Tsai, J. M. | bioRxiv Science

bioRxiv2026

Distinct Proteasomal Pathways Drive Oncogenic PPM1D Activation

PPM1D is a serine/threonine phosphatase and DNA damage response (DDR) regulator recurrently activated in cancer through amplification or C-terminal truncating mutations that increase its abundance. Here we show that truncating mutations fundamentally rewire PPM1D proteostasis, unmasking an oncogenic function of an alternative protein degradation pathway. While full-length PPM1D undergoes rapid ubiquitin-independent proteasomal degradation via a C-terminal degron, truncating mutations redirect degradation to a slower UBR5-mediated ubiquitin-dependent pathway. The resulting accumulation of PPM1D suppresses DDR signaling and enhances cellular fitness under genotoxic stress, which is further amplified by UBR5 loss. Consistent with selective pressure on this axis, cancers harboring PPM1D truncating mutations are enriched for UBR5 loss-of-function mutations. Together, these findings identify escape from ubiquitin-independent proteasomal degradation as a mechanism of oncogenic adaptation and establish proteostatic routing as a regulatory layer linking protein degradation, DDR signaling, and cancer evolution.

cell biology↗

bioRxiv2023

Template-assisted covalent modification of DCAF16 underlies activity of BRD4 molecular glue degraders

Small molecules that induce protein-protein interactions to exert proximity-driven pharmacology such as targeted protein degradation are a powerful class of therapeutics1-3. Molecular glues are of particular interest given their favorable size and chemical properties and represent the only clinically approved degrader drugs4-6. The discovery and development of molecular glues for novel targets, however, remains challenging. Covalent strategies could in principle facilitate molecular glue discovery by stabilizing the neo-protein interfaces. Here, we present structural and mechanistic studies that define a trans-labeling covalent molecular glue mechanism, which we term "template-assisted covalent modification". We found that a novel series of BRD4 molecular glue degraders act by recruiting the CUL4DCAF16 ligase to the second bromodomain of BRD4 (BRD4BD2). BRD4BD2, in complex with DCAF16, serves as a structural template to facilitate covalent modification of DCAF16, which stabilizes the BRD4-degrader-DCAF16 ternary complex formation and facilitates BRD4 degradation. A 2.2 [A] cryo-electron microscopy structure of the ternary complex demonstrates that DCAF16 and BRD4BD2 have pre-existing structural complementarity which optimally orients the reactive moiety of the degrader for DCAF16Cys58 covalent modification. Systematic mutagenesis of both DCAF16 and BRD4BD2 revealed that the loop conformation around BRD4His437, rather than specific side chains, is critical for stable interaction with DCAF16 and BD2 selectivity. Together our work establishes "template-assisted covalent modification" as a mechanism for covalent molecular glues, which opens a new path to proximity driven pharmacology.

biochemistry↗