4.8 Article

FAX-RIC enables robust profiling of dynamic RNP complex formation in multicellular organisms in vivo

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NUCLEIC ACIDS RESEARCH
卷 49, 期 5, 页码 -

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OXFORD UNIV PRESS
DOI: 10.1093/nar/gkaa1194

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  1. Ministry of Science and ICT of Korea [IBS-R008-D1]
  2. BK 21 Research Fellowships from the Ministry of Education of Korea
  3. College of Natural Sciences, Seoul National University

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RNA-protein interaction is crucial for gene regulation post-transcription, and formaldehyde crosslinking (FAX) has been introduced as an alternative method for efficient capture of RNA-protein interactions, expanding the coverage of the study. FAX-RIC technology provided a comprehensive and unbiased RNA interactome in Xenopus laevis oocytes and embryos, revealing dynamic remodeling of RNA-protein complexes. This study demonstrates the applicability of FAX-RIC in mammalian tissue samples, extending the profiling of RNA interactome into multicellular organisms.
RNA-protein interaction is central to post-transcriptional gene regulation. Identification of RNA-binding proteins relies mainly on UV-induced crosslinking (UVX) followed by the enrichment of RNA-protein conjugates and LC-MS/MS analysis. However, UVX has limited applicability in tissues of multicellular organisms due to its low penetration depth. Here, we introduce formaldehyde crosslinking (FAX) as an alternative chemical crosslinking for RNA interactome capture (RIC). Mild FAX captures RNA-protein interaction with high specificity and efficiency in cell culture. Unlike UVX-RIC, FAX-RIC robustly detects proteins that bind to structured RNAs or uracil-poor RNAs (e.g. AGO1, STAU1, UPF1, NCBP2, EIF4E, YTHDF proteins and PABP), broadening the coverage. Applied to Xenopus laevis oocytes and embryos, FAX-RIC provided comprehensive and unbiased RNA interactome, revealing dynamic remodeling of RNA-protein complexes. Notably, translation machinery changes during oocyte-to-embryo transition, for instance, from canonical eIF4E to noncanonical eIF4E3. Furthermore, using Mus musculus liver, we demonstrate that FAX-RIC is applicable to mammalian tissue samples. Taken together, we report that FAX can extend the RNA interactome profiling into multicellular organisms.

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