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General and MicroRNA-Mediated mRNA Degradation Occurs on Ribosome Complexes in Drosophila Cells
Sanja Antic1, Michael T Wolfinger2, Anna Skucha1
1Department of Microbiology, Immunobiology and Genetics, Max F. Perutz Laboratories, University of Vienna, Vienna, Austria.
Abstract:
The translation and degradation of mRNAs are two key steps in gene expression that are highly regulated and targeted by many factors, including microRNAs (miRNAs). While it is well established that translation and mRNA degradation are tightly coupled, it is still not entirely clear where in the cell mRNA degradation takes place. In this study, we investigated the possibility of mRNA degradation on the ribosome in Drosophila cells. Using polysome profiles and ribosome affinity purification, we could demonstrate the copurification of various deadenylation and decapping factors with ribosome complexes. Also, AGO1 and GW182, two key factors in the miRNA-mediated mRNA degradation pathway, were associated with ribosome complexes. Their copurification was dependent on intact mRNAs, suggesting the association of these factors with the mRNA rather than the ribosome itself. Furthermore, we isolated decapped mRNA degradation intermediates from ribosome complexes and performed high-throughput sequencing analysis. Interestingly, 93% of the decapped mRNA fragments (approximately 12,000) could be detected at the same relative abundance on ribosome complexes and in cell lysates. In summary, our findings strongly indicate the association of the majority of bulk mRNAs as well as mRNAs targeted by miRNAs with the ribosome during their degradation.
Insights
Messenger RNA (mRNA) degradation occurs on ribosomes in Drosophila cells. This study shows that key degradation factors and miRNA-targeted mRNAs associate with ribosomes during this process.
Area of Science:
- Molecular Biology
- Gene Expression Regulation
- Cell Biology
Background:
- mRNA translation and degradation are crucial, regulated steps in gene expression.
- MicroRNAs (miRNAs) influence both translation and mRNA degradation.
- The precise cellular location of mRNA degradation remains unclear.
Purpose of the Study:
- To investigate mRNA degradation occurring on ribosomes in Drosophila cells.
- To identify factors involved in mRNA degradation associated with ribosome complexes.
- To determine the role of ribosomes in miRNA-mediated mRNA decay.
Main Methods:
- Utilized polysome profiling and ribosome affinity purification.
- Investigated the association of deadenylation, decapping factors, AGO1, and GW182 with ribosome complexes.
- Performed high-throughput sequencing of decapped mRNA fragments from ribosome complexes.
Main Results:
- Deadenylation and decapping factors, along with AGO1 and GW182, copurified with ribosome complexes.
- Association of these factors with ribosomes was dependent on intact mRNAs.
- 93% of decapped mRNA fragments showed similar abundance on ribosomes and in cell lysates.
Conclusions:
- Findings strongly suggest that mRNA degradation, including miRNA-targeted mRNAs, predominantly occurs on ribosomes.
- Ribosomes are a key site for the degradation of bulk and targeted mRNAs.
- This study elucidates the spatial coupling of translation and mRNA decay on the ribosome.
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