Co-evolution in a putative bundling signal of bluetongue and epizootic hemorrhagic disease viruses

Abstract

Bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) possess a genome of 10 segmented RNAs (S1-S10), one copy of each of which is considered to be packaged in a virion. This selective packaging is thought to be mediated by supramolecular complex formation of the 10 RNAs, through intermolecular base pairing of complementary nucleotide sequences termed the bundling signal. Here, the whole genomic sequences of BTV and EHDV isolates were analyzed to identify co-evolving pairs of complementary nucleotide sequences within and between genomic segments. One co-evolving pair was identified within S5 and another between S5 and S10. The co-evolving pair between S5 and S10, consisting of six sites in each segment, was a candidate for a bundling signal and was identical to one of two putative bundling signals reported in a previous experimental study, validating the effectiveness of the method used in the present study. The six sites in S10 were confirmed to be located in a loop at the end of a stable stem. Although the six sites in S5 were located in a loop at the end of a stem of only four sites long, the complementary nucleotide sequences constituting this stem were, remarkably, the co-evolving pair within S5. These results highlight the importance not only of loops but also of stems in the intermolecular base pairing of bundling signals.