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Updated: Jun 23, 2025
A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells
Published on: June 16, 2022
m6A modification inhibits miRNAs' intracellular function, favoring their extracellular export for intercellular
Sabrina Garbo1, Daniel D'Andrea2, Alessio Colantoni3
1Istituto Pasteur Italia-Fondazione Cenci Bolognetti, Department of Molecular Medicine, Sapienza University of Rome, Viale Regina Elena 324, 00161 Rome, Italy.
Abstract:
Epitranscriptomics represents a further layer of gene expression regulation. Specifically, N6-methyladenosine (m6A) regulates RNA maturation, stability, degradation, and translation. Regarding microRNAs (miRNAs), while it has been reported that m6A impacts their biogenesis, the functional effects on mature miRNAs remain unclear. Here, we show that m6A modification on specific miRNAs weakens their coupling to AGO2, impairs their function on target mRNAs, determines their delivery into extracellular vesicles (EVs), and provides functional information to receiving cells. Mechanistically, the intracellular functional impairment is caused by m6A-mediated inhibition of AGO2/miRNA interaction, the EV loading is favored by m6A-mediated recognition by the RNA-binding protein (RBP) hnRNPA2B1, and the EV-miRNA function in the receiving cell requires their FTO-mediated demethylation. Consequently, cells express specific miRNAs that do not impact endogenous transcripts but provide regulatory information for cell-to-cell communication. This highlights that a further level of complexity should be considered when relating cellular dynamics to specific miRNAs.
Insights
N6-methyladenosine (m6A) modification in microRNAs (miRNAs) impairs their function and promotes their packaging into extracellular vesicles (EVs) for cell-to-cell communication.
Area of Science:
- Molecular Biology
- Epigenetics
- Cell Biology
Background:
- Epitranscriptomics, including N6-methyladenosine (m6A) modification, adds a layer to gene expression regulation.
- While m6A's impact on microRNA (miRNA) biogenesis is known, its effects on mature miRNAs are unclear.
Purpose of the Study:
- To investigate the functional consequences of m6A modification on mature miRNAs.
- To elucidate the mechanisms by which m6A affects miRNA function, localization, and intercellular transfer.
Main Methods:
- Analysis of m6A modification in specific miRNAs.
- Assessment of miRNA-AGO2 interaction and target mRNA regulation.
- Investigation of miRNA loading into extracellular vesicles (EVs) via RNA-binding proteins.
- Functional studies of EV-mediated miRNA transfer and demethylation by FTO.
Main Results:
- m6A modification weakens miRNA coupling to AGO2, impairing intracellular function.
- m6A promotes miRNA loading into EVs through hnRNPA2B1 recognition.
- EV-delivered miRNAs require FTO-mediated demethylation for function in recipient cells.
- Modified miRNAs act as intercellular signals rather than regulating endogenous targets.
Conclusions:
- m6A modification regulates mature miRNA function and intercellular communication.
- EV-packaged miRNAs carrying m6A modifications serve as regulatory signals between cells.
- This highlights a novel mechanism of cell-to-cell communication mediated by epitranscriptomic modifications of miRNAs.
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