A design principle for a single-stranded RNA genome that replicates with less double-strand formation

Usui, Kimihito; Ichihashi, Norikazu; Yomo, Tetsuya

In a previous study, Axelrod et al. reported that the dsRNA ratio of the replication product decreases with insertions of sequences that form a stem structure, but not with sequences that do not form any particular structure (i.e. a loop structure) (26) . In another study, dsRNA formation was reported to occur during replication primarily between a template RNA and the complementary newly synthesized RNA using the template RNA (28) . Based on these results, we constructed a model of dsRNA formation during replication ( Figure 1 ). In this model, as shown in the figure, (A) a template and a newly synthesized strand are extruded as ssRNAs from the replicase (34, 35) , and each strand forms intra-molecular secondary structures, (B) if both strands have loop structures at the same position, these loops are likely to associate to form a partial dsRNA (36) , (C) the partial dsRNA propagates to other regions of the strand and (D) full-length dsRNA is generated after replication is complete. A key event in the formation of dsRNA in this model is the partial formation of dsRNA shown in (B); therefore, the rate of formation and the stability of the partial dsRNA should influence the formation of the full-length dsRNA. The rate of partial dsRNA formation should depend on the sizes of the associated loops in both the template and the newly synthesized strands. The stability of the partial dsRNA should depend on the GC ratio in the loop region because hybridization strength is greater for GC pairs than AU pairs. In this study, we first attempted to verify this model by examining these predictions using RNAs with various structures and nucleotide compositions.