Base Editing Screens Uncover Phosphorylation Site Dynamics

The research employed a pioneering CRISPR base editing strategy to alter specific phosphorylation sites and evaluate their functionality through high-throughput screens. By merging quantitative phosphoproteomics, which maps phosphorylation dynamics, with base editing, the study pinpointed crucial phosphorylation sites that impact NFAT transcriptional activity, a pivotal pathway in T cell activation.

Importantly, this approach opens up the possibility of investigating previously unexplored phosphorylation sites, providing a comprehensive understanding of how these modifications influence cell signalling pathways.

In practical terms, the researchers identified approximately 19,000 phosphorylation sites, of which well over 7,000 were targetable by SpCas9-mediated base editors. This extensive targeting revealed that specific phosphosite mutations can either enhance or inhibit T cell activation markers, demonstrating the potential to fine-tune immune responses.

The study also identified PHLPP1, a phosphatase, as a new regulator of T cell activation, highlighting its role in NFAT and NFκB signaling pathways.

Patrick Kennedy and Samuel Myers led the study at the La Jolla Institute for Immunology, and it was published in Nature Methods this Monday.