Affinage

NEK4

Serine/threonine-protein kinase Nek4 · UniProt P51957

Length
841 aa
Mass
94.6 kDa
Annotated
2026-06-10
17 papers in source corpus 10 papers cited in narrative 10 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

NEK4 is a serine/threonine kinase that operates at several cellular control points spanning ciliogenesis, the DNA damage response, microtubule and cell-cycle regulation, and mitochondrial dynamics (PMID:21685204, PMID:22851694, PMID:20103636, PMID:34986513). At the cilium, NEK4 is a stable component of the RPGRIP1- and RPGRIP1L-associated complexes, localizes to basal bodies and the ciliary rootlet, and is required for efficient cilium assembly (PMID:21685204). In the genotoxic stress response, NEK4 assembles with the DNA-PKcs/Ku70/Ku80 complex and promotes recruitment of DNA-PKcs to double-strand breaks, supporting p53 activation, H2AX phosphorylation, and entry into replicative senescence; its loss attenuates these responses (PMID:22851694, PMID:41001016). NEK4 also promotes microtubule outgrowth and Taxol-induced G2/M arrest while sensitizing cells to microtubule-destabilizing drugs upon depletion (PMID:20103636), and it regulates mitochondrial respiration and fission through phosphorylation of DRP1, with DRP1 inhibition abolishing its respiratory effects (PMID:34986513). Additional roles include positive regulation of EMT through Zeb1/Smad signaling in lung adenocarcinoma (PMID:30247800), isoform-specific engagement of RNA splicing machinery (PMID:25798074), and modulation of synaptic structure and circadian phosphoproteomic profiles upon neuronal overexpression (PMID:39448584). Several of NEK4's cell-cycle and proliferative phenotypes are context-dependent, differing across cancer cell lines (PMID:41001016).

Mechanistic history

Synthesis pass · year-by-year structured walk · 9 steps
  1. 2010 Medium

    Established the first cellular role for NEK4 by showing it governs microtubule dynamics and the cell-cycle checkpoint response to microtubule poisons, distinguishing stabilizing from destabilizing drug responses.

    Evidence RNAi loss-of-function genetic screen with microtubule outgrowth, G2/M arrest, and mitotic aster assays after Taxol/vincristine

    PMID:20103636

    Open questions at the time
    • No direct substrate on microtubules or the mitotic apparatus identified
    • Kinase-dependence of the phenotype not established
  2. 2011 Medium

    Placed NEK4 at the basal body/ciliary rootlet as a partner of ciliopathy proteins RPGRIP1/RPGRIP1L required for cilium assembly, linking the kinase to ciliogenesis.

    Evidence Tandem affinity purification/MS complex identification, immunolocalization in ciliated cells/organs, and siRNA knockdown with cilium assembly readout

    PMID:21685204

    Open questions at the time
    • No ciliary substrate of NEK4 identified
    • Mechanism by which NEK4 promotes assembly unresolved
    • Requirement for kinase activity untested
  3. 2012 High

    Defined NEK4 as a DNA damage response factor by showing it forms a complex with DNA-PKcs/Ku70/Ku80 and is needed for DNA-PKcs recruitment to double-strand breaks, p53 activation, and timely replicative senescence.

    Evidence RNAi screen, Co-IP/MS of NEK4 immune complexes, senescence and cell-cycle assays, and γH2AX/p53 phosphorylation assays after DSB induction

    PMID:22851694

    Open questions at the time
    • Whether NEK4 phosphorylates DNA-PKcs or Ku subunits not determined
    • Direct recruitment mechanism not structurally resolved
  4. 2015 Medium

    Mapped the NEK4 interactome and revealed isoform-specific functions, showing isoform 1 engages splicing factors and RNA processing whereas the two isoforms can act oppositely.

    Evidence IP-MS of two NEK4 isoforms in human cells with functional assays of splicing differences

    PMID:25798074

    Open questions at the time
    • Direct phosphorylation of splicing factors not demonstrated
    • Functional consequence of most interactors untested
    • Structural basis of isoform divergence unknown
  5. 2018 Medium

    Identified NEK4 as a positive regulator of EMT and metastasis acting upstream of Zeb1/Smad signaling in lung adenocarcinoma.

    Evidence siRNA reporter screen, knockdown/overexpression Western blot, migration/invasion assays, and in vivo tail-vein metastasis model

    PMID:30247800

    Open questions at the time
    • Direct substrate linking NEK4 to Zeb1/Smad not identified
    • Whether effect is kinase-dependent untested
  6. 2022 Medium

    Connected NEK4 to mitochondrial physiology, showing it drives respiration and fission via DRP1 phosphorylation, with DRP1 inhibition epistatically blocking the respiratory effect.

    Evidence Overexpression/siRNA with Seahorse respiration, membrane potential and mtDNA assays, DRP1/Erk1/2 phospho-Western blots, and DRP1 inhibitor epistasis

    PMID:34986513

    Open questions at the time
    • Direct phosphorylation of DRP1 by NEK4 not shown in vitro
    • Relationship to Erk1/2 phosphorylation not ordered
  7. 2024 Medium

    Demonstrated an in vivo neuronal role, with NEK4 overexpression altering circadian behavior, synaptic structure, and diurnal phosphoproteomes, rescuable by lithium.

    Evidence Conditional transgenic and AAV overexpression in pyramidal neurons, anxiety behavior assays, confocal spine imaging, and phosphoproteomics with lithium treatment

    PMID:39448584

    Open questions at the time
    • Direct neuronal substrates not identified
    • Loss-of-function phenotype not tested
    • Mechanism of lithium rescue unresolved
  8. 2025 Medium

    Showed NEK4's cell-cycle and proliferative roles are context-dependent, with depletion in TNBC cells stimulating proliferation and shifting toward G2/M while still impairing the etoposide-induced DNA damage response.

    Evidence siRNA depletion with live-cell proliferation imaging, RNA-seq enrichment, flow-cytometry cell-cycle analysis, and γH2AX foci after etoposide

    PMID:41001016

    Open questions at the time
    • Basis of cell-line-dependent opposite phenotypes unexplained
    • p21 regulation inconsistent across contexts
  9. 2026 Low

    Positioned NEK4 downstream of TXNDC15 in G2/M checkpoint control through a physical interaction in lung adenocarcinoma.

    Evidence Co-immunoprecipitation of TXNDC15-NEK4 with proliferation and cell-cycle distribution assays

    PMID:42214920

    Open questions at the time
    • Single Co-IP without reciprocal validation
    • No direction of signaling established beyond correlation
    • Kinase-dependence untested

Open questions

Synthesis pass · forward-looking unresolved questions
  • The direct phosphorylation substrates of NEK4 across its many proposed roles remain undefined, and whether each phenotype requires its kinase activity is unresolved.
  • No validated in vitro NEK4 substrate reported
  • No structural model of NEK4 in any complex
  • Reconciliation of cilia, DNA-repair, mitochondrial, EMT and neuronal roles into a unifying mechanism absent

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016740 transferase activity 2 GO:0140096 catalytic activity, acting on a protein 1
Localization
GO:0005739 mitochondrion 1 GO:0005815 microtubule organizing center 1 GO:0005929 cilium 1
Pathway
R-HSA-1640170 Cell Cycle 2 R-HSA-73894 DNA Repair 2 R-HSA-1852241 Organelle biogenesis and maintenance 1
Partners
DNM1LPRKDCRPGRIP1RPGRIP1LSRSF1TXNDC15XRCC5XRCC6
Complex memberships
DNA-PKcs/Ku70/Ku80 complexRPGRIP1/RPGRIP1L-associated complex

Evidence

Reading pass · 10 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2011 NEK4 serine/threonine kinase was identified as a prominent component of both the RPGRIP1- and RPGRIP1L-associated protein complexes by tandem affinity purification/mass spectrometry. In ciliated cells, NEK4 localizes to basal bodies, and in ciliated organs it is detected at the ciliary rootlet. Knockdown of NEK4 in ciliated cells caused a significant decrease in cilium assembly, establishing a role for NEK4 in cilium dynamics. Tandem affinity purification + mass spectrometry (complex identification); immunolocalization in ciliated cells/organs; siRNA knockdown with cilium assembly readout Human molecular genetics Medium 21685204
2012 NEK4 suppression extended replicative lifespan in human fibroblasts and reduced p21 transcription. NEK4 forms a complex with DNA-PKcs, Ku70, and Ku80 (identified by MS of immune complexes). NEK4 suppression impaired DNA-PKcs recruitment to double-strand breaks, resulting in reduced p53 activation and H2AX phosphorylation, thereby implicating NEK4 in the DNA damage response pathway. RNAi loss-of-function screen; mass spectrometric analysis of Nek4 immune complexes; co-immunoprecipitation; cell cycle and senescence assays; γH2AX and p53 phosphorylation assays after DSB induction Molecular and cellular biology High 22851694
2010 NEK4 promotes microtubule outgrowth after Taxol treatment and is required for G2/M arrest and mitotic aster formation. Nek4 deficiency impaired G2/M arrest in response to the microtubule-stabilizing drug Taxol but sensitized cells to the microtubule-destabilizing drug vincristine, establishing a role for NEK4 in microtubule dynamics and differential response to microtubule poisons. Genetic screen (RNAi/loss-of-function); microtubule outgrowth assays; cell cycle analysis (G2/M arrest); mitotic aster formation assays after drug treatment Cancer research Medium 20103636
2015 Immunoprecipitation-mass spectrometry of two NEK4 isoforms identified 474 interacting proteins for isoform 1 and 149 for isoform 2, with ~68% overlap. Key interactors include DNA-PKcs complex members (consistent with DNA damage role), PCNA, 14-3-3ε, splicing factors SRSF1, SRSF2, SRPK1, and hnRNP proteins. Isoform 1 (but not isoform 2) participates in RNA splicing control, and the two isoforms showed opposite effects in some processes, indicating isoform-specific functional differences. Immunoprecipitation followed by mass spectrometry of two NEK4 isoforms expressed in human cells; functional assays distinguishing isoform effects on RNA splicing Proteome science Medium 25798074
2018 NEK4 acts as a positive regulator of EMT in lung adenocarcinoma cells. NEK4 knockdown increased E-cadherin and ZO1 expression, decreased Zeb1 and Smad proteins, and inhibited cell migration/invasion. NEK4 overexpression decreased E-cadherin and increased Smad3 expression. In vivo tail-vein injection of NEK4-knockdown cells showed reduced tumor formation and metastasis, placing NEK4 upstream of Zeb1 and Smad signaling in EMT. siRNA library screen with E-cadherin promoter-luciferase reporter; siRNA knockdown and overexpression with Western blot; migration/invasion assays; in vivo tail-vein injection mouse model Journal of cellular and molecular medicine Medium 30247800
2022 NEK4 overexpression activates mitochondrial respiration coupled to ATP production, increases mitochondrial membrane potential, and confers resistance to mitochondrial DNA damage. NEK4 depletion reduced mitochondrial respiration and mtDNA integrity. NEK4 regulates mitochondrial morphology through the fission protein DRP1: NEK4 overexpression increased DRP1 phosphorylation and Erk1/2 phosphorylation and promoted mitochondrial fission, while NEK4 deficiency caused mitochondrial elongation. The mitochondrial respiration effect of NEK4 overexpression was abolished by a DRP1 inhibitor. NEK4 overexpression and siRNA depletion; mitochondrial respiration assays (Seahorse); mitochondrial membrane potential assays; mtDNA damage assays; DRP1 and Erk1/2 phosphorylation by Western blot; DRP1 inhibitor epistasis The FEBS journal Medium 34986513
2021 Novel quinazolinamine analogues derived from Spautin-1 were identified as potent NEK4 inhibitors with IC50 ~1 µM, demonstrating that NEK4 kinase activity can be pharmacologically inhibited and that its inhibition is at least partially responsible for anti-proliferative activity in EGFR-mutant NSCLC cells. Structure-activity relationship study; kinase inhibition assays (IC50 determination); anti-proliferative cell assays International journal of molecular sciences Low 33435251
2024 NEK4 overexpression in pyramidal neurons (transgenic mice or AAV) caused circadian shifts in anxiety behavior and affected diurnal fluctuations in dendritic spine morphology and synaptic structure. Phosphoproteomic analyses revealed that NEK4 overexpression alters diurnal and nocturnal phosphoproteomic profiles. Lithium treatment ameliorated the behavioral effects of NEK4 overexpression. Conditional transgenic NEK4 overexpression in forebrain pyramidal neurons; AAV-mediated hippocampal overexpression; behavioral assays (anxiety); confocal imaging of dendritic spines; phosphoproteomic analysis; lithium treatment Nature communications Medium 39448584
2025 NEK4 depletion in triple-negative breast cancer (BT20) cells upregulated cell cycle and mitosis genes and downregulated mRNA splicing genes (by RNA-seq), shifted cell cycle distribution toward G2/M, and stimulated proliferation — opposite to effects in other cell lines. NEK4 depletion decreased γH2AX foci after etoposide treatment but did not affect basal DNA damage levels, confirming a role in DNA damage response. NEK4 depletion did not consistently alter p21 expression across cell lines. NEK4 siRNA depletion; live-cell imaging of proliferation; RNA-seq with enrichment analysis; cell cycle distribution by flow cytometry; γH2AX foci after etoposide Frontiers in oncology Medium 41001016
2026 TXNDC15 interacts with NEK4 and suppresses lung adenocarcinoma cell proliferation by inducing G2/M phase arrest, placing NEK4 downstream of TXNDC15 in G2/M checkpoint regulation. Co-immunoprecipitation (TXNDC15-NEK4 interaction); functional assays (proliferation, cell cycle distribution); G2/M checkpoint readouts Biochemical and biophysical research communications Low 42214920

Source papers

Stage 0 corpus · 17 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2011 The ciliopathy-associated protein homologs RPGRIP1 and RPGRIP1L are linked to cilium integrity through interaction with Nek4 serine/threonine kinase. Human molecular genetics 58 21685204
2012 Nek4 regulates entry into replicative senescence and the response to DNA damage in human fibroblasts. Molecular and cellular biology 47 22851694
2018 NEK4 kinase regulates EMT to promote lung cancer metastasis. Journal of cellular and molecular medicine 34 30247800
2010 Nek4 status differentially alters sensitivity to distinct microtubule poisons. Cancer research 33 20103636
2015 New interaction partners for Nek4.1 and Nek4.2 isoforms: from the DNA damage response to RNA splicing. Proteome science 25 25798074
1999 NIMA-related kinases: isolation and characterization of murine nek3 and nek4 cDNAs, and chromosomal localization of nek1, nek2 and nek3. Gene 25 10393247
2022 Nek4 regulates mitochondrial respiration and morphology. The FEBS journal 16 34986513
2021 Structure-Activity Relationship (SAR) Study of Spautin-1 to Entail the Discovery of Novel NEK4 Inhibitors. International journal of molecular sciences 8 33435251
2024 Unraveling NEK4 as a Potential Drug Target in Schizophrenia and Bipolar I Disorder: A Proteomic and Genomic Approach. Schizophrenia bulletin 7 38869147
2022 Anesthetic propofol suppresses growth and metastasis of lung adenocarcinoma in vitro through downregulating circ-MEMO1-miR-485-3p-NEK4 ceRNA axis. Histology and histopathology 6 35521898
2024 NEK4 modulates circadian fluctuations of emotional behaviors and synaptogenesis in male mice. Nature communications 3 39448584
2025 NEK4 suppresses cell proliferation in BT20 triple-negative breast cancer cells by diminishing expression of cell cycle genes, while its depletion mitigates proliferation in other cell lines. Frontiers in oncology 1 41001016
2026 A divergent Plasmodium NEK4 acts as a key regulator driving the early events of meiosis. Nature communications 0 42120403
2026 TXNDC15 suppresses lung adenocarcinoma progression by inducing NEK4-dependent G2/M entry blockade. Biochemical and biophysical research communications 0 42214920
2025 NEK4: prediction of available drug targets and common genetic linkages in bipolar disorder and major depressive disorder. Frontiers in psychiatry 0 39950180
2025 A divergent Plasmodium NEK4 acts as a key regulator driving the early events of meiosis. bioRxiv : the preprint server for biology 0 41332573
2025 A divergent Plasmodium NEK4 acts as a key regulator driving the early events of meiosis. Research square 0 41510253

Missed literature

Know a paper Affinage missed for NEK4? Flag it for the maintainers and the community.

No submissions yet.