Corley M, Flynn RA, Lee B, Blue SM, Chang HY, Yeo GW. 2020. Footprinting SHAPE-eCLIP Reveals Transcriptome-wide Hydrogen Bonds at RNA-Protein Interfaces. Molecular cell. 80(5):903-914.e8. Pubmed: 33242392 DOI:S1097-2765(20)30792-9


Discovering the interaction mechanism and location of RNA-binding proteins (RBPs) on RNA is critical for understanding gene expression regulation. Here, we apply selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) on in vivo transcripts compared to protein-absent transcripts in four human cell lines to identify transcriptome-wide footprints (fSHAPE) on RNA. Structural analyses indicate that fSHAPE precisely detects nucleobases that hydrogen bond with protein. We demonstrate that fSHAPE patterns predict binding sites of known RBPs, such as iron response elements in both known loci and previously unknown loci in CDC34, SLC2A4RG, COASY, and H19. Furthermore, by integrating SHAPE and fSHAPE with crosslinking and immunoprecipitation (eCLIP) of desired RBPs, we interrogate specific RNA-protein complexes, such as histone stem-loop elements and their nucleotides that hydrogen bond with stem-loop-binding proteins. Together, these technologies greatly expand our ability to study and understand specific cellular RNA interactions in RNA-protein complexes.
Copyright © 2020 Elsevier Inc. All rights reserved.

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Ryan Flynn’s laboratory is focused on the exploration and discovery of how biopolymers like RNA and glycans work together to control cellular processes in the context of human disease.

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