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Measuring A-to-I RNA editing and gene expression signatures of neuronal populations within the Drosophila brain

ABSTRACT: We used an improved INTACT (Isolation of Nuclei Tagged in A specific Cell Type) technique to isolate RNA from purified nuclei from different neuronal populations of the Drosophila brain. Using RNA-seq, we determined gene expression and A-to-I RNA editing levels at editing sites across nine distinct neuronal sub-populations and a pan-neuronal control.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE113661 | GEO | 2019/01/18

REPOSITORIES: GEO

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Measuring A-to-I RNA editing signatures of neuronal populations within the Drosophila brain

Project description:We used an improved INTACT (Isolation of Nuclei Tagged in A specific Cell Type) technique to isolate RNA from purified nuclei from different neuronal populations of the Drosophila brain. Using microfluidic multiplex PCR and sequencing (mmPCR-seq), we determined gene expression and A-to-I RNA editing levels at editing sites across nine distinct neuronal sub-populations and a pan-neuronal control.

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Project description:Studies in epitranscriptomics indicates that RNA is modified by a variety of enzymes. Among these RNA modifications, A-to-I RNA editing occurs frequently in the mammalian transcriptome. These RNA editing sites can be detected directly from RNA-seq data by examining nucleotide changes from adenosine (A) to guanine (G), which substitutes for inosine (I). However, a careful investigation of such nucleotide changes must be conducted to distinguish sequencing errors and genomic mutations from the genuine editing sites. Building upon our recent introduction of an easy-to-use bioinformatics tool, RNAEditor, to detect RNA editing events from RNA-seq data, we examined the extent by which RNA editing events affects the binding of RNA-binding proteins (RBP). Through employing bioinformatic techniques, we uncover that RNA editing sites occur frequently in RBP-bound regions. Moreover, the presence of RNA editing sites are more frequent when RNA editing islands are examined, which are regions in which RNA editing sites are present in clusters. When the binding of one RBP, HuR, was quantified experimentally, its binding was reduced upon silencing of the RNA editing enzyme ADAR compared to the control—suggesting that the presence of RNA editing islands influences HuR binding to its target regions. These data indicate RNA editing as an important mediator of RBP-RNA interactions—a mechanism which likely constitutes an additional mode of post-transcription gene regulation in biological systems.

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Measuring gene expression and RNA editing in Drosophila adapting to divergent microclimates

Project description:We performed RNA-seq to profile gene expression in the heads and whole bodies of 32 isofemale fly lines from two divergent microclimates at 'Evolution Canyon' in Israel (16 fly lines from each microclimate). We also measured RNA editing levels in the head tissue of these flies.

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Alu sequences in undifferentiated human embryonic stem cells display high levels of A-to-I RNA editing

Project description:Adenosine to Inosine (A-to-I) RNA editing is a site-specific modification of RNA transcripts, catalyzed by members of the ADAR (Adenosine Deaminase Acting on RNA) protein family. RNA editing occurs in human RNA in thousands of different sites. Some of the sites are located in protein-coding regions but the majority is found in non-coding regions, such as 3âUTRs, 5âUTRs and introns - mainly in Alu elements. While editing is found in all tissues, the highest levels of editing are found in the brain. It was shown that editing levels within protein-coding regions are increased during embryogenesis and after birth and that RNA editing is crucial for organism viability as well as for normal development. In this study we characterized the A-to-I RNA editing phenomenon during neuronal and spontaneous differentiation of human embryonic stem cells (hESCs). We identified high editing levels of Alu repetitive elements in hESCs and demonstrated a global decrease in editing levels of non-coding Alu sites when hESCs are differentiating, particularly into the neural lineage. Using RNA interference, we showed that the elevated editing levels of Alu elements in undifferentiated hESCs are highly dependent on ADAR1. DNA microarray analysis showed that ADAR1 knockdown has a global effect on gene expression in hESCs and leads to a significant increase in RNA expression levels of genes involved in differentiation and development processes, including neurogenesis. Taken together, our data suggest that A-to-I editing of Alu sequences plays a role in the regulation of hESC early differentiation decisions. Two samples, One control and the second treated

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