Nucleotide-amino acid π-stacking interactions initiate photo cross-linking in RNA-protein complexes
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Date
2022-05-17
Publication Type
Journal Article
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yes
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Abstract
Photo-induced cross-linking is a mainstay technique to characterize RNA-protein interactions. However, UV-induced cross-linking between RNA and proteins at “zero-distance” is poorly understood. Here, we investigate cross-linking of the RBFOX alternative splicing factor with its hepta-ribonucleotide binding element as a model system. We examine the influence of nucleobase, nucleotide position and amino acid composition using CLIR-MS technology (crosslinking-of-isotope-labelled-RNA-and-tandem-mass-spectrometry), that locates cross-links on RNA and protein with site-specific resolution. Surprisingly, cross-linking occurs only at nucleotides that are π-stacked to phenylalanines. Notably, this π-stacking interaction is also necessary for the amino-acids flanking phenylalanines to partake in UV-cross-linking. We confirmed these observations in several published datasets where cross-linking sites could be mapped to a high resolution structure. We hypothesize that π-stacking to aromatic amino acids activates cross-linking in RNA-protein complexes, whereafter nucleotide and peptide radicals recombine. These findings will facilitate interpretation of cross-linking data from structural studies and from genome-wide datasets generated using CLIP (cross-linking-and -immunoprecipitation) methods.
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published
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Journal / series
Volume
13 (1)
Pages / Article No.
2719
Publisher
Nature
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Date collected
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Subject
RNA; RNA-binding proteins
Organisational unit
03591 - Allain, Frédéric / Allain, Frédéric
03760 - Hall, Jonathan / Hall, Jonathan
03927 - Picotti, Paola / Picotti, Paola
Notes
Funding
115766 - Unrestricted Leveraging of Targets for Research Advancement and Drug Discovery (EC)
182880 - NCCR RNA#38;Disease (51NF40-182880): Flexibility Grant (SNF)
ETH-24 16-2 - A new technology for the structural analysis of protein-RNA interactions at single residue resolution and its application to understand pre-miRNA processing in vitro and in vivo. (ETHZ)
182880 - NCCR RNA#38;Disease (51NF40-182880): Flexibility Grant (SNF)
ETH-24 16-2 - A new technology for the structural analysis of protein-RNA interactions at single residue resolution and its application to understand pre-miRNA processing in vitro and in vivo. (ETHZ)