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Paramyxovirus RNA editing and the requirement for hexamer genome length
S Hausmann1, J P Jacques, D Kolakofsky
1Department of Genetics and Microbiology, University of Geneva School of Medicine, Switzerland.
Abstract:
Paramyxoviruses cotranscriptionally edit their P gene mRNA by the programmed insertion of G residues into a short G run contained within a larger purine run, via pseudo-templated transcription. The templates for paramyxovirus transcription are genome nucleocapsids in which each nucleoprotein subunit is associated with 6 nt, and only genomes whose lengths are multiples of 6 are found naturally or are replicated efficiently in transfected cell systems. We have examined the effect of varying total genome length on the frequency and number of insertions into the mRNA editing site in a transfected cell system, using constructs that generate mini-genome analogues. We found that, as long as the purine run sequence and the region immediately upstream were unaltered, editing occurred during mRNA synthesis independent of the precise length of the minigenome. However, when mini-genome constructs whose lengths were not multiples of 6 were used, insertions (or deletions) occurred during antigenome synthesis within the purine run, which strikingly restored the hexamer length. Genome length correction due to changes in the antigenome purine run length occurred only when the mini-genome was not a multiple of 6, and these changes were only poorly affected by mutations in the mRNA editing site and the region immediately upstream. Our results suggest that the mRNA editing site is a natural hotspot for viral polymerase slippage during genome replication, and that this site serves the dual and complementary function of maintaining hexamer genome length. The unusual requirement of paramyxoviruses for genomes of precise hexamer length may have evolved to maintain genome stability against insertions in the mRNA editing site during replication.
Insights
Paramyxoviruses edit their P gene mRNA by inserting Gs into a G run. Genome length affects this editing, with non-hexamer lengths triggering corrections during replication to maintain viral stability.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- Paramyxoviruses exhibit programmed RNA editing in their P gene mRNA.
- This editing involves G residue insertions into a G run within a purine run.
- Paramyxovirus transcription relies on genome nucleocapsids with subunits associated with 6 nucleotides (nt).
Purpose of the Study:
- To investigate the impact of varying genome lengths on P gene mRNA editing frequency and insertion numbers.
- To understand the role of genome length in paramyxovirus RNA editing and replication.
Main Methods:
- Utilized transfected cell systems with mini-genome analogues to vary total genome length.
- Examined editing frequency and insertion events in mRNA during synthesis.
- Analyzed insertions/deletions during antigenome synthesis for non-hexamer length mini-genomes.
Main Results:
- mRNA editing occurred independently of minigenome length when the purine run sequence was preserved.
- Non-hexamer minigenomes triggered insertions/deletions in the antigenome's purine run, restoring hexamer length.
- These length corrections were minimally affected by mutations in the mRNA editing site.
Conclusions:
- The mRNA editing site acts as a viral polymerase slippage hotspot during replication.
- This site plays a dual role in editing and maintaining precise hexamer genome length.
- The hexamer genome length requirement likely evolved to ensure genome stability against editing-associated insertions.