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CRISPR Gene Editing Tool for MicroRNA Cluster Network Analysis
Published on: April 25, 2022
Frequency and fate of microRNA editing in human brain
Yukio Kawahara1, Molly Megraw, Edward Kreider
1The Wistar Institute.
Abstract:
Primary transcripts of certain microRNA (miRNA) genes (pri-miRNAs) are subject to RNA editing that converts adenosine to inosine (A-->I RNA editing). However, the frequency of the pri-miRNA editing and the fate of edited pri-miRNAs remain largely to be determined. Examination of already known pri-miRNA editing sites indicated that adenosine residues of the UAG triplet sequence might be edited more frequently. In the present study, therefore, we conducted a large-scale survey of human pri-miRNAs containing the UAG triplet sequence. By direct sequencing of RT-PCR products corresponding to pri-miRNAs, we examined 209 pri-miRNAs and identified 43 UAG and also 43 non-UAG editing sites in 47 pri-miRNAs, which were highly edited in human brain. In vitro miRNA processing assay using recombinant Drosha-DGCR8 and Dicer-TRBP (the human immuno deficiency virus transactivating response RNA-binding protein) complexes revealed that a majority of pri-miRNA editing is likely to interfere with the miRNA processing steps. In addition, four new edited miRNAs with altered seed sequences were identified by targeted cloning and sequencing of the miRNAs that would be processed from edited pri-miRNAs. Our studies predict that approximately 16% of human pri-miRNAs are subject to A-->I editing and, thus, miRNA editing could have a large impact on the miRNA-mediated gene silencing.
Insights
This study reveals that adenosine-to-inosine RNA editing frequently occurs in human pri-miRNAs, impacting microRNA processing and gene silencing. Approximately 16% of pri-miRNAs undergo this editing, affecting microRNA function.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- MicroRNA (miRNA) genes produce primary transcripts (pri-miRNAs) that can undergo RNA editing.
- Adenosine-to-inosine (A-->I) RNA editing is a known post-transcriptional modification.
- The frequency and functional consequences of pri-miRNA editing are not fully understood.
Purpose of the Study:
- To investigate the frequency of A-->I RNA editing in human pri-miRNAs.
- To determine the impact of pri-miRNA editing on miRNA processing.
- To identify novel edited miRNAs and their potential effects on gene silencing.
Main Methods:
- Large-scale sequencing of RT-PCR products from 209 human pri-miRNAs, focusing on UAG triplet sequences.
- In vitro miRNA processing assays using recombinant Drosha-DGCR8 and Dicer-TRBP complexes.
- Targeted cloning and sequencing of miRNAs processed from edited pri-miRNAs.
Main Results:
- Identified 43 UAG and 43 non-UAG editing sites in 47 pri-miRNAs, with high editing frequency in the human brain.
- Demonstrated that pri-miRNA editing interferes with miRNA processing steps in vitro.
- Discovered four new edited miRNAs with altered seed sequences, potentially affecting gene targets.
Conclusions:
- A-->I RNA editing is prevalent in human pri-miRNAs, with an estimated 16% of pri-miRNAs being edited.
- Pri-miRNA editing significantly impacts miRNA processing and can alter miRNA-mediated gene silencing.
- This widespread editing suggests a substantial role for RNA editing in regulating miRNA function and gene expression.
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