Affinage

KLHL9

Kelch-like protein 9 · UniProt Q9P2J3

Length
617 aa
Mass
69.4 kDa
Annotated
2026-06-10
18 papers in source corpus 11 papers cited in narrative 10 extracted findings
Cross-family judge vs UniProt: tie faithfulness: 6/6 claims corpus-supported (100%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

KLHL9 is a substrate-recognition adaptor for CUL3-RBX1-based E3 ubiquitin ligase complexes, capturing substrates through its Kelch domain and frequently acting together with the paralogous adaptor KLHL13 (PMID:17543862). Its founding role is in mitosis, where CUL3/KLHL9/KLHL13 directly binds and ubiquitylates Aurora B to translocate the chromosomal passenger complex from chromosomes to the central spindle during anaphase, an activity required for chromosome alignment and completion of cytokinesis (PMID:17543862, PMID:19995937); this ligase is also required to sustain spindle assembly checkpoint signaling, as its loss causes premature Cyclin B degradation and mitotic exit under microtubule poisons (PMID:18075312). Beyond mitosis, KLHL9 directs CUL3-mediated proteasomal turnover of distinct substrates in several contexts: it targets insulin receptor substrate-1 (IRS1), linking autophagy deficiency to insulin resistance (PMID:31515271), and ubiquitylates Rheb at the lysosome upon amino acid stimulation, where the complex translocates to support mTORC1 activation (PMID:39708321). KLHL9 loss de-represses the C/EBPβ/C/EBPδ master regulators that drive the glioblastoma mesenchymal signature, and restoring KLHL9 triggers proteasomal degradation of these factors and reduces tumor viability (PMID:25303533). The bacterial effector BipD of Burkholderia pseudomallei binds the BACK and Kelch domains of KLHL9 to hijack the complex toward K63-linked ubiquitination of the inner mitochondrial membrane protein IMMT, initiating mitophagy that supports bacterial survival (PMID:38834545, PMID:39265641). KLHL9 substrate recognition operates through specific degron motifs definable at scale (PMID:37735597).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 2007 High

    Established that KLHL9 is a bona fide CUL3 substrate adaptor by identifying its first substrate, defining how a Kelch-domain adaptor confers specificity to a cullin-RING ligase in mitosis.

    Evidence In vitro Kelch-domain binding, reciprocal Co-IP, in vivo and reconstituted in vitro ubiquitylation of Aurora B, RNAi mitotic phenotyping

    PMID:17543862

    Open questions at the time
    • No structure of the KLHL9 Kelch-substrate interface
    • Relative contributions of KLHL9 vs KLHL13 to substrate capture not separated
  2. 2007 Medium

    Connected the ligase to checkpoint control, showing the complex is needed to maintain SAC signaling rather than only to relocalize the CPC.

    Evidence RNAi inactivation of CUL3/KLHL9/KLHL13 with Cyclin B and mitotic-exit readouts under microtubule poisons

    PMID:18075312

    Open questions at the time
    • Direct SAC substrate of the complex not identified
    • Single method, no reconstitution
  3. 2009 Medium

    Independent replication refined the mitotic role to CPC translocation from chromosomes to spindle midzone and distinguished it from the related KLHL21 adaptor.

    Evidence RNAi knockdown with immunofluorescence of CPC localization and cytokinesis, compared with KLHL21

    PMID:19995937

    Open questions at the time
    • Molecular trigger for midzone translocation not defined
  4. 2014 Medium

    Extended KLHL9 function to tumor biology, positioning it as a tumor suppressor whose loss de-represses C/EBP master regulators in glioblastoma.

    Evidence KLHL9 rescue with proteasome inhibition, in vitro/in vivo tumor viability, regulatory network analysis

    PMID:25303533

    Open questions at the time
    • Direct ubiquitylation of C/EBPβ/δ by CUL3/KLHL9 not demonstrated
    • Whether C/EBP proteins bind the Kelch domain unknown
  5. 2017 Medium

    Clarified upstream regulation, showing CUL3KLHL9/13 activity is independent of SCCRO-mediated neddylation that controls CUL3KLHL21, evidence for selective cullin regulation.

    Evidence SCCRO knockdown/knockout with CUL3KLHL9/13 activity as internal control versus CUL3KLHL21

    PMID:28620047

    Open questions at the time
    • How CUL3KLHL9/13 is neddylated/activated not defined
    • Primarily a contrast result for KLHL9
  6. 2019 Medium

    Identified a metabolic substrate (IRS1), linking the ligase to autophagy-dependent insulin resistance.

    Evidence BioID and Co-IP interaction mapping, siRNA knockdown with IRS1 protein and insulin-signaling readouts, proteasome inhibition

    PMID:31515271

    Open questions at the time
    • Degron on IRS1 not mapped
    • Direct reconstituted ubiquitylation of IRS1 not shown
  7. 2023 Medium

    Provided systematic substrate/degron logic, showing KLHL9/KLHL13 recognize defined degron sequences at scale.

    Evidence Multiplex CRISPR screening with site-saturation degron mutagenesis on substrate pools

    PMID:37735597

    Open questions at the time
    • KLHL9-specific substrates from the screen not individually detailed
    • Physiological relevance of screened substrates untested
  8. 2023 Low

    Placed KLHL9 in cell-intrinsic antiviral immunity, showing it is required for IFNγ-mediated restriction of norovirus.

    Evidence Klhl9 and Cul3 knockdown/knockout with IFNγ-induced norovirus replication readout in mouse cells

    PMID:37905813

    Open questions at the time
    • No molecular mechanism or substrate established for the antiviral role
    • Single lab, loss-of-function only
  9. 2024 High

    Defined KLHL9 as the essential adaptor for lysosomal Rheb ubiquitination, integrating it into amino-acid-induced mTORC1 activation.

    Evidence CUL3/RBX1/KLHL9 KO lines, Rheb ubiquitination assay, S6K-based mTORC1 readout, lysosomal translocation imaging, Akt negative control

    PMID:39708321

    Open questions at the time
    • Rheb degron/Kelch contacts not structurally defined
    • Mechanism of amino-acid-induced lysosomal translocation unknown
  10. 2024 Medium

    Revealed pathogen hijacking of the complex, mapping a bacterial effector to KLHL9 domains and redirecting it to drive IMMT-dependent mitophagy.

    Evidence Domain-mapping Co-IP of BipD to BACK/Kelch domains, host ubiquitome profiling, K211 site-specific ubiquitination, mitophagy and mtROS assays, bacterial survival

    PMID:38834545 PMID:39265641

    Open questions at the time
    • Whether IMMT is a physiological KLHL9 substrate absent infection unknown
    • Structural basis of BipD-mediated substrate redirection not resolved

Open questions

Synthesis pass · forward-looking unresolved questions
  • The structural basis of Kelch-domain substrate selection and how KLHL9 versus KLHL13 partition substrates across these diverse pathways remain unresolved.
  • No co-crystal/cryo-EM of KLHL9 with any substrate
  • Tissue/context determinants of substrate choice undefined

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0060090 molecular adaptor activity 3 GO:0140096 catalytic activity, acting on a protein 3 GO:0016874 ligase activity 2
Localization
GO:0005764 lysosome 1
Pathway
R-HSA-1640170 Cell Cycle 3 R-HSA-392499 Metabolism of proteins 3 R-HSA-162582 Signal Transduction 2
Partners
AURKBBIPDCUL3IMMTIRS1KLHL13RBX1RHEB
Complex memberships
CUL3-RBX1-KLHL9/KLHL13 E3 ubiquitin ligase

Evidence

Reading pass · 10 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2007 KLHL9 and KLHL13 function as substrate-specific adaptors for a CUL3-based E3 ubiquitin ligase complex. Aurora B directly binds to the substrate-recognition (Kelch) domain of KLHL9 in vitro, coimmunoprecipitates with the CUL3 complex during mitosis, and is ubiquitylated in a CUL3-dependent manner in vivo and by reconstituted CUL3/KLHL9/KLHL13 ligase in vitro. This complex removes Aurora B (chromosomal passenger complex) from mitotic chromosomes, allowing accumulation on the central spindle during anaphase, and is required for correct chromosome alignment and completion of cytokinesis. In vitro binding assay (Aurora B binds KLHL9 Kelch domain), co-immunoprecipitation, in vivo ubiquitylation assay, reconstituted in vitro ubiquitylation with purified CUL3/KLHL9/KLHL13, RNAi loss-of-function with mitotic phenotype readout Developmental cell High 17543862
2007 Inactivation of the CUL3/KLHL9/KLHL13 ligase leads to premature degradation of Cyclin B and exit from the mitotic state in the presence of microtubule poisons, indicating this E3 ligase is required to maintain spindle assembly checkpoint (SAC) signaling in human cells. RNAi-mediated inactivation of CUL3/KLHL9/KLHL13 complex with readout of Cyclin B levels and mitotic exit under microtubule poison treatment Cell cycle (Georgetown, Tex.) Medium 18075312
2009 KLHL9 and KLHL13 regulate translocation of the chromosomal passenger complex (CPC) from chromosomes to the spindle midzone in anaphase, a function distinct from KLHL21 which localizes to midzone microtubules. KLHL9/KLHL13 are required for cytokinesis. RNAi knockdown of KLHL9/KLHL13 with immunofluorescence readout of CPC localization and cytokinesis completion; comparison with KLHL21 knockdown The Journal of cell biology Medium 19995937
2014 KLHL9 deletion acts as an upstream activator of C/EBPβ and C/EBPδ master regulators in glioblastoma mesenchymal subtype. Rescue of KLHL9 expression induced proteasomal degradation of C/EBP proteins, abrogated the mesenchymal gene expression signature, and reduced tumor viability in vitro and in vivo. KLHL9 rescue expression in KLHL9-deleted glioblastoma cells, proteasome inhibition assay, in vitro and in vivo tumor viability assays, regulatory network analysis Cell Medium 25303533
2017 The activity of the CUL3/KLHL9/KLHL13 complex during mitosis was intact in SCCRO-deficient cells, demonstrating that SCCRO selectively (rather than globally) neddylates cullins and that KLHL9/KLHL13-containing CUL3 complexes are regulated differently from CUL3KLHL21 during abscission. SCCRO knockdown/knockout with biochemical assay of CUL3KLHL9/KLHL13 activity as internal control compared with CUL3KLHL21 The Journal of biological chemistry Medium 28620047
2019 KLHL9 and KLHL13, in complex with CUL3, serve as an E3 ubiquitin ligase that targets insulin receptor substrate-1 (IRS1) for proteasomal degradation. KLHL9 and KLHL13 were identified as novel IRS1 interactors by proximity-dependent biotin identification (BioID) and co-immunoprecipitation. siRNA-mediated knockdown of Klhl9, Klhl13, or Cul3 recovered IRS1 expression, and knockdown of Klhl13 and Cul3 increased insulin signaling. Elevated expression of Klhl9 and Klhl13 in ATG16L1-deficient cells mediates IRS1 degradation and insulin resistance. BioID proximity labeling, co-immunoprecipitation, siRNA knockdown with IRS1 protein level and insulin signaling readouts, proteasome inhibition assay The Journal of biological chemistry Medium 31515271
2023 Multiplex CRISPR screening identified substrates and degron motifs for the CUL3KLHL9/13 E3 ligase complex, demonstrating that KLHL9/KLHL13 together recognize specific degron sequences on substrate proteins at scale. Multiplex CRISPR screening platform (~100 simultaneous screens), site-saturation mutagenesis of degron motifs, full-length substrate pools Nature cell biology Medium 37735597
2023 Klhl9 (together with Cul3) is required for interferon-γ-induced inhibition of murine norovirus replication in mouse cells, placing the CUL3/KLHL9 complex in an IFNγ-dependent cell-intrinsic antiviral pathway. Genetic knockdown/knockout of Klhl9 and Cul3 with readout of IFNγ-induced norovirus replication inhibition mBio Low 37905813
2024 The CUL3-RBX1-KLHL9 E3 ubiquitin ligase complex ubiquitinates Rheb on the lysosome in response to amino acid stimulation, supporting mTORC1 activation. KLHL9 serves as the essential adaptor for CUL3-RBX1 to target Rheb. Deletion of CUL3, RBX1, or KLHL9 diminishes Rheb ubiquitination and reduces amino acid-induced mTORC1 activation without affecting lysosomal mTORC1 localization or Akt activity. The complex translocates to the lysosome upon amino acid stimulation. Genetic deletion (CUL3, RBX1, KLHL9 KO), ubiquitination assay of Rheb, mTORC1 activity assay (S6K phosphorylation), lysosomal localization by fractionation/imaging, Akt activity as negative control Cell reports High 39708321
2024 The bacterial effector BipD of Burkholderia pseudomallei binds to the BACK and Kelch domains of KLHL9 (and KLHL13), hijacking the KLHL9-KLHL13-CUL3 E3 ligase complex to promote K63-linked ubiquitination of the inner mitochondrial membrane protein IMMT at K211, initiating mitophagy and reducing mitochondrial ROS to support bacterial intracellular survival. Co-IP/interaction mapping (BipD binding to BACK and Kelch domains of KLHL9), host ubiquitome profiling identifying IMMT as substrate, site-specific ubiquitination (K211) determination, mitophagy assay, mtROS measurement, bacterial survival assay Nature communications Medium 38834545 39265641

Source papers

Stage 0 corpus · 18 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2013 Update on the Kelch-like (KLHL) gene family. Human genomics 213 23676014
2007 A Cul3-based E3 ligase removes Aurora B from mitotic chromosomes, regulating mitotic progression and completion of cytokinesis in human cells. Developmental cell 196 17543862
2014 Identification of causal genetic drivers of human disease through systems-level analysis of regulatory networks. Cell 139 25303533
2009 The Cul3-KLHL21 E3 ubiquitin ligase targets aurora B to midzone microtubules in anaphase and is required for cytokinesis. The Journal of cell biology 118 19995937
2023 Defining E3 ligase-substrate relationships through multiplex CRISPR screening. Nature cell biology 53 37735597
2018 Understanding the regulatory mechanisms of milk production using integrative transcriptomic and proteomic analyses: improving inefficient utilization of crop by-products as forage in dairy industry. BMC genomics 44 29843597
2019 Deficiency of the autophagy gene ATG16L1 induces insulin resistance through KLHL9/KLHL13/CUL3-mediated IRS1 degradation. The Journal of biological chemistry 34 31515271
2008 The contribution of large genomic deletions at the CDKN2A locus to the burden of familial melanoma. British journal of cancer 24 18612309
2007 A Cul3-based E3 ligase regulates mitosis and is required to maintain the spindle assembly checkpoint in human cells. Cell cycle (Georgetown, Tex.) 20 18075312
2023 Autophagy gene-dependent intracellular immunity triggered by interferon-γ. mBio 15 37905813
2017 Squamous cell carcinoma-related oncogene (SCCRO) neddylates Cul3 protein to selectively promote midbody localization and activity of Cul3KLHL21 protein complex during abscission. The Journal of biological chemistry 15 28620047
2014 Homozygous deletions at 3p22, 5p14, 6q15, and 9p21 result in aberrant expression of tumor suppressor genes in gastric cancer. Genes, chromosomes & cancer 15 25521327
2024 Burkholderia pseudomallei BipD modulates host mitophagy to evade killing. Nature communications 13 38834545
2019 Identification of Dysregulated Competitive Endogenous RNA Networks Driven by Copy Number Variations in Malignant Gliomas. Frontiers in genetics 9 31719831
2022 Significance of Chr9p22.1-p21.3 Deletion in Cancer Development: A Pan-cancer In Silico Analysis. Anticancer research 8 36288884
2024 The Cullin3-Rbx1-KLHL9 E3 ubiquitin ligase complex ubiquitinates Rheb and supports amino acid-induced mTORC1 activation. Cell reports 3 39708321
2024 Burkholderia pseudomallei BipD initiates mitophagy to evade killing by hijacking host KLHL9-KLHL13-CUL3 E3 ligase to ubiquitinate IMMT. Autophagy 1 39265641
2025 KLHL9-linked distal myopathy: a second family suggesting broad phenotypic variability. Neuromuscular disorders : NMD 0 40818927

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