{"gene":"MKRN2","run_date":"2026-06-10T02:59:50","timeline":{"discoveries":[{"year":2017,"finding":"MKRN2 is a ubiquitin E3 ligase that binds to the p65 subunit of NF-κB, promotes its polyubiquitination and proteasome-dependent degradation through the MKRN2 RING finger domain, thereby suppressing p65-mediated NF-κB transactivation. MKRN2 and PDLIM2 synergistically promote polyubiquitination and degradation of p65. MKRN2 was identified via yeast two-hybrid screening as a PDLIM2-interacting protein. Knockdown of MKRN2 in dendritic cells resulted in increased nuclear p65 and augmented proinflammatory cytokine production.","method":"Yeast two-hybrid screening, co-immunoprecipitation, in vitro ubiquitination assay, RING domain mutagenesis, siRNA knockdown, NF-κB reporter assay","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — multiple orthogonal methods including yeast two-hybrid, Co-IP, in vitro ubiquitination, RING domain mutagenesis, and functional KD with cytokine readout in one study","pmids":["28378844"],"is_preprint":false},{"year":2018,"finding":"MKRN2 inhibits migration and invasion of non-small-cell lung cancer cells through downregulation of the PI3K/Akt pathway, as demonstrated by altered MKRN2 expression in NSCLC cell lines.","method":"siRNA knockdown and overexpression in NSCLC cell lines, migration/invasion assays, western blotting for PI3K/Akt pathway components","journal":"Journal of experimental & clinical cancer research : CR","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — KD/OE with defined cellular phenotype and pathway placement, but no direct substrate identification for PI3K/Akt regulation; single lab","pmids":["30103781"],"is_preprint":false},{"year":2020,"finding":"MKRN2 ubiquitinates IGF2BP3 (an RNA-binding protein) as a direct substrate, leading to its degradation. This MKRN2-mediated ubiquitination of IGF2BP3 regulates the expression of CD44 and PDPN (downstream IGF2BP3 targets) in neuroblastoma SHSY5Y cells. MKRN2 knockdown promotes proliferation and migration of these cells.","method":"shRNA knockdown, co-immunoprecipitation, in vitro ubiquitination assay, western blotting","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and in vitro ubiquitination to identify substrate, functional KD phenotype, single lab with two orthogonal methods","pmids":["32560817"],"is_preprint":false},{"year":2016,"finding":"Mkrn2 knockout in mice causes male infertility characterized by low sperm number, poor motility, aberrant morphology, spermiation failure, and misarrangement of ectoplasmic specialization in testes. Loss of Mkrn2 results in decreased expression of Odf2, a protein vital for spermatogenesis.","method":"Knockout mouse model, histology, sperm analysis, western blotting for Odf2 expression","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KO mouse with defined spermatogenic phenotypes and downstream molecular readout (Odf2), single lab","pmids":["28008940"],"is_preprint":false},{"year":2020,"finding":"Mkrn2 deficiency leads to abnormally high apoptosis in testes. MKRN2 inhibits expression of p53 apoptosis effector PERP, and loss of Mkrn2 upregulates PERP, implicating a Mkrn2-p53/PERP signaling axis in protecting germ cells from excessive apoptosis during spermatogenesis.","method":"Mkrn2 knockout mouse model, digital gene expression profiling, GSEA/KEGG pathway analysis, protein expression analysis","journal":"Asian journal of andrology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — pathway placement inferred from transcriptomics and KO phenotype without direct biochemical confirmation of MKRN2-p53/PERP interaction; single lab","pmids":["31489847"],"is_preprint":false},{"year":2020,"finding":"MKRN2 directly interacts with and ubiquitinates p53, promoting its degradation, thereby promoting melanoma cell proliferation in a p53-dependent manner. Downregulation of MKRN2 inhibited melanoma cell growth specifically in cells with functional p53 (rescued in p53-knockout cells).","method":"Co-immunoprecipitation, GST pulldown, in vitro ubiquitination assay, CRISPR-Cas9 p53 knockout, MTT and colony formation assays, shRNA knockdown","journal":"Oncology letters","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro ubiquitination assay, reciprocal Co-IP, GST pulldown, and genetic rescue (p53 KO) in one study confirming direct substrate relationship","pmids":["32194692"],"is_preprint":false},{"year":2020,"finding":"MKRN2 physically interacts with GLE1 (a DEAD-box helicase activator implicated in mRNA export termination), as identified by affinity-purification mass spectrometry. MKRN2 binds selectively to the 3' UTR of a subset of mRNAs, and nuclear export of MKRN2-associated mRNAs is enhanced upon MKRN2 knockdown. Genetic epistasis in zebrafish shows that morpholino knockdown or CRISPR knockout of MKRN2 partially rescues retinal developmental defects caused by GLE1 depletion, placing MKRN2 downstream of GLE1 in mRNA export regulation.","method":"Affinity-purification mass spectrometry, ribonomic approaches (RNA-binding profiling), morpholino knockdown, CRISPR/Cas9 knockout in zebrafish, genetic epistasis","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — AP-MS interaction, ribonomic binding assay, and in vivo genetic epistasis across two knockdown strategies; multiple orthogonal methods in one study","pmids":["32460013"],"is_preprint":false},{"year":2024,"finding":"MKRN2 associates with influenza A virus (IAV) mRNAs and positively regulates IAV mRNA nuclear-cytoplasmic trafficking, potentially through an association with GLE1. In the absence of MKRN2, IAV mRNAs accumulate in the nucleus of infected cells and may be degraded by the nuclear RNA exosome.","method":"RNA interactome capture (RIC), MKRN2 knockdown, RNA FISH, nuclear/cytoplasmic fractionation","journal":"PLoS pathogens","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RIC and functional KD with nuclear accumulation readout, single lab, mechanistic link to GLE1 inferred but not directly demonstrated","pmids":["38753876"],"is_preprint":false},{"year":2025,"finding":"MKRN2 selectively inhibits IL-6 expression in LPS-activated macrophages. Mechanistically, after binding to Il6 mRNA, MKRN2 attaches K29 polyubiquitin chains to the Lys179 residue of PAIP1 (a translation initiation coactivator), which blocks the PAIP1-eIF4A interaction and inhibits translational efficiency of Il6 mRNA. LysM-Cre+Mkrn2fl/fl mice showed increased serum IL-6 after LPS and increased severity of experimental colitis.","method":"Conditional knockout mouse model, in vitro ubiquitination assay, mutagenesis of PAIP1 K179, Co-immunoprecipitation, polysome profiling, RNA-binding assay","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro ubiquitination with site-specific mutagenesis (K179), Co-IP, polysome profiling, and in vivo conditional KO with defined phenotype; multiple orthogonal methods","pmids":["40524017"],"is_preprint":false},{"year":2022,"finding":"MKRN2 promotes ubiquitination-mediated degradation of PKM2 and attenuates its effect on ERK, thereby inhibiting gastric cancer cell proliferation.","method":"Overexpression and knockdown in gastric cancer cell lines, co-immunoprecipitation, ubiquitination assay, western blotting for ERK pathway, in vivo xenograft","journal":"Aging","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assay identify PKM2 as substrate, functional KD/OE phenotype with pathway placement, single lab","pmids":["35196650"],"is_preprint":false},{"year":2023,"finding":"MKRN2 directly interacts with STAT1 in testis and MEF cells, as shown by Co-IP. MKRN2 also regulates expression of SIX4 and TNC via EBF2 transcription factor in mice. Loss of Mkrn2 in knockout mice results in decreased STAT1, SIX4, and TNC expression and male infertility.","method":"Mkrn2 knockout mouse model, co-immunoprecipitation, western blotting, transcriptional analysis","journal":"Frontiers in endocrinology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP for STAT1 interaction; the SIX4/TNC/EBF2 mechanism is inferred from expression data without direct biochemical validation","pmids":["36967804"],"is_preprint":false},{"year":2025,"finding":"MKRN2 interacts with PPP2CA (Protein Phosphatase 2 Catalytic Subunit Alpha) and promotes K48-linked ubiquitination at its K41 residue, leading to proteasomal degradation of PPP2CA. This results in increased β-catenin phosphorylation and decreased β-catenin levels, causing inactivation of the Wnt signaling pathway and increased apoptosis in clear cell renal cell carcinoma cells.","method":"Co-immunoprecipitation, immunofluorescence co-localization, ubiquitination assay with K48-linkage specificity, western blotting for β-catenin and Wnt pathway, in vivo xenograft","journal":"International journal of biological sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, site-specific ubiquitination (K41), K48-linkage characterization, downstream pathway readout; single lab","pmids":["40959281"],"is_preprint":false},{"year":2025,"finding":"MKRN2 functions as an E3 ubiquitin ligase that targets NF-κB p65 for proteasomal degradation, constraining NF-κB/COX2-mediated inflammatory signaling. MKRN2 deficiency promotes M1-to-M2 macrophage polarization in tumor-associated macrophages, leading to accelerated tumor growth in MKRN2 knockout mice.","method":"MKRN2 knockout mouse model, in vivo tumor implantation, flow cytometry for macrophage phenotyping, reconstitution experiments with MKRN2 overexpression","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vivo KO model with quantitative immune profiling and functional reconstitution; p65 ubiquitination mechanism references prior work (PMID:28378844); single lab","pmids":["40925500"],"is_preprint":false},{"year":2025,"finding":"MKRN2 promotes ubiquitination and proteasomal degradation of p53 in lung epithelial cells, attenuating LPS-induced apoptosis. Co-IP confirmed the direct interaction between MKRN2 and p53, and transcriptome sequencing confirmed MKRN2 modulates apoptosis via the p53 signaling pathway.","method":"Co-immunoprecipitation, in vitro ubiquitination assay, transcriptome sequencing, adenovirus-mediated overexpression, siRNA knockdown, in vivo LPS-ARDS mouse model","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assay, transcriptomics, and in vivo model; consistent with prior p53 ubiquitination findings (PMID:32194692); single lab","pmids":["40885043"],"is_preprint":false},{"year":2026,"finding":"The covalent compound DPB directly targets MKRN2 by covalently modifying the Cys335 residue, acting as a molecular glue that recruits the ribosomal protein RPS7 to MKRN2. This induces ubiquitination and proteasomal degradation of RPS7, triggering nucleolar stress and apoptosis selectively in p53-deficient NSCLC cells. The synthetic lethal effect is entirely dependent on the MKRN2-RPS7 axis.","method":"Quantitative thiol-reactivity proteomics (QTRP), co-immunoprecipitation mass spectrometry, site-directed mutagenesis of Cys335, genetic knockout/rescue studies, biophysical binding assays, in vivo orthotopic mouse model","journal":"British journal of pharmacology","confidence":"High","confidence_rationale":"Tier 1 / Strong — covalent target identification by QTRP, mutagenesis of active site Cys335, Co-IP-MS for neo-substrate recruitment, genetic rescue studies confirming mechanism; multiple orthogonal methods","pmids":["41991154"],"is_preprint":false},{"year":2026,"finding":"MKRN2 mediates ubiquitination of the RNA-binding protein CSDE1 at four specific lysine residues (K81, K91, K208, K727), identified by mass spectrometry and validated by mutagenesis. MKRN2 and CSDE1 form co-localized condensates via liquid-liquid phase separation (LLPS), which is disrupted by functional impairment of either protein. Mkrn2 knockout mice exhibit sex-specific social abnormalities recapitulating ASD features, and CSDE1-dependent targets MARK1 and HNRNPUL2 mRNAs are regulated in a ubiquitination-dependent manner.","method":"Mass spectrometry substrate identification, mutagenesis of CSDE1 lysine residues, LLPS assays in HEK293 and SH-SY5Y cells, Mkrn2 knockout mouse behavioral testing, western blotting","journal":"Frontiers in cellular neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — MS substrate identification validated by mutagenesis, LLPS assay, and in vivo KO with behavioral phenotype; single lab","pmids":["41757349"],"is_preprint":false},{"year":2025,"finding":"MKRN2 localizes to stress granules (SGs) in a manner dependent on active ubiquitination (UBA1 activity). Within SGs, MKRN2 promotes proper SG formation and disassembly following stress recovery by preventing accumulation of defective ribosomal products (DRiPs). MKRN2 was identified as a SG-associated E3 ligase by proximity proteomics.","method":"Proximity proteomics (BioID), UBA1 inhibitor treatment, live-cell imaging of SG dynamics, loss-of-function assays","journal":"bioRxiv","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity proteomics plus functional SG assays with pharmacological inhibition; preprint, not yet peer-reviewed","pmids":["bio_10.1101_2025.10.15.682570"],"is_preprint":true},{"year":2026,"finding":"MKRN2 promotes cell cycle progression and proliferation of hepatocellular carcinoma cells through activation of the p38 MAPK signaling pathway, leading to c-Myc activation. MKRN2 depletion arrests cells at the G1/S transition.","method":"siRNA knockdown, RNA-seq, flow cytometry cell cycle analysis, western blotting for p38 MAPK and c-Myc, CCK-8/EdU/colony formation assays, in vivo xenograft","journal":"Human cell","confidence":"Low","confidence_rationale":"Tier 3 / Weak — pathway placement inferred from RNA-seq and KD phenotype without direct biochemical identification of MKRN2 substrate or binding partner in the p38/c-Myc axis; single lab","pmids":["41741886"],"is_preprint":false}],"current_model":"MKRN2 is an RNA-binding E3 ubiquitin ligase that suppresses NF-κB-mediated inflammation by ubiquitinating p65 for proteasomal degradation (cooperatively with PDLIM2), inhibits IL-6 translation by K29-polyubiquitinating PAIP1 at K179 to block PAIP1-eIF4A interaction, interacts with GLE1 at the nuclear pore to selectively regulate mRNA nuclear export (including for viral RNAs), ubiquitinates multiple substrates including p53, IGF2BP3, PKM2, PPP2CA, CSDE1, and RPS7 via its RING finger domain, localizes to stress granules in a ubiquitination-dependent manner to regulate granulostasis, and is required for normal spermatogenesis and male fertility in mice."},"narrative":{"mechanistic_narrative":"MKRN2 is a RING-finger E3 ubiquitin ligase that combines protein ubiquitination with RNA binding to control inflammation, mRNA fate, and cell survival [PMID:28378844, PMID:40524017, PMID:32460013]. As a ubiquitin ligase it directs proteasomal degradation of a range of substrates through its RING domain, including the NF-κB subunit p65—which it ubiquitinates cooperatively with PDLIM2 to suppress NF-κB transactivation and proinflammatory cytokine output [PMID:28378844, PMID:40925500]—and the tumor suppressor p53, which it targets to promote cell proliferation and limit apoptosis [PMID:32194692, PMID:40885043]. Additional substrates established by direct interaction and ubiquitination assays include the RNA-binding protein IGF2BP3 [PMID:32560817], the glycolytic enzyme PKM2 [PMID:35196650], the phosphatase catalytic subunit PPP2CA, which it modifies with K48-linked chains at K41 to inactivate Wnt/β-catenin signaling [PMID:40959281], and the RNA-binding protein CSDE1, with which it also forms co-localized phase-separated condensates [PMID:41757349]. Beyond degradative ubiquitination, MKRN2 acts as an RNA-binding translational and export regulator: it binds Il6 mRNA and attaches non-degradative K29 polyubiquitin chains to PAIP1 at K179, blocking the PAIP1–eIF4A interaction to selectively repress IL-6 translation [PMID:40524017], and it interacts with the mRNA export factor GLE1 to control nuclear export of a subset of cellular and influenza A virus mRNAs [PMID:32460013, PMID:38753876]. MKRN2 is required for normal spermatogenesis and male fertility in mice, where its loss causes spermiation failure and excess germ-cell apoptosis [PMID:28008940]. A covalent molecular-glue compound exploits MKRN2 by modifying Cys335 to recruit and degrade the ribosomal protein RPS7, producing synthetic lethality in p53-deficient lung cancer cells [PMID:41991154].","teleology":[{"year":2016,"claim":"Established the first organismal requirement for MKRN2, showing it is essential for male fertility before any substrate was known.","evidence":"Mkrn2 knockout mouse with histology, sperm analysis, and Odf2 readout","pmids":["28008940"],"confidence":"Medium","gaps":["Did not identify the direct ubiquitination substrate driving spermatogenic failure","Odf2 link is correlative, not shown to be a ubiquitination target"]},{"year":2017,"claim":"Defined MKRN2 as a functional RING E3 ligase by identifying p65 as a degradation substrate, linking it to negative regulation of NF-κB inflammation.","evidence":"Yeast two-hybrid, Co-IP, in vitro ubiquitination, RING mutagenesis, and siRNA knockdown in dendritic cells","pmids":["28378844"],"confidence":"High","gaps":["Ubiquitin chain linkage on p65 not characterized","Relative contributions of MKRN2 vs PDLIM2 to ligase activity not separated"]},{"year":2018,"claim":"Extended MKRN2 function to cancer cell behavior, placing it as a suppressor of migration/invasion via the PI3K/Akt pathway.","evidence":"Knockdown/overexpression in NSCLC lines with invasion assays and pathway blotting","pmids":["30103781"],"confidence":"Medium","gaps":["No direct ubiquitination substrate linking MKRN2 to PI3K/Akt","Mechanism of pathway downregulation unresolved"]},{"year":2020,"claim":"Identified RNA-binding proteins and p53 as direct MKRN2 substrates, broadening its substrate repertoire and connecting it to proliferation control.","evidence":"Co-IP, GST pulldown, in vitro ubiquitination, and CRISPR p53-knockout rescue in melanoma and neuroblastoma cells","pmids":["32194692","32560817"],"confidence":"High","gaps":["Chain linkage types not defined for these substrates","Whether the same RING residues govern all substrates not tested"]},{"year":2020,"claim":"Revealed an unexpected RNA-export function by placing MKRN2 downstream of GLE1, showing it binds mRNA 3' UTRs and restrains nuclear export.","evidence":"AP-MS, ribonomic RNA-binding profiling, and zebrafish morpholino/CRISPR genetic epistasis","pmids":["32460013"],"confidence":"High","gaps":["Whether export regulation requires ligase activity not established","Direct GLE1 binding interface unmapped"]},{"year":2020,"claim":"Connected MKRN2 loss to germ-cell apoptosis through a p53/PERP axis, contextualizing the fertility phenotype.","evidence":"Mkrn2 KO mouse with transcriptomics and protein expression analysis","pmids":["31489847"],"confidence":"Low","gaps":["p53/PERP axis inferred from expression, no direct biochemical interaction shown","Single lab, no rescue"]},{"year":2022,"claim":"Added PKM2 as a degradation substrate, linking MKRN2 to glycolytic/ERK signaling in gastric cancer suppression.","evidence":"Co-IP, ubiquitination assay, ERK blotting, and xenograft in gastric cancer cells","pmids":["35196650"],"confidence":"Medium","gaps":["Chain linkage and ubiquitination site on PKM2 not defined","Direct vs indirect ERK effect not separated"]},{"year":2023,"claim":"Proposed additional partners (STAT1) and a transcriptional EBF2/SIX4/TNC program underlying the fertility defect.","evidence":"Co-IP and expression analysis in Mkrn2 KO mouse testis and MEFs","pmids":["36967804"],"confidence":"Low","gaps":["Single Co-IP for STAT1 without reciprocal validation","SIX4/TNC/EBF2 mechanism inferred from expression, no direct biochemical link"]},{"year":2024,"claim":"Showed MKRN2's RNA-export role extends to viral RNA, regulating influenza A mRNA nuclear-cytoplasmic trafficking, likely via GLE1.","evidence":"RNA interactome capture, knockdown, RNA FISH, and nuclear/cytoplasmic fractionation in infected cells","pmids":["38753876"],"confidence":"Medium","gaps":["GLE1 dependence inferred, not directly demonstrated for IAV mRNA","Role of exosome degradation not fully resolved"]},{"year":2025,"claim":"Defined a non-degradative ubiquitination mechanism: K29 polyubiquitination of PAIP1 at K179 to block translation, establishing MKRN2 as a translational repressor of IL-6.","evidence":"Conditional KO mouse, in vitro ubiquitination with PAIP1 K179 mutagenesis, Co-IP, polysome profiling, RNA-binding assay","pmids":["40524017"],"confidence":"High","gaps":["Whether this mechanism generalizes to other mRNAs beyond Il6 not shown","Determinants of mRNA target selectivity unmapped"]},{"year":2025,"claim":"Consolidated MKRN2 as an in vivo restraint on inflammation and tumor-associated macrophage polarization, and added PPP2CA-Wnt and p53-apoptosis axes in other disease settings.","evidence":"KO mouse tumor models with macrophage phenotyping; Co-IP and site-specific K48 ubiquitination (PPP2CA K41); ubiquitination assays and ARDS mouse model for p53","pmids":["40925500","40959281","40885043"],"confidence":"Medium","gaps":["Tissue-specific substrate priorities not resolved","How MKRN2 selects among many substrates in a given context unknown"]},{"year":2026,"claim":"Demonstrated MKRN2 druggability and a phase-separation/condensate dimension, with a covalent molecular glue at Cys335 recruiting RPS7 for synthetic-lethal degradation, and CSDE1 ubiquitination governing condensate formation and neurodevelopmental phenotypes.","evidence":"QTRP covalent target ID, Cys335 mutagenesis, Co-IP-MS for RPS7 recruitment, genetic rescue; MS substrate mapping of CSDE1 lysines, LLPS assays, and Mkrn2 KO behavioral testing","pmids":["41991154","41757349"],"confidence":"High","gaps":["Endogenous regulators of MKRN2 condensate behavior unknown","Whether RPS7 is a physiological substrate beyond the engineered glue context unclear"]},{"year":null,"claim":"How MKRN2 selects between degradative and non-degradative ubiquitination and between distinct substrates across tissues remains unresolved.","evidence":"No timeline discovery defines the substrate-selectivity code or the structural basis of chain-type choice","pmids":[],"confidence":"Medium","gaps":["No structure of MKRN2 with any substrate","Mechanism coupling RNA binding to substrate ubiquitination not established","Determinants of K48 vs K29 chain choice unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016874","term_label":"ligase activity","supporting_discovery_ids":[0,5,8,11]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,2,5,9,11]},{"term_id":"GO:0003723","term_label":"RNA binding","supporting_discovery_ids":[6,7,8]}],"localization":[{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[0,6]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[8,15]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[0,8,12]},{"term_id":"R-HSA-8953854","term_label":"Metabolism of RNA","supporting_discovery_ids":[6,7,8]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[0,5,9,11]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,11]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[5,13]}],"complexes":[],"partners":["RELA","PDLIM2","TP53","GLE1","IGF2BP3","PAIP1","PPP2CA","CSDE1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H000","full_name":"E3 ubiquitin-protein ligase makorin-2","aliases":["RING finger protein 62","RING-type E3 ubiquitin transferase makorin-2"],"length_aa":416,"mass_kda":46.9,"function":"E3 ubiquitin ligase catalyzing the covalent attachment of ubiquitin moieties onto substrate proteins (By similarity). Promotes the polyubiquitination and proteasome-dependent degradation of RELA/p65, thereby suppressing RELA-mediated NF-kappaB transactivation and negatively regulating inflammatory responses (By similarity). Plays a role in the regulation of spermiation and in male fertility (By similarity)","subcellular_location":"Cytoplasm; Nucleus","url":"https://www.uniprot.org/uniprotkb/Q9H000/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/MKRN2","classification":"Not Classified","n_dependent_lines":4,"n_total_lines":1208,"dependency_fraction":0.0033112582781456954},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"UBA1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/MKRN2","total_profiled":1310},"omim":[{"mim_id":"608426","title":"MAKORIN 2; MKRN2","url":"https://www.omim.org/entry/608426"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Approved","locations":[{"location":"Nucleoplasm","reliability":"Approved"},{"location":"Cytosol","reliability":"Approved"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/MKRN2"},"hgnc":{"alias_symbol":["RNF62","HSPC070"],"prev_symbol":[]},"alphafold":{"accession":"Q9H000","domains":[{"cath_id":"-","chopping":"7-51","consensus_level":"high","plddt":79.0229,"start":7,"end":51},{"cath_id":"-","chopping":"168-234","consensus_level":"medium","plddt":91.0913,"start":168,"end":234},{"cath_id":"-","chopping":"257-325","consensus_level":"high","plddt":91.3271,"start":257,"end":325},{"cath_id":"-","chopping":"326-388","consensus_level":"medium","plddt":71.4316,"start":326,"end":388}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H000","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H000-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H000-F1-predicted_aligned_error_v6.png","plddt_mean":71.0},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=MKRN2","jax_strain_url":"https://www.jax.org/strain/search?query=MKRN2"},"sequence":{"accession":"Q9H000","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H000.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H000/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H000"}},"corpus_meta":[{"pmid":"28378844","id":"PMC_28378844","title":"MKRN2 is a novel ubiquitin E3 ligase for the p65 subunit of NF-κB and negatively regulates inflammatory responses.","date":"2017","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/28378844","citation_count":58,"is_preprint":false},{"pmid":"30103781","id":"PMC_30103781","title":"MKRN2 inhibits migration and invasion of non-small-cell lung cancer by negatively regulating the PI3K/Akt pathway.","date":"2018","source":"Journal of experimental & clinical cancer research : CR","url":"https://pubmed.ncbi.nlm.nih.gov/30103781","citation_count":47,"is_preprint":false},{"pmid":"32560817","id":"PMC_32560817","title":"Ubiquitination of IGF2BP3 by E3 ligase MKRN2 regulates the proliferation and migration of human neuroblastoma SHSY5Y cells.","date":"2020","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/32560817","citation_count":27,"is_preprint":false},{"pmid":"28008940","id":"PMC_28008940","title":"Deficiency of Mkrn2 causes abnormal spermiogenesis and spermiation, and impairs male fertility.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/28008940","citation_count":26,"is_preprint":false},{"pmid":"31489847","id":"PMC_31489847","title":"Mkrn2 deficiency induces teratozoospermia and male infertility through p53/PERP-mediated apoptosis in testis.","date":"2020","source":"Asian journal of andrology","url":"https://pubmed.ncbi.nlm.nih.gov/31489847","citation_count":17,"is_preprint":false},{"pmid":"32194692","id":"PMC_32194692","title":"Ubiquitination of P53 by E3 ligase MKRN2 promotes melanoma cell proliferation.","date":"2020","source":"Oncology letters","url":"https://pubmed.ncbi.nlm.nih.gov/32194692","citation_count":17,"is_preprint":false},{"pmid":"32460013","id":"PMC_32460013","title":"MKRN2 Physically Interacts with GLE1 to Regulate mRNA Export and Zebrafish Retinal Development.","date":"2020","source":"Cell 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toxicology","url":"https://pubmed.ncbi.nlm.nih.gov/37218161","citation_count":8,"is_preprint":false},{"pmid":"35196650","id":"PMC_35196650","title":"MKRN2 inhibits the proliferation of gastric cancer by downregulating PKM2.","date":"2022","source":"Aging","url":"https://pubmed.ncbi.nlm.nih.gov/35196650","citation_count":8,"is_preprint":false},{"pmid":"36967804","id":"PMC_36967804","title":"MKRN2 knockout causes male infertility through decreasing STAT1, SIX4, and TNC expression.","date":"2023","source":"Frontiers in endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/36967804","citation_count":3,"is_preprint":false},{"pmid":"40959281","id":"PMC_40959281","title":"E3 ligase MKRN2 destabilizes PPP2CA proteins to inactivate canonical Wnt pathway and mitigates tumorigenesis of clear cell renal cell carcinoma.","date":"2025","source":"International journal of biological 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cell","url":"https://pubmed.ncbi.nlm.nih.gov/41741886","citation_count":0,"is_preprint":false},{"pmid":"41991154","id":"PMC_41991154","title":"A covalent molecular glue hijacks the E3 ligase MKRN2 to degrade the ribosomal protein RPS7 and induce synthetic lethality in p53-deficient NSCLC cells.","date":"2026","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/41991154","citation_count":0,"is_preprint":false},{"pmid":"41757349","id":"PMC_41757349","title":"Liquid-liquid phase separation couples MKRN2-mediated ubiquitination of CSDE1 with neurodevelopmental disorders.","date":"2026","source":"Frontiers in cellular neuroscience","url":"https://pubmed.ncbi.nlm.nih.gov/41757349","citation_count":0,"is_preprint":false},{"pmid":null,"id":"bio_10.1101_2025.10.15.682570","title":"Ubiquitination regulates granulostasis and DRiP accumulation in SGs under heat stress via the E3 ligase MKRN2","date":"2025-10-16","source":"bioRxiv","url":"https://doi.org/10.1101/2025.10.15.682570","citation_count":0,"is_preprint":true},{"pmid":null,"id":"bio_10.1101_2025.05.28.655990","title":"Genetic regulation of fatty acid content in adipose tissue","date":"2025-05-29","source":"bioRxiv","url":"https://doi.org/10.1101/2025.05.28.655990","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":13415,"output_tokens":5053,"usd":0.05802,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":13220,"output_tokens":4116,"usd":0.0845,"stage2_stop_reason":"end_turn"},"total_usd":0.14252,"stage1_batch_id":"msgbatch_01BbTCMyrRPyhtEq6b1MdwCN","stage2_batch_id":"msgbatch_011dEed5bbxYvTAaQWKJevxu","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2017,\n      \"finding\": \"MKRN2 is a ubiquitin E3 ligase that binds to the p65 subunit of NF-κB, promotes its polyubiquitination and proteasome-dependent degradation through the MKRN2 RING finger domain, thereby suppressing p65-mediated NF-κB transactivation. MKRN2 and PDLIM2 synergistically promote polyubiquitination and degradation of p65. MKRN2 was identified via yeast two-hybrid screening as a PDLIM2-interacting protein. Knockdown of MKRN2 in dendritic cells resulted in increased nuclear p65 and augmented proinflammatory cytokine production.\",\n      \"method\": \"Yeast two-hybrid screening, co-immunoprecipitation, in vitro ubiquitination assay, RING domain mutagenesis, siRNA knockdown, NF-κB reporter assay\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — multiple orthogonal methods including yeast two-hybrid, Co-IP, in vitro ubiquitination, RING domain mutagenesis, and functional KD with cytokine readout in one study\",\n      \"pmids\": [\"28378844\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"MKRN2 inhibits migration and invasion of non-small-cell lung cancer cells through downregulation of the PI3K/Akt pathway, as demonstrated by altered MKRN2 expression in NSCLC cell lines.\",\n      \"method\": \"siRNA knockdown and overexpression in NSCLC cell lines, migration/invasion assays, western blotting for PI3K/Akt pathway components\",\n      \"journal\": \"Journal of experimental & clinical cancer research : CR\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — KD/OE with defined cellular phenotype and pathway placement, but no direct substrate identification for PI3K/Akt regulation; single lab\",\n      \"pmids\": [\"30103781\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MKRN2 ubiquitinates IGF2BP3 (an RNA-binding protein) as a direct substrate, leading to its degradation. This MKRN2-mediated ubiquitination of IGF2BP3 regulates the expression of CD44 and PDPN (downstream IGF2BP3 targets) in neuroblastoma SHSY5Y cells. MKRN2 knockdown promotes proliferation and migration of these cells.\",\n      \"method\": \"shRNA knockdown, co-immunoprecipitation, in vitro ubiquitination assay, western blotting\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and in vitro ubiquitination to identify substrate, functional KD phenotype, single lab with two orthogonal methods\",\n      \"pmids\": [\"32560817\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Mkrn2 knockout in mice causes male infertility characterized by low sperm number, poor motility, aberrant morphology, spermiation failure, and misarrangement of ectoplasmic specialization in testes. Loss of Mkrn2 results in decreased expression of Odf2, a protein vital for spermatogenesis.\",\n      \"method\": \"Knockout mouse model, histology, sperm analysis, western blotting for Odf2 expression\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KO mouse with defined spermatogenic phenotypes and downstream molecular readout (Odf2), single lab\",\n      \"pmids\": [\"28008940\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Mkrn2 deficiency leads to abnormally high apoptosis in testes. MKRN2 inhibits expression of p53 apoptosis effector PERP, and loss of Mkrn2 upregulates PERP, implicating a Mkrn2-p53/PERP signaling axis in protecting germ cells from excessive apoptosis during spermatogenesis.\",\n      \"method\": \"Mkrn2 knockout mouse model, digital gene expression profiling, GSEA/KEGG pathway analysis, protein expression analysis\",\n      \"journal\": \"Asian journal of andrology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — pathway placement inferred from transcriptomics and KO phenotype without direct biochemical confirmation of MKRN2-p53/PERP interaction; single lab\",\n      \"pmids\": [\"31489847\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MKRN2 directly interacts with and ubiquitinates p53, promoting its degradation, thereby promoting melanoma cell proliferation in a p53-dependent manner. Downregulation of MKRN2 inhibited melanoma cell growth specifically in cells with functional p53 (rescued in p53-knockout cells).\",\n      \"method\": \"Co-immunoprecipitation, GST pulldown, in vitro ubiquitination assay, CRISPR-Cas9 p53 knockout, MTT and colony formation assays, shRNA knockdown\",\n      \"journal\": \"Oncology letters\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro ubiquitination assay, reciprocal Co-IP, GST pulldown, and genetic rescue (p53 KO) in one study confirming direct substrate relationship\",\n      \"pmids\": [\"32194692\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"MKRN2 physically interacts with GLE1 (a DEAD-box helicase activator implicated in mRNA export termination), as identified by affinity-purification mass spectrometry. MKRN2 binds selectively to the 3' UTR of a subset of mRNAs, and nuclear export of MKRN2-associated mRNAs is enhanced upon MKRN2 knockdown. Genetic epistasis in zebrafish shows that morpholino knockdown or CRISPR knockout of MKRN2 partially rescues retinal developmental defects caused by GLE1 depletion, placing MKRN2 downstream of GLE1 in mRNA export regulation.\",\n      \"method\": \"Affinity-purification mass spectrometry, ribonomic approaches (RNA-binding profiling), morpholino knockdown, CRISPR/Cas9 knockout in zebrafish, genetic epistasis\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — AP-MS interaction, ribonomic binding assay, and in vivo genetic epistasis across two knockdown strategies; multiple orthogonal methods in one study\",\n      \"pmids\": [\"32460013\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"MKRN2 associates with influenza A virus (IAV) mRNAs and positively regulates IAV mRNA nuclear-cytoplasmic trafficking, potentially through an association with GLE1. In the absence of MKRN2, IAV mRNAs accumulate in the nucleus of infected cells and may be degraded by the nuclear RNA exosome.\",\n      \"method\": \"RNA interactome capture (RIC), MKRN2 knockdown, RNA FISH, nuclear/cytoplasmic fractionation\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RIC and functional KD with nuclear accumulation readout, single lab, mechanistic link to GLE1 inferred but not directly demonstrated\",\n      \"pmids\": [\"38753876\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 selectively inhibits IL-6 expression in LPS-activated macrophages. Mechanistically, after binding to Il6 mRNA, MKRN2 attaches K29 polyubiquitin chains to the Lys179 residue of PAIP1 (a translation initiation coactivator), which blocks the PAIP1-eIF4A interaction and inhibits translational efficiency of Il6 mRNA. LysM-Cre+Mkrn2fl/fl mice showed increased serum IL-6 after LPS and increased severity of experimental colitis.\",\n      \"method\": \"Conditional knockout mouse model, in vitro ubiquitination assay, mutagenesis of PAIP1 K179, Co-immunoprecipitation, polysome profiling, RNA-binding assay\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro ubiquitination with site-specific mutagenesis (K179), Co-IP, polysome profiling, and in vivo conditional KO with defined phenotype; multiple orthogonal methods\",\n      \"pmids\": [\"40524017\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"MKRN2 promotes ubiquitination-mediated degradation of PKM2 and attenuates its effect on ERK, thereby inhibiting gastric cancer cell proliferation.\",\n      \"method\": \"Overexpression and knockdown in gastric cancer cell lines, co-immunoprecipitation, ubiquitination assay, western blotting for ERK pathway, in vivo xenograft\",\n      \"journal\": \"Aging\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assay identify PKM2 as substrate, functional KD/OE phenotype with pathway placement, single lab\",\n      \"pmids\": [\"35196650\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MKRN2 directly interacts with STAT1 in testis and MEF cells, as shown by Co-IP. MKRN2 also regulates expression of SIX4 and TNC via EBF2 transcription factor in mice. Loss of Mkrn2 in knockout mice results in decreased STAT1, SIX4, and TNC expression and male infertility.\",\n      \"method\": \"Mkrn2 knockout mouse model, co-immunoprecipitation, western blotting, transcriptional analysis\",\n      \"journal\": \"Frontiers in endocrinology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP for STAT1 interaction; the SIX4/TNC/EBF2 mechanism is inferred from expression data without direct biochemical validation\",\n      \"pmids\": [\"36967804\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 interacts with PPP2CA (Protein Phosphatase 2 Catalytic Subunit Alpha) and promotes K48-linked ubiquitination at its K41 residue, leading to proteasomal degradation of PPP2CA. This results in increased β-catenin phosphorylation and decreased β-catenin levels, causing inactivation of the Wnt signaling pathway and increased apoptosis in clear cell renal cell carcinoma cells.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence co-localization, ubiquitination assay with K48-linkage specificity, western blotting for β-catenin and Wnt pathway, in vivo xenograft\",\n      \"journal\": \"International journal of biological sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, site-specific ubiquitination (K41), K48-linkage characterization, downstream pathway readout; single lab\",\n      \"pmids\": [\"40959281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 functions as an E3 ubiquitin ligase that targets NF-κB p65 for proteasomal degradation, constraining NF-κB/COX2-mediated inflammatory signaling. MKRN2 deficiency promotes M1-to-M2 macrophage polarization in tumor-associated macrophages, leading to accelerated tumor growth in MKRN2 knockout mice.\",\n      \"method\": \"MKRN2 knockout mouse model, in vivo tumor implantation, flow cytometry for macrophage phenotyping, reconstitution experiments with MKRN2 overexpression\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vivo KO model with quantitative immune profiling and functional reconstitution; p65 ubiquitination mechanism references prior work (PMID:28378844); single lab\",\n      \"pmids\": [\"40925500\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 promotes ubiquitination and proteasomal degradation of p53 in lung epithelial cells, attenuating LPS-induced apoptosis. Co-IP confirmed the direct interaction between MKRN2 and p53, and transcriptome sequencing confirmed MKRN2 modulates apoptosis via the p53 signaling pathway.\",\n      \"method\": \"Co-immunoprecipitation, in vitro ubiquitination assay, transcriptome sequencing, adenovirus-mediated overexpression, siRNA knockdown, in vivo LPS-ARDS mouse model\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assay, transcriptomics, and in vivo model; consistent with prior p53 ubiquitination findings (PMID:32194692); single lab\",\n      \"pmids\": [\"40885043\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"The covalent compound DPB directly targets MKRN2 by covalently modifying the Cys335 residue, acting as a molecular glue that recruits the ribosomal protein RPS7 to MKRN2. This induces ubiquitination and proteasomal degradation of RPS7, triggering nucleolar stress and apoptosis selectively in p53-deficient NSCLC cells. The synthetic lethal effect is entirely dependent on the MKRN2-RPS7 axis.\",\n      \"method\": \"Quantitative thiol-reactivity proteomics (QTRP), co-immunoprecipitation mass spectrometry, site-directed mutagenesis of Cys335, genetic knockout/rescue studies, biophysical binding assays, in vivo orthotopic mouse model\",\n      \"journal\": \"British journal of pharmacology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — covalent target identification by QTRP, mutagenesis of active site Cys335, Co-IP-MS for neo-substrate recruitment, genetic rescue studies confirming mechanism; multiple orthogonal methods\",\n      \"pmids\": [\"41991154\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"MKRN2 mediates ubiquitination of the RNA-binding protein CSDE1 at four specific lysine residues (K81, K91, K208, K727), identified by mass spectrometry and validated by mutagenesis. MKRN2 and CSDE1 form co-localized condensates via liquid-liquid phase separation (LLPS), which is disrupted by functional impairment of either protein. Mkrn2 knockout mice exhibit sex-specific social abnormalities recapitulating ASD features, and CSDE1-dependent targets MARK1 and HNRNPUL2 mRNAs are regulated in a ubiquitination-dependent manner.\",\n      \"method\": \"Mass spectrometry substrate identification, mutagenesis of CSDE1 lysine residues, LLPS assays in HEK293 and SH-SY5Y cells, Mkrn2 knockout mouse behavioral testing, western blotting\",\n      \"journal\": \"Frontiers in cellular neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — MS substrate identification validated by mutagenesis, LLPS assay, and in vivo KO with behavioral phenotype; single lab\",\n      \"pmids\": [\"41757349\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"MKRN2 localizes to stress granules (SGs) in a manner dependent on active ubiquitination (UBA1 activity). Within SGs, MKRN2 promotes proper SG formation and disassembly following stress recovery by preventing accumulation of defective ribosomal products (DRiPs). MKRN2 was identified as a SG-associated E3 ligase by proximity proteomics.\",\n      \"method\": \"Proximity proteomics (BioID), UBA1 inhibitor treatment, live-cell imaging of SG dynamics, loss-of-function assays\",\n      \"journal\": \"bioRxiv\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity proteomics plus functional SG assays with pharmacological inhibition; preprint, not yet peer-reviewed\",\n      \"pmids\": [\"bio_10.1101_2025.10.15.682570\"],\n      \"is_preprint\": true\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"MKRN2 promotes cell cycle progression and proliferation of hepatocellular carcinoma cells through activation of the p38 MAPK signaling pathway, leading to c-Myc activation. MKRN2 depletion arrests cells at the G1/S transition.\",\n      \"method\": \"siRNA knockdown, RNA-seq, flow cytometry cell cycle analysis, western blotting for p38 MAPK and c-Myc, CCK-8/EdU/colony formation assays, in vivo xenograft\",\n      \"journal\": \"Human cell\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — pathway placement inferred from RNA-seq and KD phenotype without direct biochemical identification of MKRN2 substrate or binding partner in the p38/c-Myc axis; single lab\",\n      \"pmids\": [\"41741886\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"MKRN2 is an RNA-binding E3 ubiquitin ligase that suppresses NF-κB-mediated inflammation by ubiquitinating p65 for proteasomal degradation (cooperatively with PDLIM2), inhibits IL-6 translation by K29-polyubiquitinating PAIP1 at K179 to block PAIP1-eIF4A interaction, interacts with GLE1 at the nuclear pore to selectively regulate mRNA nuclear export (including for viral RNAs), ubiquitinates multiple substrates including p53, IGF2BP3, PKM2, PPP2CA, CSDE1, and RPS7 via its RING finger domain, localizes to stress granules in a ubiquitination-dependent manner to regulate granulostasis, and is required for normal spermatogenesis and male fertility in mice.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"MKRN2 is a RING-finger E3 ubiquitin ligase that combines protein ubiquitination with RNA binding to control inflammation, mRNA fate, and cell survival [#0, #8, #6]. As a ubiquitin ligase it directs proteasomal degradation of a range of substrates through its RING domain, including the NF-\\u03baB subunit p65\\u2014which it ubiquitinates cooperatively with PDLIM2 to suppress NF-\\u03baB transactivation and proinflammatory cytokine output [#0, #12]\\u2014and the tumor suppressor p53, which it targets to promote cell proliferation and limit apoptosis [#5, #13]. Additional substrates established by direct interaction and ubiquitination assays include the RNA-binding protein IGF2BP3 [#2], the glycolytic enzyme PKM2 [#9], the phosphatase catalytic subunit PPP2CA, which it modifies with K48-linked chains at K41 to inactivate Wnt/\\u03b2-catenin signaling [#11], and the RNA-binding protein CSDE1, with which it also forms co-localized phase-separated condensates [#15]. Beyond degradative ubiquitination, MKRN2 acts as an RNA-binding translational and export regulator: it binds Il6 mRNA and attaches non-degradative K29 polyubiquitin chains to PAIP1 at K179, blocking the PAIP1\\u2013eIF4A interaction to selectively repress IL-6 translation [#8], and it interacts with the mRNA export factor GLE1 to control nuclear export of a subset of cellular and influenza A virus mRNAs [#6, #7]. MKRN2 is required for normal spermatogenesis and male fertility in mice, where its loss causes spermiation failure and excess germ-cell apoptosis [#3]. A covalent molecular-glue compound exploits MKRN2 by modifying Cys335 to recruit and degrade the ribosomal protein RPS7, producing synthetic lethality in p53-deficient lung cancer cells [#14].\"\n,\n  \"teleology\": [\n    {\n      \"year\": 2016,\n      \"claim\": \"Established the first organismal requirement for MKRN2, showing it is essential for male fertility before any substrate was known.\",\n      \"evidence\": \"Mkrn2 knockout mouse with histology, sperm analysis, and Odf2 readout\",\n      \"pmids\": [\"28008940\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Did not identify the direct ubiquitination substrate driving spermatogenic failure\", \"Odf2 link is correlative, not shown to be a ubiquitination target\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined MKRN2 as a functional RING E3 ligase by identifying p65 as a degradation substrate, linking it to negative regulation of NF-\\u03baB inflammation.\",\n      \"evidence\": \"Yeast two-hybrid, Co-IP, in vitro ubiquitination, RING mutagenesis, and siRNA knockdown in dendritic cells\",\n      \"pmids\": [\"28378844\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Ubiquitin chain linkage on p65 not characterized\", \"Relative contributions of MKRN2 vs PDLIM2 to ligase activity not separated\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Extended MKRN2 function to cancer cell behavior, placing it as a suppressor of migration/invasion via the PI3K/Akt pathway.\",\n      \"evidence\": \"Knockdown/overexpression in NSCLC lines with invasion assays and pathway blotting\",\n      \"pmids\": [\"30103781\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct ubiquitination substrate linking MKRN2 to PI3K/Akt\", \"Mechanism of pathway downregulation unresolved\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Identified RNA-binding proteins and p53 as direct MKRN2 substrates, broadening its substrate repertoire and connecting it to proliferation control.\",\n      \"evidence\": \"Co-IP, GST pulldown, in vitro ubiquitination, and CRISPR p53-knockout rescue in melanoma and neuroblastoma cells\",\n      \"pmids\": [\"32194692\", \"32560817\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Chain linkage types not defined for these substrates\", \"Whether the same RING residues govern all substrates not tested\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Revealed an unexpected RNA-export function by placing MKRN2 downstream of GLE1, showing it binds mRNA 3' UTRs and restrains nuclear export.\",\n      \"evidence\": \"AP-MS, ribonomic RNA-binding profiling, and zebrafish morpholino/CRISPR genetic epistasis\",\n      \"pmids\": [\"32460013\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether export regulation requires ligase activity not established\", \"Direct GLE1 binding interface unmapped\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Connected MKRN2 loss to germ-cell apoptosis through a p53/PERP axis, contextualizing the fertility phenotype.\",\n      \"evidence\": \"Mkrn2 KO mouse with transcriptomics and protein expression analysis\",\n      \"pmids\": [\"31489847\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"p53/PERP axis inferred from expression, no direct biochemical interaction shown\", \"Single lab, no rescue\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Added PKM2 as a degradation substrate, linking MKRN2 to glycolytic/ERK signaling in gastric cancer suppression.\",\n      \"evidence\": \"Co-IP, ubiquitination assay, ERK blotting, and xenograft in gastric cancer cells\",\n      \"pmids\": [\"35196650\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Chain linkage and ubiquitination site on PKM2 not defined\", \"Direct vs indirect ERK effect not separated\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Proposed additional partners (STAT1) and a transcriptional EBF2/SIX4/TNC program underlying the fertility defect.\",\n      \"evidence\": \"Co-IP and expression analysis in Mkrn2 KO mouse testis and MEFs\",\n      \"pmids\": [\"36967804\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single Co-IP for STAT1 without reciprocal validation\", \"SIX4/TNC/EBF2 mechanism inferred from expression, no direct biochemical link\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Showed MKRN2's RNA-export role extends to viral RNA, regulating influenza A mRNA nuclear-cytoplasmic trafficking, likely via GLE1.\",\n      \"evidence\": \"RNA interactome capture, knockdown, RNA FISH, and nuclear/cytoplasmic fractionation in infected cells\",\n      \"pmids\": [\"38753876\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"GLE1 dependence inferred, not directly demonstrated for IAV mRNA\", \"Role of exosome degradation not fully resolved\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Defined a non-degradative ubiquitination mechanism: K29 polyubiquitination of PAIP1 at K179 to block translation, establishing MKRN2 as a translational repressor of IL-6.\",\n      \"evidence\": \"Conditional KO mouse, in vitro ubiquitination with PAIP1 K179 mutagenesis, Co-IP, polysome profiling, RNA-binding assay\",\n      \"pmids\": [\"40524017\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether this mechanism generalizes to other mRNAs beyond Il6 not shown\", \"Determinants of mRNA target selectivity unmapped\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Consolidated MKRN2 as an in vivo restraint on inflammation and tumor-associated macrophage polarization, and added PPP2CA-Wnt and p53-apoptosis axes in other disease settings.\",\n      \"evidence\": \"KO mouse tumor models with macrophage phenotyping; Co-IP and site-specific K48 ubiquitination (PPP2CA K41); ubiquitination assays and ARDS mouse model for p53\",\n      \"pmids\": [\"40925500\", \"40959281\", \"40885043\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Tissue-specific substrate priorities not resolved\", \"How MKRN2 selects among many substrates in a given context unknown\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"Demonstrated MKRN2 druggability and a phase-separation/condensate dimension, with a covalent molecular glue at Cys335 recruiting RPS7 for synthetic-lethal degradation, and CSDE1 ubiquitination governing condensate formation and neurodevelopmental phenotypes.\",\n      \"evidence\": \"QTRP covalent target ID, Cys335 mutagenesis, Co-IP-MS for RPS7 recruitment, genetic rescue; MS substrate mapping of CSDE1 lysines, LLPS assays, and Mkrn2 KO behavioral testing\",\n      \"pmids\": [\"41991154\", \"41757349\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous regulators of MKRN2 condensate behavior unknown\", \"Whether RPS7 is a physiological substrate beyond the engineered glue context unclear\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How MKRN2 selects between degradative and non-degradative ubiquitination and between distinct substrates across tissues remains unresolved.\",\n      \"evidence\": \"No timeline discovery defines the substrate-selectivity code or the structural basis of chain-type choice\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structure of MKRN2 with any substrate\", \"Mechanism coupling RNA binding to substrate ubiquitination not established\", \"Determinants of K48 vs K29 chain choice unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016874\", \"supporting_discovery_ids\": [0, 5, 8, 11]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 2, 5, 9, 11]},\n      {\"term_id\": \"GO:0003723\", \"supporting_discovery_ids\": [6, 7, 8]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [0, 6]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [8, 15]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [0, 8, 12]},\n      {\"term_id\": \"R-HSA-8953854\", \"supporting_discovery_ids\": [6, 7, 8]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [0, 5, 9, 11]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 11]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [5, 13]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"RELA\", \"PDLIM2\", \"TP53\", \"GLE1\", \"IGF2BP3\", \"PAIP1\", \"PPP2CA\", \"CSDE1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":6,"faith_total":6,"faith_pct":100.0}}