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Nat Commun. 2019 Nov 22;10(1):5317. doi: 10.1038/s41467-019-13235-w.

In-cell identification and measurement of RNA-protein interactions.

Author information

1
Institut Curie, PSL Research University, CNRS UMR3215, INSERM U934, Paris, 75005, France.
2
Whitehead Institute for Biomedical Research, Cambridge, MA, 02142, USA.
3
Department of Biological Regulation, Weizmann Institute of Science, Rehovot, 76100, Israel.
4
Biomolecular Screening and Protein Technologies Unit, Centre de Regulació Genòmica (CRG), The Barcelona Institute of Science and Technology, Barcelona, Spain.
5
Medical System Biology, UCC, Medical Faculty Carl Gustav Carus, TU Dresden, Dresden, Germany.
6
Max Planck Institute for Molecular Cell Biology and Genetics, Dresden, Germany.
7
Donnelly Centre for Cellular and Biomolecular Research, Department of Molecular Genetics, Toronto, Canada.
8
Institut Curie, PSL Research University, CNRS UMR3215, INSERM U934, Paris, 75005, France. alena.shkumatava@curie.fr.

Abstract

Regulatory RNAs exert their cellular functions through RNA-binding proteins (RBPs). Identifying RNA-protein interactions is therefore key for a molecular understanding of regulatory RNAs. To date, RNA-bound proteins have been identified primarily through RNA purification followed by mass spectrometry. Here, we develop incPRINT (in cell protein-RNA interaction), a high-throughput method to identify in-cell RNA-protein interactions revealed by quantifiable luminescence. Applying incPRINT to long noncoding RNAs (lncRNAs), we identify RBPs specifically interacting with the lncRNA Firre and three functionally distinct regions of the lncRNA Xist. incPRINT confirms previously known lncRNA-protein interactions and identifies additional interactions that had evaded detection with other approaches. Importantly, the majority of the incPRINT-defined interactions are specific to individual functional regions of the large Xist transcript. Thus, we present an RNA-centric method that enables reliable identification of RNA-region-specific RBPs and is applicable to any RNA of interest.

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