FBXL3

FBXL3 is the substrate-recognition subunit of an SCF (SKP1–CUL1–F-box) E3 ubiquitin ligase whose central, conserved role is to set the period of the mammalian circadian clock by driving ubiquitin-mediated degradation of the cryptochrome repressors CRY1 and CRY2 (PMID:17463252, PMID:35216494). Structurally, FBXL3 captures CRY by inserting its C-terminal tail into the cryptochrome FAD-binding pocket while burying the PER-binding surface, so that FAD and PER proteins competitively oppose FBXL3 engagement and thereby gate the timing of CRY turnover (PMID:23503662). SCF(FBXL3) assembly is itself substrate-driven: the C-terminal LRR domain autoinhibits SKP1 binding until CRY binding relieves this inhibition, so the ligase forms productively only in the presence of its substrate (PMID:24085301). Beyond promoting CRY degradation through the E-box loop, FBXL3 also acts genetically through inactivation of the REV-ERBα:HDAC3 corepressor complex on RRE-driven transcription, the two arms together accounting for its long-period phenotype (PMID:23471982). FBXL3 additionally repurposes cryptochromes as substrate-recruiting cofactors for non-clock targets: CRY2 is an essential cofactor for ubiquitylation of T58-phosphorylated c-MYC (PMID:27840026), and CRY1/CRY2 recruit the active cell-cycle kinase TLK2 for degradation, linking the clock to proliferation (PMID:30655559). In muscle satellite cells FBXL3 degrades the transcription factor TCF12 to suppress the MEF2C–myogenin axis, acting as a negative regulator of myogenic differentiation and muscle repair (PMID:40755783, PMID:40554051). In a distinct nuclear, non-degradative role downstream of EGFR/SRC signaling, SRC-mediated phosphorylation of FBXL3 at Y306 enables K63-linked polyubiquitination of GLDC, promoting GLDC-driven repression of MHC-I transcription and tumor immune evasion (PMID:41728086).

1. 2007 High

   Established that a previously uncharacterized F-box protein governs circadian period, defining FBXL3 as a core clock component by controlling CRY protein stability.

   Evidence ENU mutagenesis screen with the Afh (Cys358Ser) allele, in vivo wheel-running, Per2::Luciferase tissue bioluminescence, and in vitro degradation assays in mouse

   PMID:17463252

   Open questions at the time

   • Did not resolve the structural basis of CRY recognition
   • Did not address non-clock substrates
2. 2013 High

   Resolved how FBXL3 recognizes its substrate and how that recognition is regulated, showing tail insertion into the CRY FAD pocket and competition by FAD and PER.

   Evidence X-ray crystallography of CRY2 apo, FAD-bound, and FBXL3–SKP1-complexed forms, plus FAD competition and structural mutagenesis

   PMID:23503662

   Open questions at the time

   • Did not quantify in-cell competition kinetics between FAD/PER and FBXL3
   • Structure of the full SCF holocomplex on CRY not determined
3. 2013 High

   Showed that SCF(FBXL3) assembly is substrate-gated, with LRR autoinhibition of SKP1 binding relieved by CRY engagement — explaining how ligase activity is coupled to substrate availability.

   Evidence Co-IP in transfected cells, in vitro reconstitution with recombinant proteins, and domain-swap mutagenesis

   PMID:24085301

   Open questions at the time

   • The inferred autoinhibition-relieving interactor was not identified
   • Single-lab biochemistry
4. 2013 High

   Separated FBXL3's clock action into two genetically distinct arms — CRY degradation and REV-ERBα:HDAC3 inactivation — broadening its role beyond the E-box loop.

   Evidence Double-mutant mouse genetics (Fbxl3-KO × Rev-erbα-KO; Fbxl3 × Cry1) with circadian behavioral and reporter assays

   PMID:23471982

   Open questions at the time

   • Molecular mechanism by which FBXL3 inactivates REV-ERBα:HDAC3 not defined
   • Whether the RRE arm involves direct FBXL3 substrate ubiquitination unclear
5. 2013 High

   Confirmed that FBXL3's circadian function is exclusively CRY-dependent and that CRY1 and CRY2 are non-redundant period determinants.

   Evidence SCN bioluminescence and wheel-running in Cry-deficient mice carrying Fbxl3(Afh); genetic epistasis

   PMID:23616524

   Open questions at the time

   • Did not address tissue-specific CRY targeting outside the SCN
6. 2016 High

   Revealed a non-clock function in which CRY2 serves as a cofactor enabling FBXL3 to degrade oncogenic c-MYC, connecting the ligase to growth control.

   Evidence Reciprocal Co-IP, ubiquitylation assays, CRY1/CRY2 substitution, and siRNA knockdown with protein stability measurements

   PMID:27840026

   Open questions at the time

   • In vivo tumor relevance of c-MYC targeting not established
   • Basis of CRY2-specific (vs CRY1) cofactor function not structurally defined
7. 2019 Medium

   Extended the cofactor model to TLK2, linking circadian and cell-cycle oscillators through CRY-dependent FBXL3 substrate recruitment.

   Evidence APMS, Co-IP, CRY overexpression/deletion with TLK2 quantification, and kinase-dead TLK2 mutant

   PMID:30655559

   Open questions at the time

   • Physiological consequences of TLK2 turnover not characterized
   • Single-lab data
8. 2022 Medium

   Demonstrated evolutionary conservation of FBXL3's circadian role in a non-mammalian vertebrate.

   Evidence Zebrafish fbxl3a loss-of-function with circadian promoter, mRNA oscillation, and sleep-wake behavioral assays

   PMID:35216494

   Open questions at the time

   • Did not test non-clock substrates in fish
   • Single-lab study
9. 2025 Medium

   Identified TCF12 as an FBXL3 degradation target in satellite cells, defining FBXL3 as a negative regulator of myogenic differentiation via the MEF2C–myogenin axis.

   Evidence Satellite-cell-specific Fbxl3 KO, RNA-seq/GSEA, ChIP-PCR, dual-luciferase, ubiquitination assays, and AAV silencing

   PMID:40755783

   Open questions at the time

   • Whether TCF12 targeting requires a CRY cofactor not tested
   • Single-lab study
10. 2025 Medium

    Showed the muscle-repair relevance of FBXL3 loss in a dystrophic model, supporting it as a brake on regeneration.

    Evidence Satellite-cell-specific Fbxl3 deletion in mdx mice with grip/endurance tests, histology, and AAV silencing

    PMID:40554051

    Open questions at the time

    • Long-term therapeutic durability not assessed
    • Single-lab study
11. 2025 Medium

    Placed Fbxl3 as a transcriptional output of YAP-TEAD mechanotransduction that feeds back to suppress clock oscillations.

    Evidence ATAC-seq, ChIP, RNA-seq, verteporfin inhibition, and circadian bioluminescence in iPSC embryoid bodies

    PMID:40413171

    Open questions at the time

    • Direct YAP-TEAD binding at the Fbxl3 promoter in vivo not fully resolved
    • Single-lab study
12. 2026 Medium

    Uncovered a non-degradative, phosphorylation-gated nuclear function in which FBXL3 mediates K63-linked GLDC ubiquitination to repress MHC-I and drive immune evasion.

    Evidence Co-IP, K63-linkage-specific ubiquitination assays, Y306F/K636R mutagenesis, SRC inhibition, and CD8+ T cell assays

    PMID:41728086

    Open questions at the time

    • Whether this requires SCF holocomplex or a CRY cofactor not addressed
    • In vivo tumor immunity validation limited
    • Single-lab study

• It remains unknown how FBXL3 selects between its degradative clock/cell-cycle substrates and its non-degradative K63-ubiquitination nuclear targets, and what governs cofactor-dependent versus cofactor-independent substrate recruitment across tissues.
• No unified model integrating CRY-cofactor recruitment with phospho-gated targeting
• The autoinhibition-relieving interactor of the LRR domain is unidentified
• Mechanism of REV-ERBα:HDAC3 inactivation undefined