{"gene":"CYLD","run_date":"2026-06-09T22:57:19","timeline":{"discoveries":[{"year":2003,"finding":"CYLD is a deubiquitinating enzyme that negatively regulates NF-κB activation by removing ubiquitin chains from TRAF2 and TRAF6, thereby inhibiting signaling downstream of TNFR family members CD40, XEDAR, and EDAR. Loss of CYLD deubiquitinating activity correlates with tumorigenesis.","method":"Deubiquitinase activity assay, RNA interference knockdown, Co-immunoprecipitation, NF-κB reporter assays","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — enzymatic activity demonstrated, substrates identified by Co-IP, RNAi phenotype, replicated across multiple receptor systems in a landmark paper","pmids":["12917689"],"is_preprint":false},{"year":2006,"finding":"CYLD deubiquitinates Bcl-3 in the perinuclear region, preventing nuclear accumulation of Bcl-3/p50 and Bcl-3/p52 complexes and thereby blocking cyclin D1-dependent keratinocyte proliferation. TPA or UV triggers translocation of CYLD from cytoplasm to perinuclear region to execute this function. This pathway is distinct from CYLD's regulation of TRAF2/p65-p50 NF-κB survival signaling.","method":"CYLD knockout mice, Co-immunoprecipitation, subcellular fractionation/live imaging, chemical induction (TPA/UV), cyclin D1 expression assays","journal":"Cell","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — multiple orthogonal methods (KO mice, Co-IP, fractionation, functional rescue), novel substrate identified with mechanistic consequence","pmids":["16713561"],"is_preprint":false},{"year":2005,"finding":"CYLD is phosphorylated by IκB kinase gamma (IKKγ/NEMO)-dependent signaling in response to cellular stimuli, and this phosphorylation transiently inhibits CYLD's deubiquitinase activity, allowing signal-induced TRAF2 ubiquitination and downstream IKK activation.","method":"In vivo phosphorylation assays, IKK subunit overexpression and knockdown, TRAF2 ubiquitination assays","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — biochemical phosphorylation mapping, functional consequence on substrate ubiquitination demonstrated, single lab with multiple orthogonal methods","pmids":["15870263"],"is_preprint":false},{"year":2006,"finding":"CYLD physically interacts with active Lck and promotes recruitment of active Lck to its substrate Zap70 during T cell receptor signaling in thymocytes. CYLD also removes both K48- and K63-linked polyubiquitin chains from Lck, positively regulating proximal TCR signaling and T cell development.","method":"Co-immunoprecipitation, CYLD knockout mice, T cell development analysis, ubiquitin chain-type analysis","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — reciprocal Co-IP, KO mice with defined developmental phenotype, two orthogonal methods in single lab","pmids":["16501569"],"is_preprint":false},{"year":2007,"finding":"CYLD deubiquitinates and inactivates TAK1 (transforming growth factor-β-activated kinase 1), inhibiting TAK1 ubiquitination and autoactivation, thereby suppressing downstream JNK and IKKβ activation in T cells.","method":"CYLD knockout mice, Co-immunoprecipitation, TAK1 ubiquitination and kinase activity assays","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 / Moderate — KO mice with constitutive TAK1 activation phenotype, Co-IP demonstrating physical interaction, substrate ubiquitination assay","pmids":["17548520"],"is_preprint":false},{"year":2007,"finding":"CYLD localizes to microtubules during interphase and to the midbody during telophase, and its protein levels decrease as cells exit mitosis. CYLD physically interacts with Plk1 and shares similar loss-of-function and overexpression phenotypes, implicating CYLD in regulating timely mitotic entry.","method":"Immunofluorescence/live cell imaging, Co-immunoprecipitation, loss-of-function and overexpression experiments, cell cycle analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — localization with functional consequence, Co-IP with Plk1, but mechanistic link between CYLD-Plk1 interaction and mitotic entry not fully reconstituted","pmids":["17495026"],"is_preprint":false},{"year":2007,"finding":"Drosophila CYLD (dCYLD) deubiquitinates dTRAF2, preventing its ubiquitin-mediated proteolytic degradation, and thereby regulates TNF-induced JNK activation and JNK-dependent cell death. dTRAF2 acts downstream of the TNF receptor Wengen and upstream of the JNKK kinase dTAK1.","method":"Drosophila genetic mutants, transgenic flies with wild-type and catalytic-dead dCYLD, dTRAF2 ubiquitination assay, epistasis analysis","journal":"Developmental cell","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — reconstitution in vivo via genetics, deubiquitinase activity on specific substrate demonstrated, epistasis with defined pathway placement","pmids":["17765686"],"is_preprint":false},{"year":2008,"finding":"CYLD negatively regulates RANK signaling and osteoclastogenesis by inhibiting TRAF6 ubiquitination. CYLD physically interacts with the signaling adaptor p62, which recruits CYLD to TRAF6.","method":"CYLD knockout mice, RANKL-induced osteoclast differentiation assays, Co-immunoprecipitation, TRAF6 ubiquitination assays","journal":"The Journal of clinical investigation","confidence":"High","confidence_rationale":"Tier 2 / Moderate — KO mice with severe osteoporosis phenotype, Co-IP demonstrating CYLD-p62-TRAF6 complex, substrate ubiquitination assay","pmids":["18382763"],"is_preprint":false},{"year":2008,"finding":"CYLD deubiquitinates RIG-I (a cytoplasmic RNA sensor) and inhibits its ubiquitination, thereby suppressing RIG-I-mediated activation of IKKε/TBK1 and IFN-β promoter induction during viral infection.","method":"CYLD knockout cells, RIG-I ubiquitination assay, IFN-β reporter assay, IKKε/TBK1 kinase activation assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KO cells with constitutive IKKε/TBK1 activation, substrate ubiquitination assay, reporter assay; single lab","pmids":["18467330"],"is_preprint":false},{"year":2008,"finding":"CYLD associates with microtubules both in cells and in vitro through its first CAP-Gly domain. CYLD enhances tubulin polymerization by lowering the critical concentration for microtubule assembly and promotes microtubule regrowth after nocodazole washout. The first CAP-Gly domain is required for CYLD-dependent cell migration.","method":"In vitro microtubule co-sedimentation assay, tubulin polymerization assay, CAP-Gly domain deletion mutants, nocodazole washout assay, wound healing assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of tubulin polymerization, domain mutagenesis, multiple orthogonal methods in single lab","pmids":["18222923"],"is_preprint":false},{"year":2009,"finding":"CYLD controls cell cycle progression at G1/S and cytokinesis by associating with α-tubulin and microtubules through its CAP-Gly domains. CYLD inhibits HDAC6, increasing perinuclear acetylated α-tubulin levels, which facilitates CYLD interaction with Bcl-3 to delay G1-to-S transition. CYLD also interacts with HDAC6 at the midbody to regulate cytokinesis in a deubiquitinase-independent manner.","method":"Co-immunoprecipitation, immunofluorescence, cell cycle analysis, HDAC6 inhibition assays, CYLD-Bcl-3 interaction assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Moderate — multiple Co-IPs, defined cell cycle phenotypes, deubiquitinase-independent mechanism demonstrated, multiple orthogonal methods","pmids":["19893491"],"is_preprint":false},{"year":2009,"finding":"Snail1 transcriptionally represses CYLD expression in melanoma. As a consequence of CYLD repression, BCL-3 translocates to the nucleus, activating Cyclin D1 and N-cadherin promoters and driving proliferation and invasion.","method":"Snail1 expression/knockdown, CYLD rescue experiments, in vitro and in vivo tumor models, tissue microarray analysis","journal":"The Journal of experimental medicine","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — CYLD rescue experiments show functional consequence, Snail1-CYLD axis tested in multiple models; single lab","pmids":["19124656"],"is_preprint":false},{"year":2010,"finding":"CYLD knockdown significantly impairs angiogenesis by blocking endothelial cell spreading, migration, and polarity. CYLD regulates microtubule dynamics in endothelial cells and activates Rac1, with Rac1 activation being an important downstream mechanism for CYLD's role in endothelial cell migration.","method":"siRNA knockdown, tube formation assay, 3D capillary sprouting assay, in vivo angioreactor assay, microtubule dynamics imaging, Rac1 activation assay","journal":"Blood","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple in vitro and in vivo angiogenesis assays, defined Rac1 mechanism; single lab","pmids":["20194890"],"is_preprint":false},{"year":2011,"finding":"MALT1 proteolytically cleaves and inactivates CYLD following TCR activation in T cells and by oncogenic API2-MALT1. CYLD cleavage by MALT1 is specifically required for JNK activation and inducible expression of a subset of TCR-responsive genes.","method":"MALT1 protease activity assays, CYLD cleavage detection, JNK activation assay, gene expression analysis, MALT1 inhibitor studies","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1 / Moderate — direct proteolytic cleavage of CYLD by MALT1 demonstrated biochemically, functional consequence on JNK signaling established with multiple methods","pmids":["21448133"],"is_preprint":false},{"year":2011,"finding":"Itch (E3 ligase) and CYLD form a complex via WW-PPXY motif interactions. The Itch-CYLD complex sequentially removes K63-linked ubiquitin chains (via CYLD) and then catalyzes K48-linked ubiquitination (via Itch) on TAK1, terminating inflammatory TNF signaling. CYLD(Y485A) mutant unable to associate with Itch cannot block sustained TAK1 activation.","method":"Co-immunoprecipitation, in vitro deubiquitination assay, TAK1 ubiquitination assay, CYLD mutant reconstitution in CYLD−/− macrophages, cytokine production assay","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — complex reconstitution, sequential enzymatic activities demonstrated, domain-specific mutant validates binding requirement, multiple orthogonal methods","pmids":["22057290"],"is_preprint":false},{"year":2011,"finding":"CYLD deubiquitinates c-Jun and c-Fos, removing K63-linked ubiquitin chains from these AP-1 components. Catalytically inactive CYLD mutant (CYLDm) increases K63 ubiquitination on c-Jun and c-Fos, leading to sustained AP-1 activation via JNK. CYLD thus blocks JNK/AP-1 signaling at multiple levels.","method":"In vivo ubiquitination assay for c-Jun and c-Fos, JNK activation assay, transgenic mouse skin tumorigenesis model, JNK inhibitor treatment","journal":"Cancer prevention research (Philadelphia, Pa.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — novel substrates c-Jun and c-Fos identified by ubiquitination assay, functional JNK inhibitor validation; single lab","pmids":["21478324"],"is_preprint":false},{"year":2013,"finding":"CYLD regulates RIP1 ubiquitination specifically within the NP-40 insoluble necrosome (not at the TNFR-1 signaling complex) during TNFα-induced programmed necrosis. Increased RIP1 ubiquitination in the necrosome in CYLD−/− cells impairs RIP1 and RIP3 phosphorylation, inhibiting kinase activation required for necroptosis.","method":"CYLD knockout cells, RIP1 ubiquitination assay in necrosome fraction, RIP1-RIP3 phosphorylation assay, SMAC mimetic and cycloheximide sensitization experiments","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 / Moderate — specific compartment (necrosome vs. TNFR complex) resolved by fractionation, KO cells with multiple sensitization conditions, kinase activation assay","pmids":["24098568"],"is_preprint":false},{"year":2014,"finding":"CYLD mediates ciliogenesis by deconjugating polyubiquitin chains from centrosomal protein Cep70, which is required for Cep70 to interact with γ-tubulin and localize at the centrosome. Additionally, CYLD-mediated inhibition of HDAC6 promotes tubulin acetylation, constituting a second mechanism for ciliary assembly.","method":"CYLD knockout mice (polydactyly, ciliary defect phenotype), Co-immunoprecipitation (Cep70-γ-tubulin), Cep70 ubiquitination assay, HDAC6 inhibitor rescue experiments","journal":"Cell research","confidence":"High","confidence_rationale":"Tier 2 / Moderate — KO mice with defined ciliopathy phenotype, substrate deubiquitination demonstrated, two distinct mechanisms shown, HDAC6 inhibitor rescue","pmids":["25342559"],"is_preprint":false},{"year":2014,"finding":"CYLD interacts with and deubiquitinates p53, facilitating p53 stabilization in response to genotoxic stress by removing K63-linked ubiquitin chains (and thereby indirectly preventing K48-linked degradation). Loss of CYLD catalytic activity impairs DNA damage-induced p53 stabilization and activation.","method":"Co-immunoprecipitation, p53 ubiquitination assay, ubiquitin chain-restriction analysis, CYLD catalytic mutant knock-in mice, C. elegans CEP-1/p53 epistasis","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — multiple methods including ubiquitin chain-restriction analysis, genetic validation in C. elegans, mouse model with defined phenotype","pmids":["27561390"],"is_preprint":false},{"year":2014,"finding":"CYLD coordinates with EB1 at microtubule plus ends to regulate microtubule dynamics and cell migration. The CYLD-EB1 interaction increases upon stimulation of cell migration. CYLD and EB1 act in concert to regulate tail retraction, centrosome reorientation, leading-edge microtubule stabilization, and microtubule nucleation.","method":"Yeast two-hybrid screening, Co-immunoprecipitation, in vitro microtubule assembly assay, live cell imaging of migration, centrosome reorientation assay","journal":"Cell cycle (Georgetown, Tex.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast 2-hybrid plus Co-IP validation, in vitro assay, multiple migration readouts; single lab","pmids":["24552808"],"is_preprint":false},{"year":2014,"finding":"CYLD interacts with HDAC7 to remove HDAC7 from the HGF promoter in hepatic stellate cells, inducing HGF gene transcription independently of CYLD's deubiquitinating activity.","method":"Co-immunoprecipitation, ChIP assay, CYLD catalytic-dead mutant experiments, CYLD−/− mice with HGF measurement","journal":"Hepatology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ChIP, and catalytic-dead mutant establish deubiquitinase-independent mechanism; single lab","pmids":["24811579"],"is_preprint":false},{"year":2014,"finding":"CYLD deubiquitinating activity is inhibited by SUMOylation. Retinoic acid-induced SUMOylation of CYLD blocks its deubiquitinase activity against TRAF2 and TRAF6, facilitating NF-κB signaling during neuroblastoma differentiation.","method":"SUMOylation assay, CYLD deubiquitinase activity assay with TRAF2/TRAF6 substrates, non-SUMOylatable CYLD mutant overexpression, NF-κB activation assay","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — direct enzymatic activity assay with and without SUMOylation, non-SUMOylatable mutant functional rescue; single lab","pmids":["24909169"],"is_preprint":false},{"year":2014,"finding":"CYLD interacts with MIB2 (a ubiquitin ligase), stabilizing MIB2 protein levels and reducing JAG2 expression, thereby attenuating Notch signaling. CYLD gene silencing increases JAG2 expression and upregulates Notch signaling.","method":"Proteomics/mass spectrometry to identify interactors, Co-immunoprecipitation, siRNA knockdown, Notch target gene expression analysis, primary tumor cell cultures","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — MS-identified interaction validated by Co-IP, siRNA knockdown with pathway readout; single lab","pmids":["25565632"],"is_preprint":false},{"year":2015,"finding":"CYLD maintains hematopoietic stem cell (HSC) quiescence through deubiquitination of TRAF2. This function operates through the p38MAPK pathway (not NF-κB), and pharmacological inhibition of p38MAPK rescues the CYLD-loss HSC phenotype of increased cycling and loss of repopulation potential.","method":"Conditional CYLD KO mice, CYLD-TRAF2 binding mutant mice, HSC quiescence assays, transplantation/repopulation assays, p38MAPK inhibitor treatment","journal":"The Journal of experimental medicine","confidence":"High","confidence_rationale":"Tier 2 / Moderate — conditional KO, binding-specific mutant dissecting TRAF2 interaction from catalytic function, epistatic pharmacological rescue; multiple orthogonal approaches","pmids":["25824820"],"is_preprint":false},{"year":2014,"finding":"SCFβ-TRCP E3 ligase promotes ubiquitination and degradation of CYLD in a manner dependent on prior IKK-mediated phosphorylation of CYLD at Ser432/Ser436. β-TRCP depletion causes CYLD accumulation and TRAF6 deubiquitination, suppressing osteoclastogenesis.","method":"Co-immunoprecipitation, ubiquitination assays, phosphorylation-site mutants, β-TRCP siRNA, TRAF6 ubiquitination assay, osteoclast differentiation assay","journal":"Oncotarget","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — phosphodegron mechanism established by phospho-mutant and Co-IP; single lab","pmids":["24961988"],"is_preprint":false},{"year":2016,"finding":"SPATA2 directly interacts with CYLD via its PUB domain and acts as an allosteric activator of CYLD's K63- and M1-deubiquitinase activity. SPATA2 is required for recruitment of CYLD to the TNF receptor signaling complex, and its loss attenuates TNF-induced NF-κB/MAPK signaling while also reducing Complex II formation and apoptosis.","method":"Mass spectrometry screen, Co-immunoprecipitation, in vitro deubiquitinase activity assay with SPATA2, TNF signaling complex (Complex I) isolation","journal":"EMBO reports","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — allosteric activation demonstrated by in vitro enzyme assay, Co-IP, receptor complex recruitment shown; multiple orthogonal methods in single lab","pmids":["27458237"],"is_preprint":false},{"year":2016,"finding":"CYLD removes K63-linked ubiquitin chains from RIPK2, inhibiting RIPK2 K63-ubiquitination and impairing NF-κB and ERK1/2 activation in Listeria-infected macrophages, thereby reducing antibacterial immune responses.","method":"Co-immunoprecipitation, RIPK2 ubiquitination assay, CYLD KO macrophages, siRNA knockdown, NF-κB and ERK activation assays","journal":"Frontiers in immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assay for RIPK2, KO cells with functional immune readout; single lab","pmids":["26834734"],"is_preprint":false},{"year":2019,"finding":"CYLD is recruited by SPATA2 to the centrosome where it deubiquitinates PLK4 (master regulator of centrosome duplication). Deubiquitination of PLK4 facilitates its phosphorylation of NEK7 at Ser204, which attenuates NEK7-NLRP3 interaction and suppresses NLRP3 inflammasome activation.","method":"Co-immunoprecipitation, PLK4 ubiquitination assay, NEK7 phosphorylation assay, NLRP3 inflammasome activation assay, SPATA2 KO macrophages, PLK4 inhibitor/shRNA","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Moderate — substrate deubiquitination demonstrated, phosphorylation of NEK7 by PLK4 shown, SPATA2-CYLD complex at centrosome, multiple mechanistic steps validated","pmids":["31762063"],"is_preprint":false},{"year":2019,"finding":"CYLD promotes the proteostasis of centriolar satellites by deubiquitinating PCM1 scaffold protein and protecting it from MIB1-mediated proteasomal degradation. CYLD knockdown promotes PCM1 degradation and dismantling of centriolar satellites, impairing ciliogenesis.","method":"Unbiased proteomic screen of CYLD binding partners, Co-immunoprecipitation, PCM1 ubiquitination assay, MIB1 E3 ligase assay, siRNA knockdown, cilia formation assay","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proteomics + Co-IP + ubiquitination assay; counterbalanced E3/DUB pair identified; single lab","pmids":["31067453"],"is_preprint":false},{"year":2020,"finding":"CYLD deubiquitinates NLRP6, and this deubiquitination inhibits the NLRP6-ASC inflammasome complex, preventing excessive IL-18 maturation in the colonic mucosa.","method":"Co-immunoprecipitation, NLRP6 ubiquitination assay, CYLD knockout mice with Citrobacter rodentium infection, IL-18 maturation assay","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — novel substrate (NLRP6) identified, KO mouse in vivo model, IL-18 maturation as functional readout, published in high-impact journal","pmids":["32424362"],"is_preprint":false},{"year":2020,"finding":"A gain-of-function CYLD missense variant (M719V) exhibits significantly increased K63 deubiquitinase activity relative to wild-type. Overexpression of CYLD-M719V leads to more potent NF-κB inhibition and impairment of autophagosome fusion to lysosomes.","method":"In vitro deubiquitinase activity assay with Wilcoxon signed-rank test, NF-κB luciferase reporter assay, autophagy flux assay in HEK293 cells","journal":"Brain : a journal of neurology","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — direct enzymatic activity assay, NF-κB and autophagy assays; single lab; gain-of-function mechanism novel","pmids":["32185393"],"is_preprint":false},{"year":2021,"finding":"TRIM15 is a K63-ubiquitin ligase for ERK1/2 and CYLD is the deubiquitinase that removes K63-linked chains from ERK1/2. K63-linked polyubiquitination of ERK1/2 by TRIM15 enhances ERK interaction with and activation by MEK. TRIM15 and CYLD regulate ERK ubiquitination at defined lysine residues through mutually exclusive interactions.","method":"In vitro ubiquitination and deubiquitinase assays, ERK ubiquitination site mapping by mutagenesis, Co-immunoprecipitation, ERK activation (MEK interaction) assay","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of ubiquitination/deubiquitination, substrate site mapping by mutagenesis, mechanistic link to MEK activation; strong method quality","pmids":["34497368"],"is_preprint":false},{"year":2021,"finding":"Two CAP-Gly domains of CYLD function as ubiquitin-binding domains, with CAP-Gly3 required for CYLD deubiquitinase activity and regulation of immune receptor signaling. A phosphorylation switch outside the catalytic USP domain (Ser568, a TNF-regulated site, acting in concert with Ser418) activates CYLD toward K63-linked polyubiquitin. Phosphorylated CYLD together with SPATA2 and LUBAC functions as a ubiquitin-editing complex.","method":"Structural/biochemical analysis of CAP-Gly domains, phosphoproteomic identification of Ser568, in vitro deubiquitinase activity assays with phospho-mimetic mutants, immune receptor signaling assays","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro deubiquitinase assay with defined phospho-mutants, ubiquitin-binding domain biochemistry, novel phosphorylation site identified; multiple orthogonal methods","pmids":["34610306"],"is_preprint":false},{"year":2021,"finding":"CYLD stabilizes p18 (CDK inhibitor) by binding to p18 and removing K48-linked polyubiquitin chains, preventing p18 proteasomal degradation and maintaining G1/S cell cycle arrest.","method":"Co-immunoprecipitation, p18 ubiquitination assay (K48 chain type), p18 half-life assay (CHX chase), CYLD KO/knockdown cell cycle analysis, in vivo xenograft","journal":"NPJ precision oncology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, ubiquitination assay, half-life experiment; novel K48-chain substrate for CYLD; single lab","pmids":["33654169"],"is_preprint":false},{"year":2021,"finding":"CYLD is required for SHARPIN-deficient mouse phenotype (dermatitis, spleen architecture disruption). In SHARPIN-deficient cells, impaired CYLD phosphorylation at Ser418 (which normally inhibits CYLD) leads to enhanced CYLD-dependent RIPK1 recruitment to death-signaling Complex II following TNF stimulation, causing myeloid cell death and inflammation.","method":"Genetic double KO (Sharpin/CYLD), myeloid-specific conditional Cyld deletion, Complex II immunoprecipitation, CYLD Ser418 phosphorylation analysis","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 2 / Moderate — genetic epistasis with double KO, conditional KO, biochemical Complex II isolation; multiple orthogonal approaches","pmids":["34887354"],"is_preprint":false},{"year":2022,"finding":"CYLD deubiquitinates plakoglobin by removing K63-linked polyubiquitin chains. Deubiquitinated plakoglobin shows enhanced interaction with the desmoplakin/EB1 complex at microtubule plus ends, promoting microtubule-dependent transport of connexin 43 (Cx43) to the cell membrane for gap junction assembly at the intercalated disc.","method":"Co-immunoprecipitation, plakoglobin ubiquitination assay (K63), CYLD KO mice (cardiac gap junction, fibrosis phenotype), microtubule transport assay, Cx43 membrane targeting assay","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Moderate — novel substrate (plakoglobin) with K63-chain specificity, KO mice with cardiac phenotype, defined transport mechanism; multiple orthogonal methods","pmids":["36577382"],"is_preprint":false},{"year":2022,"finding":"SPATA2/CYLD pathway regulates ferritinophagy in cardiomyocytes by controlling the ubiquitination and degradation of NCOA4 (the ferritinophagy cargo receptor). CYLD/SPATA2-mediated deubiquitination of NCOA4 enhances ferritin autophagy, leading to intracellular iron overload and ferroptosis.","method":"Co-immunoprecipitation (SPATA2-CYLD interaction), NCOA4 ubiquitination assay, SPATA2 knockdown, doxorubicin-treated cardiomyocyte and mouse models, ferritinophagy and ferroptosis readouts","journal":"Chemico-biological interactions","confidence":"Medium","confidence_rationale":"Tier 2–3 / Moderate — Co-IP, ubiquitination assay, SPATA2 KD with mechanistic readout; single lab","pmids":["36195186"],"is_preprint":false},{"year":2019,"finding":"CYLD promotes dendritic growth in neurons through regulation of α-tubulin acetylation. CYLD also promotes postsynaptic spine formation through a mechanism dependent on its first microtubule-binding domain but independent of tubulin acetylation, indicating distinct molecular mechanisms for dendritic growth vs. spine formation.","method":"CYLD overexpression and knockdown in hippocampal neurons, tubulin acetylation-site mutants (co-expression rescue), CYLD domain truncation/mutation analysis, live cell imaging of dendrites and spines","journal":"The European journal of neuroscience","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — domain mutagenesis, mechanistic dissection of two cellular processes; single lab","pmids":["31001844"],"is_preprint":false},{"year":2020,"finding":"CYLD phosphorylation (inhibitory modification) is mediated by TBK1/IKKε and IKKβ, and its reversal by IKK inhibitors restores CYLD deubiquitinase activity toward RIPK1, reducing RIPK1 K63-ubiquitination and triggering RIPK1 recruitment to the DISC and cell death in ATLL cells.","method":"IKK inhibitors (MRT67307, TPCA), kinase-inactive TBK1 overexpression, CYLD phosphorylation assay, RIPK1 ubiquitination assay, DISC immunoprecipitation, CYLD KO controls","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological and genetic manipulation of IKK, CYLD phosphorylation and substrate assay, mechanistic DISC readout; single lab","pmids":["32024820"],"is_preprint":false},{"year":2021,"finding":"CYLD overexpression promotes K48-linked ubiquitination and degradation of NoxO1 (NADPH oxidase organizer 1), reducing NoxO1 protein half-life and suppressing excessive ROS generation. CYLD-mediated NoxO1 destabilization suppresses prostate cancer cell proliferation and tumor growth.","method":"CRISPR/Cas9 DUB-KO library screen, Co-immunoprecipitation, NoxO1 ubiquitination assay, CHX half-life assay, CYLD CRISPR KO in prostate cancer cells, xenograft tumor assay","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genome-wide screen validated by Co-IP and ubiquitination assay, half-life experiment; novel K48-chain substrate; single lab","pmids":["34742871"],"is_preprint":false}],"current_model":"CYLD is a lysine 63 (and K48/M1)-deubiquitinase that functions as a tumor suppressor by removing non-degradative ubiquitin chains from a broad set of signaling substrates—including TRAF2, TRAF6, TAK1, RIG-I, RIPK1, RIPK2, RIP3, PLK4, Cep70, plakoglobin, Bcl-3, p53, ERK1/2, p18, NoxO1, and NLRP6—thereby attenuating NF-κB, JNK, MAPK, necroptotic, and inflammasome signaling; its activity is regulated by IKK-dependent phosphorylation (inhibitory at Ser418/Ser568 and degradation-targeting at Ser432/436 via SCFβ-TRCP), MALT1-mediated proteolytic cleavage, SUMOylation, and allosteric activation by the adaptor SPATA2, while it also regulates microtubule dynamics and cell cycle progression through its CAP-Gly ubiquitin-binding domains and HDAC6 inhibition, independently of its catalytic activity in some contexts."},"narrative":{"mechanistic_narrative":"CYLD is a deubiquitinase that acts as a central negative regulator of inflammatory and survival signaling, removing predominantly K63-linked (and in some contexts K48- and M1-linked) ubiquitin chains from a broad repertoire of signaling proteins to function as a tumor suppressor [PMID:12917689]. In the canonical NF-κB and MAPK axes it deubiquitinates the TRAF/TAK1 module—TRAF2 and TRAF6 to dampen TNFR-family and RANK signaling [PMID:12917689, PMID:18382763] and TAK1 to limit JNK and IKKβ activation [PMID:17548520]—and extends this control to RIG-I antiviral signaling, RIPK2, and the AP-1 components c-Jun and c-Fos [PMID:18467330, PMID:26834734, PMID:21478324]. CYLD partitions inflammatory versus death outcomes by editing RIPK1 ubiquitination within the necrosome and at death-signaling Complex II, governing necroptosis and TNF-induced cell death [PMID:24098568, PMID:34887354]. Beyond cytokine signaling it restrains inflammasome activity by deubiquitinating NLRP6 and, via centrosomal PLK4, the NEK7–NLRP3 axis [PMID:32424362, PMID:31762063]. A second, frequently catalysis-independent arm of CYLD function operates on the cytoskeleton: through its CAP-Gly ubiquitin-binding domains it associates with α-tubulin and microtubule plus-end factor EB1, enhances microtubule assembly, and inhibits HDAC6 to raise tubulin acetylation, thereby controlling cell migration, cell-cycle progression, ciliogenesis (via Cep70 and PCM1), and gap-junction assembly (via plakoglobin) [PMID:18222923, PMID:19893491, PMID:24552808, PMID:25342559, PMID:31067453, PMID:36577382]. CYLD activity is tightly regulated by IKK-family phosphorylation that is inhibitory at Ser418/Ser568 and degradation-targeting at Ser432/436 through SCFβ-TRCP, by MALT1 proteolytic cleavage, by SUMOylation, and by allosteric activation through the adaptor SPATA2, which recruits CYLD to the TNF receptor complex and the centrosome [PMID:15870263, PMID:34610306, PMID:24961988, PMID:21448133, PMID:24909169, PMID:27458237]. It additionally controls p53 stabilization, the CDK inhibitor p18, and Bcl-3 nuclear accumulation, linking its deubiquitinase activity to growth-suppressive and proliferative checkpoints [PMID:27561390, PMID:33654169, PMID:16713561].","teleology":[{"year":2003,"claim":"Established CYLD's foundational identity as a deubiquitinase that restrains NF-κB signaling, explaining how its loss drives tumorigenesis.","evidence":"Deubiquitinase activity assays, RNAi, and Co-IP across CD40/XEDAR/EDAR receptor systems with NF-κB reporters","pmids":["12917689"],"confidence":"High","gaps":["Did not resolve ubiquitin chain-type specificity in vivo","Direct enzymatic action on TRAF2/TRAF6 chains versus indirect effects not fully separated"]},{"year":2005,"claim":"Defined the first regulatory feedback on CYLD, showing IKKγ/NEMO-dependent phosphorylation transiently silences its activity to permit signal-induced TRAF2 ubiquitination.","evidence":"In vivo phosphorylation assays with IKK subunit overexpression/knockdown and TRAF2 ubiquitination readout","pmids":["15870263"],"confidence":"High","gaps":["Precise phospho-sites and how phosphorylation alters catalysis not mapped","Kinase responsible for direct phosphorylation versus complex effects unclear"]},{"year":2006,"claim":"Revealed context-dependent and even positive roles for CYLD—deubiquitinating Bcl-3 to block keratinocyte proliferation and deubiquitinating Lck to promote TCR signaling—beyond a uniform negative regulator.","evidence":"CYLD knockout mice, subcellular fractionation/live imaging, Co-IP, and T cell development analysis","pmids":["16713561","16501569"],"confidence":"High","gaps":["Molecular basis of stimulus-induced CYLD perinuclear translocation unresolved","How CYLD distinguishes positive vs negative substrate outcomes not defined"]},{"year":2007,"claim":"Extended CYLD substrate control to TAK1 and uncovered a deubiquitinase-independent cytoskeletal role, broadening its mechanism into both signaling and microtubule biology.","evidence":"Knockout mice and ubiquitination/kinase assays for TAK1; immunofluorescence, Plk1 Co-IP, and Drosophila dTRAF2 genetics for cytoskeletal/developmental roles","pmids":["17548520","17765686","17495026"],"confidence":"High","gaps":["Mechanistic link between CYLD-Plk1 interaction and mitotic entry not reconstituted","How a single enzyme coordinates signaling and microtubule functions unclear"]},{"year":2008,"claim":"Mapped CYLD's cytoskeletal mechanism to its first CAP-Gly domain that binds microtubules and enhances tubulin polymerization, and added RIG-I antiviral and TRAF6/RANK osteoclast substrates.","evidence":"In vitro microtubule co-sedimentation and polymerization assays with CAP-Gly mutants; knockout cells/mice with RIG-I and TRAF6 ubiquitination assays","pmids":["18222923","18467330","18382763"],"confidence":"High","gaps":["Whether CAP-Gly microtubule binding requires ubiquitin recognition not resolved","Recruitment adaptor p62 mechanism for TRAF6 not generalized"]},{"year":2009,"claim":"Integrated CYLD's cytoskeletal and proliferation control by showing CAP-Gly-mediated HDAC6 inhibition raises acetylated tubulin to gate G1/S and cytokinesis, and linked Snail1 repression of CYLD to Bcl-3-driven oncogenesis.","evidence":"Co-IP, immunofluorescence, cell-cycle analysis, HDAC6 inhibition assays; Snail1 expression/knockdown with CYLD rescue in tumor models","pmids":["19893491","19124656"],"confidence":"High","gaps":["Whether HDAC6 is a direct CYLD binding partner vs indirect target not fully defined","Catalytic-independent vs dependent contributions to each phenotype not cleanly separated"]},{"year":2011,"claim":"Established multilayered regulation and substrate logic—MALT1 proteolytic cleavage inactivates CYLD to permit JNK activation, the Itch-CYLD complex sequentially edits TAK1 from K63 to K48, and CYLD blocks AP-1 by deubiquitinating c-Jun/c-Fos.","evidence":"MALT1 protease assays, complex reconstitution with domain-specific CYLD mutants, in vitro and in vivo ubiquitination assays","pmids":["21448133","22057290","21478324"],"confidence":"High","gaps":["Physiological balance between cleavage and editing not quantified","How CYLD partner choice (Itch vs others) is determined unclear"]},{"year":2014,"claim":"Greatly expanded CYLD's substrate and regulatory landscape—p53 stabilization, ciliogenesis via Cep70, EB1 plus-end coordination, HDAC7-dependent transcription, MIB2/Notch, SUMOylation and β-TRCP phosphodegron control—cementing it as a hub of both catalytic and non-catalytic functions.","evidence":"Knockout/catalytic-mutant mice, ubiquitin chain-restriction analysis, ChIP, yeast two-hybrid, SUMOylation and phospho-mutant assays","pmids":["27561390","25342559","24552808","24811579","25565632","24909169","24961988"],"confidence":"High","gaps":["Relative in vivo importance of the many substrates unranked","Some interactions rest on single-lab Co-IP without reciprocal validation"]},{"year":2015,"claim":"Demonstrated pathway-selective output by showing CYLD-TRAF2 deubiquitination maintains hematopoietic stem cell quiescence through p38MAPK rather than NF-κB.","evidence":"Conditional and TRAF2-binding-mutant knockout mice with quiescence/repopulation assays and p38MAPK inhibitor rescue","pmids":["25824820"],"confidence":"High","gaps":["Mechanism directing TRAF2 deubiquitination to p38 vs NF-κB output not defined","Direct p38 substrate connection not established"]},{"year":2016,"claim":"Identified SPATA2 as the direct allosteric activator and recruitment adaptor for CYLD at the TNF receptor complex, explaining how CYLD is positioned and switched on.","evidence":"Mass spectrometry, Co-IP, in vitro deubiquitinase assay with SPATA2, and Complex I isolation","pmids":["27458237"],"confidence":"High","gaps":["Structural basis of PUB-domain allosteric activation not solved here","Whether SPATA2 governs all CYLD substrate engagements unknown"]},{"year":2019,"claim":"Connected the SPATA2-CYLD module to centrosome and inflammasome biology via PLK4-NEK7-NLRP3 and PCM1-dependent centriolar satellite proteostasis, and extended cytoskeletal roles to neuronal dendrite and spine formation.","evidence":"Co-IP, substrate ubiquitination and phosphorylation assays, SPATA2/PCM1 knockout/knockdown, and neuronal domain-mutant imaging","pmids":["31762063","31067453","31001844"],"confidence":"High","gaps":["Coordination of centrosomal vs receptor-complex CYLD pools unclear","PCM1/Cep70 regulation rest on single-lab evidence"]},{"year":2020,"claim":"Linked CYLD regulation to disease through NLRP6 inflammasome control, IKK-reversible RIPK1 editing in leukemic cell death, and a gain-of-function M719V variant tied to neurological disease and autophagy.","evidence":"Knockout mouse infection models with IL-18 readout, IKK inhibitor/DISC studies, and in vitro deubiquitinase plus NF-κB/autophagy assays of the variant","pmids":["32424362","32024820","32185393"],"confidence":"High","gaps":["How elevated catalytic activity of M719V perturbs autophagosome-lysosome fusion mechanistically unclear","In vivo consequences of IKK-reversal in normal tissue not addressed"]},{"year":2021,"claim":"Refined the activation logic—defining CAP-Gly3 as essential for catalysis, a Ser418/Ser568 phosphorylation switch activating CYLD toward K63 chains, and a CYLD-SPATA2-LUBAC ubiquitin-editing complex—and added ERK1/2, p18, plakoglobin, and NoxO1 as substrates with K63 or K48 specificity.","evidence":"Biochemical CAP-Gly/phospho-mutant deubiquitinase assays, in vitro ubiquitination/site-mapping with TRIM15, half-life and ubiquitination assays, and KO mouse cardiac phenotyping","pmids":["34610306","34497368","33654169","34742871","36577382"],"confidence":"High","gaps":["Structural model of phospho-activated CYLD not provided","Switching between K63 and K48 chain editing on different substrates mechanistically unresolved"]},{"year":2022,"claim":"Tied the SPATA2-CYLD axis to cell-death and metabolic pathologies, regulating NCOA4-dependent ferritinophagy/ferroptosis and SHARPIN-dependent Complex II death signaling.","evidence":"Co-IP, NCOA4 ubiquitination assays, SPATA2 knockdown in cardiomyocyte/mouse models; genetic double-knockout and Complex II immunoprecipitation","pmids":["36195186","34887354"],"confidence":"High","gaps":["NCOA4 ferritinophagy axis rests on single-lab evidence","Tissue-specific determinants of pro-death versus pro-survival CYLD output not defined"]},{"year":null,"claim":"How CYLD's catalytic deubiquitinase functions and its catalysis-independent CAP-Gly/microtubule and transcriptional roles are integrated, and which of its many substrates dominate in any given tissue, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified structural model linking phospho-regulation, SPATA2 binding, and chain-type selectivity","Substrate hierarchy across tissues unranked","Mechanistic basis of catalysis-independent functions incompletely defined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,4,8,18,26,29,31,35]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,14,21,32]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[9,10,19]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[10,20]}],"localization":[{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[9,10,19]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[1,16]},{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[17,27,28]},{"term_id":"GO:0005929","term_label":"cilium","supporting_discovery_ids":[17,28]}],"pathway":[{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[0,4,8,31]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[3,7,26,29]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[16,34]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[5,10,33]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[17,27,28]}],"complexes":["CYLD-SPATA2-LUBAC ubiquitin-editing complex","Itch-CYLD complex","TNF receptor signaling complex (Complex I)"],"partners":["SPATA2","TRAF2","TRAF6","TAK1","EB1","HDAC6","ITCH","PLK1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9NQC7","full_name":"Ubiquitin carboxyl-terminal hydrolase CYLD","aliases":["Deubiquitinating enzyme CYLD","Ubiquitin thioesterase CYLD","Ubiquitin-specific-processing protease CYLD"],"length_aa":956,"mass_kda":107.3,"function":"Deubiquitinase that specifically cleaves 'Lys-63'- and linear 'Met-1'-linked polyubiquitin chains and is involved in NF-kappa-B activation and TNF-induced necroptosis (PubMed:18313383, PubMed:18636086, PubMed:26670046, PubMed:26997266, PubMed:27458237, PubMed:27591049, PubMed:27746020, PubMed:29291351, PubMed:32185393). Negatively regulates NF-kappa-B activation by deubiquitinating upstream signaling factors (PubMed:12917689, PubMed:12917691, PubMed:32185393). Contributes to the regulation of cell survival, proliferation and differentiation via its effects on NF-kappa-B activation (PubMed:12917690). Negative regulator of Wnt signaling (PubMed:20227366). Inhibits HDAC6 and thereby promotes acetylation of alpha-tubulin and stabilization of microtubules (PubMed:19893491). Plays a role in the regulation of microtubule dynamics, and thereby contributes to the regulation of cell proliferation, cell polarization, cell migration, and angiogenesis (PubMed:18222923, PubMed:20194890). Required for normal cell cycle progress and normal cytokinesis (PubMed:17495026, PubMed:19893491). Inhibits nuclear translocation of NF-kappa-B (PubMed:18636086). Plays a role in the regulation of inflammation and the innate immune response, via its effects on NF-kappa-B activation (PubMed:18636086). Dispensable for the maturation of intrathymic natural killer cells, but required for the continued survival of immature natural killer cells (By similarity). Negatively regulates TNFRSF11A signaling and osteoclastogenesis (By similarity). Involved in the regulation of ciliogenesis, allowing ciliary basal bodies to migrate and dock to the plasma membrane; this process does not depend on NF-kappa-B activation (By similarity). Ability to remove linear ('Met-1'-linked) polyubiquitin chains regulates innate immunity and TNF-induced necroptosis: recruited to the LUBAC complex via interaction with SPATA2 and restricts linear polyubiquitin formation on target proteins (PubMed:26670046, PubMed:26997266, PubMed:27458237, PubMed:27591049). Regulates innate immunity by restricting linear polyubiquitin formation on RIPK2 in response to NOD2 stimulation (PubMed:26997266). Involved in TNF-induced necroptosis by removing linear ('Met-1'-linked) polyubiquitin chains from RIPK1, thereby regulating the kinase activity of RIPK1 (By similarity). Negatively regulates intestinal inflammation by removing 'Lys-63' linked polyubiquitin chain of NLRP6, thereby reducing the interaction between NLRP6 and PYCARD/ASC and formation of the NLRP6 inflammasome (By similarity). Does not catalyze deubiquitination of heterotypic 'Lys-63'-/'Lys-48'-linked branched ubiquitin chains (PubMed:27746020). Removes 'Lys-63' linked polyubiquitin chain of MAP3K7, which inhibits phosphorylation and blocks downstream activation of the JNK-p38 kinase cascades (PubMed:29291351). Also removes 'Lys-63'-linked polyubiquitin chains of MAP3K1 and MA3P3K3, which inhibit their interaction with MAP2K1 and MAP2K2 (PubMed:34497368)","subcellular_location":"Cytoplasm; Cytoplasm, perinuclear region; Cytoplasm, cytoskeleton; Cell membrane; Cytoplasm, cytoskeleton, microtubule organizing center, centrosome; Cytoplasm, cytoskeleton, spindle; Cytoplasm, cytoskeleton, cilium basal body","url":"https://www.uniprot.org/uniprotkb/Q9NQC7/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CYLD","classification":"Not Classified","n_dependent_lines":34,"n_total_lines":1208,"dependency_fraction":0.028145695364238412},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/CYLD","total_profiled":1310},"omim":[{"mim_id":"621119","title":"ENKURIN DOMAIN-CONTAINING PROTEIN 1; ENKD1","url":"https://www.omim.org/entry/621119"},{"mim_id":"619132","title":"FRONTOTEMPORAL DEMENTIA AND/OR AMYOTROPHIC LATERAL SCLEROSIS 8; FTDALS8","url":"https://www.omim.org/entry/619132"},{"mim_id":"612099","title":"TRICHOEPITHELIOMA, MULTIPLE FAMILIAL, 2","url":"https://www.omim.org/entry/612099"},{"mim_id":"611885","title":"SHANK-ASSOCIATED RH DOMAIN INTERACTOR; SHARPIN","url":"https://www.omim.org/entry/611885"},{"mim_id":"609888","title":"LEPROSY, SUSCEPTIBILITY TO, 1; LPRS1","url":"https://www.omim.org/entry/609888"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Centriolar satellite","reliability":"Supported"},{"location":"Nucleoplasm","reliability":"Additional"},{"location":"Primary cilium transition zone","reliability":"Additional"},{"location":"Basal body","reliability":"Additional"},{"location":"Principal piece","reliability":"Additional"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"bone marrow","ntpm":89.0}],"url":"https://www.proteinatlas.org/search/CYLD"},"hgnc":{"alias_symbol":["KIAA0849","USPL2"],"prev_symbol":["CYLD1"]},"alphafold":{"accession":"Q9NQC7","domains":[{"cath_id":"2.30.30.190","chopping":"124-207","consensus_level":"medium","plddt":87.1311,"start":124,"end":207},{"cath_id":"2.30.30.190","chopping":"233-303","consensus_level":"high","plddt":86.0241,"start":233,"end":303},{"cath_id":"2.30.30.190","chopping":"473-540","consensus_level":"high","plddt":87.5121,"start":473,"end":540},{"cath_id":"-","chopping":"785-854","consensus_level":"medium","plddt":84.8767,"start":785,"end":854}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NQC7","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NQC7-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9NQC7-F1-predicted_aligned_error_v6.png","plddt_mean":74.44},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CYLD","jax_strain_url":"https://www.jax.org/strain/search?query=CYLD"},"sequence":{"accession":"Q9NQC7","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9NQC7.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9NQC7/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9NQC7"}},"corpus_meta":[{"pmid":"12917689","id":"PMC_12917689","title":"CYLD 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2003,\n      \"finding\": \"CYLD is a deubiquitinating enzyme that negatively regulates NF-κB activation by removing ubiquitin chains from TRAF2 and TRAF6, thereby inhibiting signaling downstream of TNFR family members CD40, XEDAR, and EDAR. Loss of CYLD deubiquitinating activity correlates with tumorigenesis.\",\n      \"method\": \"Deubiquitinase activity assay, RNA interference knockdown, Co-immunoprecipitation, NF-κB reporter assays\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — enzymatic activity demonstrated, substrates identified by Co-IP, RNAi phenotype, replicated across multiple receptor systems in a landmark paper\",\n      \"pmids\": [\"12917689\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"CYLD deubiquitinates Bcl-3 in the perinuclear region, preventing nuclear accumulation of Bcl-3/p50 and Bcl-3/p52 complexes and thereby blocking cyclin D1-dependent keratinocyte proliferation. TPA or UV triggers translocation of CYLD from cytoplasm to perinuclear region to execute this function. This pathway is distinct from CYLD's regulation of TRAF2/p65-p50 NF-κB survival signaling.\",\n      \"method\": \"CYLD knockout mice, Co-immunoprecipitation, subcellular fractionation/live imaging, chemical induction (TPA/UV), cyclin D1 expression assays\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — multiple orthogonal methods (KO mice, Co-IP, fractionation, functional rescue), novel substrate identified with mechanistic consequence\",\n      \"pmids\": [\"16713561\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"CYLD is phosphorylated by IκB kinase gamma (IKKγ/NEMO)-dependent signaling in response to cellular stimuli, and this phosphorylation transiently inhibits CYLD's deubiquitinase activity, allowing signal-induced TRAF2 ubiquitination and downstream IKK activation.\",\n      \"method\": \"In vivo phosphorylation assays, IKK subunit overexpression and knockdown, TRAF2 ubiquitination assays\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — biochemical phosphorylation mapping, functional consequence on substrate ubiquitination demonstrated, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"15870263\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"CYLD physically interacts with active Lck and promotes recruitment of active Lck to its substrate Zap70 during T cell receptor signaling in thymocytes. CYLD also removes both K48- and K63-linked polyubiquitin chains from Lck, positively regulating proximal TCR signaling and T cell development.\",\n      \"method\": \"Co-immunoprecipitation, CYLD knockout mice, T cell development analysis, ubiquitin chain-type analysis\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — reciprocal Co-IP, KO mice with defined developmental phenotype, two orthogonal methods in single lab\",\n      \"pmids\": [\"16501569\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"CYLD deubiquitinates and inactivates TAK1 (transforming growth factor-β-activated kinase 1), inhibiting TAK1 ubiquitination and autoactivation, thereby suppressing downstream JNK and IKKβ activation in T cells.\",\n      \"method\": \"CYLD knockout mice, Co-immunoprecipitation, TAK1 ubiquitination and kinase activity assays\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO mice with constitutive TAK1 activation phenotype, Co-IP demonstrating physical interaction, substrate ubiquitination assay\",\n      \"pmids\": [\"17548520\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"CYLD localizes to microtubules during interphase and to the midbody during telophase, and its protein levels decrease as cells exit mitosis. CYLD physically interacts with Plk1 and shares similar loss-of-function and overexpression phenotypes, implicating CYLD in regulating timely mitotic entry.\",\n      \"method\": \"Immunofluorescence/live cell imaging, Co-immunoprecipitation, loss-of-function and overexpression experiments, cell cycle analysis\",\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 — localization with functional consequence, Co-IP with Plk1, but mechanistic link between CYLD-Plk1 interaction and mitotic entry not fully reconstituted\",\n      \"pmids\": [\"17495026\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"Drosophila CYLD (dCYLD) deubiquitinates dTRAF2, preventing its ubiquitin-mediated proteolytic degradation, and thereby regulates TNF-induced JNK activation and JNK-dependent cell death. dTRAF2 acts downstream of the TNF receptor Wengen and upstream of the JNKK kinase dTAK1.\",\n      \"method\": \"Drosophila genetic mutants, transgenic flies with wild-type and catalytic-dead dCYLD, dTRAF2 ubiquitination assay, epistasis analysis\",\n      \"journal\": \"Developmental cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — reconstitution in vivo via genetics, deubiquitinase activity on specific substrate demonstrated, epistasis with defined pathway placement\",\n      \"pmids\": [\"17765686\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"CYLD negatively regulates RANK signaling and osteoclastogenesis by inhibiting TRAF6 ubiquitination. CYLD physically interacts with the signaling adaptor p62, which recruits CYLD to TRAF6.\",\n      \"method\": \"CYLD knockout mice, RANKL-induced osteoclast differentiation assays, Co-immunoprecipitation, TRAF6 ubiquitination assays\",\n      \"journal\": \"The Journal of clinical investigation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO mice with severe osteoporosis phenotype, Co-IP demonstrating CYLD-p62-TRAF6 complex, substrate ubiquitination assay\",\n      \"pmids\": [\"18382763\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"CYLD deubiquitinates RIG-I (a cytoplasmic RNA sensor) and inhibits its ubiquitination, thereby suppressing RIG-I-mediated activation of IKKε/TBK1 and IFN-β promoter induction during viral infection.\",\n      \"method\": \"CYLD knockout cells, RIG-I ubiquitination assay, IFN-β reporter assay, IKKε/TBK1 kinase activation assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO cells with constitutive IKKε/TBK1 activation, substrate ubiquitination assay, reporter assay; single lab\",\n      \"pmids\": [\"18467330\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"CYLD associates with microtubules both in cells and in vitro through its first CAP-Gly domain. CYLD enhances tubulin polymerization by lowering the critical concentration for microtubule assembly and promotes microtubule regrowth after nocodazole washout. The first CAP-Gly domain is required for CYLD-dependent cell migration.\",\n      \"method\": \"In vitro microtubule co-sedimentation assay, tubulin polymerization assay, CAP-Gly domain deletion mutants, nocodazole washout assay, wound healing assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of tubulin polymerization, domain mutagenesis, multiple orthogonal methods in single lab\",\n      \"pmids\": [\"18222923\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"CYLD controls cell cycle progression at G1/S and cytokinesis by associating with α-tubulin and microtubules through its CAP-Gly domains. CYLD inhibits HDAC6, increasing perinuclear acetylated α-tubulin levels, which facilitates CYLD interaction with Bcl-3 to delay G1-to-S transition. CYLD also interacts with HDAC6 at the midbody to regulate cytokinesis in a deubiquitinase-independent manner.\",\n      \"method\": \"Co-immunoprecipitation, immunofluorescence, cell cycle analysis, HDAC6 inhibition assays, CYLD-Bcl-3 interaction assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple Co-IPs, defined cell cycle phenotypes, deubiquitinase-independent mechanism demonstrated, multiple orthogonal methods\",\n      \"pmids\": [\"19893491\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Snail1 transcriptionally represses CYLD expression in melanoma. As a consequence of CYLD repression, BCL-3 translocates to the nucleus, activating Cyclin D1 and N-cadherin promoters and driving proliferation and invasion.\",\n      \"method\": \"Snail1 expression/knockdown, CYLD rescue experiments, in vitro and in vivo tumor models, tissue microarray analysis\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — CYLD rescue experiments show functional consequence, Snail1-CYLD axis tested in multiple models; single lab\",\n      \"pmids\": [\"19124656\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"CYLD knockdown significantly impairs angiogenesis by blocking endothelial cell spreading, migration, and polarity. CYLD regulates microtubule dynamics in endothelial cells and activates Rac1, with Rac1 activation being an important downstream mechanism for CYLD's role in endothelial cell migration.\",\n      \"method\": \"siRNA knockdown, tube formation assay, 3D capillary sprouting assay, in vivo angioreactor assay, microtubule dynamics imaging, Rac1 activation assay\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple in vitro and in vivo angiogenesis assays, defined Rac1 mechanism; single lab\",\n      \"pmids\": [\"20194890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"MALT1 proteolytically cleaves and inactivates CYLD following TCR activation in T cells and by oncogenic API2-MALT1. CYLD cleavage by MALT1 is specifically required for JNK activation and inducible expression of a subset of TCR-responsive genes.\",\n      \"method\": \"MALT1 protease activity assays, CYLD cleavage detection, JNK activation assay, gene expression analysis, MALT1 inhibitor studies\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — direct proteolytic cleavage of CYLD by MALT1 demonstrated biochemically, functional consequence on JNK signaling established with multiple methods\",\n      \"pmids\": [\"21448133\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Itch (E3 ligase) and CYLD form a complex via WW-PPXY motif interactions. The Itch-CYLD complex sequentially removes K63-linked ubiquitin chains (via CYLD) and then catalyzes K48-linked ubiquitination (via Itch) on TAK1, terminating inflammatory TNF signaling. CYLD(Y485A) mutant unable to associate with Itch cannot block sustained TAK1 activation.\",\n      \"method\": \"Co-immunoprecipitation, in vitro deubiquitination assay, TAK1 ubiquitination assay, CYLD mutant reconstitution in CYLD−/− macrophages, cytokine production assay\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — complex reconstitution, sequential enzymatic activities demonstrated, domain-specific mutant validates binding requirement, multiple orthogonal methods\",\n      \"pmids\": [\"22057290\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CYLD deubiquitinates c-Jun and c-Fos, removing K63-linked ubiquitin chains from these AP-1 components. Catalytically inactive CYLD mutant (CYLDm) increases K63 ubiquitination on c-Jun and c-Fos, leading to sustained AP-1 activation via JNK. CYLD thus blocks JNK/AP-1 signaling at multiple levels.\",\n      \"method\": \"In vivo ubiquitination assay for c-Jun and c-Fos, JNK activation assay, transgenic mouse skin tumorigenesis model, JNK inhibitor treatment\",\n      \"journal\": \"Cancer prevention research (Philadelphia, Pa.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — novel substrates c-Jun and c-Fos identified by ubiquitination assay, functional JNK inhibitor validation; single lab\",\n      \"pmids\": [\"21478324\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"CYLD regulates RIP1 ubiquitination specifically within the NP-40 insoluble necrosome (not at the TNFR-1 signaling complex) during TNFα-induced programmed necrosis. Increased RIP1 ubiquitination in the necrosome in CYLD−/− cells impairs RIP1 and RIP3 phosphorylation, inhibiting kinase activation required for necroptosis.\",\n      \"method\": \"CYLD knockout cells, RIP1 ubiquitination assay in necrosome fraction, RIP1-RIP3 phosphorylation assay, SMAC mimetic and cycloheximide sensitization experiments\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — specific compartment (necrosome vs. TNFR complex) resolved by fractionation, KO cells with multiple sensitization conditions, kinase activation assay\",\n      \"pmids\": [\"24098568\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD mediates ciliogenesis by deconjugating polyubiquitin chains from centrosomal protein Cep70, which is required for Cep70 to interact with γ-tubulin and localize at the centrosome. Additionally, CYLD-mediated inhibition of HDAC6 promotes tubulin acetylation, constituting a second mechanism for ciliary assembly.\",\n      \"method\": \"CYLD knockout mice (polydactyly, ciliary defect phenotype), Co-immunoprecipitation (Cep70-γ-tubulin), Cep70 ubiquitination assay, HDAC6 inhibitor rescue experiments\",\n      \"journal\": \"Cell research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KO mice with defined ciliopathy phenotype, substrate deubiquitination demonstrated, two distinct mechanisms shown, HDAC6 inhibitor rescue\",\n      \"pmids\": [\"25342559\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD interacts with and deubiquitinates p53, facilitating p53 stabilization in response to genotoxic stress by removing K63-linked ubiquitin chains (and thereby indirectly preventing K48-linked degradation). Loss of CYLD catalytic activity impairs DNA damage-induced p53 stabilization and activation.\",\n      \"method\": \"Co-immunoprecipitation, p53 ubiquitination assay, ubiquitin chain-restriction analysis, CYLD catalytic mutant knock-in mice, C. elegans CEP-1/p53 epistasis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — multiple methods including ubiquitin chain-restriction analysis, genetic validation in C. elegans, mouse model with defined phenotype\",\n      \"pmids\": [\"27561390\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD coordinates with EB1 at microtubule plus ends to regulate microtubule dynamics and cell migration. The CYLD-EB1 interaction increases upon stimulation of cell migration. CYLD and EB1 act in concert to regulate tail retraction, centrosome reorientation, leading-edge microtubule stabilization, and microtubule nucleation.\",\n      \"method\": \"Yeast two-hybrid screening, Co-immunoprecipitation, in vitro microtubule assembly assay, live cell imaging of migration, centrosome reorientation assay\",\n      \"journal\": \"Cell cycle (Georgetown, Tex.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast 2-hybrid plus Co-IP validation, in vitro assay, multiple migration readouts; single lab\",\n      \"pmids\": [\"24552808\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD interacts with HDAC7 to remove HDAC7 from the HGF promoter in hepatic stellate cells, inducing HGF gene transcription independently of CYLD's deubiquitinating activity.\",\n      \"method\": \"Co-immunoprecipitation, ChIP assay, CYLD catalytic-dead mutant experiments, CYLD−/− mice with HGF measurement\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ChIP, and catalytic-dead mutant establish deubiquitinase-independent mechanism; single lab\",\n      \"pmids\": [\"24811579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD deubiquitinating activity is inhibited by SUMOylation. Retinoic acid-induced SUMOylation of CYLD blocks its deubiquitinase activity against TRAF2 and TRAF6, facilitating NF-κB signaling during neuroblastoma differentiation.\",\n      \"method\": \"SUMOylation assay, CYLD deubiquitinase activity assay with TRAF2/TRAF6 substrates, non-SUMOylatable CYLD mutant overexpression, NF-κB activation assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct enzymatic activity assay with and without SUMOylation, non-SUMOylatable mutant functional rescue; single lab\",\n      \"pmids\": [\"24909169\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"CYLD interacts with MIB2 (a ubiquitin ligase), stabilizing MIB2 protein levels and reducing JAG2 expression, thereby attenuating Notch signaling. CYLD gene silencing increases JAG2 expression and upregulates Notch signaling.\",\n      \"method\": \"Proteomics/mass spectrometry to identify interactors, Co-immunoprecipitation, siRNA knockdown, Notch target gene expression analysis, primary tumor cell cultures\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — MS-identified interaction validated by Co-IP, siRNA knockdown with pathway readout; single lab\",\n      \"pmids\": [\"25565632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"CYLD maintains hematopoietic stem cell (HSC) quiescence through deubiquitination of TRAF2. This function operates through the p38MAPK pathway (not NF-κB), and pharmacological inhibition of p38MAPK rescues the CYLD-loss HSC phenotype of increased cycling and loss of repopulation potential.\",\n      \"method\": \"Conditional CYLD KO mice, CYLD-TRAF2 binding mutant mice, HSC quiescence assays, transplantation/repopulation assays, p38MAPK inhibitor treatment\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — conditional KO, binding-specific mutant dissecting TRAF2 interaction from catalytic function, epistatic pharmacological rescue; multiple orthogonal approaches\",\n      \"pmids\": [\"25824820\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"SCFβ-TRCP E3 ligase promotes ubiquitination and degradation of CYLD in a manner dependent on prior IKK-mediated phosphorylation of CYLD at Ser432/Ser436. β-TRCP depletion causes CYLD accumulation and TRAF6 deubiquitination, suppressing osteoclastogenesis.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assays, phosphorylation-site mutants, β-TRCP siRNA, TRAF6 ubiquitination assay, osteoclast differentiation assay\",\n      \"journal\": \"Oncotarget\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — phosphodegron mechanism established by phospho-mutant and Co-IP; single lab\",\n      \"pmids\": [\"24961988\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"SPATA2 directly interacts with CYLD via its PUB domain and acts as an allosteric activator of CYLD's K63- and M1-deubiquitinase activity. SPATA2 is required for recruitment of CYLD to the TNF receptor signaling complex, and its loss attenuates TNF-induced NF-κB/MAPK signaling while also reducing Complex II formation and apoptosis.\",\n      \"method\": \"Mass spectrometry screen, Co-immunoprecipitation, in vitro deubiquitinase activity assay with SPATA2, TNF signaling complex (Complex I) isolation\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — allosteric activation demonstrated by in vitro enzyme assay, Co-IP, receptor complex recruitment shown; multiple orthogonal methods in single lab\",\n      \"pmids\": [\"27458237\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"CYLD removes K63-linked ubiquitin chains from RIPK2, inhibiting RIPK2 K63-ubiquitination and impairing NF-κB and ERK1/2 activation in Listeria-infected macrophages, thereby reducing antibacterial immune responses.\",\n      \"method\": \"Co-immunoprecipitation, RIPK2 ubiquitination assay, CYLD KO macrophages, siRNA knockdown, NF-κB and ERK activation assays\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assay for RIPK2, KO cells with functional immune readout; single lab\",\n      \"pmids\": [\"26834734\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CYLD is recruited by SPATA2 to the centrosome where it deubiquitinates PLK4 (master regulator of centrosome duplication). Deubiquitination of PLK4 facilitates its phosphorylation of NEK7 at Ser204, which attenuates NEK7-NLRP3 interaction and suppresses NLRP3 inflammasome activation.\",\n      \"method\": \"Co-immunoprecipitation, PLK4 ubiquitination assay, NEK7 phosphorylation assay, NLRP3 inflammasome activation assay, SPATA2 KO macrophages, PLK4 inhibitor/shRNA\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — substrate deubiquitination demonstrated, phosphorylation of NEK7 by PLK4 shown, SPATA2-CYLD complex at centrosome, multiple mechanistic steps validated\",\n      \"pmids\": [\"31762063\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CYLD promotes the proteostasis of centriolar satellites by deubiquitinating PCM1 scaffold protein and protecting it from MIB1-mediated proteasomal degradation. CYLD knockdown promotes PCM1 degradation and dismantling of centriolar satellites, impairing ciliogenesis.\",\n      \"method\": \"Unbiased proteomic screen of CYLD binding partners, Co-immunoprecipitation, PCM1 ubiquitination assay, MIB1 E3 ligase assay, siRNA knockdown, cilia formation assay\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proteomics + Co-IP + ubiquitination assay; counterbalanced E3/DUB pair identified; single lab\",\n      \"pmids\": [\"31067453\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CYLD deubiquitinates NLRP6, and this deubiquitination inhibits the NLRP6-ASC inflammasome complex, preventing excessive IL-18 maturation in the colonic mucosa.\",\n      \"method\": \"Co-immunoprecipitation, NLRP6 ubiquitination assay, CYLD knockout mice with Citrobacter rodentium infection, IL-18 maturation assay\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — novel substrate (NLRP6) identified, KO mouse in vivo model, IL-18 maturation as functional readout, published in high-impact journal\",\n      \"pmids\": [\"32424362\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"A gain-of-function CYLD missense variant (M719V) exhibits significantly increased K63 deubiquitinase activity relative to wild-type. Overexpression of CYLD-M719V leads to more potent NF-κB inhibition and impairment of autophagosome fusion to lysosomes.\",\n      \"method\": \"In vitro deubiquitinase activity assay with Wilcoxon signed-rank test, NF-κB luciferase reporter assay, autophagy flux assay in HEK293 cells\",\n      \"journal\": \"Brain : a journal of neurology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — direct enzymatic activity assay, NF-κB and autophagy assays; single lab; gain-of-function mechanism novel\",\n      \"pmids\": [\"32185393\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"TRIM15 is a K63-ubiquitin ligase for ERK1/2 and CYLD is the deubiquitinase that removes K63-linked chains from ERK1/2. K63-linked polyubiquitination of ERK1/2 by TRIM15 enhances ERK interaction with and activation by MEK. TRIM15 and CYLD regulate ERK ubiquitination at defined lysine residues through mutually exclusive interactions.\",\n      \"method\": \"In vitro ubiquitination and deubiquitinase assays, ERK ubiquitination site mapping by mutagenesis, Co-immunoprecipitation, ERK activation (MEK interaction) assay\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of ubiquitination/deubiquitination, substrate site mapping by mutagenesis, mechanistic link to MEK activation; strong method quality\",\n      \"pmids\": [\"34497368\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"Two CAP-Gly domains of CYLD function as ubiquitin-binding domains, with CAP-Gly3 required for CYLD deubiquitinase activity and regulation of immune receptor signaling. A phosphorylation switch outside the catalytic USP domain (Ser568, a TNF-regulated site, acting in concert with Ser418) activates CYLD toward K63-linked polyubiquitin. Phosphorylated CYLD together with SPATA2 and LUBAC functions as a ubiquitin-editing complex.\",\n      \"method\": \"Structural/biochemical analysis of CAP-Gly domains, phosphoproteomic identification of Ser568, in vitro deubiquitinase activity assays with phospho-mimetic mutants, immune receptor signaling assays\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro deubiquitinase assay with defined phospho-mutants, ubiquitin-binding domain biochemistry, novel phosphorylation site identified; multiple orthogonal methods\",\n      \"pmids\": [\"34610306\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CYLD stabilizes p18 (CDK inhibitor) by binding to p18 and removing K48-linked polyubiquitin chains, preventing p18 proteasomal degradation and maintaining G1/S cell cycle arrest.\",\n      \"method\": \"Co-immunoprecipitation, p18 ubiquitination assay (K48 chain type), p18 half-life assay (CHX chase), CYLD KO/knockdown cell cycle analysis, in vivo xenograft\",\n      \"journal\": \"NPJ precision oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, ubiquitination assay, half-life experiment; novel K48-chain substrate for CYLD; single lab\",\n      \"pmids\": [\"33654169\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CYLD is required for SHARPIN-deficient mouse phenotype (dermatitis, spleen architecture disruption). In SHARPIN-deficient cells, impaired CYLD phosphorylation at Ser418 (which normally inhibits CYLD) leads to enhanced CYLD-dependent RIPK1 recruitment to death-signaling Complex II following TNF stimulation, causing myeloid cell death and inflammation.\",\n      \"method\": \"Genetic double KO (Sharpin/CYLD), myeloid-specific conditional Cyld deletion, Complex II immunoprecipitation, CYLD Ser418 phosphorylation analysis\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis with double KO, conditional KO, biochemical Complex II isolation; multiple orthogonal approaches\",\n      \"pmids\": [\"34887354\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CYLD deubiquitinates plakoglobin by removing K63-linked polyubiquitin chains. Deubiquitinated plakoglobin shows enhanced interaction with the desmoplakin/EB1 complex at microtubule plus ends, promoting microtubule-dependent transport of connexin 43 (Cx43) to the cell membrane for gap junction assembly at the intercalated disc.\",\n      \"method\": \"Co-immunoprecipitation, plakoglobin ubiquitination assay (K63), CYLD KO mice (cardiac gap junction, fibrosis phenotype), microtubule transport assay, Cx43 membrane targeting assay\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — novel substrate (plakoglobin) with K63-chain specificity, KO mice with cardiac phenotype, defined transport mechanism; multiple orthogonal methods\",\n      \"pmids\": [\"36577382\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"SPATA2/CYLD pathway regulates ferritinophagy in cardiomyocytes by controlling the ubiquitination and degradation of NCOA4 (the ferritinophagy cargo receptor). CYLD/SPATA2-mediated deubiquitination of NCOA4 enhances ferritin autophagy, leading to intracellular iron overload and ferroptosis.\",\n      \"method\": \"Co-immunoprecipitation (SPATA2-CYLD interaction), NCOA4 ubiquitination assay, SPATA2 knockdown, doxorubicin-treated cardiomyocyte and mouse models, ferritinophagy and ferroptosis readouts\",\n      \"journal\": \"Chemico-biological interactions\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2–3 / Moderate — Co-IP, ubiquitination assay, SPATA2 KD with mechanistic readout; single lab\",\n      \"pmids\": [\"36195186\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"CYLD promotes dendritic growth in neurons through regulation of α-tubulin acetylation. CYLD also promotes postsynaptic spine formation through a mechanism dependent on its first microtubule-binding domain but independent of tubulin acetylation, indicating distinct molecular mechanisms for dendritic growth vs. spine formation.\",\n      \"method\": \"CYLD overexpression and knockdown in hippocampal neurons, tubulin acetylation-site mutants (co-expression rescue), CYLD domain truncation/mutation analysis, live cell imaging of dendrites and spines\",\n      \"journal\": \"The European journal of neuroscience\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — domain mutagenesis, mechanistic dissection of two cellular processes; single lab\",\n      \"pmids\": [\"31001844\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CYLD phosphorylation (inhibitory modification) is mediated by TBK1/IKKε and IKKβ, and its reversal by IKK inhibitors restores CYLD deubiquitinase activity toward RIPK1, reducing RIPK1 K63-ubiquitination and triggering RIPK1 recruitment to the DISC and cell death in ATLL cells.\",\n      \"method\": \"IKK inhibitors (MRT67307, TPCA), kinase-inactive TBK1 overexpression, CYLD phosphorylation assay, RIPK1 ubiquitination assay, DISC immunoprecipitation, CYLD KO controls\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological and genetic manipulation of IKK, CYLD phosphorylation and substrate assay, mechanistic DISC readout; single lab\",\n      \"pmids\": [\"32024820\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CYLD overexpression promotes K48-linked ubiquitination and degradation of NoxO1 (NADPH oxidase organizer 1), reducing NoxO1 protein half-life and suppressing excessive ROS generation. CYLD-mediated NoxO1 destabilization suppresses prostate cancer cell proliferation and tumor growth.\",\n      \"method\": \"CRISPR/Cas9 DUB-KO library screen, Co-immunoprecipitation, NoxO1 ubiquitination assay, CHX half-life assay, CYLD CRISPR KO in prostate cancer cells, xenograft tumor assay\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genome-wide screen validated by Co-IP and ubiquitination assay, half-life experiment; novel K48-chain substrate; single lab\",\n      \"pmids\": [\"34742871\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CYLD is a lysine 63 (and K48/M1)-deubiquitinase that functions as a tumor suppressor by removing non-degradative ubiquitin chains from a broad set of signaling substrates—including TRAF2, TRAF6, TAK1, RIG-I, RIPK1, RIPK2, RIP3, PLK4, Cep70, plakoglobin, Bcl-3, p53, ERK1/2, p18, NoxO1, and NLRP6—thereby attenuating NF-κB, JNK, MAPK, necroptotic, and inflammasome signaling; its activity is regulated by IKK-dependent phosphorylation (inhibitory at Ser418/Ser568 and degradation-targeting at Ser432/436 via SCFβ-TRCP), MALT1-mediated proteolytic cleavage, SUMOylation, and allosteric activation by the adaptor SPATA2, while it also regulates microtubule dynamics and cell cycle progression through its CAP-Gly ubiquitin-binding domains and HDAC6 inhibition, independently of its catalytic activity in some contexts.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"CYLD is a deubiquitinase that acts as a central negative regulator of inflammatory and survival signaling, removing predominantly K63-linked (and in some contexts K48- and M1-linked) ubiquitin chains from a broad repertoire of signaling proteins to function as a tumor suppressor [#0]. In the canonical NF-\\u03baB and MAPK axes it deubiquitinates the TRAF/TAK1 module\\u2014TRAF2 and TRAF6 to dampen TNFR-family and RANK signaling [#0, #7] and TAK1 to limit JNK and IKK\\u03b2 activation [#4]\\u2014and extends this control to RIG-I antiviral signaling, RIPK2, and the AP-1 components c-Jun and c-Fos [#8, #26, #15]. CYLD partitions inflammatory versus death outcomes by editing RIPK1 ubiquitination within the necrosome and at death-signaling Complex II, governing necroptosis and TNF-induced cell death [#16, #34]. Beyond cytokine signaling it restrains inflammasome activity by deubiquitinating NLRP6 and, via centrosomal PLK4, the NEK7\\u2013NLRP3 axis [#29, #27]. A second, frequently catalysis-independent arm of CYLD function operates on the cytoskeleton: through its CAP-Gly ubiquitin-binding domains it associates with \\u03b1-tubulin and microtubule plus-end factor EB1, enhances microtubule assembly, and inhibits HDAC6 to raise tubulin acetylation, thereby controlling cell migration, cell-cycle progression, ciliogenesis (via Cep70 and PCM1), and gap-junction assembly (via plakoglobin) [#9, #10, #19, #17, #28, #35]. CYLD activity is tightly regulated by IKK-family phosphorylation that is inhibitory at Ser418/Ser568 and degradation-targeting at Ser432/436 through SCF\\u03b2-TRCP, by MALT1 proteolytic cleavage, by SUMOylation, and by allosteric activation through the adaptor SPATA2, which recruits CYLD to the TNF receptor complex and the centrosome [#2, #32, #24, #13, #21, #25]. It additionally controls p53 stabilization, the CDK inhibitor p18, and Bcl-3 nuclear accumulation, linking its deubiquitinase activity to growth-suppressive and proliferative checkpoints [#18, #33, #1].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Established CYLD's foundational identity as a deubiquitinase that restrains NF-\\u03baB signaling, explaining how its loss drives tumorigenesis.\",\n      \"evidence\": \"Deubiquitinase activity assays, RNAi, and Co-IP across CD40/XEDAR/EDAR receptor systems with NF-\\u03baB reporters\",\n      \"pmids\": [\"12917689\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not resolve ubiquitin chain-type specificity in vivo\", \"Direct enzymatic action on TRAF2/TRAF6 chains versus indirect effects not fully separated\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Defined the first regulatory feedback on CYLD, showing IKK\\u03b3/NEMO-dependent phosphorylation transiently silences its activity to permit signal-induced TRAF2 ubiquitination.\",\n      \"evidence\": \"In vivo phosphorylation assays with IKK subunit overexpression/knockdown and TRAF2 ubiquitination readout\",\n      \"pmids\": [\"15870263\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Precise phospho-sites and how phosphorylation alters catalysis not mapped\", \"Kinase responsible for direct phosphorylation versus complex effects unclear\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Revealed context-dependent and even positive roles for CYLD\\u2014deubiquitinating Bcl-3 to block keratinocyte proliferation and deubiquitinating Lck to promote TCR signaling\\u2014beyond a uniform negative regulator.\",\n      \"evidence\": \"CYLD knockout mice, subcellular fractionation/live imaging, Co-IP, and T cell development analysis\",\n      \"pmids\": [\"16713561\", \"16501569\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of stimulus-induced CYLD perinuclear translocation unresolved\", \"How CYLD distinguishes positive vs negative substrate outcomes not defined\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Extended CYLD substrate control to TAK1 and uncovered a deubiquitinase-independent cytoskeletal role, broadening its mechanism into both signaling and microtubule biology.\",\n      \"evidence\": \"Knockout mice and ubiquitination/kinase assays for TAK1; immunofluorescence, Plk1 Co-IP, and Drosophila dTRAF2 genetics for cytoskeletal/developmental roles\",\n      \"pmids\": [\"17548520\", \"17765686\", \"17495026\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanistic link between CYLD-Plk1 interaction and mitotic entry not reconstituted\", \"How a single enzyme coordinates signaling and microtubule functions unclear\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Mapped CYLD's cytoskeletal mechanism to its first CAP-Gly domain that binds microtubules and enhances tubulin polymerization, and added RIG-I antiviral and TRAF6/RANK osteoclast substrates.\",\n      \"evidence\": \"In vitro microtubule co-sedimentation and polymerization assays with CAP-Gly mutants; knockout cells/mice with RIG-I and TRAF6 ubiquitination assays\",\n      \"pmids\": [\"18222923\", \"18467330\", \"18382763\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether CAP-Gly microtubule binding requires ubiquitin recognition not resolved\", \"Recruitment adaptor p62 mechanism for TRAF6 not generalized\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Integrated CYLD's cytoskeletal and proliferation control by showing CAP-Gly-mediated HDAC6 inhibition raises acetylated tubulin to gate G1/S and cytokinesis, and linked Snail1 repression of CYLD to Bcl-3-driven oncogenesis.\",\n      \"evidence\": \"Co-IP, immunofluorescence, cell-cycle analysis, HDAC6 inhibition assays; Snail1 expression/knockdown with CYLD rescue in tumor models\",\n      \"pmids\": [\"19893491\", \"19124656\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether HDAC6 is a direct CYLD binding partner vs indirect target not fully defined\", \"Catalytic-independent vs dependent contributions to each phenotype not cleanly separated\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Established multilayered regulation and substrate logic\\u2014MALT1 proteolytic cleavage inactivates CYLD to permit JNK activation, the Itch-CYLD complex sequentially edits TAK1 from K63 to K48, and CYLD blocks AP-1 by deubiquitinating c-Jun/c-Fos.\",\n      \"evidence\": \"MALT1 protease assays, complex reconstitution with domain-specific CYLD mutants, in vitro and in vivo ubiquitination assays\",\n      \"pmids\": [\"21448133\", \"22057290\", \"21478324\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological balance between cleavage and editing not quantified\", \"How CYLD partner choice (Itch vs others) is determined unclear\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Greatly expanded CYLD's substrate and regulatory landscape\\u2014p53 stabilization, ciliogenesis via Cep70, EB1 plus-end coordination, HDAC7-dependent transcription, MIB2/Notch, SUMOylation and \\u03b2-TRCP phosphodegron control\\u2014cementing it as a hub of both catalytic and non-catalytic functions.\",\n      \"evidence\": \"Knockout/catalytic-mutant mice, ubiquitin chain-restriction analysis, ChIP, yeast two-hybrid, SUMOylation and phospho-mutant assays\",\n      \"pmids\": [\"27561390\", \"25342559\", \"24552808\", \"24811579\", \"25565632\", \"24909169\", \"24961988\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative in vivo importance of the many substrates unranked\", \"Some interactions rest on single-lab Co-IP without reciprocal validation\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Demonstrated pathway-selective output by showing CYLD-TRAF2 deubiquitination maintains hematopoietic stem cell quiescence through p38MAPK rather than NF-\\u03baB.\",\n      \"evidence\": \"Conditional and TRAF2-binding-mutant knockout mice with quiescence/repopulation assays and p38MAPK inhibitor rescue\",\n      \"pmids\": [\"25824820\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism directing TRAF2 deubiquitination to p38 vs NF-\\u03baB output not defined\", \"Direct p38 substrate connection not established\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Identified SPATA2 as the direct allosteric activator and recruitment adaptor for CYLD at the TNF receptor complex, explaining how CYLD is positioned and switched on.\",\n      \"evidence\": \"Mass spectrometry, Co-IP, in vitro deubiquitinase assay with SPATA2, and Complex I isolation\",\n      \"pmids\": [\"27458237\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of PUB-domain allosteric activation not solved here\", \"Whether SPATA2 governs all CYLD substrate engagements unknown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Connected the SPATA2-CYLD module to centrosome and inflammasome biology via PLK4-NEK7-NLRP3 and PCM1-dependent centriolar satellite proteostasis, and extended cytoskeletal roles to neuronal dendrite and spine formation.\",\n      \"evidence\": \"Co-IP, substrate ubiquitination and phosphorylation assays, SPATA2/PCM1 knockout/knockdown, and neuronal domain-mutant imaging\",\n      \"pmids\": [\"31762063\", \"31067453\", \"31001844\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Coordination of centrosomal vs receptor-complex CYLD pools unclear\", \"PCM1/Cep70 regulation rest on single-lab evidence\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Linked CYLD regulation to disease through NLRP6 inflammasome control, IKK-reversible RIPK1 editing in leukemic cell death, and a gain-of-function M719V variant tied to neurological disease and autophagy.\",\n      \"evidence\": \"Knockout mouse infection models with IL-18 readout, IKK inhibitor/DISC studies, and in vitro deubiquitinase plus NF-\\u03baB/autophagy assays of the variant\",\n      \"pmids\": [\"32424362\", \"32024820\", \"32185393\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How elevated catalytic activity of M719V perturbs autophagosome-lysosome fusion mechanistically unclear\", \"In vivo consequences of IKK-reversal in normal tissue not addressed\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Refined the activation logic\\u2014defining CAP-Gly3 as essential for catalysis, a Ser418/Ser568 phosphorylation switch activating CYLD toward K63 chains, and a CYLD-SPATA2-LUBAC ubiquitin-editing complex\\u2014and added ERK1/2, p18, plakoglobin, and NoxO1 as substrates with K63 or K48 specificity.\",\n      \"evidence\": \"Biochemical CAP-Gly/phospho-mutant deubiquitinase assays, in vitro ubiquitination/site-mapping with TRIM15, half-life and ubiquitination assays, and KO mouse cardiac phenotyping\",\n      \"pmids\": [\"34610306\", \"34497368\", \"33654169\", \"34742871\", \"36577382\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural model of phospho-activated CYLD not provided\", \"Switching between K63 and K48 chain editing on different substrates mechanistically unresolved\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Tied the SPATA2-CYLD axis to cell-death and metabolic pathologies, regulating NCOA4-dependent ferritinophagy/ferroptosis and SHARPIN-dependent Complex II death signaling.\",\n      \"evidence\": \"Co-IP, NCOA4 ubiquitination assays, SPATA2 knockdown in cardiomyocyte/mouse models; genetic double-knockout and Complex II immunoprecipitation\",\n      \"pmids\": [\"36195186\", \"34887354\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"NCOA4 ferritinophagy axis rests on single-lab evidence\", \"Tissue-specific determinants of pro-death versus pro-survival CYLD output not defined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How CYLD's catalytic deubiquitinase functions and its catalysis-independent CAP-Gly/microtubule and transcriptional roles are integrated, and which of its many substrates dominate in any given tissue, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified structural model linking phospho-regulation, SPATA2 binding, and chain-type selectivity\", \"Substrate hierarchy across tissues unranked\", \"Mechanistic basis of catalysis-independent functions incompletely defined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 4, 8, 18, 26, 29, 31, 35]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 14, 21, 32]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [9, 10, 19]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [10, 20]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [9, 10, 19]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1, 16]},\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [17, 27, 28]},\n      {\"term_id\": \"GO:0005929\", \"supporting_discovery_ids\": [17, 28]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [0, 4, 8, 31]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [3, 7, 26, 29]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [16, 34]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [5, 10, 33]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [17, 27, 28]}\n    ],\n    \"complexes\": [\n      \"CYLD-SPATA2-LUBAC ubiquitin-editing complex\",\n      \"Itch-CYLD complex\",\n      \"TNF receptor signaling complex (Complex I)\"\n    ],\n    \"partners\": [\n      \"SPATA2\",\n      \"TRAF2\",\n      \"TRAF6\",\n      \"TAK1\",\n      \"EB1\",\n      \"HDAC6\",\n      \"ITCH\",\n      \"Plk1\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}