Cellular parameters shaping pathways of targeted protein degradation

Abstract

In recent years the development of proteolysis-targeting chimeras (PROTACs) has enhanced the field of ubiquitin signalling through advancing therapeutic targeted protein degradation (TPD) strategies and generating tools to explore the ubiquitin landscape. However, the interplay between PROTACs and their substrates, and other components of the ubiquitin proteasome system (UPS), raises fundamental questions about cellular parameters that might influence the action of PROTACs and the amenability of a given target to PROTAC-mediated degradation. In this perspective we discuss examples of cellular parameters that have been shown to influence PROTAC sensitivity and consider others likely to be important for PROTAC-mediated target degradation but not yet routinely considered in design of novel TPD strategies: Target localisation and accessibility on the one hand, and expression patterns, localisation and activity of E3 ligases, deubiquitinases (DUBs) and wider ubiquitin machinery on the other, are critical parameters in the exploitation of PROTACs, and establishing a better understanding of these parameters will facilitate the rational design of PROTACs.

Fig. 1. Schematic of PROTAC activity.

The heterobifunctional small molecule promotes ternary complex formation between E3 and a target protein (1) to bring about ubiquitination (2) then degradation (3) of the target as a ‘neo-substrate’ of the E3. Created in BioRender. Cardno, A. (2025) https://BioRender.com/7ggcsvs.

Fig. 2. Schematic showing examples of spatiotemporal influence on PROTAC activity.

A Spatial regulation of PROTAC-mediated AURKA degradation in mitotic cells. B Temporal regulation of PROTAC-mediated CDK2 degradation through the cell cycle. Created in BioRender. Cardno, A. (2025) https://BioRender.com/ud1xchr.

Fig. 3. Schematic of cellular factors influencing PROTAC activity.

Additional ubiquitin ligase activities (E3/E4) can act to promote ubiquitination and degradation of substrates in certain contexts (top) while deubiquitinases (DUBs) can remove or trim ubiquitin conjugates to stabilise or ‘rescue’ neosubstrates from degradation at the proteasome (bottom). Created in BioRender. Cardno, A. (2025) https://BioRender.com/zozyz3i.