Affinage

CBLL1

E3 ubiquitin-protein ligase Hakai · UniProt Q75N03

Length
491 aa
Mass
54.5 kDa
Annotated
2026-06-09
36 papers in source corpus 15 papers cited in narrative 17 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

CBLL1/Hakai is a RING-type E3 ubiquitin ligase that controls epithelial cell adhesion and motility by recognizing tyrosine-phosphorylated substrates and targeting them for degradation (PMID:11836526). It binds the E-cadherin complex in a phosphotyrosine-dependent manner, ubiquitinates the E-cadherin juxtamembrane domain at lysines K5 and K83 to block p120-catenin association, and drives E-cadherin endocytosis and lysosomal degradation, thereby disrupting cell-cell contacts and promoting epithelial-mesenchymal transition downstream of Slit2/Robo1 signaling (PMID:11836526, PMID:21283129, PMID:22693575). Substrate recognition is mediated by an atypical dimeric phosphotyrosine-binding (HYB) domain whose zinc-coordinated homodimerization is required to engage phosphotyrosine motifs of substrates such as E-cadherin, cortactin, and DOK1 (PMID:25074933). Beyond cadherin turnover, Hakai is an essential, mutually stabilized structural component of the m6A mRNA methyltransferase writer complex, where it associates with other writer subunits and is required for normal m6A deposition and downstream sex-specific alternative splicing (PMID:33846330). Hakai also acts independently of its ligase activity as an ERα corepressor that competes with coactivators SRC-1 and GRIP-1 (PMID:20608937) and stabilizes δ-catenin through Src (PMID:28069439), while extending its catalytic activity to additional substrates including FASN (lysosomal degradation, lipid regulation) (PMID:36266428) and the Runx2 stabilizer Smurf2 (promoting osteoblast differentiation) (PMID:39034451). Its own abundance is set post-transcriptionally by miR-203 (PMID:23285092) and by HSP90 chaperone-dependent stabilization (PMID:31952268).

Mechanistic history

Synthesis pass · year-by-year structured walk · 17 steps
  1. 2002 High

    Established Hakai as an E3 ligase that couples tyrosine phosphorylation of E-cadherin to its ubiquitination and endocytosis, providing a mechanism for dynamic regulation of cell-cell adhesion.

    Evidence Modified yeast two-hybrid, Co-IP, in vitro ubiquitination, and endocytosis assays in epithelial cells

    PMID:11836526

    Open questions at the time
    • Did not resolve the structural basis of phosphotyrosine recognition
    • Degradation route (lysosomal vs proteasomal) not fully dissected
  2. 2009 Medium

    Extended Hakai's reach beyond adhesion by linking it to post-transcriptional RNA regulation via the splicing factor PSF, hinting at a non-cadherin proliferative function.

    Evidence Co-IP, cDNA arrays, siRNA knockdown and BrdU proliferation epistasis

    PMID:19535458

    Open questions at the time
    • Direct vs indirect effect on RNA binding not separated
    • Single-lab finding
  3. 2009 Medium

    Drosophila genetics demonstrated essential in vivo roles for Hakai in epithelial integrity and cell specification that exceed simple E-cadherin ubiquitination.

    Evidence Null and maternal mutant analysis, S2-cell Co-IP, immunostaining

    PMID:19682089

    Open questions at the time
    • Molecular basis of the non-cadherin phenotypes unresolved
    • Relationship to later-defined m6A role not yet appreciated
  4. 2010 Medium

    Revealed a ligase-independent transcriptional role for Hakai as an ERα corepressor, showing it acts through protein-protein competition as well as catalysis.

    Evidence Co-IP with domain mapping, reporter assays, catalytically inactive mutant

    PMID:20608937

    Open questions at the time
    • Genomic occupancy of Hakai at ERα targets not mapped
    • Single-lab finding
  5. 2010 Medium

    Distinguished Hakai's requirement for bacterial endocytosis from flavivirus entry, defining the specificity of its membrane-trafficking contribution.

    Evidence Multiple siRNAs with bacterial internalization and viral infection assays

    PMID:21191016

    Open questions at the time
    • Substrate or trafficking step mediating Listeria entry not identified
  6. 2011 Medium

    Placed Hakai downstream of Slit2/Robo1 signaling as the effector routing E-cadherin to lysosomal degradation during EMT.

    Evidence siRNA knockdown rescue, recombinant ligand treatment, xenograft model

    PMID:21283129

    Open questions at the time
    • How Robo1 signaling recruits/activates Hakai mechanistically unknown
  7. 2012 Medium

    Mapped the specific E-cadherin lysines ubiquitinated by Hakai and showed ubiquitination directly competes with p120-catenin binding.

    Evidence Mitochondrial JMD targeting, site-directed mutagenesis, Co-IP, proteasome inhibition

    PMID:22693575

    Open questions at the time
    • Polyubiquitin chain topology not defined
  8. 2012 Medium

    Identified miR-203 as a direct post-transcriptional repressor of Hakai, defining an upstream control on its proliferative output.

    Evidence Luciferase 3'-UTR reporter, miRNA mimic/inhibitor, siRNA epistasis, BrdU assays

    PMID:23285092

    Open questions at the time
    • Physiological contexts where miR-203 regulates Hakai not established
  9. 2014 High

    Solved the structural basis of substrate recognition, showing the HYB domain must homodimerize via zinc coordination to bind phosphotyrosine motifs.

    Evidence NMR structure with ITC, AUC, SEC, DLS, and CD validation

    PMID:25074933

    Open questions at the time
    • Full-length ligase architecture and catalytic coupling not resolved
  10. 2017 Medium

    Showed Hakai can stabilize rather than degrade a partner (δ-catenin) by acting through Src, expanding its repertoire beyond destructive ubiquitination.

    Evidence Co-IP, catalytically inactive mutant, Src pharmacological inhibition

    PMID:28069439

    Open questions at the time
    • Whether Src is a direct Hakai target unclear
    • Single-lab finding
  11. 2018 Medium

    Demonstrated Hakai can route a substrate to neddylation rather than proteasomal ubiquitination, diversifying its modification outputs.

    Evidence Co-IP, domain mapping, MLN4924 vs MG132 inhibitor dissection

    PMID:30041665

    Open questions at the time
    • Mechanism by which Hakai promotes neddylation not defined
  12. 2021 High

    Established Hakai as an essential, mutually stabilizing subunit of the m6A writer complex, defining a non-ligase structural role in mRNA methylation and splicing control.

    Evidence Drosophila mutants, Co-IP of writer components, MeRIP-seq, splicing RT-PCR, behavioral assays

    PMID:33846330

    Open questions at the time
    • Precise structural contribution of Hakai within the writer complex unresolved
    • Generality to mammalian writer complex not directly shown here
  13. 2020 Medium

    Identified HSP90 as a chaperone that maintains Hakai stability, linking proteostasis to E-cadherin and motility outcomes.

    Evidence Co-IP, geldanamycin treatment, lysosome/proteasome inhibitor dissection, motility assays

    PMID:31952268

    Open questions at the time
    • Co-chaperones and recognition determinants not identified
  14. 2022 Medium

    Expanded the catalytic substrate set to FASN, connecting Hakai to lysosomal control of lipid metabolism in an inflammatory model.

    Evidence Interactome analysis, Co-IP, ubiquitination assay, lysosome inhibition, IHC in AOM/DSS mice

    PMID:36266428

    Open questions at the time
    • Ubiquitin linkage and physiological lipid impact not fully defined
  15. 2024 Medium

    Showed Hakai stabilizes Runx2 by degrading its antagonist Smurf2 in a catalysis-dependent manner, demonstrating indirect substrate stabilization driving osteoblast differentiation.

    Evidence Affinity proteomics, Co-IP, WT vs C109A mutant, shRNA, in vitro/in vivo differentiation (ovariectomized rat)

    PMID:39034451

    Open questions at the time
    • Direct Smurf2 ubiquitination by Hakai not shown biochemically
  16. 2025 Medium

    Positioned Hakai within a UBE3C-controlled axis governing m6A levels during cortical neurogenesis, linking its turnover to brain development.

    Evidence Proteomics of UBE3C-deficient mouse forebrain and human organoids, genetic complementation, METTL3-inhibitor rescue (preprint)

    Open questions at the time
    • Preprint not yet peer-reviewed
    • Direct UBE3C-mediated ubiquitination of Hakai not biochemically reconstituted
  17. 2025 Medium

    Implicated Hakai-mediated LRP4 degradation in Wnt/β-catenin hyperactivation and validated HYB-domain inhibition as a pharmacological strategy in colorectal cancer.

    Evidence Co-IP, ubiquitination assay, TOPFlash reporter, inducible shRNA, Hakin-1 HYB inhibitor (preprint)

    Open questions at the time
    • Preprint not yet peer-reviewed
    • LRP4 ubiquitin linkage and selectivity not defined

Open questions

Synthesis pass · forward-looking unresolved questions
  • How Hakai's distinct activities — catalytic phosphotyrosine-dependent ubiquitination versus its structural role in the m6A writer and its ligase-independent transcriptional/stabilizing functions — are partitioned and coordinated within the cell remains unresolved.
  • No structure of full-length Hakai engaged with the writer complex
  • Determinants selecting degradative vs stabilizing vs neddylation outcomes unknown
  • Mammalian counterparts of Drosophila m6A findings not directly demonstrated in the corpus

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0016874 ligase activity 4 GO:0140096 catalytic activity, acting on a protein 4 GO:0005198 structural molecule activity 1 GO:0140110 transcription regulator activity 1
Pathway
R-HSA-392499 Metabolism of proteins 3 R-HSA-1266738 Developmental Biology 2 R-HSA-162582 Signal Transduction 1 R-HSA-8953854 Metabolism of RNA 1
Partners
AJUBACDH1FASNLRP4RUNX2SRCUBE3C
Complex memberships
m6A mRNA writer complex

Evidence

Reading pass · 17 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2002 Hakai (CBLL1) is an E3 ubiquitin ligase that interacts with E-cadherin in a tyrosine phosphorylation-dependent manner, induces ubiquitination of the E-cadherin complex, and promotes its endocytosis, disrupting cell-cell contacts and enhancing cell motility. Hakai contains SH2, RING, zinc-finger and proline-rich domains. Modified yeast 2-hybrid, co-immunoprecipitation, ubiquitination assays, endocytosis assays, overexpression in epithelial cells Nature cell biology High 11836526
2009 Hakai interacts with PTB-associated splicing factor (PSF), an RNA-binding protein, and enhances PSF's ability to bind cancer-related mRNAs; knockdown of PSF suppresses Hakai-induced cell proliferation, placing Hakai upstream of PSF-mediated post-transcriptional regulation. Co-immunoprecipitation, cDNA arrays, siRNA knockdown, BrdU proliferation assays Molecular biology of the cell Medium 19535458
2009 Drosophila Hakai forms a complex with E-cadherin (Shotgun) in a manner distinct from the mammalian interaction; maternal Hakai mutants show stochastic loss of E-cadherin expression and reduction of aPKC, epithelial integrity defects, and defects in cell specification and migration, demonstrating essential in vivo roles beyond E-cadherin ubiquitination alone. Drosophila genetics (null mutants, maternal mutants), co-immunoprecipitation in S2 cells, immunostaining Genes to cells : devoted to molecular & cellular mechanisms Medium 19682089
2010 Hakai acts as a corepressor of estrogen receptor alpha (ERα) in breast cancer cells by binding directly to the DNA-binding domain of ERα and competing with coactivators SRC-1 and GRIP-1; this activity does not require Hakai's ubiquitin-ligase activity and results in inhibited ERα transcriptional activity, cell proliferation, and migration. Co-immunoprecipitation, reporter gene assays, domain mapping, overexpression/knockdown, proliferation assays Cancer science Medium 20608937
2011 Slit2/Robo1 signaling recruits Hakai to ubiquitinate E-cadherin, leading to its lysosomal degradation and EMT; knockdown of Hakai rescues E-cadherin levels and reverses EMT induced by Slit2/Robo1. siRNA knockdown, recombinant protein treatment, immunoblot, xenograft model Cell research Medium 21283129
2012 Ubiquitination of the E-cadherin juxtamembrane domain (JMD) at lysines K5 and K83 by Hakai inhibits p120-catenin binding and targets E-cadherin for degradation; mutation of these lysines stabilizes JMD and allows p120-catenin binding, establishing competitive regulation between ubiquitination and p120-catenin association. Mitochondrial targeting of JMD, site-directed mutagenesis, co-immunoprecipitation, proteasome inhibitor treatment, immunofluorescence PloS one Medium 22693575
2012 miR-203 post-transcriptionally represses Hakai expression by binding two sites in the Hakai mRNA 3'-UTR; inhibition of miR-203 elevates Hakai levels and increases cell proliferation in a Hakai-dependent manner. miRNA precursor/inhibitor overexpression, luciferase reporter assays, siRNA knockdown epistasis, BrdU incorporation PloS one Medium 23285092
2014 The Hakai phosphotyrosine-binding (HYB) domain forms an atypical zinc-coordinated tight homodimer essential for recognizing phosphotyrosine motifs of substrates including E-cadherin, cortactin, and DOK1; a C-terminal truncation mutant (HYB-ΔC) is monomeric but dimerizes upon phosphotyrosine substrate binding, demonstrating that the dimeric architecture is required for phosphotyrosine binding. NMR structure determination, isothermal titration calorimetry, analytical ultracentrifugation, size-exclusion chromatography, dynamic light scattering, circular dichroism The Journal of biological chemistry High 25074933
2017 Hakai stabilizes δ-catenin independently of its E3 ligase activity by stabilizing Src kinase; Src in turn inhibits binding between GSK-3β and δ-catenin, reducing proteasomal degradation of δ-catenin. Hakai and Src act synergistically to increase δ-catenin stability. Co-immunoprecipitation, overexpression/knockdown, immunoblot, pharmacological inhibition of Src Cellular signalling Medium 28069439
2018 Hakai interacts with Ajuba LIM domain protein via its HYB domain and induces Ajuba neddylation (not proteasomal ubiquitination), destabilizing Ajuba; this is blocked by neddylation inhibitor MLN4924 but not proteasome inhibitor MG132. Co-immunoprecipitation, confocal microscopy, siRNA knockdown, pharmacological inhibition (MLN4924, MG132), immunoblot Journal of experimental & clinical cancer research : CR Medium 30041665
2020 Hakai is a client protein of the HSP90 chaperone complex; pharmacological inhibition of Hsp90 with geldanamycin causes lysosome-dependent degradation of Hakai, accompanied by increased E-cadherin and Annexin A2 expression and suppressed cell motility. Co-immunoprecipitation, geldanamycin treatment, lysosome/proteasome inhibitor dissection, immunoblot, cell motility assays Cancers Medium 31952268
2021 Drosophila Hakai colocalizes and physically interacts with other m6A writer complex components (Vir, Fl(2)d, Flacc); Hakai mutants display reduced m6A levels in mRNA, aberrant Sxl alternative splicing, and wing/behavior defects. Disruption of Hakai, Vir, or Fl(2)d causes degradation of the other complex components, indicating mutual stabilization. m6A modification is deposited on Sxl mRNA in a sex-specific, writer-dependent manner. Drosophila genetics (Hakai mutants), co-immunoprecipitation, MeRIP-seq, RT-PCR for alternative splicing, behavioral assays Nature communications High 33846330
2022 Hakai interacts with Fatty Acid Synthase (FASN) via interactome analysis and induces FASN ubiquitination and lysosomal degradation, thereby regulating FASN-mediated lipid accumulation; inverse expression of FASN and Hakai was observed in an inflammatory AOM/DSS mouse model. Hakai interactome/proteomic analysis, co-immunoprecipitation, ubiquitination assay, lysosome inhibitor treatment, immunohistochemistry in mouse colitis model Scientific reports Medium 36266428
2024 Hakai physically interacts with Runx2 transcription factor and rescues it from Smurf2-mediated proteasomal degradation by inducing proteasome-dependent degradation of Smurf2 itself; catalytically inactive Hakai-C109A mutant has minimal effect, indicating dependence on E3 ligase activity. This promotes osteoblast differentiation. Affinity pulldown-based proteomics, co-immunoprecipitation, overexpression of wild-type vs. C109A mutant Hakai, shRNA knockdown, osteoblast differentiation assays in vitro and in vivo (ovariectomized rat model) Journal of cellular physiology Medium 39034451
2025 CBLL1/Hakai is identified as a substrate of UBE3C ubiquitin ligase; the UBE3C-Cbll1 axis drives N6-methyladenosine (m6A) mRNA methylation in neural progenitors, and hyperactivation of m6A writers in UBE3C-deficient forebrains impairs cell cycle exit during cortical neurogenesis. Proteomic profiling of UBE3C-deficient mouse forebrains and human brain organoids, genetic complementation, METTL3 inhibitor (STM2457) rescue in vivo bioRxivpreprint Medium
2025 Hakai interacts with LRP4 (a negative regulator of Wnt/β-catenin signaling), promotes LRP4 ubiquitination and degradation, and thereby induces hyperactivation of Wnt/β-catenin signaling; pharmacological inhibition of Hakai's HYB domain with Hakin-1 restores LRP4 levels and attenuates Wnt/β-catenin activity in colorectal cancer tumourspheres. Co-immunoprecipitation, ubiquitination assay, TOPFlash Wnt reporter assay, inducible shRNA knockdown, immunofluorescence, Western blot, RT-qPCR, Hakin-1 inhibitor treatment bioRxivpreprint Medium
2010 CBLL1 knockdown using specific siRNAs strongly inhibits endocytosis of Listeria monocytogenes, confirming a role for CBLL1 in bacterial invasion; however, the same CBLL1 knockdown does not impair WNV, dengue virus, or yellow fever virus infection, indicating CBLL1 is not required for flavivirus entry. siRNA knockdown, viral infection assays, bacterial internalization assay Journal of virology Medium 21191016

Source papers

Stage 0 corpus · 36 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2002 Hakai, a c-Cbl-like protein, ubiquitinates and induces endocytosis of the E-cadherin complex. Nature cell biology 700 11836526
2011 Slit-Robo signaling induces malignant transformation through Hakai-mediated E-cadherin degradation during colorectal epithelial cell carcinogenesis. Cell research 125 21283129
2010 Appraising the roles of CBLL1 and the ubiquitin/proteasome system for flavivirus entry and replication. Journal of virology 64 21191016
2009 Novel roles of hakai in cell proliferation and oncogenesis. Molecular biology of the cell 63 19535458
2021 Role of Hakai in m6A modification pathway in Drosophila. Nature communications 51 33846330
2020 Circ_0072083 interference enhances growth-inhibiting effects of cisplatin in non-small-cell lung cancer cells via miR-545-3p/CBLL1 axis. Cancer cell international 49 32190002
2018 Ajuba inhibits hepatocellular carcinoma cell growth via targeting of β-catenin and YAP signaling and is regulated by E3 ligase Hakai through neddylation. Journal of experimental & clinical cancer research : CR 45 30041665
2012 Competitive regulation of E-cadherin juxtamembrane domain degradation by p120-catenin binding and Hakai-mediated ubiquitination. PloS one 39 22693575
2012 miR-203 regulates cell proliferation through its influence on Hakai expression. PloS one 35 23285092
2023 Pepino mosaic virus antagonizes plant m6A modification by promoting the autophagic degradation of the m6A writer HAKAI. aBIOTECH 31 37581026
2009 Hacking RNA: Hakai promotes tumorigenesis by enhancing the RNA-binding function of PSF. Cell cycle (Georgetown, Tex.) 31 19855157
2009 Essential requirement for RING finger E3 ubiquitin ligase Hakai in early embryonic development of Drosophila. Genes to cells : devoted to molecular & cellular mechanisms 29 19682089
2019 CBLL1 is highly expressed in non-small cell lung cancer and promotes cell proliferation and invasion. Thoracic cancer 27 31124298
2018 Hakai overexpression effectively induces tumour progression and metastasis in vivo. Scientific reports 27 29472634
2020 Hakin-1, a New Specific Small-Molecule Inhibitor for the E3 Ubiquitin-Ligase Hakai, Inhibits Carcinoma Growth and Progression. Cancers 25 32456234
2011 Hakai reduces cell-substratum adhesion and increases epithelial cell invasion. BMC cancer 25 22051109
2019 Downregulation of long non-coding RNA XIST inhibits cell proliferation, migration, invasion and EMT by regulating miR-212-3p/CBLL1 axis in non-small cell lung cancer cells. European review for medical and pharmacological sciences 23 31646569
2010 Hakai acts as a coregulator of estrogen receptor alpha in breast cancer cells. Cancer science 23 20608937
2018 E3 ubiquitin ligase Hakai regulates cell growth and invasion, and increases the chemosensitivity to cisplatin in non‑small‑cell lung cancer cells. International journal of molecular medicine 20 29786107
2014 Dimeric switch of Hakai-truncated monomers during substrate recognition: insights from solution studies and NMR structure. The Journal of biological chemistry 15 25074933
2009 Expression of E-cadherin, Snail and Hakai in epithelial cells isolated from the primary tumor and from peritumoral tissue of invasive ductal breast carcinomas. Brazilian journal of medical and biological research = Revista brasileira de pesquisas medicas e biologicas 15 19893985
2017 Hakai, an E3-ligase for E-cadherin, stabilizes δ-catenin through Src kinase. Cellular signalling 14 28069439
2020 Heat Shock Protein 90 Chaperone Regulates the E3 Ubiquitin-Ligase Hakai Protein Stability. Cancers 13 31952268
2022 Role of the E3 ubiquitin-ligase Hakai in intestinal inflammation and cancer bowel disease. Scientific reports 11 36266428
2017 Proteomic Analysis of the E3 Ubiquitin-Ligase Hakai Highlights a Role in Plasticity of the Cytoskeleton Dynamics and in the Proteasome System. Journal of proteome research 10 28675930
2012 Upregulation of CBLL1 in rat brain cortex after lipopolysaccharide treated. Journal of molecular histology 9 23160791
2025 Beyond destruction: emerging roles of the E3 ubiquitin ligase Hakai. Cellular & molecular biology letters 6 39833727
2024 CBLL1 promotes endometrial stromal cell senescence via inhibiting PTEN in recurrent spontaneous abortion. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 6 39012313
2024 Hakai, a novel Runx2 interacting protein, augments osteoblast differentiation by rescuing Runx2 from Smurf2-mediated proteasome degradation. Journal of cellular physiology 6 39034451
2022 Daurisoline Inhibiting Tumor Angiogenesis and Epithelial-Mesenchymal Transition in Bladder Cancer by Mediating HAKAI Protein Stability. Iranian journal of pharmaceutical research : IJPR 6 36937208
2023 Ethoxysanguinarine Induces Apoptosis, Inhibits Metastasis and Sensitizes cells to Docetaxel in Breast Cancer Cells through Inhibition of Hakai. Chemistry & biodiversity 5 36633334
2024 Stratification of Colorectal Patients Based on Survival Analysis Shows the Value of Consensus Molecular Subtypes and Reveals the CBLL1 Gene as a Biomarker of CMS2 Tumours. International journal of molecular sciences 2 38339195
2025 Steroid sulfatase suppresses keratinization by inducing proteasomal degradation of E-cadherin via Hakai regulation. Biochimica et biophysica acta. Molecular cell research 1 39764917
2023 CBLL1 is hypomethylated and correlates with cortical thickness in transgender men before gender affirming hormone treatment. Scientific reports 1 38062063
2026 Chromones from Saposhnikovia divaricata modulate m6A RNA methylation-mediated macrophage polarization by targeting CBLL1 to ameliorate RA. Phytomedicine : international journal of phytotherapy and phytopharmacology 0 42001838
2024 DNA barcoding aids in generating a preliminary checklist of the lichens and allied fungi of Calvert Island, British Columbia: Results from the 2018 Hakai Terrestrial BioBlitz. Biodiversity data journal 0 38469225

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