TRIBE: Hijacking an RNA-Editing Enzyme to Identify Cell-Specific Targets of RNA-Binding Proteins

• Published in: Cell Vol. 165; no. 3; pp. 742 - 753
• Main Authors: McMahon, Aoife C., Rahman, Reazur, Jin, Hua, Shen, James L., Fieldsend, Allegra, Luo, Weifei, Rosbash, Michael
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 21.04.2016
• Subjects: 3' Untranslated Regions; Adenosine Deaminase - metabolism; Animals; Drosophila; Drosophila melanogaster - enzymology; Drosophila Proteins - metabolism; Genetic Techniques; Heterogeneous-Nuclear Ribonucleoproteins - metabolism; RNA Editing; RNA-Binding Proteins
• ISSN: 0092-8674; 1097-4172
• DOI: 10.1016/j.cell.2016.03.007

RNA transcripts are bound and regulated by RNA-binding proteins (RBPs). Current methods for identifying in vivo targets of an RBP are imperfect and not amenable to examining small numbers of cells. To address these issues, we developed TRIBE (targets of RNA-binding proteins identified by editing), a technique that couples an RBP to the catalytic domain of the Drosophila RNA-editing enzyme ADAR and expresses the fusion protein in vivo. RBP targets are marked with novel RNA editing events and identified by sequencing RNA. We have used TRIBE to identify the targets of three RBPs (Hrp48, dFMR1, and NonA). TRIBE compares favorably to other methods, including CLIP, and we have identified RBP targets from as little as 150 specific fly neurons. TRIBE can be performed without an antibody and in small numbers of specific cells. [Display omitted] •TRIBE is a genetic tool that identifies in vivo targets of RNA-binding proteins (RBPs)•An RBP of interest is fused to the catalytic domain of an RNA-editing enzyme•RBP targets are marked by novel RNA-editing events•Cell-specific targets can be identified from tiny amounts of RNA A technique called TRIBE identifies cell-specific targets of RNA-binding proteins, even in small cell populations, via the detection of RNA-editing events conferred by a genetically encoded enzymatic fusion to the RNA-binding protein of interest.