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

Cited 10 time in Web of Science Cited 10 time in Scopus
Authors

Na, Yongwoo; Kim, Hyunjoon; Choi, Yeon; Shin, Sanghee; Jung, Jae Hun; Kwon, S. Chul; Kim, V. NarryKim, Jong-Seo

Issue Date
2021-03
Publisher
Oxford University Press
Citation
Nucleic Acids Research, Vol.49 No.5
Abstract
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.
ISSN
0305-1048
URI
https://hdl.handle.net/10371/178031
DOI
https://doi.org/10.1093/nar/gkaa1194
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Research Area Molecular Biology & Genetics

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