Photocatalytic Chemical Crosslinking for Profiling RNA–Protein Interactions in Living Cells

ABSTRACT

The dynamic interactions between RNAs and proteins play crucial roles in regulating diverse cellular processes. Proteome‐wide characterization of these interactions in their native cellular context remains desirable but challenging. Herein, we developed a photocatalytic crosslinking (PhotoCAX) strategy coupled with mass spectrometry (PhotoCAX‐MS) and RNA sequencing (PhotoCAX‐seq) for the study of the composition and dynamics of protein‐RNA interactions. By integrating the blue light‐triggered photocatalyst with a dual‐functional RNA–protein crosslinker (RP‐linker) and the phase separation‐based enrichment strategy, PhotoCAX‐MS revealed a total of 2044 RBPs in human HEK293 cells. We further employed PhotoCAX to investigate the dynamic change of RBPome in macrophage cells upon LPS‐stimulation, as well as the identification of RBPs interacting directly with the 5′ untranslated regions of SARS‐CoV‐2 RNA. [ FROM AUTHOR] Copyright of Angewandte Chemie International Edition is the property of John Wiley & Sons, Inc. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)