Proteolysis-targeting chimeras (PROTACs) present a potentially effective strategy against various diseases via selective proteolysis. How to increase the efficacy of PROTACs remains challenging. Here, we explore the necessity of the linker, which has been deemed as an integral part of heterobifunctional PROTACs. Adopting single amino acid-based degradation signals, we find that the linker is not a required feature of the PROTACs. Notably, the linker-free PROTAC, Pro-BA, exhibits superior efficacy over its linker-bearing counterparts in degrading EML4-ALK and inhibiting lung cancer cell growth, as Pro-BA induces a stronger interaction between the target and the E3 ubiquitin ligase. Pro-BA is a water-soluble, orally administered degrader that significantly inhibits the tumor growth in a xenograft mouse model. The broad applicability of this linker-free PROTAC strategy is further validated through the development of BCR-ABL degrader. Our study introduces a design paradigm for PROTACs, potentially facilitating the advancement of more efficient therapeutic degraders. Linkers are traditionally seen as important for PROTAC activity. Here, the authors demonstrate that linker-free PROTACs can outperform traditional designs, marking a paradigm shift in PROTAC development for targeted protein degradation.
Autophagy maintains cellular homeostasis by degrading and recycling intracellular components, while the selective clearance of damaged mitochondria, known as mitophagy, ensures mitochondrial quality control. Protein complexes orchestrate these processes, yet their dynamic regulation remains incompletely understood. Here, we integrate thermal proteome profiling with co-aggregation analysis to monitor protein thermal stability and complex assembly dynamics in response to two canonical inducers: Torin 1, which activates autophagy via mTOR inhibition, and CCCP, a mitochondrial uncoupler that triggers mitophagy. This endeavor provides a global view of the dynamic variations of known autophagy- and mitophagy-associated complexes, revealing their assembly state at various stages. Notably, we identify previously uncharacterized complexes containing the eukaryotic elongation factor 1 A1 (EEF1A1) that exhibit enhanced aggregation under both treatments. Functional analyses show that EEF1A1 depletion impairs autophagosome maturation and mitophagic degradation, while pulsed-SILAC demonstrates that EEF1A1 directly regulates the synthesis of core autophagy proteins. Together, these findings map the dynamic landscape of protein complex regulation during autophagy and mitophagy and uncover EEF1A1-mediated translational control as a previously unrecognized regulatory mechanism.
Shuang Zhang, Ya Zeng, Fengming Li et al.· Cell Communication and Signa...· 0 citations