Programmable C-to-U editing to track endogenous proteins

Protein labeling techniques provide robust tools for studying protein localization, structure, and function. Nonetheless, the challenge persists in labeling endogenous proteins within live cells under their native conditions. Here, we present a universal approach by combining programmable cytidine to uridine (C-to-U) RNA editing with non-canonical amino acids (ncAAs) to achieve site-specific labeling of a diverse array of endogenous proteins using minimal amino acid side-chain tags in living cells. This innovative system, termed RNA Editing mediated ncAAs Protein Tagging (RENAPT), allows for real-time tracking of endogenous proteins by integrating ncAAs into target proteins at specific sites. RENAPT thus emerges as a promising platform with broad applicability for tagging endogenous proteins in live cells, facilitating investigations into their localization and functions.