HERC4

HERC4 is a HECT-domain E3 ubiquitin ligase that targets a diverse set of substrates for proteasomal degradation and non-degradative ubiquitin signaling, functioning in spermatogenesis, developmental signaling, and oncogenic pathways. HERC4 catalyzes K48-linked polyubiquitination of c-Maf (at K85/K297), LATS1, Smoothened, SAV1, and KRT19, directing these substrates to proteasomal degradation, thereby modulating Hippo, Hedgehog, and myeloma-relevant transcriptional programs (PMID:26825710, PMID:30710319, PMID:30925584, PMID:26125558, PMID:41192149). HERC4 also mediates K63-linked polyubiquitination of MafA at K33, which does not trigger degradation but instead blocks GSK3β-dependent phosphorylation and suppresses MafA transcriptional activity and downstream STAT3 signaling (PMID:37028761). In vivo, HERC4 is required for cytoplasmic droplet removal during spermiogenesis, and male mice lacking functional Herc4 exhibit approximately 50% reduced fertility with morphologically abnormal spermatozoa (PMID:17967448).

1. 2007 High

   The first in vivo function of HERC4 was established: it is essential for spermatozoon maturation, resolving the question of whether this HECT-domain ligase has a physiological role beyond generic ubiquitin conjugation.

   Evidence Herc4-mutant male mice showed ~50% reduced fertility with cytoplasmic droplet retention and reduced sperm motility

   PMID:17967448

   Open questions at the time

   • The spermatogenesis substrate(s) of HERC4 remain unidentified
   • Whether HERC4 catalytic activity (versus scaffolding) is required for droplet removal is untested
   • Female reproductive or other developmental phenotypes not examined
2. 2015 Medium

   HERC4 was linked to Hippo pathway regulation through ubiquitination and destabilization of the scaffold protein Salvador (Sav), revealing that Hippo kinase activity creates a positive feedback loop by antagonizing the Sav–Herc4 interaction.

   Evidence RNAi screen, Co-IP, ubiquitination assay, and genetic epistasis in Drosophila

   PMID:26125558

   Open questions at the time

   • Mammalian conservation of Hippo-dependent regulation of HERC4–SAV1 interaction not demonstrated at this time
   • Ubiquitin chain type on Sav not determined
   • Direct HECT-domain catalytic requirement not shown via catalytic-dead mutant
3. 2016 High

   HERC4 was identified as the E3 ligase responsible for K48-linked polyubiquitination and proteasomal degradation of the myeloma oncoprotein c-Maf, establishing a direct tumor-suppressive mechanism and identifying specific ubiquitination sites (K85, K297).

   Evidence Affinity chromatography/MS identification of HERC4–c-Maf interaction, Co-IP, in vitro ubiquitination, site-directed mutagenesis, and xenograft validation

   PMID:26825710

   Open questions at the time

   • How HERC4 activity toward c-Maf is regulated remains unclear
   • Whether USP5-mediated antagonism is the sole deubiquitinase counterbalancing HERC4 is unresolved
4. 2019 Medium

   HERC4 was shown to ubiquitinate and degrade Smoothened (Smo), establishing it as a negative regulator of Hedgehog signaling in both Drosophila and mammalian (NSCLC) contexts, with regulation by PKA-primed phosphorylation.

   Evidence Drosophila genetic modifier screen, Co-IP and ubiquitination assays in fly and mammalian cells, Hedgehog reporter and proliferation readouts

   PMID:30925584 PMID:31010679

   Open questions at the time

   • Whether HERC4-mediated Smo degradation is relevant in non-cancer Hedgehog-dependent tissues (e.g. limb, neural tube) is untested
   • Ubiquitin chain linkage type on Smo not characterized
5. 2019 Medium

   HERC4 was identified as an E3 ligase for the Hippo pathway kinase LATS1, broadening its role as a multi-substrate regulator of Hippo signaling beyond Salvador.

   Evidence Co-IP, ubiquitination assay, knockdown/overexpression in breast cancer cells, xenograft

   PMID:30710319

   Open questions at the time

   • Whether HERC4 targets LATS1 and SAV1 simultaneously or in distinct contexts is unknown
   • The ubiquitin chain type on LATS1 was not determined
6. 2021 Medium

   HERC4-mediated SAV1 downregulation was confirmed in mammalian hepatocellular carcinoma cells, extending the Drosophila Salvador finding to a human cancer context.

   Evidence Co-IP, western blot, functional overexpression/knockdown assays in HCC cells, xenograft

   PMID:35116265

   Open questions at the time

   • Direct ubiquitination of SAV1 by HERC4 was not formally demonstrated in this study
   • Whether Hippo kinase-dependent feedback seen in Drosophila operates in HCC is unknown
7. 2023 High

   HERC4 was shown to catalyze a non-degradative K63-linked polyubiquitination of MafA at K33, establishing that HERC4 can regulate substrate activity without promoting degradation — by blocking GSK3β phosphorylation and suppressing STAT3 signaling.

   Evidence Co-IP, K63-linkage-specific ubiquitination assay, K33R mutagenesis, transcriptional reporter, xenograft

   PMID:37028761

   Open questions at the time

   • Structural basis for how K63-ubiquitination at K33 sterically blocks GSK3β access is unknown
   • Whether HERC4 uses K63-linkage on other substrates beyond MafA is untested
8. 2024 High

   HERC4 was shown to be co-optable by a molecular glue (AK59) for targeted degradation of STING, demonstrating its pharmacological tractability as an E3 ligase for induced proximity approaches.

   Evidence Targeted protein degradation screen, proteomic identification, functional ubiquitination and STING degradation assays

   PMID:38811577

   Open questions at the time

   • Whether HERC4 ubiquitinates STING under physiological (non-drug) conditions is unknown
   • Structural basis for AK59-mediated HERC4–STING proximity not resolved
9. 2025 Medium

   HERC4 was identified as the E3 ligase for KRT19, linking its ubiquitin ligase activity to epithelial-mesenchymal transition and metastasis in lung adenocarcinoma.

   Evidence Co-IP, GST pull-down, ubiquitination assay, EMT/invasion readouts, mouse model

   PMID:41192149

   Open questions at the time

   • Whether KRT19 ubiquitination by HERC4 is K48-linked and proteasomal was not explicitly determined
   • Regulation of HERC4 expression or activity in the EMT context is not characterized

• Key unresolved questions include the structural basis for HERC4's broad substrate selectivity, the identity of its spermatogenesis substrate(s), and whether its K63-linked ubiquitination activity extends beyond MafA.
• No crystal or cryo-EM structure of HERC4 or HERC4–substrate complex exists
• The spermatogenesis substrate(s) remain unknown despite clear in vivo phenotype
• Mechanisms regulating HERC4 expression, stability, or catalytic activation are largely uncharacterized