Profiling dynamic RNA–protein interactions using small-molecule-induced RNA editing

RNA-binding proteins (RBPs) play an important role in biology, and characterizing dynamic RNA–protein interactions is essential for understanding RBP function. In this study, we developed targets of RBPs identified by editing induced through dimerization (TRIBE-ID), a facile strategy for quantifying state-specific RNA–protein interactions upon rapamycin-mediated chemically induced dimerization and RNA editing. We performed TRIBE-ID with G3BP1 and YBX1 to study RNA–protein interactions during normal conditions and upon oxidative stress-induced biomolecular condensate formation. We quantified editing kinetics to infer interaction persistence and show that stress granule formation strengthens pre-existing RNA–protein interactions and induces new RNA–protein binding events. Furthermore, we demonstrate that G3BP1 stabilizes its targets under normal and oxidative stress conditions independent of stress granule formation. Finally, we apply our method to characterize small-molecule modulators of G3BP1–RNA binding. Taken together, our work provides a general approach to profile dynamic RNA–protein interactions in cellular contexts with temporal control. Seo and Kleiner developed a small-molecule-dependent RNA editing platform termed TRIBE-ID to profile RNA–protein interactions in cells with temporal control and to study substrates of the stress granule protein G3BP1 during biomolecular condensation.