Development of NanoLuc-targeting protein degraders and a universal reporter system to benchmark tag-targeted degradation platforms

Christoph Grohmann, Charlene M. Magtoto, Joel R. Walker, Ngee Kiat Chua, Anna Gabrielyan, Mary Hall, Simon A. Cobbold, Stephen Mieruszynski, Martin Brzozowski, Daniel S. Simpson, Hao Dong, Bridget Dorizzi, Annette V. Jacobsen, Emma Morrish, Natasha Silke, James M. Murphy, Joan K. Heath, Andrea Testa, Chiara Maniaci, Alessio CiulliGuillaume Lessene, John Silke, Rebecca Feltham

Research output: Contribution to journalArticleResearchpeer-review

11 Citations (Scopus)

Abstract

Modulation of protein abundance using tag-Targeted Protein Degrader (tTPD) systems targeting FKBP12F36V (dTAGs) or HaloTag7 (HaloPROTACs) are powerful approaches for preclinical target validation. Interchanging tags and tag-targeting degraders is important to achieve efficient substrate degradation, yet limited degrader/tag pairs are available and side-by-side comparisons have not been performed. To expand the tTPD repertoire we developed catalytic NanoLuc-targeting PROTACs (NanoTACs) to hijack the CRL4CRBN complex and degrade NanoLuc tagged substrates, enabling rapid luminescence-based degradation screening. To benchmark NanoTACs against existing tTPD systems we use an interchangeable reporter system to comparatively test optimal degrader/tag pairs. Overall, we find the dTAG system exhibits superior degradation. To align tag-induced degradation with physiology we demonstrate that NanoTACs limit MLKL-driven necroptosis. In this work we extend the tTPD platform to include NanoTACs adding flexibility to tTPD studies, and benchmark each tTPD system to highlight the importance of comparing each system against each substrate.

Original languageEnglish
Article number2073
Number of pages13
JournalNature Communications
Volume13
Issue number1
DOIs
Publication statusPublished - Dec 2022
Externally publishedYes

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