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Trendel, Jakob; Schwarzl, Thomas; Horos, Rastislav; Prakash, Ananth; Bateman, Alex; Hentze, Matthias W.; Krijgsveld, Jeroen
Cell, 01/2019, Volume: 176, Issue: 1-2Journal Article
Proteins and RNA functionally and physically intersect in multiple biological processes, however, currently no universal method is available to purify protein-RNA complexes. Here, we introduce XRNAX, a method for the generic purification of protein-crosslinked RNA, and demonstrate its versatility to study the composition and dynamics of protein-RNA interactions by various transcriptomic and proteomic approaches. We show that XRNAX captures all RNA biotypes and use this to characterize the sub-proteomes that interact with coding and non-coding RNAs (ncRNAs) and to identify hundreds of protein-RNA interfaces. Exploiting the quantitative nature of XRNAX, we observe drastic remodeling of the RNA-bound proteome during arsenite-induced stress, distinct from autophagy-related changes in the total proteome. In addition, we combine XRNAX with crosslinking immunoprecipitation sequencing (CLIP-seq) to validate the interaction of ncRNA with lamin B1 and EXOSC2. Thus, XRNAX is a resourceful approach to study structural and compositional aspects of protein-RNA interactions to address fundamental questions in RNA-biology. Display omitted •XRNAX purifies protein-crosslinked RNA of all biotypes from UV-crosslinked cells•Discovery of the WKF RNA-binding domain•Discovery of more than 700 proteins interacting with non-polyadenylated RNA•Profiling of stress-induced changes in RNA-binding proteomes A general approach for characterizing cellular RNA-protein interactions allows examination of dynamic changes to the RNA-bound proteome.
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