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Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells

ABSTRACT: This SuperSeries is composed of the SubSeries listed below.

ORGANISM(S): Homo sapiens Drosophila melanogaster

PROVIDER: GSE223557 | GEO | 2023/02/03

REPOSITORIES: GEO

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Comparison of TRIBE and STAMP for identifying targets of RNA binding proteins in human and <i>Drosophila</i> cells.

Project description:RNA binding proteins (RBPs) perform a myriad of functions and are implicated in numerous neurological diseases. To identify the targets of RBPs in small numbers of cells, we developed TRIBE, in which the catalytic domain of the RNA editing enzyme ADAR (ADARcd) is fused to an RBP. When the RBP binds to an mRNA, ADAR catalyzes A to G modifications in the target mRNA that can be easily identified in standard RNA sequencing. In STAMP, the concept is the same except the ADARcd is replaced by the RNA editing enzyme APOBEC. Here we compared TRIBE and STAMP side-by-side in human and Drosophila cells. The goal is to learn the pros and cons of each method so that researchers can choose the method best suited to their RBP and system. In human cells, TRIBE and STAMP were performed using the RBP TDP-43. Although they both identified TDP-43 target mRNAs, combining the two methods more successfully identified high-confidence targets. In Drosophila cells, RBP-APOBEC fusions generated only low numbers of editing sites, comparable to the level of control editing. This was true for two different RBPs, Hrp48 and Thor (Drosophila EIF4E-BP), indicating that STAMP does not work well in Drosophila.

| S-EPMC10351885 | biostudies-literature

Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [Drosophila]

Project description:RNA binding proteins (RBPs) perform a myriad of functions and are implicated in numerous neurological diseases. To identify the targets of RBPs in small numbers of cells, we developed TRIBE, in which the catalytic domain of the RNA editing enzyme ADAR (ADARcd) is fused to a RBP. In STAMP, the ADARcd is replaced by the RNA editing enzyme Apobec (REF). Here we compared the two enzymes fused to the RBP TDP43 in human cells. Although they both identified TDP43 target mRNAs, combining the two methods more successfully identified high confidence targets. We also assayed the two enzymes in Drosophila cells in which RBP-Apobec fusions generated only low numbers of editing sites comparable to the level of control editing. This was true for two different RBPs, Hrp48 and Thor (Drosophila EIF4E-BP), and contrasted with successful RBP-ADARcd fusions. The results indicate that TRIBE is the method of choice in Drosophila.

2023-02-05 | GSE223555 | GEO

Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [human]

Project description:RNA binding proteins (RBPs) perform a myriad of functions and are implicated in numerous neurological diseases. To identify the targets of RBPs in small numbers of cells, we developed TRIBE, in which the catalytic domain of the RNA editing enzyme ADAR (ADARcd) is fused to a RBP. In STAMP, the ADARcd is replaced by the RNA editing enzyme Apobec (REF). Here we compared the two enzymes fused to the RBP TDP43 in human cells. Although they both identified TDP43 target mRNAs, combining the two methods more successfully identified high confidence targets. We also assayed the two enzymes in Drosophila cells in which RBP-Apobec fusions generated only low numbers of editing sites comparable to the level of control editing. This was true for two different RBPs, Hrp48 and Thor (Drosophila EIF4E-BP), and contrasted with successful RBP-ADARcd fusions. The results indicate that TRIBE is the method of choice in Drosophila.

2023-02-05 | GSE223556 | GEO

Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [Drosophila]

Project description:Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [Drosophila]

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Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells

Project description:Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells

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Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [human]

Project description:Comparison of STAMP and TRIBE for identifying the targets of RNA-binding proteins in human and Drosophila cells [human]

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A quantitative analysis of CLIP methods for identifying binding sites of RNA-binding proteins

Project description:This SuperSeries is composed of the SubSeries listed below.

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

Project description: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.

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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series

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Quaking isoform depletion in C2C12 myoblasts; direct binding targets of Quaking RNA binding proteins

Project description:This SuperSeries is composed of the SubSeries listed below.

2018-07-11 | GSE102615 | GEO