{"gene":"NEK7","run_date":"2026-06-10T05:19:52","timeline":{"discoveries":[{"year":2016,"finding":"NEK7 is an essential mediator of NLRP3 inflammasome activation downstream of potassium efflux. NEK7 binds directly to the catalytic domain of NLRP3, and this interaction is promoted by K+ efflux. NEK7 loss abolishes caspase-1 activation and IL-1β release in response to NLRP3 stimuli but not AIM2 or NLRC4 stimuli. Notably, the catalytic activity of NEK7 is dispensable for NLRP3 inflammasome activation; its scaffolding role is sufficient. NEK7 is required for high-molecular-mass NLRP3 complex formation and ASC oligomerization/speck formation.","method":"Co-immunoprecipitation, genetic knockout (Nek7-/- macrophages and mouse chimeras), caspase-1 activation assay, IL-1β release assay, ASC speck imaging, kinase-dead mutant rescue","journal":"Nature","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, genetic KO in cells and in vivo chimeras, multiple orthogonal functional readouts, independently replicated by at least two contemporaneous studies","pmids":["26814970"],"is_preprint":false},{"year":2015,"finding":"NEK7 binds the leucine-rich repeat (LRR) domain of NLRP3 in a kinase-independent manner downstream of mitochondrial ROS induction, and this interaction is necessary for NLRP3-ASC complex formation, ASC oligomerization, and caspase-1 activation. NLRP3 inflammasome activation is restricted to interphase because NEK7 is sequestered during mitosis, making mitosis and inflammasome activation mutually exclusive.","method":"Co-immunoprecipitation, domain mapping, siRNA knockdown, cell-cycle synchronization, in vivo peritonitis and EAE models","journal":"Nature immunology","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP with domain mapping, genetic KD with functional readouts, multiple orthogonal methods, independent replication by He et al. 2016","pmids":["26642356"],"is_preprint":false},{"year":2015,"finding":"Genome-wide CRISPR/Cas9 screen in macrophages identified NEK7 as an essential and specific component of NLRP3 inflammasome activation; Nek7-deficient macrophages show blunted NLRP3 response while AIM2-mediated inflammasome activation is intact.","method":"Genome-wide CRISPR/Cas9 screen, FACS-based pyroptosis assay, targeted validation in Nek7-KO macrophages","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 2 / Strong — unbiased genome-wide genetic screen plus targeted validation, independently replicated by two contemporaneous studies","pmids":["26553871"],"is_preprint":false},{"year":2019,"finding":"Cryo-EM structure of inactive human NLRP3 in complex with NEK7 at 3.8 Å resolution shows the C-terminal lobe of NEK7 nestles against both the LRR and NACHT domains of NLRP3. Mutagenesis of the NLRP3-NEK7 interface abolishes activation in vitro and in cells. Modeling predicts NEK7 bridges adjacent NLRP3 subunits in the active oligomer, and mutations to this secondary interface also abolish activation, suggesting NEK7 mediates NLRP3 oligomerization via bipartite interactions.","method":"Cryo-electron microscopy (3.8 Å), mutagenesis in vitro and in NEK7-KO / NLRP3-KO cell rescue assays, structural modeling based on NLRC4 inflammasome","journal":"Nature","confidence":"High","confidence_rationale":"Tier 1 / Strong — cryo-EM structure with mutagenesis validation in cells, multiple orthogonal approaches in one rigorous study","pmids":["31189953"],"is_preprint":false},{"year":2003,"finding":"Nercc1/NEK9 directly phosphorylates NEK7 (and NEK6) on the activation loop in vitro, leading to activation of NEK7 kinase activity (~20–25-fold for NEK6; similar for NEK7). NEK7 binds the C-terminal non-catalytic tail of NEK9. This establishes a mitotic kinase cascade: NEK9 → NEK7.","method":"In vitro kinase assay, co-expression with activated NEK9 mutant, recombinant protein phosphorylation, mass spectrometry of phosphorylation site","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Strong — direct in vitro phosphorylation reconstitution, activation loop site identified, replicated in multiple assay formats","pmids":["12840024"],"is_preprint":false},{"year":2009,"finding":"Crystal structure of NEK7 reveals an autoinhibited conformation in which Tyr97 protrudes into the active site, interacts with the activation loop, and blocks the αC helix. Mutation of Tyr97 to phenylalanine renders NEK7 constitutively active. Binding of the NEK9 non-catalytic C-terminal domain increases NEK6/NEK7 activity but not the Tyr97Phe mutant activity, indicating NEK9 activates NEK7 by releasing the autoinhibitory tyrosine.","method":"X-ray crystallography, site-directed mutagenesis, in vitro kinase assay","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure plus mutagenesis with functional kinase assay validation","pmids":["19941817"],"is_preprint":false},{"year":2015,"finding":"Self-association of the NEK9 C-terminal domain (CTD) is required for NEK7 activation. Crystal structure of NEK7(Y97F) bound to NEK9(810-828) reveals a binding site on the C-lobe of NEK7. NEK7(Y97F) crystallizes as a back-to-back dimer between N-lobes; this dimer interface is coupled to the conformation of residue 97. NEK9-CTD activates NEK7 by promoting back-to-back dimerization that releases the autoinhibitory Tyr97.","method":"X-ray crystallography, mutagenesis, in vitro kinase assay, binding mapping","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure with mutagenesis and functional validation, mechanistically extends prior structural work","pmids":["26522158"],"is_preprint":false},{"year":2009,"finding":"NEK6 and NEK7 are both activated in mitosis and are required for robust mitotic spindle formation. Depletion or expression of reduced-activity mutants causes metaphase arrest with fragile spindles; hypomorphic conditions cause cytokinesis defects. NEK6 localizes to spindle microtubules in metaphase/anaphase and to the midbody; NEK7 localizes primarily to spindle poles. Despite high similarity, NEK6 and NEK7 are non-redundant for mitotic progression.","method":"siRNA depletion, dominant-negative/hypomorphic mutant expression, immunofluorescence localization, cell cycle analysis","journal":"Molecular and cellular biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — systematic KD plus mutant analysis with multiple orthogonal readouts, establishes non-redundancy","pmids":["19414596"],"is_preprint":false},{"year":2006,"finding":"Endogenous NEK7 is enriched at the centrosome throughout the cell cycle in a microtubule-independent manner, and transiently localizes to the midbody during cytokinesis. siRNA-mediated knockdown of NEK7 causes a significant increase in cells with multipolar spindles, indicating a role in spindle assembly and mitotic progression.","method":"Immunofluorescence microscopy (endogenous protein), siRNA knockdown, microtubule depolymerization experiments","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct localization experiment with functional KD phenotype, single lab, single main method per claim","pmids":["17101132"],"is_preprint":false},{"year":2007,"finding":"NEK7 knockdown by RNAi causes prometaphase arrest with monopolar or disorganized spindles, reduces centrosomal γ-tubulin levels, and impairs microtubule re-growth activity at the centrosome, indicating NEK7 is required for centrosomal microtubule nucleation. NEK7 also directly participates in cytokinesis, as evidenced by its midbody localization.","method":"RNAi knockdown, γ-tubulin immunofluorescence, microtubule re-growth assay, live-cell imaging","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — KD with defined cellular phenotypes and mechanistic readout (γ-tubulin reduction), single lab","pmids":["17586473"],"is_preprint":false},{"year":2010,"finding":"Nek7-deficient mice die in late embryogenesis or early postnatal stages with severe growth retardation. MEFs from Nek7-/- embryos show chromosomal lagging, micronuclei, cytokinesis failure, tetraploidy, aneuploidy, increased multicentrosomal cells, and altered primary cilia (reduced frequency of primary cilia, appearance of two-cilia cells), confirming NEK7 as an essential regulator of cell division in vivo.","method":"Genetic knockout mouse generation, MEF cytogenetic analysis, FISH, immunofluorescence for centrosomes and cilia","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic KO with multiple orthogonal cellular phenotype readouts","pmids":["20473324"],"is_preprint":false},{"year":2011,"finding":"NEK7 is essential for centriole duplication: NEK7 depletion inhibits centriole duplication, and centrosome-targeted NEK7 overexpression drives extra centriole formation in a kinase-activity-dependent manner. NEK7 depletion prevents pericentriolar material (PCM) protein accumulation at centrosomes in a cell-cycle-stage-specific manner (particularly at G1/G2), indicating a role beyond mitosis in centrosome maturation.","method":"siRNA depletion, centrosome-directed overexpression constructs, kinase-dead mutant, immunofluorescence for centriole markers and PCM proteins","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD and OE with kinase-dead controls, two orthogonal readouts, single lab","pmids":["22100915"],"is_preprint":false},{"year":2017,"finding":"NEK7 is recruited to telomeres after oxidative DNA damage in an ATM-activation-dependent manner. NEK7 phosphorylates TRF1 at Ser114, preventing TRF1 binding to Fbx4 (an SCF E3 ligase subunit) and thereby blocking TRF1 proteasomal degradation, stabilizing the TRF1-Tin2 shelterin complex at telomeres. NEK7 deficiency leads to telomere aberrations and sustained DNA damage foci after oxidative damage.","method":"Co-immunoprecipitation, in vitro kinase assay with mutagenesis (Ser114), mass spectrometry, ChIP, immunofluorescence, NEK7 KO cells","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro phosphorylation with site-specific mutagenesis, Co-IP, ChIP, and KO phenotype, multiple orthogonal methods in one study","pmids":["28216227"],"is_preprint":false},{"year":2019,"finding":"NEK7 regulates dendrite morphogenesis and spine formation in neurons. NEK7 kinase activity is required for dendrite growth and branching. Mechanistically, NEK7 phosphorylates the kinesin Eg5/KIF11, promoting its accumulation on microtubules in distal dendrites where Eg5 limits retrograde microtubule polymerization (inhibitory to dendrite growth), acting through microtubule stabilization independent of Eg5 motor activity.","method":"In vitro and in vivo RNAi, kinase-dead rescue, phosphomimetic/phosphodead Eg5 mutants, live-cell microtubule imaging, in vivo cortical neuron analysis","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — kinase activity requirement established by mutagenesis, substrate phosphorylation (Eg5) validated by phosphomimetic rescue, multiple orthogonal methods in vitro and in vivo","pmids":["29899413"],"is_preprint":false},{"year":2019,"finding":"NEK6 and NEK7 phosphorylate the N-terminal domain of the microtubule-associated protein EML4 at Ser144 and Ser146 in vitro during mitosis. This phosphorylation reduces EML4 affinity for microtubule acidic C-terminal tails. Depletion of NEK6/NEK7 increases EML4 binding to mitotic spindle microtubules, stabilizes them, and impairs chromosome congression. An S144A-S146A double mutant phenocopies kinase depletion.","method":"In vitro kinase assay, cryo-EM with 3D reconstruction (microtubule sedimentation), siRNA depletion, phosphomimetic/phosphodead mutants, chromosome congression imaging","journal":"Science signaling","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay with site-specific mutagenesis, structural microtubule binding data, and cell-based rescue experiments","pmids":["31409757"],"is_preprint":false},{"year":2019,"finding":"GSTO1-1 (glutathione transferase omega 1-1) deglutathionylates NEK7 at cysteine 253, promoting NEK7 activity and NLRP3 inflammasome activation. Inhibition or knockout of GSTO1-1 blocks NLRP3 activation, identifying glutathionylation/deglutathionylation of NEK7 Cys253 as a regulatory post-translational modification.","method":"Small molecule GSTO1-1 inhibitor (C1-27), siRNA knockdown, GSTO1-1-/- mice, site-directed mutagenesis (Cys253), biochemical deglutathionylation assay","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 1–2 / Moderate — PTM identified with site-specific mutagenesis and KO mouse, single lab","pmids":["31577945"],"is_preprint":false},{"year":2019,"finding":"PLK4, the master regulator of centrosome duplication, phosphorylates NEK7 at Ser204. This phosphorylation attenuates NEK7-NLRP3 interaction and suppresses NLRP3 inflammasome activation. The deubiquitinase CYLD is recruited to the centrosome by Spata2 to deubiquitinate PLK4, facilitating PLK4-mediated NEK7 phosphorylation. Mutation of NEK7 Ser204 augments NEK7-NLRP3 interaction and NLRP3 activation.","method":"Co-immunoprecipitation, in vitro kinase assay, phospho-site mutagenesis (Ser204), shRNA knockdown, PLK4 pharmacological inhibition, Spata2-deficient macrophages and peritonitis model","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro kinase assay with site-specific mutagenesis, Co-IP, genetic KD, in vivo model, multiple orthogonal methods","pmids":["31762063"],"is_preprint":false},{"year":2023,"finding":"ZDHHC5 palmitoylates NLRP3 at the LRR domain; this S-palmitoylation promotes NLRP3-NEK7 interaction, NLRP3 oligomerization, and inflammasome assembly. The depalmitoylase ABHD17A reverses this modification. ZDHHC5 silencing blocks NLRP3-NEK7 interaction and ASC speck formation, and Zdhhc5-/- mice show defective NLRP3 activation in vivo.","method":"Acyl-RAC palmitoylation assay, Co-immunoprecipitation, ZDHHC5 siRNA/KO, ABHD17A functional assay, mutagenesis of palmitoylation sites, Zdhhc5-/- mice","journal":"Molecular cell","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — PTM biochemically defined with multiple orthogonal methods (acyl-RAC, Co-IP, mutagenesis), in vivo KO mouse, single rigorous study","pmids":["38092000"],"is_preprint":false},{"year":2023,"finding":"MARCH5, a mitochondria-associated E3 ligase, interacts with the NACHT domain of NLRP3 and promotes K27-linked polyubiquitination of NLRP3 at K324 and K430. This ubiquitination is required for NLRP3 to bind NEK7 and form NLRP3 oligomers; ubiquitination-defective NLRP3 mutants (K324R, K430R) cannot bind NEK7 or form ASC specks. March5 conditional KO mice fail to produce IL-1β/IL-18 upon infection.","method":"Co-immunoprecipitation, ubiquitination assay with linkage-specific analysis, site-directed mutagenesis (K324R, K430R), myeloid-specific conditional KO mice, macrophage infection models","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — ubiquitination biochemically characterized with site-specific mutagenesis and linkage analysis, genetic KO in vivo, multiple orthogonal methods","pmids":["37575012"],"is_preprint":false},{"year":2025,"finding":"NEK7 is rapidly phosphorylated at threonine-190/191 by JNK1 downstream of K+ efflux and gasdermin D (GSDMD) pore formation after NLRP3 activation. This phosphorylation enhances NEK7-NLRP3 binding and further promotes inflammasome assembly. Knock-in mice with Thr190/191Val substitutions show impaired NEK7 phosphorylation, reduced NLRP3 inflammasome activation, and decreased IL-1β secretion.","method":"Phospho-proteomics, in vitro JNK1 kinase assay, knock-in mutant mice (T190V/T191V), Co-immunoprecipitation, NLRP3 activation assays","journal":"Science immunology","confidence":"High","confidence_rationale":"Tier 1–2 / Strong — in vitro kinase assay identifying writer (JNK1), phospho-site knock-in mice, Co-IP, multiple orthogonal methods in one rigorous study","pmids":["39752537"],"is_preprint":false},{"year":2024,"finding":"A20 (TNFAIP3) directly binds NEK7 and mediates K48-linked ubiquitination of NEK7 at K189 and K293, targeting NEK7 for proteasomal degradation. A20 deficiency increases NEK7 protein levels. A20 also disrupts NEK7-NLRP3 association, potentially via its OTU domain and/or ZnF4/ZnF7 motifs. NEK7 deletion attenuates NLRP3 inflammasome activation in A20-deficient conditions both in vitro and in vivo.","method":"Co-immunoprecipitation, ubiquitination assay (K48-linkage specific), site-directed mutagenesis (K189, K293), A20-deficient macrophages and in vivo models","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP, site-specific ubiquitination mutagenesis, in vitro and in vivo KO, single lab","pmids":["38865260"],"is_preprint":false},{"year":2024,"finding":"METTL3-dependent m6A modification of NEK7 mRNA increases its stability (recognized by IGF2BP2), promoting NEK7 expression and NLRP3 inflammasome activation. Silencing METTL3 reduces A20/TNFAIP3 m6A modification, stabilizes TNFAIP3 mRNA (increasing A20 protein), which then ubiquitinates NEK7 to impair NLRP3 assembly. In a separate study, silencing METTL3 inhibits m6A methylation of NEK7, reducing NEK7 mRNA stability and blocking chondrocyte pyroptosis.","method":"m6A-RIP, RNA immunoprecipitation, dual-luciferase reporter, METTL3 KO/KD, TNFAIP3 ubiquitination assay, in vivo periodontitis model","journal":"Advanced science (Weinheim, Baden-Wurttemberg, Germany)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — m6A-RIP, RIP, luciferase reporter, and KO experiments establish PTM-mRNA stability link, single lab","pmids":["38696610"],"is_preprint":false},{"year":2024,"finding":"OGT-mediated O-GlcNAcylation of NEK7 at S260 suppresses NEK7 phosphorylation at S260, which promotes NEK7-NLRP3 interaction and chondrocyte pyroptosis. OGT silencing enhances NEK7 phosphorylation and blocks the NEK7-NLRP3 complex, reducing NLRP3 activation.","method":"Co-immunoprecipitation, western blot for O-GlcNAc and phosphorylation, OGT knockout/knockdown, site-specific mutagenesis (S260), in vivo OA model","journal":"Autoimmunity","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with site-specific analysis, OGT KO in vivo, single lab, moderate mechanistic depth","pmids":["38389178"],"is_preprint":false},{"year":2024,"finding":"SIRT5 desuccinylates NEK7 at K81, downregulating NEK7 expression/activity. Electroacupuncture in a MCAO mouse model of ischemia-reperfusion injury activates SIRT5, leading to decreased NEK7 succinylation at K81 and reduced neuronal pyroptosis. NEK7 overexpression reverses these protective effects.","method":"MCAO mouse model, succinylation assay, SIRT5 KD, NEK7 overexpression rescue, western blot, brain infarct analysis","journal":"Brain research bulletin","confidence":"Low","confidence_rationale":"Tier 3 / Weak — in vivo KD/OE with defined PTM site, but indirect evidence for SIRT5 directly desuccinylating NEK7 K81 (no in vitro biochemical reconstitution shown in abstract), single lab","pmids":["39694147"],"is_preprint":false},{"year":2023,"finding":"TRIM32 (E3 ubiquitin ligase) promotes ubiquitylation of NEK7 at K64, leading to NEK7 downregulation and suppression of NLRP3-dependent microglial pyroptosis. Inhibiting NEK7 ubiquitylation reverses TRIM32-mediated pyroptosis suppression.","method":"Co-immunoprecipitation, ubiquitylation assay, site-directed mutagenesis (K64), siRNA/overexpression, in vivo SCI mouse model","journal":"Molecular biotechnology","confidence":"Low","confidence_rationale":"Tier 3 / Weak — Co-IP and K64 mutagenesis establish ubiquitylation site, but in vitro reconstitution not demonstrated in abstract, single lab","pmids":["38030945"],"is_preprint":false},{"year":2020,"finding":"EML4-ALK V3 oncogenic fusion protein recruits NEK9 and NEK7 to microtubules via the EML4 N-terminal microtubule-binding region. This complex promotes microtubule stabilization, extended cytoplasmic protrusions, and increased cell migration. Constitutively active NEK9 accelerates migration in a microtubule-dependent manner requiring downstream NEK7 but not ALK activity.","method":"Co-immunoprecipitation, siRNA depletion, dominant-active NEK9 expression, cell morphology and migration assays, microtubule imaging","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP with NEK9/NEK7, functional epistasis (NEK9→NEK7) established by depletion rescue, two orthogonal methods, single lab","pmids":["32184261"],"is_preprint":false},{"year":2024,"finding":"In the context of EML4-ALK V3, NEK7 phosphorylates the kinesin Eg5 at Ser1033 on interphase microtubules. Eg5 phosphorylation at S1033 by NEK7 drives the mesenchymal morphology of EML4-ALK V3 cells. NEK7 depletion reduces Eg5 recruitment to microtubules, and a phosphomimetic Eg5-S1033D mutant rescues mesenchymal morphology in NEK7-depleted cells.","method":"siRNA depletion of NEK7, phosphomimetic/phosphodead Eg5 mutants, immunofluorescence for Eg5 microtubule association, cell morphology analysis","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — substrate phosphorylation site (S1033) validated by phosphomimetic rescue, mechanistic epistasis established, single lab","pmids":["38458397"],"is_preprint":false},{"year":2017,"finding":"NEK7 depletion inhibits G1 progression in human U2OS cells by downregulating cyclins and CDKs. NEK7 loss also inhibits the earliest stages of procentriole formation (STIL degradation by APC/CCdh1) and induces primary cilia formation in RPE1 cells. Abnormal APC/CCdh1 accumulation at centrioles and continuous STIL degradation are the mechanistic basis for centriole assembly failure.","method":"siRNA depletion, cell cycle analysis, immunofluorescence for centriole markers (STIL, Cdh1), quantitative microscopy","journal":"Molecular biology of the cell","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with multiple mechanistic readouts (cyclin levels, Cdh1 localization, STIL degradation), single lab","pmids":["28539406"],"is_preprint":false},{"year":2013,"finding":"NEK7 regulates microtubule dynamic instability: siRNA-mediated NEK7 knockdown reduces speeds of MT growth and catastrophe, reduces relative time in catastrophe, and lowers overall MT dynamicity. Ectopic overexpression has inverse effects. These phenotypes are recapitulated in Nek7-/- MEFs, establishing NEK7 as a direct regulator of MT dynamics.","method":"siRNA knockdown, NEK7-/- MEFs, PlusTipTracker live-cell MT imaging, rescue by re-expression","journal":"Biochimica et biophysica acta","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD and KO with quantitative live-imaging MT dynamics, genetic KO recapitulates KD phenotype, single lab","pmids":["23313050"],"is_preprint":false},{"year":2015,"finding":"Anks3 interacts with NEK7 (but not NEK6) and retains NEK7 in the cytoplasm, preventing NEK7 nuclear localization. Anks3 undergoes an ~20 kDa molecular weight increase upon NEK7 co-expression that is not caused by NEK7-dependent phosphorylation (occurs with kinase-dead NEK7 mutant), indicating an alternative modification mechanism.","method":"Co-immunoprecipitation, subcellular fractionation, immunofluorescence, kinase-dead mutant, mass spectrometry","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — Co-IP with kinase-dead control establishes binding but modification mechanism not fully resolved; single lab","pmids":["26188091"],"is_preprint":false},{"year":2019,"finding":"Licochalcone B (LicoB) directly binds NEK7 and inhibits the NEK7-NLRP3 interaction, specifically suppressing NLRP3 inflammasome activation without affecting AIM2 or NLRC4 inflammasomes. This is the first direct small-molecule targeting of the NEK7-NLRP3 interface as a therapeutic approach.","method":"Direct binding assay (drug-protein interaction), Co-immunoprecipitation, macrophage inflammasome activation assays, in vivo sepsis/peritonitis/NASH models","journal":"EMBO reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct NEK7 binding shown, mechanistic Co-IP confirms disruption of NEK7-NLRP3 interface, in vivo validation, single lab","pmids":["34882936"],"is_preprint":false},{"year":2020,"finding":"Berberine directly targets NEK7 via a hydrogen bond between its 2,3-methylenedioxy group and Arg121 of NEK7. Because R121 is located within the key domain mediating NEK7-NLRP3 interaction, berberine specifically blocks the NEK7-NLRP3 interaction and inhibits IL-1β release independently of NF-κB and TLR4 signaling. Anti-inflammatory efficacy in vivo is NEK7-dependent.","method":"Activity-based protein profiling (direct target identification), site-directed mutagenesis (R121), Co-immunoprecipitation, NEK7 KD in vivo rescue","journal":"Journal of medicinal chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct target identification by ABPP plus R121 mutagenesis validates binding site, in vivo NEK7-dependence confirmed, single lab","pmids":["33440945"],"is_preprint":false},{"year":2022,"finding":"A single amino acid difference—Arg121 in NEK7 versus Gln132 in NEK6—accounts for NEK7's ability to bind NLRP3 and support inflammasome activation, while NEK6 cannot. Substituting Gln132 with Arg in NEK6 confers NLRP3 binding and inflammasome activation activity in macrophages.","method":"Site-directed mutagenesis (R121/Q132), Co-immunoprecipitation, caspase-1 activation assay in macrophages, NEK7-KO rescue","journal":"Journal of immunology (Baltimore, Md. : 1950)","confidence":"High","confidence_rationale":"Tier 2 / Strong — single residue swap in both directions (gain and loss of function) with biochemical and cell-based validation","pmids":["35354613"],"is_preprint":false},{"year":2019,"finding":"PAF (platelet-activating factor) activates the NLRP3 inflammasome in a NEK7-dependent (and NLRP3, ASC, caspase-1-dependent) manner, requiring potassium efflux, independently of PAFR signaling. NEK7 is thus required for a lipid-triggered, PAFR-independent pathway of NLRP3 activation.","method":"Genetic KO macrophages (NEK7, NLRP3, ASC, caspase-1), K+ efflux measurement, in vivo peritonitis model","journal":"The Journal of experimental medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO panel establishes NEK7 requirement in this pathway, in vivo validation, single lab","pmids":["31558613"],"is_preprint":false},{"year":2019,"finding":"Co-chaperone UNC45A is required for expression of NEK7 in cancer cells. UNC45A localizes to the cancer cell nucleus, upregulates glucocorticoid receptor transcriptional activity, and thereby promotes NEK7 transcription. UNC45A-deficient cancer cells show pericentrosomal material disorganization, centrosomal separation defects, and metaphase/cytokinesis stalling; these phenotypes are rescued by heterologous NEK7 expression.","method":"siRNA knockdown, immunofluorescence, gene microarray, RT-qPCR, glucocorticoid receptor activity assay, NEK7 rescue expression","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis by rescue, transcriptional mechanism defined, multiple cell-based readouts, single lab","pmids":["30737284"],"is_preprint":false},{"year":2025,"finding":"NEK7 directly binds EGFR and phosphorylates it at serine 1070, activating MAPK and PI3K/AKT signaling pathways to drive acquired lenvatinib resistance in hepatocellular carcinoma. An inhibitory peptide targeting the NEK7-binding domain (EGFR aa 979–1099) blocks EGFR S1070 phosphorylation and suppresses resistance.","method":"Kinase CRISPR-Cas9 genetic screen, Co-IP (NEK7-EGFR binding), site-specific phosphorylation analysis (S1070), inhibitory peptide design, organoid and xenograft models","journal":"Hepatology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR screen plus Co-IP and site-specific phosphorylation evidence, organoid/in vivo validation, single lab","pmids":["40694824"],"is_preprint":false},{"year":2025,"finding":"Isocyanic acid (produced by LACC1 enzyme) carbamoylates NLRP3 at lysine-593, disrupting NLRP3-NEK7 interaction and limiting NLRP3 inflammasome activation. LACC1 KO or K593 carbamoylation-deficient NLRP3 mutant promotes inflammatory responses in vitro and in vivo.","method":"Carbamoylation mass spectrometry, site-directed mutagenesis (K593), Co-immunoprecipitation, LACC1 KO/Lacc1-/- mice, macrophage inflammasome assays","journal":"Science advances","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — PTM site defined by MS and mutagenesis, disruption of NEK7-NLRP3 interface shown by Co-IP, KO in vivo, single lab","pmids":["40053593"],"is_preprint":false},{"year":2018,"finding":"NEK7 is required for proper wiring of cortical parvalbumin (PV+) interneurons. NEK7-deficient PV+ interneurons show altered microtubule dynamics, impaired axon growth cone steering, reduced axon length, lower arbor complexity, and fewer synaptic contacts with pyramidal cells in vivo.","method":"Conditional/targeted NEK7 ablation in PV interneurons, in vitro and in vivo morphology analysis, microtubule dynamics live imaging, synapse quantification","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-type-specific KO with multiple in vitro and in vivo readouts, single lab","pmids":["30067978"],"is_preprint":false},{"year":2021,"finding":"NLRP3 inflammasome activation by NEK7 is restricted to interphase; the NEK7-NLRP3 interaction is required for NLRP3 activation, and the interaction is enhanced by K+ efflux. The Influenza A virus accessory protein PB1-F2 limits the conformational transition of NLRP3 from its auto-repressed closed conformation to its active state, thereby diminishing NEK7-NLRP3 interaction and blocking inflammasome assembly in human macrophages.","method":"IAV mutant infection (PB1-F2 deficient), Co-immunoprecipitation (NEK7-NLRP3), caspase-1 activation assay, gasdermin D cleavage, LDH/IL-1β release","journal":"EMBO reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — viral protein loss-of-function with Co-IP readout of NEK7-NLRP3 interaction, mechanistic specificity established, single lab","pmids":["33180976"],"is_preprint":false},{"year":2022,"finding":"In HEK293 cells reconstituted with NLRP3 and ASC (but not NEK7), ASC speck polymerization occurs independently of NEK7, suggesting a NEK7-independent mode of NLRP3 activation exists in this cell system. Evidence supports a stacked-torus hexameric NLRP3 oligomer in this context.","method":"HEK293 cell reconstitution of NLRP3 inflammasome without NEK7, ASC speck assay, interface mutagenesis based on structural models","journal":"International journal of molecular sciences","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single heterologous reconstitution system, negative result (NEK7-independent activation) may reflect cell-system artifact, single lab","pmids":["36142182"],"is_preprint":false},{"year":2025,"finding":"A cereblon (CRBN) glue degrader of NEK7 (NK7-902) potently and selectively degrades NEK7 in human immune cells. Full NEK7 degradation completely blocked NLRP3-dependent IL-1β release in vitro in some but not all donors/conditions, and in mice. In cynomolgus monkeys, NK7-902 caused long-lasting NEK7 degradation but only transiently blocked IL-1β, suggesting NEK7 contributes to but may not be absolutely required for NLRP3 activation in primates.","method":"Targeted protein degradation (CRBN molecular glue), NEK7 protein level measurement in primary human cells and whole blood, NLRP3 activation assays, in vivo mouse and monkey models","journal":"Cell chemical biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — selective degrader with cross-species comparison, multiple biological systems tested, single lab; highlights context-dependence of NEK7 requirement","pmids":["40639372"],"is_preprint":false}],"current_model":"NEK7 is a dual-function NIMA-related serine/threonine kinase that (1) drives mitotic progression by localizing to centrosomes, where—activated via NEK9-induced dimerization and release of autoinhibitory Tyr97—it phosphorylates substrates such as EML4 and Eg5 to regulate microtubule dynamics, spindle assembly, centriole duplication, and cytokinesis; and (2) acts as an essential scaffolding co-activator of the NLRP3 inflammasome in interphase cells, binding the LRR and NACHT domains of NLRP3 (via its C-terminal lobe, with critical contact at Arg121) downstream of K+ efflux to bridge adjacent NLRP3 subunits and license oligomerization, ASC speck formation, and caspase-1 activation, a process regulated by multiple PTMs on NEK7 (JNK1-mediated T190/T191 phosphorylation amplifying NLRP3 binding; PLK4-mediated S204 phosphorylation suppressing it; GSTO1-1-mediated Cys253 deglutathionylation activating it; A20/TRIM32/TNFAIP3-mediated ubiquitination degrading it) and on NLRP3 itself (MARCH5-mediated K27 ubiquitination required for NEK7 recruitment; ZDHHC5-mediated palmitoylation promoting the interaction; isocyanic acid carbamoylation at K593 and NLRP3 LRR mutations altering the interface); in post-mitotic neurons NEK7 additionally regulates dendrite morphogenesis through Eg5 phosphorylation, and protects telomere integrity by phosphorylating TRF1-Ser114 to prevent its proteasomal degradation."},"narrative":{"mechanistic_narrative":"NEK7 is a NIMA-related serine/threonine kinase with two largely separable cellular roles: an enzymatic role in mitotic cell division and a non-catalytic scaffolding role in innate immune signaling [PMID:19414596, PMID:26814970]. As a kinase, NEK7 is activated late in a mitotic cascade in which NEK9 binds its non-catalytic C-terminal tail and phosphorylates the NEK7 activation loop [PMID:12840024]; structurally, this activation works by promoting a back-to-back NEK7 dimer that displaces an autoinhibitory Tyr97 from the active site [PMID:19941817, PMID:26522158]. Active NEK7 localizes to centrosomes and spindle poles and is required for centriole duplication, centrosomal microtubule nucleation, spindle assembly, and cytokinesis, with loss causing multipolar/monopolar spindles, aneuploidy, and embryonic lethality in mice [PMID:17101132, PMID:17586473, PMID:20473324, PMID:22100915]. It controls microtubule dynamics directly and through substrate phosphorylation, including EML4 at Ser144/Ser146 to reduce microtubule affinity during chromosome congression and the kinesin Eg5/KIF11 to govern dendrite morphogenesis and cortical interneuron wiring [PMID:31409757, PMID:29899413, PMID:30067978, PMID:23313050]. NEK7 also phosphorylates TRF1-Ser114 to block its proteasomal degradation and protect telomere integrity after oxidative damage [PMID:28216227]. Independently of its kinase activity, NEK7 is an essential co-activator of the NLRP3 inflammasome, acting downstream of K+ efflux: it binds the LRR and NACHT domains of NLRP3 through its C-terminal lobe and bridges adjacent NLRP3 subunits to license oligomerization, ASC speck formation, and caspase-1 activation [PMID:26814970, PMID:26642356, PMID:31189953]. A single residue, Arg121, distinguishes NEK7 from NEK6 and is required for NLRP3 binding [PMID:35354613], and small molecules engaging this interface (e.g., berberine at Arg121) selectively block NLRP3 activation [PMID:33440945, PMID:34882936]. This interaction is gated by post-translational modifications on both partners—JNK1 phosphorylation of NEK7 Thr190/191 and GSTO1-1 deglutathionylation of Cys253 promote it, PLK4 phosphorylation of Ser204 suppresses it, and MARCH5-mediated K27 ubiquitination and ZDHHC5 palmitoylation of NLRP3 are required for NEK7 recruitment [PMID:39752537, PMID:31577945, PMID:31762063, PMID:37575012, PMID:38092000].","teleology":[{"year":2003,"claim":"Established that NEK7 sits downstream in a kinase cascade, answering how this kinase becomes activated.","evidence":"In vitro kinase reconstitution with activated NEK9 and mass spectrometry of the activation-loop site","pmids":["12840024"],"confidence":"High","gaps":["Did not define the structural basis of activation","Physiological triggers of NEK9 activity not addressed"]},{"year":2009,"claim":"Defined the autoinhibition mechanism, showing how NEK7 is held inactive and how NEK9 relieves it.","evidence":"X-ray crystallography of NEK7 with Tyr97Phe mutagenesis and in vitro kinase assays","pmids":["19941817"],"confidence":"High","gaps":["Did not capture the active conformation","Mechanism coupling NEK9 binding to Tyr97 release unresolved at this stage"]},{"year":2009,"claim":"Placed NEK7 in mitotic spindle formation and distinguished it functionally from the related NEK6.","evidence":"siRNA depletion, hypomorphic mutants, and immunofluorescence localization across mitosis","pmids":["19414596"],"confidence":"High","gaps":["Spindle-pole substrates not identified","Basis of NEK6/NEK7 non-redundancy unexplained"]},{"year":2011,"claim":"Extended NEK7's mitotic role to centriole duplication and centrosome maturation, requiring kinase activity.","evidence":"siRNA depletion, centrosome-targeted overexpression, kinase-dead mutants, PCM marker imaging","pmids":["22100915","20473324"],"confidence":"High","gaps":["Centrosomal substrates not identified","Link between kinase activity and PCM accumulation mechanistic detail missing"]},{"year":2015,"claim":"Resolved how NEK9 activates NEK7 structurally, linking dimerization to Tyr97 displacement.","evidence":"Crystal structure of NEK7(Y97F)–NEK9 peptide complex with mutagenesis and kinase assays","pmids":["26522158"],"confidence":"High","gaps":["In-cell relevance of the back-to-back dimer not directly visualized","Does not address scaffolding (kinase-independent) functions"]},{"year":2015,"claim":"Identified NEK7 as an essential, specific NLRP3 inflammasome component and revealed its mitosis–inflammasome mutual exclusivity.","evidence":"Genome-wide CRISPR screen, Co-IP with domain mapping, siRNA, cell-cycle synchronization, in vivo models","pmids":["26553871","26642356"],"confidence":"High","gaps":["Whether kinase activity is required not fully settled at this point","Structural basis of the NEK7–NLRP3 interface unknown"]},{"year":2016,"claim":"Established that NEK7's scaffolding role, not its catalytic activity, mediates NLRP3 activation downstream of K+ efflux.","evidence":"Reciprocal Co-IP, Nek7-/- macrophages and chimeras, kinase-dead rescue, ASC speck imaging","pmids":["26814970"],"confidence":"High","gaps":["Structural mechanism of subunit bridging not yet resolved","How K+ efflux promotes the interaction unknown"]},{"year":2017,"claim":"Revealed a telomere-protective kinase function, showing NEK7 phosphorylates TRF1 to block its degradation.","evidence":"In vitro kinase assay with Ser114 mutagenesis, Co-IP, ChIP, mass spectrometry, KO cells","pmids":["28216227"],"confidence":"High","gaps":["Signal coupling ATM activation to NEK7 telomere recruitment incompletely defined","Relationship to NEK7's centrosomal pool unclear"]},{"year":2019,"claim":"Provided the structural basis for NEK7-mediated NLRP3 oligomerization via bipartite interactions.","evidence":"3.8 Å cryo-EM of inactive NLRP3–NEK7 with interface mutagenesis and cell rescue","pmids":["31189953"],"confidence":"High","gaps":["Structure of the active oligomer was modeled, not directly resolved","Conformational steps from binding to oligomerization not captured"]},{"year":2019,"claim":"Identified mitotic and neuronal substrates of NEK7 kinase activity that control microtubule behavior.","evidence":"In vitro kinase assays with phosphosite mutants for EML4 (S144/S146) and Eg5, microtubule and morphology imaging in vitro and in vivo","pmids":["31409757","29899413","30067978","23313050"],"confidence":"High","gaps":["Full substrate repertoire not defined","How NEK7 selects spindle vs dendritic targets unknown"]},{"year":2019,"claim":"Began defining the PTM code on NEK7 and NLRP3 that tunes inflammasome assembly, including suppressive PLK4 phosphorylation and activating deglutathionylation.","evidence":"In vitro kinase and deglutathionylation assays, site-specific mutagenesis (S204, Cys253), Co-IP, KO mice and macrophages","pmids":["31762063","31577945"],"confidence":"High","gaps":["Hierarchy and interplay among NEK7 PTMs unresolved","Spatial coordination with centrosomal PLK4 in immune cells unclear"]},{"year":2022,"claim":"Pinpointed Arg121 as the determinant of NLRP3 binding specificity that explains why NEK7, not NEK6, supports the inflammasome.","evidence":"Reciprocal single-residue swap (R121/Q132) with Co-IP and caspase-1 assays in macrophages","pmids":["35354613"],"confidence":"High","gaps":["Does not address whether other contacts contribute under physiological conditions"]},{"year":2023,"claim":"Showed that NLRP3-side modifications—K27 ubiquitination and palmitoylation—are prerequisites for NEK7 recruitment.","evidence":"Linkage-specific ubiquitination and acyl-RAC assays, site mutagenesis (K324/K430; LRR palmitoylation), Co-IP, conditional/KO mice","pmids":["37575012","38092000"],"confidence":"High","gaps":["Temporal ordering relative to K+ efflux not fully resolved","Whether these modifications act on the same NLRP3 pool simultaneously unknown"]},{"year":2024,"claim":"Expanded the regulatory layer with additional NEK7 PTMs and mRNA-level control governing its abundance and NLRP3 binding.","evidence":"Co-IP, K48-ubiquitination and O-GlcNAc assays, site mutagenesis (K189/K293; S260), m6A-RIP/luciferase, KO macrophages and disease models","pmids":["38865260","38389178","38696610"],"confidence":"Medium","gaps":["Many sites from single labs without cross-validation","Quantitative contribution of each PTM in vivo unclear"]},{"year":2024,"claim":"Identified non-canonical NEK7 kinase functions in cancer, including EML4-ALK-associated migration and EGFR-driven drug resistance.","evidence":"CRISPR screens, Co-IP, phosphosite analysis (Eg5 S1033; EGFR S1070), inhibitory peptide, organoid/xenograft models","pmids":["32184261","38458397","40694824"],"confidence":"Medium","gaps":["Single-lab findings","Generality across tumor types and direct kinase-substrate relationships need broader validation"]},{"year":2025,"claim":"Tested NEK7's necessity for NLRP3 activation pharmacologically and revealed context- and species-dependence.","evidence":"JNK1 phosphosite knock-in mice (T190V/T191V), carbamoylation MS, and a CRBN glue degrader across human cells, mice, and monkeys","pmids":["39752537","40053593","40639372"],"confidence":"Medium","gaps":["Why NEK7 requirement varies across primates/conditions unresolved","Existence of NEK7-independent NLRP3 activation modes not fully defined"]},{"year":null,"claim":"How NEK7's two roles—mitotic kinase and inflammasome scaffold—are coordinated spatially and temporally within a single cell, and whether NEK7 is absolutely required for NLRP3 activation across species, remains open.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No unified model linking cell-cycle sequestration of NEK7 to inflammasome timing at the molecular level","Species-dependent dispensability suggests redundancy that is uncharacterized"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[12,13,14,26,35]},{"term_id":"GO:0140657","term_label":"ATP-dependent activity","supporting_discovery_ids":[4,5]},{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[0,3,32]},{"term_id":"GO:0008092","term_label":"cytoskeletal protein binding","supporting_discovery_ids":[14,28]}],"localization":[{"term_id":"GO:0005815","term_label":"microtubule organizing center","supporting_discovery_ids":[8,9,11]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[7,28]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[29]},{"term_id":"GO:0005694","term_label":"chromosome","supporting_discovery_ids":[12]}],"pathway":[{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[0,1,2,3]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[7,9,10,11]},{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[0,19,33]},{"term_id":"R-HSA-1852241","term_label":"Organelle biogenesis and maintenance","supporting_discovery_ids":[11,27]}],"complexes":["NLRP3 inflammasome"],"partners":["NLRP3","NEK9","EML4","KIF11","TRF1","EGFR","ANKS3","A20"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q8TDX7","full_name":"Serine/threonine-protein kinase Nek7","aliases":["Never in mitosis A-related kinase 7","NimA-related protein kinase 7"],"length_aa":302,"mass_kda":34.6,"function":"Protein kinase which plays an important role in mitotic cell cycle progression (PubMed:17101132, PubMed:19941817, PubMed:31409757). Required for microtubule nucleation activity of the centrosome, robust mitotic spindle formation and cytokinesis (PubMed:17586473, PubMed:19414596, PubMed:19941817, PubMed:26522158, PubMed:31409757). Phosphorylates EML4 at 'Ser-146', promoting its dissociation from microtubules during mitosis which is required for efficient chromosome congression (PubMed:31409757). Phosphorylates RPS6KB1 (By similarity). Acts as an essential activator of the NLRP3 inflammasome assembly independently of its kinase activity (PubMed:26642356, PubMed:36442502, PubMed:39173637). Acts by unlocking NLRP3 following NLRP3 tranlocation into the microtubule organizing center (MTOC), relieving NLRP3 autoinhibition and promoting formation of the NLRP3:PYCARD complex, and activation of CASP1 (PubMed:26642356, PubMed:31189953, PubMed:36442502, PubMed:39173637, PubMed:40450990). Serves as a cellular switch that enforces mutual exclusivity of the inflammasome response and cell division: interaction with NEK9 prevents interaction with NLRP3 and activation of the inflammasome during mitosis (PubMed:26642356, PubMed:31189953)","subcellular_location":"Nucleus; Cytoplasm; Cytoplasm, cytoskeleton, spindle pole; Cytoplasm, cytoskeleton, microtubule organizing center, centrosome","url":"https://www.uniprot.org/uniprotkb/Q8TDX7/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/NEK7","classification":"Not Classified","n_dependent_lines":9,"n_total_lines":1208,"dependency_fraction":0.0074503311258278145},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"DYNLL1","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/NEK7","total_profiled":1310},"omim":[{"mim_id":"618726","title":"NIMA-RELATED KINASE 10; NEK10","url":"https://www.omim.org/entry/618726"},{"mim_id":"609798","title":"NIMA-RELATED KINASE 9; NEK9","url":"https://www.omim.org/entry/609798"},{"mim_id":"609720","title":"CRUMBS CELL POLARITY COMPLEX COMPONENT 2; CRB2","url":"https://www.omim.org/entry/609720"},{"mim_id":"606848","title":"NIMA-RELATED KINASE 7; NEK7","url":"https://www.omim.org/entry/606848"},{"mim_id":"606416","title":"NLR FAMILY, PYRIN DOMAIN-CONTAINING 3; NLRP3","url":"https://www.omim.org/entry/606416"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Nucleoplasm","reliability":"Supported"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"heart muscle","ntpm":131.4}],"url":"https://www.proteinatlas.org/search/NEK7"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"Q8TDX7","domains":[{"cath_id":"3.30.200.20","chopping":"26-111","consensus_level":"high","plddt":90.7022,"start":26,"end":111},{"cath_id":"1.10.510.10","chopping":"118-185_199-300","consensus_level":"high","plddt":92.8976,"start":118,"end":300}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TDX7","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TDX7-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q8TDX7-F1-predicted_aligned_error_v6.png","plddt_mean":87.31},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=NEK7","jax_strain_url":"https://www.jax.org/strain/search?query=NEK7"},"sequence":{"accession":"Q8TDX7","fasta_url":"https://rest.uniprot.org/uniprotkb/Q8TDX7.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q8TDX7/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q8TDX7"}},"corpus_meta":[{"pmid":"26814970","id":"PMC_26814970","title":"NEK7 is an essential mediator of NLRP3 activation downstream of potassium efflux.","date":"2016","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/26814970","citation_count":1081,"is_preprint":false},{"pmid":"31189953","id":"PMC_31189953","title":"Structural mechanism for NEK7-licensed activation of NLRP3 inflammasome.","date":"2019","source":"Nature","url":"https://pubmed.ncbi.nlm.nih.gov/31189953","citation_count":653,"is_preprint":false},{"pmid":"26642356","id":"PMC_26642356","title":"NLRP3 activation and mitosis are mutually exclusive events coordinated by NEK7, a new inflammasome component.","date":"2015","source":"Nature immunology","url":"https://pubmed.ncbi.nlm.nih.gov/26642356","citation_count":632,"is_preprint":false},{"pmid":"26553871","id":"PMC_26553871","title":"A Genome-wide CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) Screen Identifies NEK7 as an Essential Component of NLRP3 Inflammasome Activation.","date":"2015","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/26553871","citation_count":384,"is_preprint":false},{"pmid":"31787755","id":"PMC_31787755","title":"NEK7 interacts with NLRP3 to modulate the pyroptosis in inflammatory bowel disease via NF-κB signaling.","date":"2019","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/31787755","citation_count":326,"is_preprint":false},{"pmid":"12840024","id":"PMC_12840024","title":"A mitotic cascade of NIMA family kinases. 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NEK7 binds directly to the catalytic domain of NLRP3, and this interaction is promoted by K+ efflux. NEK7 loss abolishes caspase-1 activation and IL-1β release in response to NLRP3 stimuli but not AIM2 or NLRC4 stimuli. Notably, the catalytic activity of NEK7 is dispensable for NLRP3 inflammasome activation; its scaffolding role is sufficient. NEK7 is required for high-molecular-mass NLRP3 complex formation and ASC oligomerization/speck formation.\",\n      \"method\": \"Co-immunoprecipitation, genetic knockout (Nek7-/- macrophages and mouse chimeras), caspase-1 activation assay, IL-1β release assay, ASC speck imaging, kinase-dead mutant rescue\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, genetic KO in cells and in vivo chimeras, multiple orthogonal functional readouts, independently replicated by at least two contemporaneous studies\",\n      \"pmids\": [\"26814970\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"NEK7 binds the leucine-rich repeat (LRR) domain of NLRP3 in a kinase-independent manner downstream of mitochondrial ROS induction, and this interaction is necessary for NLRP3-ASC complex formation, ASC oligomerization, and caspase-1 activation. NLRP3 inflammasome activation is restricted to interphase because NEK7 is sequestered during mitosis, making mitosis and inflammasome activation mutually exclusive.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, siRNA knockdown, cell-cycle synchronization, in vivo peritonitis and EAE models\",\n      \"journal\": \"Nature immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP with domain mapping, genetic KD with functional readouts, multiple orthogonal methods, independent replication by He et al. 2016\",\n      \"pmids\": [\"26642356\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Genome-wide CRISPR/Cas9 screen in macrophages identified NEK7 as an essential and specific component of NLRP3 inflammasome activation; Nek7-deficient macrophages show blunted NLRP3 response while AIM2-mediated inflammasome activation is intact.\",\n      \"method\": \"Genome-wide CRISPR/Cas9 screen, FACS-based pyroptosis assay, targeted validation in Nek7-KO macrophages\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — unbiased genome-wide genetic screen plus targeted validation, independently replicated by two contemporaneous studies\",\n      \"pmids\": [\"26553871\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Cryo-EM structure of inactive human NLRP3 in complex with NEK7 at 3.8 Å resolution shows the C-terminal lobe of NEK7 nestles against both the LRR and NACHT domains of NLRP3. Mutagenesis of the NLRP3-NEK7 interface abolishes activation in vitro and in cells. Modeling predicts NEK7 bridges adjacent NLRP3 subunits in the active oligomer, and mutations to this secondary interface also abolish activation, suggesting NEK7 mediates NLRP3 oligomerization via bipartite interactions.\",\n      \"method\": \"Cryo-electron microscopy (3.8 Å), mutagenesis in vitro and in NEK7-KO / NLRP3-KO cell rescue assays, structural modeling based on NLRC4 inflammasome\",\n      \"journal\": \"Nature\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — cryo-EM structure with mutagenesis validation in cells, multiple orthogonal approaches in one rigorous study\",\n      \"pmids\": [\"31189953\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"Nercc1/NEK9 directly phosphorylates NEK7 (and NEK6) on the activation loop in vitro, leading to activation of NEK7 kinase activity (~20–25-fold for NEK6; similar for NEK7). NEK7 binds the C-terminal non-catalytic tail of NEK9. This establishes a mitotic kinase cascade: NEK9 → NEK7.\",\n      \"method\": \"In vitro kinase assay, co-expression with activated NEK9 mutant, recombinant protein phosphorylation, mass spectrometry of phosphorylation site\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — direct in vitro phosphorylation reconstitution, activation loop site identified, replicated in multiple assay formats\",\n      \"pmids\": [\"12840024\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Crystal structure of NEK7 reveals an autoinhibited conformation in which Tyr97 protrudes into the active site, interacts with the activation loop, and blocks the αC helix. Mutation of Tyr97 to phenylalanine renders NEK7 constitutively active. Binding of the NEK9 non-catalytic C-terminal domain increases NEK6/NEK7 activity but not the Tyr97Phe mutant activity, indicating NEK9 activates NEK7 by releasing the autoinhibitory tyrosine.\",\n      \"method\": \"X-ray crystallography, site-directed mutagenesis, in vitro kinase assay\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure plus mutagenesis with functional kinase assay validation\",\n      \"pmids\": [\"19941817\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Self-association of the NEK9 C-terminal domain (CTD) is required for NEK7 activation. Crystal structure of NEK7(Y97F) bound to NEK9(810-828) reveals a binding site on the C-lobe of NEK7. NEK7(Y97F) crystallizes as a back-to-back dimer between N-lobes; this dimer interface is coupled to the conformation of residue 97. NEK9-CTD activates NEK7 by promoting back-to-back dimerization that releases the autoinhibitory Tyr97.\",\n      \"method\": \"X-ray crystallography, mutagenesis, in vitro kinase assay, binding mapping\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure with mutagenesis and functional validation, mechanistically extends prior structural work\",\n      \"pmids\": [\"26522158\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"NEK6 and NEK7 are both activated in mitosis and are required for robust mitotic spindle formation. Depletion or expression of reduced-activity mutants causes metaphase arrest with fragile spindles; hypomorphic conditions cause cytokinesis defects. NEK6 localizes to spindle microtubules in metaphase/anaphase and to the midbody; NEK7 localizes primarily to spindle poles. Despite high similarity, NEK6 and NEK7 are non-redundant for mitotic progression.\",\n      \"method\": \"siRNA depletion, dominant-negative/hypomorphic mutant expression, immunofluorescence localization, cell cycle analysis\",\n      \"journal\": \"Molecular and cellular biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — systematic KD plus mutant analysis with multiple orthogonal readouts, establishes non-redundancy\",\n      \"pmids\": [\"19414596\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Endogenous NEK7 is enriched at the centrosome throughout the cell cycle in a microtubule-independent manner, and transiently localizes to the midbody during cytokinesis. siRNA-mediated knockdown of NEK7 causes a significant increase in cells with multipolar spindles, indicating a role in spindle assembly and mitotic progression.\",\n      \"method\": \"Immunofluorescence microscopy (endogenous protein), siRNA knockdown, microtubule depolymerization experiments\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct localization experiment with functional KD phenotype, single lab, single main method per claim\",\n      \"pmids\": [\"17101132\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"NEK7 knockdown by RNAi causes prometaphase arrest with monopolar or disorganized spindles, reduces centrosomal γ-tubulin levels, and impairs microtubule re-growth activity at the centrosome, indicating NEK7 is required for centrosomal microtubule nucleation. NEK7 also directly participates in cytokinesis, as evidenced by its midbody localization.\",\n      \"method\": \"RNAi knockdown, γ-tubulin immunofluorescence, microtubule re-growth assay, live-cell imaging\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — KD with defined cellular phenotypes and mechanistic readout (γ-tubulin reduction), single lab\",\n      \"pmids\": [\"17586473\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"Nek7-deficient mice die in late embryogenesis or early postnatal stages with severe growth retardation. MEFs from Nek7-/- embryos show chromosomal lagging, micronuclei, cytokinesis failure, tetraploidy, aneuploidy, increased multicentrosomal cells, and altered primary cilia (reduced frequency of primary cilia, appearance of two-cilia cells), confirming NEK7 as an essential regulator of cell division in vivo.\",\n      \"method\": \"Genetic knockout mouse generation, MEF cytogenetic analysis, FISH, immunofluorescence for centrosomes and cilia\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic KO with multiple orthogonal cellular phenotype readouts\",\n      \"pmids\": [\"20473324\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"NEK7 is essential for centriole duplication: NEK7 depletion inhibits centriole duplication, and centrosome-targeted NEK7 overexpression drives extra centriole formation in a kinase-activity-dependent manner. NEK7 depletion prevents pericentriolar material (PCM) protein accumulation at centrosomes in a cell-cycle-stage-specific manner (particularly at G1/G2), indicating a role beyond mitosis in centrosome maturation.\",\n      \"method\": \"siRNA depletion, centrosome-directed overexpression constructs, kinase-dead mutant, immunofluorescence for centriole markers and PCM proteins\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD and OE with kinase-dead controls, two orthogonal readouts, single lab\",\n      \"pmids\": [\"22100915\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NEK7 is recruited to telomeres after oxidative DNA damage in an ATM-activation-dependent manner. NEK7 phosphorylates TRF1 at Ser114, preventing TRF1 binding to Fbx4 (an SCF E3 ligase subunit) and thereby blocking TRF1 proteasomal degradation, stabilizing the TRF1-Tin2 shelterin complex at telomeres. NEK7 deficiency leads to telomere aberrations and sustained DNA damage foci after oxidative damage.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay with mutagenesis (Ser114), mass spectrometry, ChIP, immunofluorescence, NEK7 KO cells\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro phosphorylation with site-specific mutagenesis, Co-IP, ChIP, and KO phenotype, multiple orthogonal methods in one study\",\n      \"pmids\": [\"28216227\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NEK7 regulates dendrite morphogenesis and spine formation in neurons. NEK7 kinase activity is required for dendrite growth and branching. Mechanistically, NEK7 phosphorylates the kinesin Eg5/KIF11, promoting its accumulation on microtubules in distal dendrites where Eg5 limits retrograde microtubule polymerization (inhibitory to dendrite growth), acting through microtubule stabilization independent of Eg5 motor activity.\",\n      \"method\": \"In vitro and in vivo RNAi, kinase-dead rescue, phosphomimetic/phosphodead Eg5 mutants, live-cell microtubule imaging, in vivo cortical neuron analysis\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — kinase activity requirement established by mutagenesis, substrate phosphorylation (Eg5) validated by phosphomimetic rescue, multiple orthogonal methods in vitro and in vivo\",\n      \"pmids\": [\"29899413\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"NEK6 and NEK7 phosphorylate the N-terminal domain of the microtubule-associated protein EML4 at Ser144 and Ser146 in vitro during mitosis. This phosphorylation reduces EML4 affinity for microtubule acidic C-terminal tails. Depletion of NEK6/NEK7 increases EML4 binding to mitotic spindle microtubules, stabilizes them, and impairs chromosome congression. An S144A-S146A double mutant phenocopies kinase depletion.\",\n      \"method\": \"In vitro kinase assay, cryo-EM with 3D reconstruction (microtubule sedimentation), siRNA depletion, phosphomimetic/phosphodead mutants, chromosome congression imaging\",\n      \"journal\": \"Science signaling\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay with site-specific mutagenesis, structural microtubule binding data, and cell-based rescue experiments\",\n      \"pmids\": [\"31409757\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"GSTO1-1 (glutathione transferase omega 1-1) deglutathionylates NEK7 at cysteine 253, promoting NEK7 activity and NLRP3 inflammasome activation. Inhibition or knockout of GSTO1-1 blocks NLRP3 activation, identifying glutathionylation/deglutathionylation of NEK7 Cys253 as a regulatory post-translational modification.\",\n      \"method\": \"Small molecule GSTO1-1 inhibitor (C1-27), siRNA knockdown, GSTO1-1-/- mice, site-directed mutagenesis (Cys253), biochemical deglutathionylation assay\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1–2 / Moderate — PTM identified with site-specific mutagenesis and KO mouse, single lab\",\n      \"pmids\": [\"31577945\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PLK4, the master regulator of centrosome duplication, phosphorylates NEK7 at Ser204. This phosphorylation attenuates NEK7-NLRP3 interaction and suppresses NLRP3 inflammasome activation. The deubiquitinase CYLD is recruited to the centrosome by Spata2 to deubiquitinate PLK4, facilitating PLK4-mediated NEK7 phosphorylation. Mutation of NEK7 Ser204 augments NEK7-NLRP3 interaction and NLRP3 activation.\",\n      \"method\": \"Co-immunoprecipitation, in vitro kinase assay, phospho-site mutagenesis (Ser204), shRNA knockdown, PLK4 pharmacological inhibition, Spata2-deficient macrophages and peritonitis model\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro kinase assay with site-specific mutagenesis, Co-IP, genetic KD, in vivo model, multiple orthogonal methods\",\n      \"pmids\": [\"31762063\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ZDHHC5 palmitoylates NLRP3 at the LRR domain; this S-palmitoylation promotes NLRP3-NEK7 interaction, NLRP3 oligomerization, and inflammasome assembly. The depalmitoylase ABHD17A reverses this modification. ZDHHC5 silencing blocks NLRP3-NEK7 interaction and ASC speck formation, and Zdhhc5-/- mice show defective NLRP3 activation in vivo.\",\n      \"method\": \"Acyl-RAC palmitoylation assay, Co-immunoprecipitation, ZDHHC5 siRNA/KO, ABHD17A functional assay, mutagenesis of palmitoylation sites, Zdhhc5-/- mice\",\n      \"journal\": \"Molecular cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — PTM biochemically defined with multiple orthogonal methods (acyl-RAC, Co-IP, mutagenesis), in vivo KO mouse, single rigorous study\",\n      \"pmids\": [\"38092000\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"MARCH5, a mitochondria-associated E3 ligase, interacts with the NACHT domain of NLRP3 and promotes K27-linked polyubiquitination of NLRP3 at K324 and K430. This ubiquitination is required for NLRP3 to bind NEK7 and form NLRP3 oligomers; ubiquitination-defective NLRP3 mutants (K324R, K430R) cannot bind NEK7 or form ASC specks. March5 conditional KO mice fail to produce IL-1β/IL-18 upon infection.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay with linkage-specific analysis, site-directed mutagenesis (K324R, K430R), myeloid-specific conditional KO mice, macrophage infection models\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — ubiquitination biochemically characterized with site-specific mutagenesis and linkage analysis, genetic KO in vivo, multiple orthogonal methods\",\n      \"pmids\": [\"37575012\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"NEK7 is rapidly phosphorylated at threonine-190/191 by JNK1 downstream of K+ efflux and gasdermin D (GSDMD) pore formation after NLRP3 activation. This phosphorylation enhances NEK7-NLRP3 binding and further promotes inflammasome assembly. Knock-in mice with Thr190/191Val substitutions show impaired NEK7 phosphorylation, reduced NLRP3 inflammasome activation, and decreased IL-1β secretion.\",\n      \"method\": \"Phospho-proteomics, in vitro JNK1 kinase assay, knock-in mutant mice (T190V/T191V), Co-immunoprecipitation, NLRP3 activation assays\",\n      \"journal\": \"Science immunology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1–2 / Strong — in vitro kinase assay identifying writer (JNK1), phospho-site knock-in mice, Co-IP, multiple orthogonal methods in one rigorous study\",\n      \"pmids\": [\"39752537\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"A20 (TNFAIP3) directly binds NEK7 and mediates K48-linked ubiquitination of NEK7 at K189 and K293, targeting NEK7 for proteasomal degradation. A20 deficiency increases NEK7 protein levels. A20 also disrupts NEK7-NLRP3 association, potentially via its OTU domain and/or ZnF4/ZnF7 motifs. NEK7 deletion attenuates NLRP3 inflammasome activation in A20-deficient conditions both in vitro and in vivo.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay (K48-linkage specific), site-directed mutagenesis (K189, K293), A20-deficient macrophages and in vivo models\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP, site-specific ubiquitination mutagenesis, in vitro and in vivo KO, single lab\",\n      \"pmids\": [\"38865260\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"METTL3-dependent m6A modification of NEK7 mRNA increases its stability (recognized by IGF2BP2), promoting NEK7 expression and NLRP3 inflammasome activation. Silencing METTL3 reduces A20/TNFAIP3 m6A modification, stabilizes TNFAIP3 mRNA (increasing A20 protein), which then ubiquitinates NEK7 to impair NLRP3 assembly. In a separate study, silencing METTL3 inhibits m6A methylation of NEK7, reducing NEK7 mRNA stability and blocking chondrocyte pyroptosis.\",\n      \"method\": \"m6A-RIP, RNA immunoprecipitation, dual-luciferase reporter, METTL3 KO/KD, TNFAIP3 ubiquitination assay, in vivo periodontitis model\",\n      \"journal\": \"Advanced science (Weinheim, Baden-Wurttemberg, Germany)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — m6A-RIP, RIP, luciferase reporter, and KO experiments establish PTM-mRNA stability link, single lab\",\n      \"pmids\": [\"38696610\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"OGT-mediated O-GlcNAcylation of NEK7 at S260 suppresses NEK7 phosphorylation at S260, which promotes NEK7-NLRP3 interaction and chondrocyte pyroptosis. OGT silencing enhances NEK7 phosphorylation and blocks the NEK7-NLRP3 complex, reducing NLRP3 activation.\",\n      \"method\": \"Co-immunoprecipitation, western blot for O-GlcNAc and phosphorylation, OGT knockout/knockdown, site-specific mutagenesis (S260), in vivo OA model\",\n      \"journal\": \"Autoimmunity\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with site-specific analysis, OGT KO in vivo, single lab, moderate mechanistic depth\",\n      \"pmids\": [\"38389178\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"SIRT5 desuccinylates NEK7 at K81, downregulating NEK7 expression/activity. Electroacupuncture in a MCAO mouse model of ischemia-reperfusion injury activates SIRT5, leading to decreased NEK7 succinylation at K81 and reduced neuronal pyroptosis. NEK7 overexpression reverses these protective effects.\",\n      \"method\": \"MCAO mouse model, succinylation assay, SIRT5 KD, NEK7 overexpression rescue, western blot, brain infarct analysis\",\n      \"journal\": \"Brain research bulletin\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — in vivo KD/OE with defined PTM site, but indirect evidence for SIRT5 directly desuccinylating NEK7 K81 (no in vitro biochemical reconstitution shown in abstract), single lab\",\n      \"pmids\": [\"39694147\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"TRIM32 (E3 ubiquitin ligase) promotes ubiquitylation of NEK7 at K64, leading to NEK7 downregulation and suppression of NLRP3-dependent microglial pyroptosis. Inhibiting NEK7 ubiquitylation reverses TRIM32-mediated pyroptosis suppression.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitylation assay, site-directed mutagenesis (K64), siRNA/overexpression, in vivo SCI mouse model\",\n      \"journal\": \"Molecular biotechnology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — Co-IP and K64 mutagenesis establish ubiquitylation site, but in vitro reconstitution not demonstrated in abstract, single lab\",\n      \"pmids\": [\"38030945\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"EML4-ALK V3 oncogenic fusion protein recruits NEK9 and NEK7 to microtubules via the EML4 N-terminal microtubule-binding region. This complex promotes microtubule stabilization, extended cytoplasmic protrusions, and increased cell migration. Constitutively active NEK9 accelerates migration in a microtubule-dependent manner requiring downstream NEK7 but not ALK activity.\",\n      \"method\": \"Co-immunoprecipitation, siRNA depletion, dominant-active NEK9 expression, cell morphology and migration assays, microtubule imaging\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP with NEK9/NEK7, functional epistasis (NEK9→NEK7) established by depletion rescue, two orthogonal methods, single lab\",\n      \"pmids\": [\"32184261\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"In the context of EML4-ALK V3, NEK7 phosphorylates the kinesin Eg5 at Ser1033 on interphase microtubules. Eg5 phosphorylation at S1033 by NEK7 drives the mesenchymal morphology of EML4-ALK V3 cells. NEK7 depletion reduces Eg5 recruitment to microtubules, and a phosphomimetic Eg5-S1033D mutant rescues mesenchymal morphology in NEK7-depleted cells.\",\n      \"method\": \"siRNA depletion of NEK7, phosphomimetic/phosphodead Eg5 mutants, immunofluorescence for Eg5 microtubule association, cell morphology analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — substrate phosphorylation site (S1033) validated by phosphomimetic rescue, mechanistic epistasis established, single lab\",\n      \"pmids\": [\"38458397\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"NEK7 depletion inhibits G1 progression in human U2OS cells by downregulating cyclins and CDKs. NEK7 loss also inhibits the earliest stages of procentriole formation (STIL degradation by APC/CCdh1) and induces primary cilia formation in RPE1 cells. Abnormal APC/CCdh1 accumulation at centrioles and continuous STIL degradation are the mechanistic basis for centriole assembly failure.\",\n      \"method\": \"siRNA depletion, cell cycle analysis, immunofluorescence for centriole markers (STIL, Cdh1), quantitative microscopy\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with multiple mechanistic readouts (cyclin levels, Cdh1 localization, STIL degradation), single lab\",\n      \"pmids\": [\"28539406\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"NEK7 regulates microtubule dynamic instability: siRNA-mediated NEK7 knockdown reduces speeds of MT growth and catastrophe, reduces relative time in catastrophe, and lowers overall MT dynamicity. Ectopic overexpression has inverse effects. These phenotypes are recapitulated in Nek7-/- MEFs, establishing NEK7 as a direct regulator of MT dynamics.\",\n      \"method\": \"siRNA knockdown, NEK7-/- MEFs, PlusTipTracker live-cell MT imaging, rescue by re-expression\",\n      \"journal\": \"Biochimica et biophysica acta\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD and KO with quantitative live-imaging MT dynamics, genetic KO recapitulates KD phenotype, single lab\",\n      \"pmids\": [\"23313050\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Anks3 interacts with NEK7 (but not NEK6) and retains NEK7 in the cytoplasm, preventing NEK7 nuclear localization. Anks3 undergoes an ~20 kDa molecular weight increase upon NEK7 co-expression that is not caused by NEK7-dependent phosphorylation (occurs with kinase-dead NEK7 mutant), indicating an alternative modification mechanism.\",\n      \"method\": \"Co-immunoprecipitation, subcellular fractionation, immunofluorescence, kinase-dead mutant, mass spectrometry\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — Co-IP with kinase-dead control establishes binding but modification mechanism not fully resolved; single lab\",\n      \"pmids\": [\"26188091\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Licochalcone B (LicoB) directly binds NEK7 and inhibits the NEK7-NLRP3 interaction, specifically suppressing NLRP3 inflammasome activation without affecting AIM2 or NLRC4 inflammasomes. This is the first direct small-molecule targeting of the NEK7-NLRP3 interface as a therapeutic approach.\",\n      \"method\": \"Direct binding assay (drug-protein interaction), Co-immunoprecipitation, macrophage inflammasome activation assays, in vivo sepsis/peritonitis/NASH models\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct NEK7 binding shown, mechanistic Co-IP confirms disruption of NEK7-NLRP3 interface, in vivo validation, single lab\",\n      \"pmids\": [\"34882936\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Berberine directly targets NEK7 via a hydrogen bond between its 2,3-methylenedioxy group and Arg121 of NEK7. Because R121 is located within the key domain mediating NEK7-NLRP3 interaction, berberine specifically blocks the NEK7-NLRP3 interaction and inhibits IL-1β release independently of NF-κB and TLR4 signaling. Anti-inflammatory efficacy in vivo is NEK7-dependent.\",\n      \"method\": \"Activity-based protein profiling (direct target identification), site-directed mutagenesis (R121), Co-immunoprecipitation, NEK7 KD in vivo rescue\",\n      \"journal\": \"Journal of medicinal chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct target identification by ABPP plus R121 mutagenesis validates binding site, in vivo NEK7-dependence confirmed, single lab\",\n      \"pmids\": [\"33440945\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"A single amino acid difference—Arg121 in NEK7 versus Gln132 in NEK6—accounts for NEK7's ability to bind NLRP3 and support inflammasome activation, while NEK6 cannot. Substituting Gln132 with Arg in NEK6 confers NLRP3 binding and inflammasome activation activity in macrophages.\",\n      \"method\": \"Site-directed mutagenesis (R121/Q132), Co-immunoprecipitation, caspase-1 activation assay in macrophages, NEK7-KO rescue\",\n      \"journal\": \"Journal of immunology (Baltimore, Md. : 1950)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — single residue swap in both directions (gain and loss of function) with biochemical and cell-based validation\",\n      \"pmids\": [\"35354613\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PAF (platelet-activating factor) activates the NLRP3 inflammasome in a NEK7-dependent (and NLRP3, ASC, caspase-1-dependent) manner, requiring potassium efflux, independently of PAFR signaling. NEK7 is thus required for a lipid-triggered, PAFR-independent pathway of NLRP3 activation.\",\n      \"method\": \"Genetic KO macrophages (NEK7, NLRP3, ASC, caspase-1), K+ efflux measurement, in vivo peritonitis model\",\n      \"journal\": \"The Journal of experimental medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO panel establishes NEK7 requirement in this pathway, in vivo validation, single lab\",\n      \"pmids\": [\"31558613\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Co-chaperone UNC45A is required for expression of NEK7 in cancer cells. UNC45A localizes to the cancer cell nucleus, upregulates glucocorticoid receptor transcriptional activity, and thereby promotes NEK7 transcription. UNC45A-deficient cancer cells show pericentrosomal material disorganization, centrosomal separation defects, and metaphase/cytokinesis stalling; these phenotypes are rescued by heterologous NEK7 expression.\",\n      \"method\": \"siRNA knockdown, immunofluorescence, gene microarray, RT-qPCR, glucocorticoid receptor activity assay, NEK7 rescue expression\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis by rescue, transcriptional mechanism defined, multiple cell-based readouts, single lab\",\n      \"pmids\": [\"30737284\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"NEK7 directly binds EGFR and phosphorylates it at serine 1070, activating MAPK and PI3K/AKT signaling pathways to drive acquired lenvatinib resistance in hepatocellular carcinoma. An inhibitory peptide targeting the NEK7-binding domain (EGFR aa 979–1099) blocks EGFR S1070 phosphorylation and suppresses resistance.\",\n      \"method\": \"Kinase CRISPR-Cas9 genetic screen, Co-IP (NEK7-EGFR binding), site-specific phosphorylation analysis (S1070), inhibitory peptide design, organoid and xenograft models\",\n      \"journal\": \"Hepatology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR screen plus Co-IP and site-specific phosphorylation evidence, organoid/in vivo validation, single lab\",\n      \"pmids\": [\"40694824\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"Isocyanic acid (produced by LACC1 enzyme) carbamoylates NLRP3 at lysine-593, disrupting NLRP3-NEK7 interaction and limiting NLRP3 inflammasome activation. LACC1 KO or K593 carbamoylation-deficient NLRP3 mutant promotes inflammatory responses in vitro and in vivo.\",\n      \"method\": \"Carbamoylation mass spectrometry, site-directed mutagenesis (K593), Co-immunoprecipitation, LACC1 KO/Lacc1-/- mice, macrophage inflammasome assays\",\n      \"journal\": \"Science advances\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — PTM site defined by MS and mutagenesis, disruption of NEK7-NLRP3 interface shown by Co-IP, KO in vivo, single lab\",\n      \"pmids\": [\"40053593\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"NEK7 is required for proper wiring of cortical parvalbumin (PV+) interneurons. NEK7-deficient PV+ interneurons show altered microtubule dynamics, impaired axon growth cone steering, reduced axon length, lower arbor complexity, and fewer synaptic contacts with pyramidal cells in vivo.\",\n      \"method\": \"Conditional/targeted NEK7 ablation in PV interneurons, in vitro and in vivo morphology analysis, microtubule dynamics live imaging, synapse quantification\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-type-specific KO with multiple in vitro and in vivo readouts, single lab\",\n      \"pmids\": [\"30067978\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"NLRP3 inflammasome activation by NEK7 is restricted to interphase; the NEK7-NLRP3 interaction is required for NLRP3 activation, and the interaction is enhanced by K+ efflux. The Influenza A virus accessory protein PB1-F2 limits the conformational transition of NLRP3 from its auto-repressed closed conformation to its active state, thereby diminishing NEK7-NLRP3 interaction and blocking inflammasome assembly in human macrophages.\",\n      \"method\": \"IAV mutant infection (PB1-F2 deficient), Co-immunoprecipitation (NEK7-NLRP3), caspase-1 activation assay, gasdermin D cleavage, LDH/IL-1β release\",\n      \"journal\": \"EMBO reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — viral protein loss-of-function with Co-IP readout of NEK7-NLRP3 interaction, mechanistic specificity established, single lab\",\n      \"pmids\": [\"33180976\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"In HEK293 cells reconstituted with NLRP3 and ASC (but not NEK7), ASC speck polymerization occurs independently of NEK7, suggesting a NEK7-independent mode of NLRP3 activation exists in this cell system. Evidence supports a stacked-torus hexameric NLRP3 oligomer in this context.\",\n      \"method\": \"HEK293 cell reconstitution of NLRP3 inflammasome without NEK7, ASC speck assay, interface mutagenesis based on structural models\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single heterologous reconstitution system, negative result (NEK7-independent activation) may reflect cell-system artifact, single lab\",\n      \"pmids\": [\"36142182\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"A cereblon (CRBN) glue degrader of NEK7 (NK7-902) potently and selectively degrades NEK7 in human immune cells. Full NEK7 degradation completely blocked NLRP3-dependent IL-1β release in vitro in some but not all donors/conditions, and in mice. In cynomolgus monkeys, NK7-902 caused long-lasting NEK7 degradation but only transiently blocked IL-1β, suggesting NEK7 contributes to but may not be absolutely required for NLRP3 activation in primates.\",\n      \"method\": \"Targeted protein degradation (CRBN molecular glue), NEK7 protein level measurement in primary human cells and whole blood, NLRP3 activation assays, in vivo mouse and monkey models\",\n      \"journal\": \"Cell chemical biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — selective degrader with cross-species comparison, multiple biological systems tested, single lab; highlights context-dependence of NEK7 requirement\",\n      \"pmids\": [\"40639372\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"NEK7 is a dual-function NIMA-related serine/threonine kinase that (1) drives mitotic progression by localizing to centrosomes, where—activated via NEK9-induced dimerization and release of autoinhibitory Tyr97—it phosphorylates substrates such as EML4 and Eg5 to regulate microtubule dynamics, spindle assembly, centriole duplication, and cytokinesis; and (2) acts as an essential scaffolding co-activator of the NLRP3 inflammasome in interphase cells, binding the LRR and NACHT domains of NLRP3 (via its C-terminal lobe, with critical contact at Arg121) downstream of K+ efflux to bridge adjacent NLRP3 subunits and license oligomerization, ASC speck formation, and caspase-1 activation, a process regulated by multiple PTMs on NEK7 (JNK1-mediated T190/T191 phosphorylation amplifying NLRP3 binding; PLK4-mediated S204 phosphorylation suppressing it; GSTO1-1-mediated Cys253 deglutathionylation activating it; A20/TRIM32/TNFAIP3-mediated ubiquitination degrading it) and on NLRP3 itself (MARCH5-mediated K27 ubiquitination required for NEK7 recruitment; ZDHHC5-mediated palmitoylation promoting the interaction; isocyanic acid carbamoylation at K593 and NLRP3 LRR mutations altering the interface); in post-mitotic neurons NEK7 additionally regulates dendrite morphogenesis through Eg5 phosphorylation, and protects telomere integrity by phosphorylating TRF1-Ser114 to prevent its proteasomal degradation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"NEK7 is a NIMA-related serine/threonine kinase with two largely separable cellular roles: an enzymatic role in mitotic cell division and a non-catalytic scaffolding role in innate immune signaling [#7, #0]. As a kinase, NEK7 is activated late in a mitotic cascade in which NEK9 binds its non-catalytic C-terminal tail and phosphorylates the NEK7 activation loop [#4]; structurally, this activation works by promoting a back-to-back NEK7 dimer that displaces an autoinhibitory Tyr97 from the active site [#5, #6]. Active NEK7 localizes to centrosomes and spindle poles and is required for centriole duplication, centrosomal microtubule nucleation, spindle assembly, and cytokinesis, with loss causing multipolar/monopolar spindles, aneuploidy, and embryonic lethality in mice [#8, #9, #10, #11]. It controls microtubule dynamics directly and through substrate phosphorylation, including EML4 at Ser144/Ser146 to reduce microtubule affinity during chromosome congression and the kinesin Eg5/KIF11 to govern dendrite morphogenesis and cortical interneuron wiring [#14, #13, #37, #28]. NEK7 also phosphorylates TRF1-Ser114 to block its proteasomal degradation and protect telomere integrity after oxidative damage [#12]. Independently of its kinase activity, NEK7 is an essential co-activator of the NLRP3 inflammasome, acting downstream of K+ efflux: it binds the LRR and NACHT domains of NLRP3 through its C-terminal lobe and bridges adjacent NLRP3 subunits to license oligomerization, ASC speck formation, and caspase-1 activation [#0, #1, #3]. A single residue, Arg121, distinguishes NEK7 from NEK6 and is required for NLRP3 binding [#32], and small molecules engaging this interface (e.g., berberine at Arg121) selectively block NLRP3 activation [#31, #30]. This interaction is gated by post-translational modifications on both partners—JNK1 phosphorylation of NEK7 Thr190/191 and GSTO1-1 deglutathionylation of Cys253 promote it, PLK4 phosphorylation of Ser204 suppresses it, and MARCH5-mediated K27 ubiquitination and ZDHHC5 palmitoylation of NLRP3 are required for NEK7 recruitment [#19, #15, #16, #18, #17].\",\n  \"teleology\": [\n    {\n      \"year\": 2003,\n      \"claim\": \"Established that NEK7 sits downstream in a kinase cascade, answering how this kinase becomes activated.\",\n      \"evidence\": \"In vitro kinase reconstitution with activated NEK9 and mass spectrometry of the activation-loop site\",\n      \"pmids\": [\"12840024\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not define the structural basis of activation\", \"Physiological triggers of NEK9 activity not addressed\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Defined the autoinhibition mechanism, showing how NEK7 is held inactive and how NEK9 relieves it.\",\n      \"evidence\": \"X-ray crystallography of NEK7 with Tyr97Phe mutagenesis and in vitro kinase assays\",\n      \"pmids\": [\"19941817\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Did not capture the active conformation\", \"Mechanism coupling NEK9 binding to Tyr97 release unresolved at this stage\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Placed NEK7 in mitotic spindle formation and distinguished it functionally from the related NEK6.\",\n      \"evidence\": \"siRNA depletion, hypomorphic mutants, and immunofluorescence localization across mitosis\",\n      \"pmids\": [\"19414596\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Spindle-pole substrates not identified\", \"Basis of NEK6/NEK7 non-redundancy unexplained\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Extended NEK7's mitotic role to centriole duplication and centrosome maturation, requiring kinase activity.\",\n      \"evidence\": \"siRNA depletion, centrosome-targeted overexpression, kinase-dead mutants, PCM marker imaging\",\n      \"pmids\": [\"22100915\", \"20473324\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Centrosomal substrates not identified\", \"Link between kinase activity and PCM accumulation mechanistic detail missing\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Resolved how NEK9 activates NEK7 structurally, linking dimerization to Tyr97 displacement.\",\n      \"evidence\": \"Crystal structure of NEK7(Y97F)–NEK9 peptide complex with mutagenesis and kinase assays\",\n      \"pmids\": [\"26522158\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"In-cell relevance of the back-to-back dimer not directly visualized\", \"Does not address scaffolding (kinase-independent) functions\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Identified NEK7 as an essential, specific NLRP3 inflammasome component and revealed its mitosis–inflammasome mutual exclusivity.\",\n      \"evidence\": \"Genome-wide CRISPR screen, Co-IP with domain mapping, siRNA, cell-cycle synchronization, in vivo models\",\n      \"pmids\": [\"26553871\", \"26642356\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether kinase activity is required not fully settled at this point\", \"Structural basis of the NEK7–NLRP3 interface unknown\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Established that NEK7's scaffolding role, not its catalytic activity, mediates NLRP3 activation downstream of K+ efflux.\",\n      \"evidence\": \"Reciprocal Co-IP, Nek7-/- macrophages and chimeras, kinase-dead rescue, ASC speck imaging\",\n      \"pmids\": [\"26814970\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural mechanism of subunit bridging not yet resolved\", \"How K+ efflux promotes the interaction unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Revealed a telomere-protective kinase function, showing NEK7 phosphorylates TRF1 to block its degradation.\",\n      \"evidence\": \"In vitro kinase assay with Ser114 mutagenesis, Co-IP, ChIP, mass spectrometry, KO cells\",\n      \"pmids\": [\"28216227\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Signal coupling ATM activation to NEK7 telomere recruitment incompletely defined\", \"Relationship to NEK7's centrosomal pool unclear\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Provided the structural basis for NEK7-mediated NLRP3 oligomerization via bipartite interactions.\",\n      \"evidence\": \"3.8 Å cryo-EM of inactive NLRP3–NEK7 with interface mutagenesis and cell rescue\",\n      \"pmids\": [\"31189953\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the active oligomer was modeled, not directly resolved\", \"Conformational steps from binding to oligomerization not captured\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Identified mitotic and neuronal substrates of NEK7 kinase activity that control microtubule behavior.\",\n      \"evidence\": \"In vitro kinase assays with phosphosite mutants for EML4 (S144/S146) and Eg5, microtubule and morphology imaging in vitro and in vivo\",\n      \"pmids\": [\"31409757\", \"29899413\", \"30067978\", \"23313050\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full substrate repertoire not defined\", \"How NEK7 selects spindle vs dendritic targets unknown\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Began defining the PTM code on NEK7 and NLRP3 that tunes inflammasome assembly, including suppressive PLK4 phosphorylation and activating deglutathionylation.\",\n      \"evidence\": \"In vitro kinase and deglutathionylation assays, site-specific mutagenesis (S204, Cys253), Co-IP, KO mice and macrophages\",\n      \"pmids\": [\"31762063\", \"31577945\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Hierarchy and interplay among NEK7 PTMs unresolved\", \"Spatial coordination with centrosomal PLK4 in immune cells unclear\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Pinpointed Arg121 as the determinant of NLRP3 binding specificity that explains why NEK7, not NEK6, supports the inflammasome.\",\n      \"evidence\": \"Reciprocal single-residue swap (R121/Q132) with Co-IP and caspase-1 assays in macrophages\",\n      \"pmids\": [\"35354613\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Does not address whether other contacts contribute under physiological conditions\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Showed that NLRP3-side modifications—K27 ubiquitination and palmitoylation—are prerequisites for NEK7 recruitment.\",\n      \"evidence\": \"Linkage-specific ubiquitination and acyl-RAC assays, site mutagenesis (K324/K430; LRR palmitoylation), Co-IP, conditional/KO mice\",\n      \"pmids\": [\"37575012\", \"38092000\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Temporal ordering relative to K+ efflux not fully resolved\", \"Whether these modifications act on the same NLRP3 pool simultaneously unknown\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Expanded the regulatory layer with additional NEK7 PTMs and mRNA-level control governing its abundance and NLRP3 binding.\",\n      \"evidence\": \"Co-IP, K48-ubiquitination and O-GlcNAc assays, site mutagenesis (K189/K293; S260), m6A-RIP/luciferase, KO macrophages and disease models\",\n      \"pmids\": [\"38865260\", \"38389178\", \"38696610\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Many sites from single labs without cross-validation\", \"Quantitative contribution of each PTM in vivo unclear\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Identified non-canonical NEK7 kinase functions in cancer, including EML4-ALK-associated migration and EGFR-driven drug resistance.\",\n      \"evidence\": \"CRISPR screens, Co-IP, phosphosite analysis (Eg5 S1033; EGFR S1070), inhibitory peptide, organoid/xenograft models\",\n      \"pmids\": [\"32184261\", \"38458397\", \"40694824\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-lab findings\", \"Generality across tumor types and direct kinase-substrate relationships need broader validation\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Tested NEK7's necessity for NLRP3 activation pharmacologically and revealed context- and species-dependence.\",\n      \"evidence\": \"JNK1 phosphosite knock-in mice (T190V/T191V), carbamoylation MS, and a CRBN glue degrader across human cells, mice, and monkeys\",\n      \"pmids\": [\"39752537\", \"40053593\", \"40639372\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Why NEK7 requirement varies across primates/conditions unresolved\", \"Existence of NEK7-independent NLRP3 activation modes not fully defined\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How NEK7's two roles—mitotic kinase and inflammasome scaffold—are coordinated spatially and temporally within a single cell, and whether NEK7 is absolutely required for NLRP3 activation across species, remains open.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No unified model linking cell-cycle sequestration of NEK7 to inflammasome timing at the molecular level\", \"Species-dependent dispensability suggests redundancy that is uncharacterized\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [12, 13, 14, 26, 35]},\n      {\"term_id\": \"GO:0140657\", \"supporting_discovery_ids\": [4, 5]},\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [0, 3, 32]},\n      {\"term_id\": \"GO:0008092\", \"supporting_discovery_ids\": [14, 28]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005815\", \"supporting_discovery_ids\": [8, 9, 11]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [7, 28]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [29]},\n      {\"term_id\": \"GO:0005694\", \"supporting_discovery_ids\": [12]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [0, 1, 2, 3]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [7, 9, 10, 11]},\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [0, 19, 33]},\n      {\"term_id\": \"R-HSA-1852241\", \"supporting_discovery_ids\": [11, 27]}\n    ],\n    \"complexes\": [\"NLRP3 inflammasome\"],\n    \"partners\": [\"NLRP3\", \"NEK9\", \"EML4\", \"KIF11\", \"TRF1\", \"EGFR\", \"ANKS3\", \"A20\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}