CRISPR-PTM and CRISPR-VEIS: Multiplexed platforms for quantitative functional analysis of endogenous phosphosites

Connecting protein post-translational modifications (PTMs) to phenotypic outcomes is a central challenge. Although phosphoproteomics has richly catalogued specific sites, reliable methods to measure the endogenous effects of individual phosphosites on cellular fitness and signaling are still lacking. Here, we introduce CRISPR-PTM and CRISPR-VEIS as complementary platforms for quantitative, endogenous phosphosite interrogation at both individual and clustered phosphorylation events. CRISPR-PTM is a multiplexed knock-in framework generating defined phosphosite variants with internal allelic markers, enabling precise relative fitness effects in pooled populations. CRISPR-VEIS (Visualisation of