Formaldehyde-crosslinking enables comprehensive profiling of in vivo RNA-interactome in UV-impermeable samples

Development of UV-crosslinking based RNA-interactome capture protocol have expanded the repertoire of RBPs. Apparent limitations of UV-crosslinking, however, have made the RBP profiling to be incomplete and inapplicable to many of the biological samples in physiological contexts. Here we developed formaldehyde-crosslinking based RNA binding protein profiling (FAX-RBP) method and quantitatively compared to UV-crosslinking based RBP profiling (UVX-RBP) in HeLa and Xenopus laevis oocyte/embryo. Quantitative comparison of FAX-RBP with UVX-RBP in HeLa cell demonstrated that FAX-RBP has greatly enhanced efficiency for profiling hundreds of RBPs in vivo. FAX-RBP was then applied to X. laevis oocyte and stage 8 embryo and profiled the dynamics of mRNP complex for over 500 RBPs. FAX-RBP thus provide the most comprehensive and unbiased RBP profile in vivo, demonstrating its potential to expand mRNP profiling into numerous physiological contexts.