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A Reporter Assay to Analyze Intronic microRNA Maturation in Mammalian Cells
Published on: June 16, 2022
Multivesicular bodies associate with components of miRNA effector complexes and modulate miRNA activity
Derrick J Gibbings1, Constance Ciaudo, Mathieu Erhardt
1IBMP-CNRS, UPR Université de Strasbourg, France.
Nature Cell Biology
|August 18, 2009
Summary
MicroRNA (miRNA) silencing involves GW-bodies congregating with endosomes and multivesicular bodies (MVBs). This suggests membrane-associated complexes are key for miRNA action and secretion via exosomes.
Area of Science:
- Cell Biology
- Molecular Biology
- RNA Biology
Background:
- P-bodies/GW-bodies are sites of microRNA (miRNA)-mediated post-transcriptional silencing.
- miRNA effector proteins like argonaute 2 (AGO2) can associate with membranes.
- miRNAs are found in exosomes derived from multivesicular bodies (MVBs), involving ESCRT machinery.
Purpose of the Study:
- To investigate the relationship between GW-bodies, endosomes, and MVBs in miRNA silencing.
- To determine the role of MVBs and ESCRT in the localization and function of miRNA silencing components.
- To explore the potential of membrane-associated complexes in miRNA loading, action, and secretion.
Main Methods:
- Microscopy to observe GW-body localization relative to endosomes and MVBs.
- Biochemical analysis of miRNA and mRNA enrichment at cellular membranes.
- Analysis of exosome-like vesicles secreted by MVBs.
- Gene silencing assays in cells with depleted ESCRT components.
Main Results:
- GW-bodies containing GW182 and AGO2 associate with endosomes and MVBs, distinct from P-bodies.
- miRNAs and miRNA-repressible mRNAs accumulate at these membrane compartments.
- Secreted exosome-like vesicles are enriched in GW182 but not P-body components.
- Depletion of ESCRT components impairs miRNA silencing and causes GW182 accumulation.
Conclusions:
- Endosomes and MVBs serve as sites for miRNA-loaded RISC (miRISC) accumulation and potentially action.
- GW182 is sorted into MVBs for secretion or lysosomal degradation, possibly with AGO2.
- This MVB-mediated sorting facilitates continuous assembly/disassembly of membrane-associated miRISCs for miRNA regulation.
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