文章基本信息

标题：MS2-TRIBE Evaluates Both Protein-RNA Interactions and Nuclear Organization of Transcription by RNA Editing
本地全文：下载
作者：Jeetayu Biswas ; Reazur Rahman ; Varun Gupta 等
期刊名称：iScience
印刷版ISSN：2589-0042
出版年度：2020
卷号：23
期号：7
页码：1-29
DOI：10.1016/j.isci.2020.101318
语种：English
出版社：Elsevier
摘要：SummaryBoth UV-cross-linking and immunoprecipitation (CLIP) and RNA editing (TRIBE) can identify the targets of RNA-binding proteins. To evaluate false-positives of CLIP and TRIBE, endogenous β-actin mRNA was tagged with MS2 stem loops, making it the only bona fide target mRNA for the MS2 capsid protein (MCP). CLIP and TRIBE detected β-actin, albeit with false-positives. False-positive CLIP signals were attributed to nonspecific antibody interactions. In contrast, putative false-positive TRIBE targets were genes spatially proximal to the β-actin gene. MCP-ADAR edited nearby nascent transcripts consistent with interchromosomal contacts observed in Hi-C. The identification of nascent contacts implies RNA regulatory proteins (e.g., splicing factors) associated with multiple nascent transcripts, forming domains of post-transcriptional activity. Repeating these results with an integrated inducible MS2 reporter indicated that MS2-TRIBE can be applied to a broad array of cells and transcripts to study spatial organization and nuclear RNA regulation.Graphical AbstractDisplay OmittedHighlights•Using the β-actin-MBS system, we identify the false-positives of TRIBE and CLIP•Mammalian TRIBE has fewer putative false-positives than CLIP•The non-MS2 targets for TRIBE are neighboring nascent transcripts•We identify a nuclear domain where nascent RNAs and physically interactMolecular Biology Experimental Approach; Transcriptomics