Affinage

NEK3

Serine/threonine-protein kinase Nek3 · UniProt P51956

Length
506 aa
Mass
57.7 kDa
Annotated
2026-06-10
13 papers in source corpus 8 papers cited in narrative 8 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 7/7 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

NEK3 is a predominantly cytoplasmic NIMA-related serine/threonine kinase that couples prolactin receptor (PRLR) signaling to actin cytoskeletal reorganization and cell motility, with no detectable role in cell cycle progression (PMID:10224116, PMID:15618286). Upon prolactin (PRL) stimulation, NEK3 associates with the PRL receptor together with the guanine-nucleotide exchange factors Vav1/Vav2, phosphorylates Vav2 on serine and tyrosine residues, and drives Vav2-dependent Rac1 activation; this axis is abolished by a kinase-inactive NEK3 mutant (PMID:15618286). Through this pathway NEK3 is required for PRL-induced cytoskeletal remodeling, paxillin phosphorylation, focal adhesion turnover, and breast cancer cell migration and invasion (PMID:17297458). NEK3 catalytic output is gated by phosphorylation: ERK1/2 phosphorylates NEK3 at Thr-165 in the activation segment in response to PRL, and loss of this site enlarges focal adhesions, promotes stress fiber formation, and impairs migration (PMID:27489110). NEK3 also phosphorylates SNAP29 at Ser-105 to direct SNAP29 membrane association and support Golgi integrity, focal adhesion formation, and cellular recycling (PMID:29454964). A second regulatory phospho-switch at Thr-475 in the C-terminal PEST domain controls NEK3 activity toward microtubule acetylation in neurons, where loss of regulation drives HDAC6-dependent microtubule deacetylation and disturbed polarity (PMID:19509051). Biallelic loss-of-function NEK3 mutations in patients cause SIRT2-mediated α-tubulin deacetylation and downregulation of inner nuclear pore components (NUP205, NUP188, NUP155), with defective ciliary ultrastructure and abnormal cardiac left-right patterning (PMID:33230144).

Mechanistic history

Synthesis pass · year-by-year structured walk · 8 steps
  1. 1999 Medium

    Established the basic cellular identity of NEK3 as a constitutively active cytoplasmic kinase, ruling out the cell-cycle role expected of NIMA-related kinases and reframing where its function should be sought.

    Evidence Subcellular fractionation, antibody microinjection, overexpression, and cell cycle staging of murine NEK3

    PMID:10224116

    Open questions at the time
    • No physiological substrate or upstream stimulus identified
    • Function in any signaling pathway undefined
  2. 2004 High

    Defined NEK3 as an effector of PRL receptor signaling by linking it to Vav2-dependent Rac1 activation, answering what pathway NEK3 acts within and identifying its first substrate.

    Evidence Yeast two-hybrid, reciprocal co-immunoprecipitation, kinase assays, kinase-dead mutant, and Rac1 activation assay in PRL-responsive cells

    PMID:15618286

    Open questions at the time
    • Direct phosphorylation of Vav2 not mapped to specific residues
    • How PRLR triggers NEK3 activation not resolved
  3. 2007 High

    Connected NEK3 kinase activity to a concrete cellular phenotype, showing it is required for PRL-driven cytoskeletal remodeling, paxillin phosphorylation, and breast cancer cell migration/invasion.

    Evidence siRNA knockdown, overexpression, paxillin co-IP, and migration/invasion assays in breast cancer cells

    PMID:17297458

    Open questions at the time
    • Whether paxillin is a direct NEK3 substrate not established
    • Mechanistic link between Rac1 and focal adhesion remodeling incomplete
  4. 2009 Medium

    Revealed a regulatory phospho-switch (Thr-475 in the PEST domain) and a new functional readout, showing NEK3 activity opposes HDAC6-dependent microtubule deacetylation and supports neuronal polarity.

    Evidence T475A/T475D site-directed mutagenesis with neuronal overexpression and HDAC6 pathway analysis

    PMID:19509051

    Open questions at the time
    • Kinase responsible for Thr-475 phosphorylation unknown
    • Direct versus indirect control of HDAC6 not resolved
  5. 2016 Medium

    Identified the upstream activating event for NEK3, showing ERK1/2 phosphorylates Thr-165 in the activation segment to control focal adhesion remodeling and migration.

    Evidence ERK1/2 pharmacological inhibition and siRNA, T165V phospho-deficient mutant, focal adhesion immunofluorescence, and migration assays

    PMID:27489110

    Open questions at the time
    • Direct ERK1/2 phosphorylation of NEK3 not shown in vitro
    • Relationship between Thr-165 and Thr-475 regulation untested
  6. 2018 Medium

    Expanded the NEK3 substrate repertoire to membrane trafficking, identifying SNAP29 Ser-105 as a phosphosite controlling Golgi integrity and recycling, with disease relevance in CEDNIK fibroblasts.

    Evidence Phosphorylation assay, S105A mutant expression, immunofluorescence, and CEDNIK patient fibroblast rescue

    PMID:29454964

    Open questions at the time
    • How SNAP29 phosphorylation integrates with the Vav2/Rac1 axis unclear
    • In vivo relevance of the NEK3-SNAP29 link not established
  7. 2020 Medium

    Established NEK3 as a human disease gene whose loss converges on tubulin acetylation control via SIRT2 and on nuclear pore composition, linking it to ciliary defects and cardiac left-right patterning.

    Evidence Whole-exome sequencing of patients plus NEK3 siRNA in RPE cells with western blot, immunofluorescence, TEM, and transcriptome analysis

    PMID:33230144

    Open questions at the time
    • Mechanism connecting NEK3 loss to SIRT2/NNMT upregulation unknown
    • Direct effect of NEK3 on nuclear pore components versus secondary consequence unresolved
  8. 2026 Low

    Extended the NEK3-microtubule acetylation link to oocyte maturation, indicating NEK3 levels support spindle assembly and developmental competence.

    Evidence In vitro oocyte maturation under BPTMC exposure with NEK3 protein analysis, spindle/acetylation immunofluorescence, and embryo development assays

    PMID:42235065

    Open questions at the time
    • Correlative knockdown inferred from chemical exposure, not a direct NEK3 perturbation
    • Substrate or pathway mediating the acetylation effect not defined

Open questions

Synthesis pass · forward-looking unresolved questions
  • How NEK3's distinct substrate-specific roles (Vav2/Rac1 migration, SNAP29 trafficking, microtubule acetylation, nuclear pore regulation) are coordinated within a single cell, and which are direct versus indirect, remains unresolved.
  • No structural model of NEK3 substrate recognition
  • Consensus phosphorylation motif undefined
  • Integration of Thr-165 and Thr-475 regulatory inputs untested

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0140096 catalytic activity, acting on a protein 4 GO:0016740 transferase activity 3
Localization
GO:0005829 cytosol 1
Pathway
R-HSA-162582 Signal Transduction 2
Partners
MAPK1PXNSNAP29VAV1VAV2

Evidence

Reading pass · 8 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2004 NEK3 physically interacts with Vav1 and Vav2 (identified by yeast two-hybrid and confirmed by co-immunoprecipitation) in a PRL-dependent manner. PRL stimulation induces NEK3 kinase activity and promotes Vav2/NEK3 co-association with the PRL receptor. NEK3 phosphorylates Vav2 on serine and tyrosine residues, and Rac1 activation requires both NEK3 and Vav2 and is blocked by a kinase-inactive NEK3 mutant. Yeast two-hybrid, co-immunoprecipitation, kinase activity assay, kinase-dead mutant overexpression, Rac1 activation assay Molecular endocrinology (Baltimore, Md.) High 15618286
2007 NEK3 is required for PRL-mediated cytoskeletal reorganization, Rac1 activation, cell migration, and invasion of breast cancer cells. PRL stimulation induces a NEK3–paxillin interaction and increases paxillin serine phosphorylation; siRNA-mediated NEK3 knockdown reduces paxillin phosphorylation and abolishes PRL-induced Rac1 activation. siRNA knockdown, overexpression in CHO transfectants, co-immunoprecipitation, Rac1 activation assay, migration/invasion assay, western blot Oncogene High 17297458
1999 Murine NEK3 is a predominantly cytoplasmic serine/threonine kinase that shows no cell cycle-dependent variation in expression or kinase activity, and neither antibody microinjection nor overexpression of wild-type or catalytically inactive NEK3 affects cell cycle progression. Subcellular fractionation, antibody microinjection, overexpression, cell cycle staging, kinase assay The Journal of biological chemistry Medium 10224116
2009 NEK3 regulates microtubule acetylation in neurons. Phosphorylation at Thr475 in the C-terminal PEST domain acts as a regulatory switch: expression of phospho-defective (T475A) or PEST-truncated NEK3 in cultured neurons causes disturbed polarity and HDAC6-dependent deacetylation of microtubules in a kinase-dependent manner, whereas wild-type or phosphomimetic (T475D) NEK3 has no discernible effect. Site-directed mutagenesis (T475A/T475D), neuronal overexpression, immunofluorescence, HDAC6 pathway analysis Journal of cell science Medium 19509051
2016 ERK1/2 phosphorylates NEK3 at Thr-165 within the activation segment in response to PRL. A phospho-deficient NEK3-T165V mutant increases focal adhesion size, promotes zyxin-positive focal adhesion formation, induces actin stress fiber formation, and impairs breast cancer cell migration, establishing Thr-165 phosphorylation as a regulatory step controlling focal adhesion remodeling. Pharmacological ERK1/2 inhibition, siRNA knockdown of ERK1/2, phospho-deficient mutant expression (T165V), immunofluorescence of focal adhesions, migration assay The Journal of biological chemistry Medium 27489110
2018 NEK3 phosphorylates SNAP29 at serine 105 (S105). This phosphorylation directs SNAP29 membrane association; a phospho-defective S105A mutant causes defective focal adhesion formation, impaired Golgi structure, and attenuated cellular recycling. Wild-type SNAP29 (but not S105A) partially rescues abnormal morphology in CEDNIK patient-derived fibroblasts. Phosphorylation assay, S105A phospho-deficient mutant expression, immunofluorescence, CEDNIK patient fibroblast rescue experiment Biochemical and biophysical research communications Medium 29454964
2020 Biallelic loss-of-function NEK3 mutations in patients cause upregulation of SIRT2 and NNMT, leading to SIRT2-mediated α-tubulin deacetylation (detected by western blot and immunofluorescence) and downregulation of inner nuclear pore complex components NUP205, NUP188, and NUP155, associated with defective ciliary ultrastructure and abnormal cardiac left-right patterning. Whole-exome sequencing, NEK3 siRNA knockdown in RPE cells, western blot, immunofluorescence, transmission electron microscopy, transcriptome analysis Cell death & disease Medium 33230144
2026 NEK3 protein levels regulate microtubule acetylation during mouse oocyte maturation; BPTMC-induced downregulation of NEK3 causes reduced microtubule acetylation and abnormal spindle assembly, impaired polar body extrusion, and defective embryonic development. In vitro oocyte maturation, NEK3 protein level analysis, immunofluorescence of spindle/microtubule acetylation, fertilization and embryo development assay Environmental science & technology Low 42235065

Source papers

Stage 0 corpus · 13 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2007 Nek3 kinase regulates prolactin-mediated cytoskeletal reorganization and motility of breast cancer cells. Oncogene 68 17297458
2004 Novel association of Vav2 and Nek3 modulates signaling through the human prolactin receptor. Molecular endocrinology (Baltimore, Md.) 61 15618286
2009 The NIMA-family kinase Nek3 regulates microtubule acetylation in neurons. Journal of cell science 59 19509051
1999 Cloning and characterization of the murine Nek3 protein kinase, a novel member of the NIMA family of putative cell cycle regulators. The Journal of biological chemistry 31 10224116
1999 NIMA-related kinases: isolation and characterization of murine nek3 and nek4 cDNAs, and chromosomal localization of nek1, nek2 and nek3. Gene 25 10393247
2016 Identification of NEK3 Kinase Threonine 165 as a Novel Regulatory Phosphorylation Site That Modulates Focal Adhesion Remodeling Necessary for Breast Cancer Cell Migration. The Journal of biological chemistry 23 27489110
2020 Biallelic loss of function NEK3 mutations deacetylate α-tubulin and downregulate NUP205 that predispose individuals to cilia-related abnormal cardiac left-right patterning. Cell death & disease 16 33230144
2001 Molecular cloning and characterization of the human NIMA-related protein kinase 3 gene (NEK3). Cytogenetics and cell genetics 11 12063396
2018 NEK3-mediated SNAP29 phosphorylation modulates its membrane association and SNARE fusion dependent processes. Biochemical and biophysical research communications 8 29454964
2006 Is there any association between nek3 and cancers with frequent 13q14 deletion? Cancer investigation 8 17118778
2026 Multi-omics profiling reveals that targeting NEK3 synergizes with cuproptosis to promote M1 macrophage polarization in cervical cancer. International immunopharmacology 0 41483620
2026 Bisphenol TMC Impaired Mouse Oocyte Maturation by Disrupting Spindle Assembly via NEK3-Mediated Microtubule Acetylation. Environmental science & technology 0 42235065
2019 Identification of NEK3 and MOK as novel targets for lithium. Chemical biology & drug design 0 30667602

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