De novo mutation of an RNA virus is increased in the presence of engineered synonymous mutations that disrupt RNA structural elements

Using a combination of methods including selective 2?-hydroxyl acylation analyzed by primer extension sequencing (SHAPE-Seq), a complete RNA structure map of the cucumber mosaic virus (CMV) RNA 3 segment was mapped (Watters et al. (2018) Nucleic Acids Research 46, 2573?2584). To explore the effect of structural perturbations on genomic stability, infectious mutants were engineered to contain changes in one of four open reading frame (ORF) stem-loop (SL) structures SL1362, SL1439, SL1745 and SL1816, or in a highly conserved structure SL2073 in the 3? untranslated region. The mutations occurred either in the loop or stem, thereby either maintaining or disrupting the predicted SL structure. The mutants were inoculated and then passaged on plants of Nicotiana tabacum. After the eighth passage, the full-length (8 kb) sequence of the virus population was sequenced by an amplicon tiling approach using the Illumina Nextera platform. All virus passages were sequenced as technical duplicates and compared against virus wild-type passages. Across the whole genome, a significant increase in de novo SNPs (synonymous and non-synonymous) occurred in some mutants (vs. wt) as minor (?10%) and dominant populations, suggestive of compensatory genome-wide epistatic changes induced by changes in SLs.

Thompson, J., Waite, D., Cha, A. Y.

Stats

Post a comment

You need to be signed in to post comments. You can sign in here.

Comments

There are no comments yet.