],"is_preprint":false},{"year":2010,"finding":"Rpn13 is involved in selective degradation of iNOS and IκB-α via the Rpn13/UCH37 complex. Knockdown of Rpn13 by siRNA does not lead to global ubiquitinated protein accumulation but impairs iNOS interaction with UCH37. IκB-α, whose proteasomal degradation activates NF-κB, is also a substrate for the Rpn13/UCH37 pathway.","method":"siRNA knockdown, co-immunoprecipitation, protein stability/degradation assays","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — co-IP and functional knockdown with specific substrate readout, single lab","pmids":["20634424"],"is_preprint":false},{"year":2014,"finding":"Uch37 oligomerizes in solution and has very low isopeptidase activity due to autoinhibition in which each catalytic domain in the homodimer blocks the other's ubiquitin-binding site. Rpn13C (residues 270–407) disrupts Uch37 oligomerization by sequestering it in a 1:1 stoichiometric complex, activating Uch37 with ~12-fold higher activity. TR-SAXS and FRET confirmed the mode of autoinhibition and activation mechanism.","method":"Mutagenesis, NMR, SAXS, TR-SAXS, FRET, biochemical activity assay","journal":"Protein & cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple orthogonal structural and biochemical methods in a single study establishing mechanism of Uch37 autoinhibition and Rpn13-mediated activation","pmids":["24752541"],"is_preprint":false},{"year":2015,"finding":"Simultaneous liver-specific deletion of both Rpn13 and Rpn10 in mice causes severe liver injury with massive ubiquitin conjugate accumulation, while single deletion causes only modest impairment. In the double knockout, shuttle proteins mHR23B and ubiquilin/Plic-1 and -4 fail to bind the proteasome, establishing Rpn10 and Rpn13 as the two main receptors for UBL-UBA substrate delivery proteins at the proteasome.","method":"Conditional knockout mouse (liver-specific), ubiquitin conjugate accumulation assay, co-immunoprecipitation, genetic rescue by re-expression","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic double-KO with defined phenotype, rescue by re-expression, multiple biochemical readouts","pmids":["26222436"],"is_preprint":false},{"year":2015,"finding":"The N-terminal ubiquitin-like (Ubl) domain of parkin binds directly to the Pru domain of Rpn13/ADRM1 via a hydrophobic patch surrounding Ile-44, with >10-fold higher affinity than ubiquitin for the Pru domain. Knockdown of Rpn13 in cells increases parkin levels, abrogates parkin recruitment to the 26S proteasome, and delays clearance of mitochondrial proteins (TIM23, TIM44, TOM20) while enhancing parkin autoubiquitination. Rpn13 knockdown did not impair parkin recruitment to mitochondria or mitophagy initiation.","method":"Co-immunoprecipitation, NMR binding analysis, mutational analysis, siRNA knockdown, mitochondrial protein clearance assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — NMR-based binding characterization, mutagenesis, and cell-based functional validation with multiple readouts","pmids":["25666615"],"is_preprint":false},{"year":2015,"finding":"The C-terminal region of Rpn13 (DEUBAD domain) binds to the tetratricopeptide repeat (TPR) domain of SGTA via a carboxylate clamp mechanism. SGTA overexpression increases steady-state mislocalised membrane protein (MLP) levels in a manner dependent on SGTA's interaction with Rpn13. Expression of the SGTA-binding region of Rpn13 or TPR domain point mutations inhibit SGTA-Rpn13 binding and substantially reduce MLP levels.","method":"Co-immunoprecipitation, NMR structural characterization (carboxylate clamp interaction), mutational analysis, protein stability assays","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — structural and functional characterization, single lab, multiple orthogonal approaches","pmids":["26169395"],"is_preprint":false},{"year":2016,"finding":"The structural basis for SGTA-Rpn13 interaction was characterized: a region of the Rpn13 C-terminal domain is necessary and sufficient for binding to the TPR domain of SGTA through a carboxylate clamp-mediated molecular recognition event, and the interaction was validated in a cellular context.","method":"NMR structural characterization, biophysical binding assays, cellular co-immunoprecipitation","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — structural and biophysical validation, single lab","pmids":["27827410"],"is_preprint":false},{"year":2014,"finding":"Although most double-capped 26S proteasomes contain Rpn10/S5a in both 19S particles, only one of the two 19S particles contains Rpn13, defining inherent asymmetry in the 26S proteasome. This was validated in both yeast and mammals using biochemical fractionation and quantitative AQUA-MS methodology.","method":"Biochemical fractionation, AQUA-MS quantitation, validation in yeast and mammalian cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — quantitative MS (AQUA) with cross-species validation, single lab","pmids":["24429290"],"is_preprint":false},{"year":2014,"finding":"SUMOylation of Psmd1/RPN2 (by PIASy E3 ligase) at a critical lysine immediately adjacent to the Adrm1-binding domain regulates the association of Adrm1 with Psmd1/RPN2. The SUMO deconjugating enzyme xSENP1 specifically interacts with Psmd1, and disruption of xSENP1 targeting delays mitotic exit, linking this SUMOylation-mediated regulation to cell division.","method":"SUMOylation site mapping, co-immunoprecipitation, Xenopus extract cell cycle assay, SENP1 targeting experiments","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical mapping with functional cell cycle readout, single lab","pmids":["24910440"],"is_preprint":false},{"year":2019,"finding":"The structure of hRpn13 Pru:hRpn2(940-953):K48-diubiquitin was solved by NMR, revealing that hRpn2-bound hRpn13 selects a dynamic, extended conformation of K48-diubiquitin distinct from previously determined closed crystal structures. hRpn13 exchanges between the two ubiquitins within 100 ms but prefers the proximal ubiquitin due to interactions with the K48 linker region.","method":"NMR structure determination, NMR dynamics experiments","journal":"Structure","confidence":"High","confidence_rationale":"Tier 1 / Moderate — NMR structure with dynamics characterization, single lab but rigorous structural method","pmids":["32160516"],"is_preprint":false},{"year":2019,"finding":"The structure of Rpn13NTD bound to K48-diubiquitin was determined by NMR; Rpn13NTD simultaneously contacts both proximal and distal Ub subunits of K48-diUb in a compact state. The proximal Ub binds similarly to Ub monomer, while distal Ub engages a largely electrostatic surface of Rpn13NTD. A charge-reversal mutation in Rpn13NTD weakens K48-linked chain binding and causes ubiquitinated protein accumulation. Single-molecule FRET showed K48-diUb fluctuates among conformational states, with the compact state selectively enriched by Rpn13NTD.","method":"NMR structure, smFRET, charge-reversal mutagenesis, cell-based ubiquitin conjugate assay","journal":"Cell discovery","confidence":"High","confidence_rationale":"Tier 1 / Strong — NMR structure plus smFRET plus mutagenesis with cellular validation, orthogonal methods","pmids":["30962947"],"is_preprint":false},{"year":2019,"finding":"Phosphorylation of RPN2 Tyr-950 enhances its binding to RPN13. Crystal structure of the RPN2-RPN13 pTyr-950-ubiquitin complex at 1.76 Å reveals specific interactions with positively charged RPN13 side chains that explain the increased affinity without conformational change. Mutagenesis and SPR/fluorescence polarization validated the crystallographic interface.","method":"Crystal structure determination (1.76 Å), SPR, fluorescence polarization, mutagenesis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — high-resolution crystal structure with quantitative biophysical validation using multiple methods","pmids":["31064842"],"is_preprint":false},{"year":2015,"finding":"An hRpn2-derived 38-amino acid peptide binds the hRpn13 Pru domain with 12 nM affinity. NMR identified the hRpn13-interacting amino acids in this fragment. The peptide immunoprecipitates endogenous Rpn13 from 293T cells and displaces it from the proteasome. Over-expression of the peptide leads to increased ubiquitinated proteins. Conserved F948 and Y950/I951 of hRpn2 are critical for this interaction.","method":"NMR binding analysis, Co-immunoprecipitation, mutational analysis, fluorescence polarization","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — NMR and cell-based validation, single lab, multiple orthogonal approaches","pmids":["26466095"],"is_preprint":false},{"year":2010,"finding":"Phospho-ΔNp63α interacts with Rpn13 protein in cisplatin-sensitive SCC cells and this interaction leads to proteasome-dependent degradation of phospho-ΔNp63α while protecting LKB1 from degradation. In SCC cells with impaired ATM-dependent ΔNp63α phosphorylation, non-phosphorylated ΔNp63α fails to form complexes with Rpn13, allowing Rpn13 to target LKB1 for proteasome-dependent degradation, modulating cisplatin-induced autophagy.","method":"Co-immunoprecipitation, siRNA knockdown, protein stability assays","journal":"Aging","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP approach, single lab, limited mechanistic detail in abstract","pmids":["21191146"],"is_preprint":false},{"year":2010,"finding":"Phospho-ΔNp63α binds to the RPN13 gene promoter through a TP63-responsive element and, together with co-factors DDIT3/CHOP, NF-Y and NF-κB, activates RPN13 gene transcription. Cisplatin-induced, phospho-ΔNp63α-dependent RPN13 transcription leads to NOS2/iNOS degradation; RPN13 siRNA knockdown rescues NOS2 from this degradation.","method":"Chromatin immunoprecipitation, promoter-binding assay, siRNA knockdown, protein degradation assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — ChIP-based transcription factor binding with functional siRNA rescue, single lab","pmids":["20959455"],"is_preprint":false},{"year":2009,"finding":"Adrm1 interacts with Atp6v0d2 (d2 isoform of vacuolar H+-ATPase V0 domain) as identified by yeast two-hybrid screening and confirmed by co-immunoprecipitation. Adrm1 is required for cell migration and osteoclast maturation.","method":"Yeast two-hybrid screen, co-immunoprecipitation, cell migration assay, osteoclast differentiation assay","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — yeast two-hybrid plus Co-IP, single lab, limited mechanistic follow-up","pmids":["19818731"],"is_preprint":false},{"year":2018,"finding":"Shigella effector IpaH4.5 (an E3 ubiquitin ligase) targets RPN13 and induces its degradation via the ubiquitin-proteasome system. IpaH4.5-mediated RPN13 degradation causes dysfunction of the 19S regulatory particle, inhibiting guidance of ubiquitinated proteins to the 20S core particle and suppressing proteasome-catalyzed peptide splicing. This reduces antigen cross-presentation to CD8+ T cells via MHC class I. In RPN13 knockout MEFs, loss of RPN13 suppressed CD8+ T cell priming during Shigella infection.","method":"Bacterial infection model, RPN13 knockout MEFs, antigen presentation assay, T cell activation assay","journal":"Cellular microbiology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with defined immune functional readout, bacterial effector mechanistic dissection, single lab","pmids":["30414351"],"is_preprint":false},{"year":2023,"finding":"ADRM1/Rpn13 upregulates UCH37 expression and binds UCH37 to activate its deubiquitination activity. Activated UCH37 enhances deubiquitination of ALK5 (activin receptor-like kinase 5), stabilizing ALK5 expression and maintaining extracellular matrix homeostasis in cartilage. Global knockout of ADRM1 accelerates cartilage degeneration in DMM-induced OA model.","method":"Global knockout mouse, conditional overexpression, co-immunoprecipitation, deubiquitination assay, ALK5 stability assay","journal":"International journal of biological macromolecules","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — in vivo KO with biochemical mechanism (UCH37 activation of ALK5 deubiquitination) validated, single lab","pmids":["37406898"],"is_preprint":false},{"year":2019,"finding":"HDAC8 interacts with the proteasome receptor ADRM1, and inhibition of HDAC8 decreases MGMT protein levels in GBM cell lines in a manner dependent on this interaction. Treatment with TMZ disrupts the HDAC8-ADRM1 interaction exclusively in TMZ-sensitive cells.","method":"Co-immunoprecipitation, HDAC8 inhibitor/shRNA knockdown, western blotting for MGMT levels","journal":"Genes & cancer","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP, single lab, limited mechanistic detail","pmids":["31798765"],"is_preprint":false},{"year":2023,"finding":"A spliced ADRM1 variant lacking exon 9 (ADRM1-ΔEx9) interacts with a different deubiquitinase partner (BAP1 instead of UCH37) via its altered C-terminus, redirecting proteasome specificity. The novel exon 8-10 fusion creates a de novo binding site for tumor suppressor FBXW7, resulting in its selective proteasome-mediated degradation.","method":"Proteome Profiler Human Ubiquitin Array, co-immunoprecipitation, patient-derived organoids, xenograft models, murine hydrodynamic transfection","journal":"Journal of hepatology","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — multiple experimental systems (organoids, mouse model, proteomic array), single lab; note this concerns a splice variant of ADRM1","pmids":["39788431"],"is_preprint":false},{"year":2023,"finding":"ADRM1/Rpn13 functions in DNA end resection as part of a proteasome variant (CDW19S) that assembles at broken chromatin. ADRM1Rpn13 inhibits DNA resection and is removed by CRL4WDR70-catalyzed ubiquitination upon commitment to extensive resection. HBx protein (from HBV) interferes with ADRM1Rpn13 degradation, leading to a resection barrier and homologous recombination deficiency.","method":"Co-immunoprecipitation, DNA damage repair assays, ubiquitination assay, HBx expression, HR assay","journal":"The Journal of clinical investigation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — biochemical and cell-based dissection of a novel non-proteasomal function for ADRM1 in DNA repair, single lab","pmids":["37815873"],"is_preprint":false},{"year":2016,"finding":"Rpn13 and Rpn10 are the main proteasomal receptors for ubiquitin-like (UBL) domain-containing shuttle proteins (mHR23B and ubiquilins/Plic-1 and -4) that deliver ubiquitinated cargo to the proteasome. In double Rpn10/Rpn13 knockout hepatocytes, these shuttle proteins fail to associate with the proteasome.","method":"Conditional double-knockout mouse, co-immunoprecipitation, genetic rescue by re-expression","journal":"PLoS genetics","confidence":"High","confidence_rationale":"Tier 2 / Strong — clean genetic double-KO with specific molecular readout (shuttle protein binding) confirmed by rescue","pmids":["26222436"],"is_preprint":false},{"year":2016,"finding":"MHC class I processing of the NY-ESO-1 antigen is governed by both Rpn10 and Rpn13 ubiquitin receptors as well as by immunoproteasomes, and is regulated by non-canonical ubiquitination on non-lysine sites rather than canonical K48-linked chains at K124.","method":"siRNA knockdown of Rpn13 and Rpn10, antigen presentation assay, ubiquitination site mutagenesis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — siRNA knockdown with specific antigen processing readout, mutagenesis of ubiquitination sites, single lab","pmids":["26903513"],"is_preprint":false},{"year":2025,"finding":"Conditional deletion of Rpn13 in primordial germ cells (PGCs) in mice causes infertility in both sexes with dramatic reduction in germ cells in testes and ovaries. Mechanistically, Rpn13 deletion disrupts 26S proteasome assembly in germ cells, reduces PGC numbers, and blocks meiosis of spermatocytes at the zygotene stage during prophase I.","method":"Conditional knockout mouse (PGC-specific), proteasome assembly assay, histological and meiosis staging analysis","journal":"Cells","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — conditional KO with specific cellular and molecular phenotypes, single lab","pmids":["40422199"],"is_preprint":false},{"year":2025,"finding":"A Protea-Tac degrader incorporating ADRM1/Rpn13 as the ubiquitin receptor component integrates into 26S proteasomes without altering their structural or functional integrity. Proximity of target proteins (c-Fos, BRD4, Flag-TDP43, HA-tau, GFP-ODC) to the proteasome via Rpn13-antibody chimera results in their ubiquitin-independent degradation.","method":"Chimeric protein reconstitution, proteasome integrity assays, targeted degradation assays for multiple substrates, in vivo xenograft mouse model","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — reconstitution approach with multiple substrates and in vivo validation, preprint not yet peer-reviewed","pmids":["bio_10.1101_2025.08.18.670774"],"is_preprint":true},{"year":2025,"finding":"PhIX-MS and cryo-EM placed the thioredoxin domain of redox sensor TXNL1 near RPN2/PSMD1 and RPN13/ADRM1 at the proteasome regulatory particle, positioning it to reduce substrates prior to proteolysis.","method":"PhIX-MS (photo-induced in situ crosslinking-mass spectrometry), cryo-electron microscopy, AlphaFold modeling","journal":"bioRxiv","confidence":"Low","confidence_rationale":"Tier 2 / Weak — structural localization by crosslinking MS/cryo-EM, preprint, TXNL1-RPN13 proximity not functionally validated","pmids":["bio_10.1101_2025.07.31.667872"],"is_preprint":true},{"year":2020,"finding":"RA190 has no measurable effect on any of the known interactions of Rpn13 in vitro, and no physical engagement of Rpn13 by RA190 was detected in cellulo on C88 or any other residue. Chemical proteomics in two cell lines showed that dozens of other proteins are heavily engaged by RA190. Increasing or reducing Rpn13 levels had no effect on RA190 sensitivity of HeLa or melanoma cells, indicating Rpn13 is NOT the physiologically relevant target of RA190.","method":"In vitro protein interaction assays, chemical proteomics (activity-based protein profiling), cell viability assays with Rpn13 modulation","journal":"Cell chemical biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — chemical proteomics plus multiple functional assays across two cell lines; this is a negative finding contradicting PMID:24332045 regarding RA190's principal target","pmids":["32857985"],"is_preprint":false}],"current_model":"ADRM1/Rpn13 is a subunit of the 19S regulatory particle of the 26S proteasome that functions as a ubiquitin receptor (via its N-terminal Pru domain, which binds K48-linked polyubiquitin chains bivalently and also docks into the proteasome through direct interaction with the C-terminus of RPN2/PSMD1), and as a deubiquitinase adaptor (via its C-terminal DEUBAD domain, which recruits and allosterically activates the deubiquitinase UCH37/UCHL5 by disrupting its autoinhibitory oligomerization); Rpn13 is present asymmetrically in only one of the two 19S caps of the double-capped proteasome, acts redundantly with Rpn10 for bulk ubiquitin conjugate recognition but is required for recruitment of specific substrates (including iNOS, IκB-α, parkin, and mislocalised membrane proteins via SGTA) and shuttle proteins (mHR23B, ubiquilins); its activity is regulated by Ube3c/Hul5-mediated autoubiquitination (autoinhibitory under proteotoxic stress) and by SUMOylation of its docking partner RPN2; additionally, ADRM1 participates in DNA double-strand break end resection where it inhibits resection and is removed by CRL4WDR70 ubiquitination, and its deletion disrupts 26S proteasome assembly and blocks meiosis in germ cells."},"narrative":{"mechanistic_narrative":"ADRM1/Rpn13 is a subunit of the 19S regulatory particle of the 26S proteasome that couples recognition of ubiquitinated substrates to their disassembly and degradation [PMID:18497817, PMID:17139257]. It is recruited to the proteasome by direct docking onto the C-terminal tail of RPN2/PSMD1, an interaction mapped to the final ~14 residues of RPN2 binding the Rpn13 Pru domain and further tuned by RPN2 modification — Tyr-950 phosphorylation enhances affinity while SUMOylation of RPN2 adjacent to the docking site regulates Adrm1 association in a manner linked to mitotic exit [PMID:28442575, PMID:31064842, PMID:24910440]. Through its N-terminal Pru domain Rpn13 acts as a proteasomal ubiquitin receptor, binding K48-linked diubiquitin with high affinity by engaging both proximal and distal ubiquitin moieties and selecting a defined chain conformation [PMID:18497817, PMID:30962947, PMID:32160516]. The Pru domain also captures the ubiquitin-like domains of UBL-UBA shuttle proteins; genetic ablation in mice shows that Rpn13 and Rpn10 are the two main receptors that recruit shuttle factors mHR23B and ubiquilins and that they act redundantly for bulk conjugate handling, with double loss causing massive conjugate accumulation [PMID:18497817, PMID:26222436]. Through its C-terminal DEUBAD domain Rpn13 functions as a deubiquitinase adaptor, recruiting UCH37/UCHL5 and allosterically activating it by disrupting an autoinhibitory homodimer and repositioning the enzyme's ULD domain and crossover loop to license substrate binding and catalysis [PMID:16906146, PMID:25702870, PMID:24752541]. Rpn13 directs the proteasome toward specific substrates and partners — including iNOS and IκB-α via the Rpn13/UCH37 axis, parkin via high-affinity Ubl-domain docking, and mislocalised membrane proteins delivered by SGTA through a carboxylate-clamp interaction with the C-terminal domain [PMID:20634424, PMID:25666615, PMID:26169395]. Its conjugate-binding capacity is itself regulated: under proteotoxic stress the proteasome-associated ligase Ube3c/Hul5 autoubiquitinates Rpn13, an autoinhibitory mechanism that selectively suppresses ubiquitin-conjugate degradation at stalled proteasomes [PMID:24811749]. Rpn13 is present asymmetrically in only one of the two 19S caps of double-capped proteasomes [PMID:24429290]. Beyond the proteasome, Rpn13 participates in DNA double-strand break end resection, where it inhibits resection and is removed by CRL4WDR70-mediated ubiquitination [PMID:37815873], and its deletion in germ cells disrupts 26S proteasome assembly and blocks meiosis, causing infertility [PMID:40422199].","teleology":[{"year":2006,"claim":"Established that Adrm1/hRpn13 is a bona fide 19S subunit and the proteasomal recruiter of the deubiquitinase Uch37, defining a physical link between a regulatory particle protein and chain disassembly.","evidence":"Affinity purification, co-IP, in vitro reconstitution and ubiquitin-AMC assays, siRNA knockdown in mammalian cells","pmids":["16906146","17139257","16815440"],"confidence":"High","gaps":["Did not resolve the structural basis of Uch37 activation","Conflicting reports on whether knockdown affects bulk degradation"]},{"year":2008,"claim":"Defined Rpn13's N-terminal Pru domain as a high-affinity K48-polyubiquitin receptor functionally linked to Rpn10, answering how the proteasome recognizes ubiquitinated cargo through a second receptor.","evidence":"NMR, affinity measurements, mutagenesis, yeast genetic epistasis","pmids":["18497817"],"confidence":"High","gaps":["Did not establish in vivo substrate specificity","Mode of bivalent diubiquitin engagement not yet resolved at atomic level"]},{"year":2010,"claim":"Connected Rpn13 to degradation of defined regulatory substrates (iNOS, IκB-α) via the Rpn13/UCH37 complex rather than bulk turnover, indicating a specialized substrate-selective role.","evidence":"siRNA knockdown, co-IP, protein stability assays; plus ChIP-based transcriptional regulation of RPN13 by phospho-ΔNp63α","pmids":["20634424","20959455","21191146"],"confidence":"Medium","gaps":["Substrate selectivity mechanism not structurally defined","ΔNp63α/LKB1 link rests on single Co-IP without reciprocal validation"]},{"year":2014,"claim":"Resolved the mechanism of UCH37 activation and revealed two layers of regulation — Rpn13-mediated disruption of UCH37 autoinhibitory oligomerization, and stress-induced autoubiquitination of Rpn13 itself that shuts down conjugate binding.","evidence":"NMR/SAXS/TR-SAXS/FRET activity reconstitution; MS site mapping and in vitro ubiquitination with purified 26S proteasomes; AQUA-MS proteasome asymmetry mapping; RPN2 SUMOylation site mapping","pmids":["24752541","24811749","24429290","24910440"],"confidence":"High","gaps":["Physiological triggers of Ube3c/Hul5 autoubiquitination incompletely defined","Functional consequence of 19S asymmetry for Rpn13 unresolved"]},{"year":2015,"claim":"Genetically established Rpn13 and Rpn10 as the two principal proteasomal receptors for UBL-UBA shuttle proteins, and broadened Rpn13's substrate-targeting repertoire to parkin and SGTA-delivered mislocalised membrane proteins.","evidence":"Liver-specific conditional double-knockout mice with rescue; NMR binding and siRNA functional assays for parkin; carboxylate-clamp NMR/co-IP for SGTA","pmids":["26222436","25666615","26169395"],"confidence":"High","gaps":["Relative contribution of Rpn13 vs Rpn10 for individual substrates not fully partitioned","SGTA-Rpn13 axis characterized in a single lab"]},{"year":2015,"claim":"Provided atomic-level mechanism for both arms of Rpn13: how the DEUBAD domain repositions UCH37 to activate it (versus INO80G's inhibitory mimicry), and how the Pru domain docks onto the RPN2 C-terminus.","evidence":"Crystal structures of UCH-L5:DEUBAD and Pru:RPN2/ubiquitin complexes with mutagenesis and SPR/FP validation","pmids":["25702870","28442575","26466095"],"confidence":"High","gaps":["Conformational coupling between RPN2 docking and ubiquitin binding within intact proteasome unresolved"]},{"year":2017,"claim":"Defined how RPN2 docking masks the Pru ubiquitin-binding surface and clarified that the RA190 inhibitor binds and inactivates UCH37 rather than disrupting Rpn13-UCH37, refining the drug-target relationship.","evidence":"NMR structure of hRpn13:hRpn2 segment, SPR/FP, CRISPR cell-based assays","pmids":["28598414"],"confidence":"High","gaps":["In-cell selectivity of RA190 not yet resolved at this stage"]},{"year":2019,"claim":"Established the structural basis of bivalent K48-diubiquitin recognition and showed that RPN2 Tyr-950 phosphorylation enhances Rpn13 docking, defining how chain selection and proteasome anchoring are tuned.","evidence":"NMR structures with dynamics, smFRET, charge-reversal mutagenesis with cellular conjugate readout; 1.76 Å crystal structure of pTyr-RPN2:Rpn13:ubiquitin","pmids":["32160516","30962947","31064842"],"confidence":"High","gaps":["Kinase responsible for RPN2 Tyr-950 phosphorylation not identified in corpus","Physiological context selecting open vs compact diubiquitin states unclear"]},{"year":2020,"claim":"Challenged Rpn13 as the physiologically relevant target of the RA190 inhibitor, showing no in-cell engagement of Rpn13 and Rpn13-independent toxicity.","evidence":"In vitro interaction assays, chemical proteomics in two cell lines, Rpn13-modulation viability assays","pmids":["32857985"],"confidence":"Medium","gaps":["Directly contradicts earlier CRISPR-based RA190 target assignment","True target(s) responsible for RA190 toxicity not defined"]},{"year":2023,"claim":"Extended Rpn13 function beyond canonical degradation — into substrate-stabilizing deubiquitination (UCH37/ALK5 in cartilage), splice-variant rewiring of deubiquitinase and substrate partners, and a non-proteasomal role restraining DNA end resection.","evidence":"Global/conditional knockout mice and DMM OA model; ubiquitin-array, organoid and xenograft profiling of ADRM1-ΔEx9; co-IP and HR/ubiquitination assays for CDW19S/CRL4WDR70 resection","pmids":["37406898","39788431","37815873"],"confidence":"Medium","gaps":["Each non-canonical role characterized by a single lab","How a degradation receptor stabilizes a substrate (ALK5) mechanistically incompletely defined"]},{"year":2025,"claim":"Demonstrated a physiological requirement for Rpn13 in 26S proteasome assembly during germ-cell development and exploited Rpn13 as the receptor module for ubiquitin-independent targeted degradation.","evidence":"PGC-specific conditional knockout with proteasome assembly and meiosis staging; Protea-Tac chimeric reconstitution with multiple substrates and xenograft validation (preprint)","pmids":["40422199","bio_10.1101_2025.08.18.670774"],"confidence":"Medium","gaps":["Molecular basis of Rpn13's assembly requirement not defined","Degrader study is a preprint not yet peer-reviewed"]},{"year":null,"claim":"How Rpn13's distinct activities — ubiquitin recognition, UCH37 activation, shuttle/substrate recruitment, and non-proteasomal resection control — are spatially and temporally coordinated within a single asymmetric cap remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated model linking 19S asymmetry to functional specialization","Regulation switching between proteasomal and DNA-repair roles unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0031386","term_label":"protein tag activity","supporting_discovery_ids":[0,18,17,12]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[1,6,10]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[1,13,29]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[3,2]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[28]}],"pathway":[{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,2,11]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[1,6]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[24,30]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[28]}],"complexes":["26S proteasome 19S regulatory particle"],"partners":["UCHL5","PSMD1","PSMD4","RAD23B","PARK2","SGTA","UBE3C"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q16186","full_name":"Proteasomal ubiquitin receptor ADRM1","aliases":["110 kDa cell membrane glycoprotein","Gp110","Adhesion-regulating molecule 1","ARM-1","Proteasome regulatory particle non-ATPase 13","hRpn13","Rpn13 homolog"],"length_aa":407,"mass_kda":42.2,"function":"Component of the 26S proteasome, a multiprotein complex involved in the ATP-dependent degradation of ubiquitinated proteins (PubMed:16815440, PubMed:16906146, PubMed:16990800, PubMed:17139257, PubMed:18497817, PubMed:24752541, PubMed:25702870, PubMed:25702872). This complex plays a key role in the maintenance of protein homeostasis by removing misfolded or damaged proteins, which could impair cellular functions, and by removing proteins whose functions are no longer required (PubMed:16815440, PubMed:16906146, PubMed:16990800, PubMed:17139257, PubMed:18497817, PubMed:24752541, PubMed:25702870, PubMed:25702872). Therefore, the proteasome participates in numerous cellular processes, including cell cycle progression, apoptosis, or DNA damage repair (PubMed:16815440, PubMed:16906146, PubMed:16990800, PubMed:17139257, PubMed:18497817, PubMed:24752541, PubMed:25702870, PubMed:25702872). Within the complex, functions as a proteasomal ubiquitin receptor (PubMed:18497817). Engages and activates 19S-associated deubiquitinases UCHL5 and PSMD14 during protein degradation (PubMed:16906146, PubMed:16990800, PubMed:17139257, PubMed:24752541). UCHL5 reversibly associate with the 19S regulatory particle whereas PSMD14 is an intrinsic subunit of the proteasome lid subcomplex (PubMed:16906146, PubMed:16990800, PubMed:17139257, PubMed:24752541)","subcellular_location":"Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q16186/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ADRM1","classification":"Not Classified","n_dependent_lines":513,"n_total_lines":1208,"dependency_fraction":0.4246688741721854},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"PSMC4","stoichiometry":10.0}],"url":"https://opencell.sf.czbiohub.org/search/ADRM1","total_profiled":1310},"omim":[{"mim_id":"617842","title":"PROTEASOME 26S SUBUNIT, NON-ATPase, 1; PSMD1","url":"https://www.omim.org/entry/617842"},{"mim_id":"610650","title":"ADHESION-REGULATING MOLECULE 1; ADRM1","url":"https://www.omim.org/entry/610650"},{"mim_id":"601648","title":"PROTEASOME 26S SUBUNIT, NON-ATPase, 4; PSMD4","url":"https://www.omim.org/entry/601648"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"},{"location":"Plasma membrane","reliability":"Supported"},{"location":"Cytosol","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"skeletal muscle","ntpm":376.2}],"url":"https://www.proteinatlas.org/search/ADRM1"},"hgnc":{"alias_symbol":["GP110","Rpn13","ARM1","PSMD16"],"prev_symbol":[]},"alphafold":{"accession":"Q16186","domains":[{"cath_id":"2.30.29.70","chopping":"23-128","consensus_level":"high","plddt":90.0587,"start":23,"end":128},{"cath_id":"1.10.2020.20","chopping":"287-309_330-384","consensus_level":"high","plddt":64.0418,"start":287,"end":384}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q16186","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q16186-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q16186-F1-predicted_aligned_error_v6.png","plddt_mean":62.28},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ADRM1","jax_strain_url":"https://www.jax.org/strain/search?query=ADRM1"},"sequence":{"accession":"Q16186","fasta_url":"https://rest.uniprot.org/uniprotkb/Q16186.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q16186/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q16186"}},"corpus_meta":[{"pmid":"18497817","id":"PMC_18497817","title":"Proteasome 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Like Rpn10/S5a, Rpn13 also binds UBL domains of UBL-UBA shuttle proteins. In yeast, combining specific ubiquitin-binding mutations in both Rpn10 and Rpn13 produces a synthetic phenotype, indicating functional linkage. Rpn13 also serves as the proteasomal receptor for deubiquitinating enzyme Uch37, suggesting coupling of chain recognition and disassembly.\",\n      \"method\": \"Biochemical binding assays, NMR, mutational analysis, yeast genetic epistasis (synthetic phenotype screen)\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (NMR, affinity measurements, mutagenesis, genetic epistasis), independently foundational, replicated by subsequent studies\",\n      \"pmids\": [\"18497817\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Uch37 (UCH-L5) is recruited to mammalian PA700 (19S) proteasomes via direct binding to Adrm1/hRpn13, which is itself bound to the S1/Rpn2 subunit of the 19S complex. The C-terminal tail of Uch37 (distinct from its UCH catalytic domain) mediates binding to Adrm1. Adrm1 binding relieves Uch37 autoinhibition and accelerates hydrolysis of ubiquitin-AMC. Full incorporation into the 19S complex (not Uch37-Adrm1 or Uch37-Adrm1-S1 alone) is required for efficient polyubiquitin chain processing. PA700 isopeptidase disassembles K48-linked polyubiquitin from the distal end of the chain.\",\n      \"method\": \"Co-immunoprecipitation, in vitro biochemical reconstitution, ubiquitin-AMC fluorogenic assay, di-ubiquitin hydrolysis assay\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution with multiple biochemical assays, replicated by independent group (PMID:17139257)\",\n      \"pmids\": [\"16906146\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"ADRM1/hRpn13 is a novel 46-kDa subunit of the 19S regulatory complex whose N-terminal half is homologous to yeast Rpn13 and incorporates into the 26S proteasome, while its C-terminal half binds directly to UCH37 and enhances its isopeptidase activity. Knockdown of hRpn13 in 293T cells increases cellular ubiquitin conjugates and decreases degradation of short-lived proteins. Overproduction of hRpn13 or transfection of the C-terminal half (dominant-negative) also reduces proteolysis and induces cell death.\",\n      \"method\": \"Affinity purification, co-immunoprecipitation, siRNA knockdown, ubiquitin conjugate accumulation assay, protein degradation assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods, replicated independently (PMID:16906146)\",\n      \"pmids\": [\"17139257\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Adrm1 is a component of the regulatory ATPase complex of the 26S proteasome, present almost exclusively in soluble 26S proteasomes at equimolar amounts with core subunit S6a (Rpn1). A small fraction is membrane-associated. No free pool of Adrm1 exists in cells, but recombinant Adrm1 can bind pre-existing 26S proteasomes in cell extracts. Knockdown of Adrm1 in HeLa cells had no effect on proteasome amount, bulk protein degradation, or polyubiquitinated protein accumulation, indicating a specialized rather than general role in proteasome function.\",\n      \"method\": \"Co-immunoprecipitation, gel-filtration chromatography, non-denaturing PAGE, siRNA knockdown, protein degradation assay\",\n      \"journal\": \"Journal of molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple biochemical methods, single lab, negative functional result also reported\",\n      \"pmids\": [\"16815440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The bis-benzylidine piperidone RA190 covalently binds to cysteine 88 (C88) of ubiquitin receptor RPN13 in the 19S regulatory particle, inhibiting proteasome function and triggering rapid accumulation of polyubiquitinated proteins, ER stress-related apoptosis. CRISPR/Cas9 Rpn13-knockout abrogates RA190-induced substrate accumulation at proteasomes.\",\n      \"method\": \"Covalent drug binding assay, cell-based proteasome activity assay, CRISPR/Cas9 knockout, western blotting for polyubiquitinated proteins\",\n      \"journal\": \"Cancer cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — covalent binding site identified, CRISPR KO validation, single lab; note: a subsequent paper (PMID:32857985) challenged Rpn13 as the physiologically relevant target of RA190\",\n      \"pmids\": [\"24332045\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"During proteotoxic stress or partial proteasome inhibition, the proteasome-associated ubiquitin ligase Ube3c/Hul5 selectively poly-ubiquitinates Rpn13 on the 26S proteasome. This autoubiquitination of Rpn13 strongly decreases the proteasome's ability to bind and degrade ubiquitin-conjugated proteins without affecting activity against peptide substrates, constituting an autoinhibitory mechanism that prevents ubiquitin conjugate binding to stalled or defective proteasomes.\",\n      \"method\": \"Mass spectrometry identification of ubiquitinated sites, in vitro ubiquitination assay with purified 26S proteasomes, cell-based stress treatments, proteasome activity assays\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro reconstitution with purified proteasomes, MS identification of sites, cell-based validation with multiple stress conditions\",\n      \"pmids\": [\"24811749\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Crystal structures of UCH-L5/Uch37 in complex with the DEUBAD domain of RPN13 reveal that RPN13 activates Uch37 by repositioning its C-terminal ULD domain and crossover loop to promote substrate binding and catalysis. The related DEUBAD domain of INO80G inhibits UCH-L5 by exploiting the same structural elements but driving a radically different conformation and employing molecular mimicry to block ubiquitin docking.\",\n      \"method\": \"Crystal structure determination, mutagenesis, enzymatic activity assays\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structures with mutagenesis and functional validation, provides atomic-level mechanism\",\n      \"pmids\": [\"25702870\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"The structure of hRpn13 bound to a segment of hRpn2 (its proteasome docking site) was solved; a proline-rich C-terminal hRpn2 extension stretches across a narrow canyon of the ubiquitin-binding hRpn13 Pru domain, blocking an RA190-binding surface. hRpn13 binds preferentially to hRpn2/proteasomes over RA190. RA190 does not affect the hRpn13-Uch37 interaction but directly binds and inactivates Uch37. hRpn13 deletion from HCT116 cells abrogates RA190-induced substrate accumulation at proteasomes.\",\n      \"method\": \"NMR structure determination, biophysical binding assays (SPR, fluorescence polarization), CRISPR/Cas9 cell-based assays\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — NMR structure, multiple biophysical methods, cell-based genetic validation\",\n      \"pmids\": [\"28598414\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Crystal structures of the RPN13 PRU domain in complex with peptides corresponding to the RPN2 C terminus and ubiquitin were determined. The RPN13-binding epitope maps to the C-terminal 14 residues of RPN2. RPN2, ubiquitin, and UCH37 each bind RPN13 with independent energetics. Mutational analysis validated the RPN2-binding interface by SPR and fluorescence polarization.\",\n      \"method\": \"Crystal structure determination, SPR, fluorescence polarization, mutational analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structures with quantitative binding validation using multiple orthogonal biophysical methods\",\n      \"pmids\": [\"28442575\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Rpn13 is involved in selective degradation of iNOS and IκB-α via the Rpn13/UCH37 complex. Knockdown of Rpn13 by siRNA does not lead to global ubiquitinated protein accumulation but impairs iNOS interaction with UCH37. IκB-α, whose proteasomal degradation activates NF-κB, is also a substrate for the Rpn13/UCH37 pathway.\",\n      \"method\": \"siRNA knockdown, co-immunoprecipitation, protein stability/degradation assays\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — co-IP and functional knockdown with specific substrate readout, single lab\",\n      \"pmids\": [\"20634424\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Uch37 oligomerizes in solution and has very low isopeptidase activity due to autoinhibition in which each catalytic domain in the homodimer blocks the other's ubiquitin-binding site. Rpn13C (residues 270–407) disrupts Uch37 oligomerization by sequestering it in a 1:1 stoichiometric complex, activating Uch37 with ~12-fold higher activity. TR-SAXS and FRET confirmed the mode of autoinhibition and activation mechanism.\",\n      \"method\": \"Mutagenesis, NMR, SAXS, TR-SAXS, FRET, biochemical activity assay\",\n      \"journal\": \"Protein & cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple orthogonal structural and biochemical methods in a single study establishing mechanism of Uch37 autoinhibition and Rpn13-mediated activation\",\n      \"pmids\": [\"24752541\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Simultaneous liver-specific deletion of both Rpn13 and Rpn10 in mice causes severe liver injury with massive ubiquitin conjugate accumulation, while single deletion causes only modest impairment. In the double knockout, shuttle proteins mHR23B and ubiquilin/Plic-1 and -4 fail to bind the proteasome, establishing Rpn10 and Rpn13 as the two main receptors for UBL-UBA substrate delivery proteins at the proteasome.\",\n      \"method\": \"Conditional knockout mouse (liver-specific), ubiquitin conjugate accumulation assay, co-immunoprecipitation, genetic rescue by re-expression\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic double-KO with defined phenotype, rescue by re-expression, multiple biochemical readouts\",\n      \"pmids\": [\"26222436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The N-terminal ubiquitin-like (Ubl) domain of parkin binds directly to the Pru domain of Rpn13/ADRM1 via a hydrophobic patch surrounding Ile-44, with >10-fold higher affinity than ubiquitin for the Pru domain. Knockdown of Rpn13 in cells increases parkin levels, abrogates parkin recruitment to the 26S proteasome, and delays clearance of mitochondrial proteins (TIM23, TIM44, TOM20) while enhancing parkin autoubiquitination. Rpn13 knockdown did not impair parkin recruitment to mitochondria or mitophagy initiation.\",\n      \"method\": \"Co-immunoprecipitation, NMR binding analysis, mutational analysis, siRNA knockdown, mitochondrial protein clearance assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — NMR-based binding characterization, mutagenesis, and cell-based functional validation with multiple readouts\",\n      \"pmids\": [\"25666615\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"The C-terminal region of Rpn13 (DEUBAD domain) binds to the tetratricopeptide repeat (TPR) domain of SGTA via a carboxylate clamp mechanism. SGTA overexpression increases steady-state mislocalised membrane protein (MLP) levels in a manner dependent on SGTA's interaction with Rpn13. Expression of the SGTA-binding region of Rpn13 or TPR domain point mutations inhibit SGTA-Rpn13 binding and substantially reduce MLP levels.\",\n      \"method\": \"Co-immunoprecipitation, NMR structural characterization (carboxylate clamp interaction), mutational analysis, protein stability assays\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — structural and functional characterization, single lab, multiple orthogonal approaches\",\n      \"pmids\": [\"26169395\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"The structural basis for SGTA-Rpn13 interaction was characterized: a region of the Rpn13 C-terminal domain is necessary and sufficient for binding to the TPR domain of SGTA through a carboxylate clamp-mediated molecular recognition event, and the interaction was validated in a cellular context.\",\n      \"method\": \"NMR structural characterization, biophysical binding assays, cellular co-immunoprecipitation\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — structural and biophysical validation, single lab\",\n      \"pmids\": [\"27827410\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Although most double-capped 26S proteasomes contain Rpn10/S5a in both 19S particles, only one of the two 19S particles contains Rpn13, defining inherent asymmetry in the 26S proteasome. This was validated in both yeast and mammals using biochemical fractionation and quantitative AQUA-MS methodology.\",\n      \"method\": \"Biochemical fractionation, AQUA-MS quantitation, validation in yeast and mammalian cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — quantitative MS (AQUA) with cross-species validation, single lab\",\n      \"pmids\": [\"24429290\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"SUMOylation of Psmd1/RPN2 (by PIASy E3 ligase) at a critical lysine immediately adjacent to the Adrm1-binding domain regulates the association of Adrm1 with Psmd1/RPN2. The SUMO deconjugating enzyme xSENP1 specifically interacts with Psmd1, and disruption of xSENP1 targeting delays mitotic exit, linking this SUMOylation-mediated regulation to cell division.\",\n      \"method\": \"SUMOylation site mapping, co-immunoprecipitation, Xenopus extract cell cycle assay, SENP1 targeting experiments\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical mapping with functional cell cycle readout, single lab\",\n      \"pmids\": [\"24910440\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"The structure of hRpn13 Pru:hRpn2(940-953):K48-diubiquitin was solved by NMR, revealing that hRpn2-bound hRpn13 selects a dynamic, extended conformation of K48-diubiquitin distinct from previously determined closed crystal structures. hRpn13 exchanges between the two ubiquitins within 100 ms but prefers the proximal ubiquitin due to interactions with the K48 linker region.\",\n      \"method\": \"NMR structure determination, NMR dynamics experiments\",\n      \"journal\": \"Structure\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — NMR structure with dynamics characterization, single lab but rigorous structural method\",\n      \"pmids\": [\"32160516\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"The structure of Rpn13NTD bound to K48-diubiquitin was determined by NMR; Rpn13NTD simultaneously contacts both proximal and distal Ub subunits of K48-diUb in a compact state. The proximal Ub binds similarly to Ub monomer, while distal Ub engages a largely electrostatic surface of Rpn13NTD. A charge-reversal mutation in Rpn13NTD weakens K48-linked chain binding and causes ubiquitinated protein accumulation. Single-molecule FRET showed K48-diUb fluctuates among conformational states, with the compact state selectively enriched by Rpn13NTD.\",\n      \"method\": \"NMR structure, smFRET, charge-reversal mutagenesis, cell-based ubiquitin conjugate assay\",\n      \"journal\": \"Cell discovery\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — NMR structure plus smFRET plus mutagenesis with cellular validation, orthogonal methods\",\n      \"pmids\": [\"30962947\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Phosphorylation of RPN2 Tyr-950 enhances its binding to RPN13. Crystal structure of the RPN2-RPN13 pTyr-950-ubiquitin complex at 1.76 Å reveals specific interactions with positively charged RPN13 side chains that explain the increased affinity without conformational change. Mutagenesis and SPR/fluorescence polarization validated the crystallographic interface.\",\n      \"method\": \"Crystal structure determination (1.76 Å), SPR, fluorescence polarization, mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — high-resolution crystal structure with quantitative biophysical validation using multiple methods\",\n      \"pmids\": [\"31064842\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"An hRpn2-derived 38-amino acid peptide binds the hRpn13 Pru domain with 12 nM affinity. NMR identified the hRpn13-interacting amino acids in this fragment. The peptide immunoprecipitates endogenous Rpn13 from 293T cells and displaces it from the proteasome. Over-expression of the peptide leads to increased ubiquitinated proteins. Conserved F948 and Y950/I951 of hRpn2 are critical for this interaction.\",\n      \"method\": \"NMR binding analysis, Co-immunoprecipitation, mutational analysis, fluorescence polarization\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — NMR and cell-based validation, single lab, multiple orthogonal approaches\",\n      \"pmids\": [\"26466095\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Phospho-ΔNp63α interacts with Rpn13 protein in cisplatin-sensitive SCC cells and this interaction leads to proteasome-dependent degradation of phospho-ΔNp63α while protecting LKB1 from degradation. In SCC cells with impaired ATM-dependent ΔNp63α phosphorylation, non-phosphorylated ΔNp63α fails to form complexes with Rpn13, allowing Rpn13 to target LKB1 for proteasome-dependent degradation, modulating cisplatin-induced autophagy.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, protein stability assays\",\n      \"journal\": \"Aging\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP approach, single lab, limited mechanistic detail in abstract\",\n      \"pmids\": [\"21191146\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Phospho-ΔNp63α binds to the RPN13 gene promoter through a TP63-responsive element and, together with co-factors DDIT3/CHOP, NF-Y and NF-κB, activates RPN13 gene transcription. Cisplatin-induced, phospho-ΔNp63α-dependent RPN13 transcription leads to NOS2/iNOS degradation; RPN13 siRNA knockdown rescues NOS2 from this degradation.\",\n      \"method\": \"Chromatin immunoprecipitation, promoter-binding assay, siRNA knockdown, protein degradation assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — ChIP-based transcription factor binding with functional siRNA rescue, single lab\",\n      \"pmids\": [\"20959455\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Adrm1 interacts with Atp6v0d2 (d2 isoform of vacuolar H+-ATPase V0 domain) as identified by yeast two-hybrid screening and confirmed by co-immunoprecipitation. Adrm1 is required for cell migration and osteoclast maturation.\",\n      \"method\": \"Yeast two-hybrid screen, co-immunoprecipitation, cell migration assay, osteoclast differentiation assay\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — yeast two-hybrid plus Co-IP, single lab, limited mechanistic follow-up\",\n      \"pmids\": [\"19818731\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Shigella effector IpaH4.5 (an E3 ubiquitin ligase) targets RPN13 and induces its degradation via the ubiquitin-proteasome system. IpaH4.5-mediated RPN13 degradation causes dysfunction of the 19S regulatory particle, inhibiting guidance of ubiquitinated proteins to the 20S core particle and suppressing proteasome-catalyzed peptide splicing. This reduces antigen cross-presentation to CD8+ T cells via MHC class I. In RPN13 knockout MEFs, loss of RPN13 suppressed CD8+ T cell priming during Shigella infection.\",\n      \"method\": \"Bacterial infection model, RPN13 knockout MEFs, antigen presentation assay, T cell activation assay\",\n      \"journal\": \"Cellular microbiology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with defined immune functional readout, bacterial effector mechanistic dissection, single lab\",\n      \"pmids\": [\"30414351\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ADRM1/Rpn13 upregulates UCH37 expression and binds UCH37 to activate its deubiquitination activity. Activated UCH37 enhances deubiquitination of ALK5 (activin receptor-like kinase 5), stabilizing ALK5 expression and maintaining extracellular matrix homeostasis in cartilage. Global knockout of ADRM1 accelerates cartilage degeneration in DMM-induced OA model.\",\n      \"method\": \"Global knockout mouse, conditional overexpression, co-immunoprecipitation, deubiquitination assay, ALK5 stability assay\",\n      \"journal\": \"International journal of biological macromolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — in vivo KO with biochemical mechanism (UCH37 activation of ALK5 deubiquitination) validated, single lab\",\n      \"pmids\": [\"37406898\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"HDAC8 interacts with the proteasome receptor ADRM1, and inhibition of HDAC8 decreases MGMT protein levels in GBM cell lines in a manner dependent on this interaction. Treatment with TMZ disrupts the HDAC8-ADRM1 interaction exclusively in TMZ-sensitive cells.\",\n      \"method\": \"Co-immunoprecipitation, HDAC8 inhibitor/shRNA knockdown, western blotting for MGMT levels\",\n      \"journal\": \"Genes & cancer\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP, single lab, limited mechanistic detail\",\n      \"pmids\": [\"31798765\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"A spliced ADRM1 variant lacking exon 9 (ADRM1-ΔEx9) interacts with a different deubiquitinase partner (BAP1 instead of UCH37) via its altered C-terminus, redirecting proteasome specificity. The novel exon 8-10 fusion creates a de novo binding site for tumor suppressor FBXW7, resulting in its selective proteasome-mediated degradation.\",\n      \"method\": \"Proteome Profiler Human Ubiquitin Array, co-immunoprecipitation, patient-derived organoids, xenograft models, murine hydrodynamic transfection\",\n      \"journal\": \"Journal of hepatology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — multiple experimental systems (organoids, mouse model, proteomic array), single lab; note this concerns a splice variant of ADRM1\",\n      \"pmids\": [\"39788431\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ADRM1/Rpn13 functions in DNA end resection as part of a proteasome variant (CDW19S) that assembles at broken chromatin. ADRM1Rpn13 inhibits DNA resection and is removed by CRL4WDR70-catalyzed ubiquitination upon commitment to extensive resection. HBx protein (from HBV) interferes with ADRM1Rpn13 degradation, leading to a resection barrier and homologous recombination deficiency.\",\n      \"method\": \"Co-immunoprecipitation, DNA damage repair assays, ubiquitination assay, HBx expression, HR assay\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — biochemical and cell-based dissection of a novel non-proteasomal function for ADRM1 in DNA repair, single lab\",\n      \"pmids\": [\"37815873\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Rpn13 and Rpn10 are the main proteasomal receptors for ubiquitin-like (UBL) domain-containing shuttle proteins (mHR23B and ubiquilins/Plic-1 and -4) that deliver ubiquitinated cargo to the proteasome. In double Rpn10/Rpn13 knockout hepatocytes, these shuttle proteins fail to associate with the proteasome.\",\n      \"method\": \"Conditional double-knockout mouse, co-immunoprecipitation, genetic rescue by re-expression\",\n      \"journal\": \"PLoS genetics\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — clean genetic double-KO with specific molecular readout (shuttle protein binding) confirmed by rescue\",\n      \"pmids\": [\"26222436\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"MHC class I processing of the NY-ESO-1 antigen is governed by both Rpn10 and Rpn13 ubiquitin receptors as well as by immunoproteasomes, and is regulated by non-canonical ubiquitination on non-lysine sites rather than canonical K48-linked chains at K124.\",\n      \"method\": \"siRNA knockdown of Rpn13 and Rpn10, antigen presentation assay, ubiquitination site mutagenesis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — siRNA knockdown with specific antigen processing readout, mutagenesis of ubiquitination sites, single lab\",\n      \"pmids\": [\"26903513\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Conditional deletion of Rpn13 in primordial germ cells (PGCs) in mice causes infertility in both sexes with dramatic reduction in germ cells in testes and ovaries. Mechanistically, Rpn13 deletion disrupts 26S proteasome assembly in germ cells, reduces PGC numbers, and blocks meiosis of spermatocytes at the zygotene stage during prophase I.\",\n      \"method\": \"Conditional knockout mouse (PGC-specific), proteasome assembly assay, histological and meiosis staging analysis\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO with specific cellular and molecular phenotypes, single lab\",\n      \"pmids\": [\"40422199\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A Protea-Tac degrader incorporating ADRM1/Rpn13 as the ubiquitin receptor component integrates into 26S proteasomes without altering their structural or functional integrity. Proximity of target proteins (c-Fos, BRD4, Flag-TDP43, HA-tau, GFP-ODC) to the proteasome via Rpn13-antibody chimera results in their ubiquitin-independent degradation.\",\n      \"method\": \"Chimeric protein reconstitution, proteasome integrity assays, targeted degradation assays for multiple substrates, in vivo xenograft mouse model\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — reconstitution approach with multiple substrates and in vivo validation, preprint not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.08.18.670774\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"PhIX-MS and cryo-EM placed the thioredoxin domain of redox sensor TXNL1 near RPN2/PSMD1 and RPN13/ADRM1 at the proteasome regulatory particle, positioning it to reduce substrates prior to proteolysis.\",\n      \"method\": \"PhIX-MS (photo-induced in situ crosslinking-mass spectrometry), cryo-electron microscopy, AlphaFold modeling\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 2 / Weak — structural localization by crosslinking MS/cryo-EM, preprint, TXNL1-RPN13 proximity not functionally validated\",\n      \"pmids\": [\"bio_10.1101_2025.07.31.667872\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"RA190 has no measurable effect on any of the known interactions of Rpn13 in vitro, and no physical engagement of Rpn13 by RA190 was detected in cellulo on C88 or any other residue. Chemical proteomics in two cell lines showed that dozens of other proteins are heavily engaged by RA190. Increasing or reducing Rpn13 levels had no effect on RA190 sensitivity of HeLa or melanoma cells, indicating Rpn13 is NOT the physiologically relevant target of RA190.\",\n      \"method\": \"In vitro protein interaction assays, chemical proteomics (activity-based protein profiling), cell viability assays with Rpn13 modulation\",\n      \"journal\": \"Cell chemical biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — chemical proteomics plus multiple functional assays across two cell lines; this is a negative finding contradicting PMID:24332045 regarding RA190's principal target\",\n      \"pmids\": [\"32857985\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ADRM1/Rpn13 is a subunit of the 19S regulatory particle of the 26S proteasome that functions as a ubiquitin receptor (via its N-terminal Pru domain, which binds K48-linked polyubiquitin chains bivalently and also docks into the proteasome through direct interaction with the C-terminus of RPN2/PSMD1), and as a deubiquitinase adaptor (via its C-terminal DEUBAD domain, which recruits and allosterically activates the deubiquitinase UCH37/UCHL5 by disrupting its autoinhibitory oligomerization); Rpn13 is present asymmetrically in only one of the two 19S caps of the double-capped proteasome, acts redundantly with Rpn10 for bulk ubiquitin conjugate recognition but is required for recruitment of specific substrates (including iNOS, IκB-α, parkin, and mislocalised membrane proteins via SGTA) and shuttle proteins (mHR23B, ubiquilins); its activity is regulated by Ube3c/Hul5-mediated autoubiquitination (autoinhibitory under proteotoxic stress) and by SUMOylation of its docking partner RPN2; additionally, ADRM1 participates in DNA double-strand break end resection where it inhibits resection and is removed by CRL4WDR70 ubiquitination, and its deletion disrupts 26S proteasome assembly and blocks meiosis in germ cells.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ADRM1/Rpn13 is a subunit of the 19S regulatory particle of the 26S proteasome that couples recognition of ubiquitinated substrates to their disassembly and degradation [#0, #2]. It is recruited to the proteasome by direct docking onto the C-terminal tail of RPN2/PSMD1, an interaction mapped to the final ~14 residues of RPN2 binding the Rpn13 Pru domain and further tuned by RPN2 modification — Tyr-950 phosphorylation enhances affinity while SUMOylation of RPN2 adjacent to the docking site regulates Adrm1 association in a manner linked to mitotic exit [#8, #19, #16]. Through its N-terminal Pru domain Rpn13 acts as a proteasomal ubiquitin receptor, binding K48-linked diubiquitin with high affinity by engaging both proximal and distal ubiquitin moieties and selecting a defined chain conformation [#0, #18, #17]. The Pru domain also captures the ubiquitin-like domains of UBL-UBA shuttle proteins; genetic ablation in mice shows that Rpn13 and Rpn10 are the two main receptors that recruit shuttle factors mHR23B and ubiquilins and that they act redundantly for bulk conjugate handling, with double loss causing massive conjugate accumulation [#0, #11, #29]. Through its C-terminal DEUBAD domain Rpn13 functions as a deubiquitinase adaptor, recruiting UCH37/UCHL5 and allosterically activating it by disrupting an autoinhibitory homodimer and repositioning the enzyme's ULD domain and crossover loop to license substrate binding and catalysis [#1, #6, #10]. Rpn13 directs the proteasome toward specific substrates and partners — including iNOS and IκB-α via the Rpn13/UCH37 axis, parkin via high-affinity Ubl-domain docking, and mislocalised membrane proteins delivered by SGTA through a carboxylate-clamp interaction with the C-terminal domain [#9, #12, #13]. Its conjugate-binding capacity is itself regulated: under proteotoxic stress the proteasome-associated ligase Ube3c/Hul5 autoubiquitinates Rpn13, an autoinhibitory mechanism that selectively suppresses ubiquitin-conjugate degradation at stalled proteasomes [#5]. Rpn13 is present asymmetrically in only one of the two 19S caps of double-capped proteasomes [#15]. Beyond the proteasome, Rpn13 participates in DNA double-strand break end resection, where it inhibits resection and is removed by CRL4WDR70-mediated ubiquitination [#28], and its deletion in germ cells disrupts 26S proteasome assembly and blocks meiosis, causing infertility [#31].\",\n  \"teleology\": [\n    {\n      \"year\": 2006,\n      \"claim\": \"Established that Adrm1/hRpn13 is a bona fide 19S subunit and the proteasomal recruiter of the deubiquitinase Uch37, defining a physical link between a regulatory particle protein and chain disassembly.\",\n      \"evidence\": \"Affinity purification, co-IP, in vitro reconstitution and ubiquitin-AMC assays, siRNA knockdown in mammalian cells\",\n      \"pmids\": [\"16906146\", \"17139257\", \"16815440\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve the structural basis of Uch37 activation\", \"Conflicting reports on whether knockdown affects bulk degradation\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Defined Rpn13's N-terminal Pru domain as a high-affinity K48-polyubiquitin receptor functionally linked to Rpn10, answering how the proteasome recognizes ubiquitinated cargo through a second receptor.\",\n      \"evidence\": \"NMR, affinity measurements, mutagenesis, yeast genetic epistasis\",\n      \"pmids\": [\"18497817\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not establish in vivo substrate specificity\", \"Mode of bivalent diubiquitin engagement not yet resolved at atomic level\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Connected Rpn13 to degradation of defined regulatory substrates (iNOS, IκB-α) via the Rpn13/UCH37 complex rather than bulk turnover, indicating a specialized substrate-selective role.\",\n      \"evidence\": \"siRNA knockdown, co-IP, protein stability assays; plus ChIP-based transcriptional regulation of RPN13 by phospho-ΔNp63α\",\n      \"pmids\": [\"20634424\", \"20959455\", \"21191146\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Substrate selectivity mechanism not structurally defined\", \"ΔNp63α/LKB1 link rests on single Co-IP without reciprocal validation\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Resolved the mechanism of UCH37 activation and revealed two layers of regulation — Rpn13-mediated disruption of UCH37 autoinhibitory oligomerization, and stress-induced autoubiquitination of Rpn13 itself that shuts down conjugate binding.\",\n      \"evidence\": \"NMR/SAXS/TR-SAXS/FRET activity reconstitution; MS site mapping and in vitro ubiquitination with purified 26S proteasomes; AQUA-MS proteasome asymmetry mapping; RPN2 SUMOylation site mapping\",\n      \"pmids\": [\"24752541\", \"24811749\", \"24429290\", \"24910440\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological triggers of Ube3c/Hul5 autoubiquitination incompletely defined\", \"Functional consequence of 19S asymmetry for Rpn13 unresolved\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Genetically established Rpn13 and Rpn10 as the two principal proteasomal receptors for UBL-UBA shuttle proteins, and broadened Rpn13's substrate-targeting repertoire to parkin and SGTA-delivered mislocalised membrane proteins.\",\n      \"evidence\": \"Liver-specific conditional double-knockout mice with rescue; NMR binding and siRNA functional assays for parkin; carboxylate-clamp NMR/co-IP for SGTA\",\n      \"pmids\": [\"26222436\", \"25666615\", \"26169395\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative contribution of Rpn13 vs Rpn10 for individual substrates not fully partitioned\", \"SGTA-Rpn13 axis characterized in a single lab\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Provided atomic-level mechanism for both arms of Rpn13: how the DEUBAD domain repositions UCH37 to activate it (versus INO80G's inhibitory mimicry), and how the Pru domain docks onto the RPN2 C-terminus.\",\n      \"evidence\": \"Crystal structures of UCH-L5:DEUBAD and Pru:RPN2/ubiquitin complexes with mutagenesis and SPR/FP validation\",\n      \"pmids\": [\"25702870\", \"28442575\", \"26466095\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Conformational coupling between RPN2 docking and ubiquitin binding within intact proteasome unresolved\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined how RPN2 docking masks the Pru ubiquitin-binding surface and clarified that the RA190 inhibitor binds and inactivates UCH37 rather than disrupting Rpn13-UCH37, refining the drug-target relationship.\",\n      \"evidence\": \"NMR structure of hRpn13:hRpn2 segment, SPR/FP, CRISPR cell-based assays\",\n      \"pmids\": [\"28598414\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In-cell selectivity of RA190 not yet resolved at this stage\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established the structural basis of bivalent K48-diubiquitin recognition and showed that RPN2 Tyr-950 phosphorylation enhances Rpn13 docking, defining how chain selection and proteasome anchoring are tuned.\",\n      \"evidence\": \"NMR structures with dynamics, smFRET, charge-reversal mutagenesis with cellular conjugate readout; 1.76 Å crystal structure of pTyr-RPN2:Rpn13:ubiquitin\",\n      \"pmids\": [\"32160516\", \"30962947\", \"31064842\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Kinase responsible for RPN2 Tyr-950 phosphorylation not identified in corpus\", \"Physiological context selecting open vs compact diubiquitin states unclear\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Challenged Rpn13 as the physiologically relevant target of the RA190 inhibitor, showing no in-cell engagement of Rpn13 and Rpn13-independent toxicity.\",\n      \"evidence\": \"In vitro interaction assays, chemical proteomics in two cell lines, Rpn13-modulation viability assays\",\n      \"pmids\": [\"32857985\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Directly contradicts earlier CRISPR-based RA190 target assignment\", \"True target(s) responsible for RA190 toxicity not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Extended Rpn13 function beyond canonical degradation — into substrate-stabilizing deubiquitination (UCH37/ALK5 in cartilage), splice-variant rewiring of deubiquitinase and substrate partners, and a non-proteasomal role restraining DNA end resection.\",\n      \"evidence\": \"Global/conditional knockout mice and DMM OA model; ubiquitin-array, organoid and xenograft profiling of ADRM1-ΔEx9; co-IP and HR/ubiquitination assays for CDW19S/CRL4WDR70 resection\",\n      \"pmids\": [\"37406898\", \"39788431\", \"37815873\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Each non-canonical role characterized by a single lab\", \"How a degradation receptor stabilizes a substrate (ALK5) mechanistically incompletely defined\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Demonstrated a physiological requirement for Rpn13 in 26S proteasome assembly during germ-cell development and exploited Rpn13 as the receptor module for ubiquitin-independent targeted degradation.\",\n      \"evidence\": \"PGC-specific conditional knockout with proteasome assembly and meiosis staging; Protea-Tac chimeric reconstitution with multiple substrates and xenograft validation (preprint)\",\n      \"pmids\": [\"40422199\", \"bio_10.1101_2025.08.18.670774\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Molecular basis of Rpn13's assembly requirement not defined\", \"Degrader study is a preprint not yet peer-reviewed\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How Rpn13's distinct activities — ubiquitin recognition, UCH37 activation, shuttle/substrate recruitment, and non-proteasomal resection control — are spatially and temporally coordinated within a single asymmetric cap remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated model linking 19S asymmetry to functional specialization\", \"Regulation switching between proteasomal and DNA-repair roles unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0031386\", \"supporting_discovery_ids\": [0, 18, 17, 12]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [1, 6, 10]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [1, 13, 29]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [3, 2]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [28]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 2, 11]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [1, 6]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [24, 30]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [28]}\n    ],\n    \"complexes\": [\n      \"26S proteasome 19S regulatory particle\"\n    ],\n    \"partners\": [\n      \"UCHL5\",\n      \"PSMD1\",\n      \"PSMD4\",\n      \"RAD23B\",\n      \"PARK2\",\n      \"SGTA\",\n      \"UBE3C\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":9,"faith_total":9,"faith_pct":100.0}}