KBTBD4

KBTBD4 is the substrate-recognition subunit of a CULLIN3-RING E3 ubiquitin ligase (CRL3-KBTBD4) that controls epigenetic state by directing CoREST corepressor components for proteasomal degradation (PMID:33417871). Through its KELCH-repeat β-propeller domain, KBTBD4 engages the HDAC1/2-LSD1-CoREST complex, recognizing substrates via a conserved motif in the ELM2 domain whose N-terminal residues are required for degradation; targets include RCOR1/LSD1 as well as RCOR3, RREB1, ZNF217, and MIER2 (PMID:33417871, PMID:36997086). The small molecule UM171 functions as a molecular glue that bridges a KBTBD4 homodimer asymmetrically onto the HDAC1/2 catalytic domain, with both KELCH propellers clamping HDAC1 and the endogenous cofactor inositol hexakisphosphate (IP6) acting as a cooperative second glue that buttresses the interface (PMID:40175372, PMID:39939763, PMID:38798619). Recurrent in-frame insertion (indel) mutations at a KELCH-domain hotspot in group 3/4 medulloblastoma confer neomorphic gain-of-function: the mutation reshapes the same KBTBD4 surface engaged by UM171, inserting a bulky side chain into the HDAC1 active-site pocket to stabilize the interface and constitutively recruit HDAC1/2 and CoREST as neo-substrates, driving aberrant CoREST degradation, epigenetic reprogramming, and cancer cell stemness; HDAC1/2 inhibitors block this interface and the proliferation of mutant cells (PMID:35379950, PMID:38798357, PMID:40175372, PMID:39939763). Beyond chromatin regulators, UM171-activated CRL3-KBTBD4 also targets chromatin-bound MYC for degradation in hematopoietic stem cells, restraining cell cycle entry to preserve reconstitution capacity and multipotency (PMID:38207291).

1. 2021 High

   Established that KBTBD4 is the substrate receptor of a CRL3 E3 ligase and links it to a pharmacological mechanism by showing UM171 potentiates degradation of CoREST corepressor components, explaining how a small molecule reprograms HSC epigenetics.

   Evidence Co-IP, ubiquitylation and proteasome-inhibitor assays with UM171 perturbation and epigenetic mark quantification in HSCs

   PMID:33417871

   Open questions at the time

   • Structural basis of UM171-induced substrate engagement not defined
   • Full substrate repertoire not mapped
2. 2022 High

   Showed that medulloblastoma KELCH-domain indel mutations are neomorphic gain-of-function, converting CoREST/RCOR1 into a neo-substrate and linking KBTBD4 directly to oncogenic epigenetic reprogramming.

   Evidence In-cell ubiquitylation, mutant vs. wild-type Co-IP, proteasome-inhibitor rescue, and RNA-seq across >200 medulloblastoma samples

   PMID:35379950

   Open questions at the time

   • Molecular basis by which the mutation reshapes substrate binding unresolved
   • Whether HDAC1/2 rather than CoREST is the direct neo-target not yet distinguished
3. 2023 Medium

   Defined the substrate recognition determinant by mapping the targeted proteome and identifying a conserved ELM2-domain motif required for degradation, broadening the known target set beyond RCOR1/LSD1.

   Evidence Global proteomics of the UM171-targeted proteome with site-directed mutagenesis of ELM2 residues and degradation assays in a single lab

   PMID:36997086

   Open questions at the time

   • Single-lab proteomics not independently replicated
   • Functional significance of each newly named target not individually validated
4. 2024 Medium

   Extended CRL3-KBTBD4 function to MYC turnover, showing UM171-activated degradation of chromatin-bound MYC controls HSC cell-cycle entry independently of CoREST degradation.

   Evidence MYC overexpression epistasis in UM171-treated HSCs with reconstitution and multipotency assays, single lab

   PMID:38207291

   Open questions at the time

   • Direct ubiquitylation of MYC by CRL3-KBTBD4 not structurally demonstrated
   • Relationship between MYC and CoREST target selection unclear
5. 2024 High

   Resolved the structural neomorphic mechanism: cryo-EM and deep mutational scanning showed mutant KBTBD4 homodimers asymmetrically clamp HDAC1 via two KELCH propellers, with the inserted side chain entering the HDAC1 active-site pocket, identifying HDAC1/2 as the direct neo-target and HDAC inhibitors as a therapeutic vulnerability.

   Evidence Cryo-EM of two cancer mutants bound to LSD1-HDAC1-CoREST, deep mutational scanning, in-cell ubiquitylation, and HDAC-inhibitor proliferation assays (preprint, later published in Nature)

   PMID:38798357

   Open questions at the time

   • In vivo efficacy of HDAC inhibition against KBTBD4-mutant tumors not established
   • Generality across all hotspot indel variants not fully mapped
6. 2025 High

   Unified the small-molecule and genetic mechanisms by demonstrating UM171 is a bona fide molecular glue acting across the KBTBD4-HDAC1-CoREST interface and that the indel mutation is a natural molecular-glue mimic reshaping the same surface, with IP6 as a cooperative endogenous second glue.

   Evidence High-resolution cryo-EM (2.7–3.3 Å) of multiple wild-type and mutant KBTBD4-HDAC2(-CoREST1) complexes, UM171 analog SAR, and in situ base-editor scanning of KBTBD4 and HDAC1

   PMID:38798619 PMID:39939763 PMID:40175372

   Open questions at the time

   • Physiological role and regulation of IP6 cofactor occupancy not defined
   • How the same surface discriminates among the multiple ELM2-motif substrates remains open

• It remains unknown how substrate selectivity is governed across the full KBTBD4 target set and whether endogenous signals other than UM171 or IP6 modulate CRL3-KBTBD4 activity in vivo.
• No mechanism connecting ELM2-motif recognition to substrate prioritization
• Endogenous physiological trigger for wild-type CRL3-KBTBD4 activation undefined