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Identification of proteins and miRNAs that specifically bind an mRNA in vivo

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Item Type:Article
Title:Identification of proteins and miRNAs that specifically bind an mRNA in vivo
Creators Name:Theil, K. and Imami, K. and Rajewsky, N.
Abstract:Understanding regulation of an mRNA requires knowledge of its regulators. However, methods for reliable de-novo identification of proteins binding to a particular RNA are scarce and were thus far only successfully applied to abundant noncoding RNAs in cell culture. Here, we present vIPR, an RNA-protein crosslink, RNA pulldown, and shotgun proteomics approach to identify proteins bound to selected mRNAs in C. elegans. Applying vIPR to the germline-specific transcript gld-1 led to enrichment of known and novel interactors. By comparing enrichment upon gld-1 and lin-41 pulldown, we demonstrate that vIPR recovers both common and specific RNA-binding proteins, and we validate DAZ-1 as a specific gld-1 regulator. Finally, combining vIPR with small RNA sequencing, we recover known and biologically important transcript-specific miRNA interactions, and we identify miR-84 as a specific interactor of the gld-1 transcript. We envision that vIPR will provide a platform for investigating RNA in vivo regulation in diverse biological systems.
Keywords:Caenorhabditis elegans Proteins, Messenger RNA, MicroRNAs, Protein Binding, RNA-Binding Proteins, Transcription Factors, Animals, Caenorhabditis elegans
Source:Nature Communications
Publisher:Nature Publishing Group
Page Range:4205
Date:16 September 2019
Official Publication:https://doi.org/10.1038/s41467-019-12050-7
PubMed:View item in PubMed

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