Stimulation of stop codon readthrough: frequent presence of an extended 3' RNA structural element

Firth, Andrew E.; Wills, Norma M.; Gesteland, Raymond F.; Atkins, John F.

Curiously this phenomenon was not just limited to the alphaviruses. The potential to form an extended stemloop structure 3 0 -adjacent to a RT stop codon-phylogenetically conserved and supported by a pair of peaks in synonymous site conservation-was also found in a number of plant virus RT cases, for example, in the replicase gene in the genera (Figures 3 and 4) . Further, the predicted stem is well-supported by a large number of compensatory substitutions-i.e. paired substitutions that preserve the predicted base pairings-between the different species ( Figure 3 and Supplementary Data). The furoviruses and pomoviruses have a second RT site in the CP gene. Here, however, there is a marked dichotomy between the two genera in the RT context. In the furoviruses, the RT context is generally UGA-CGG (UGA-UGG in the highly divergent sorghum chlorotic spot virus, AB033692) and there was evidence for tandem synonymous site conservation peaks and an associated stem-loop structure that, together with a 7 nt 'spacer', covered 96 nt 3 0 -adjacent to the stop codon (Figures 3 and 4) . In the pomoviruses, however, the RT context is generally A-UAG-CAA-UYA (A-UAA-CAA-UUA in the highly divergent broad bean necrosis virus, D86637) and the synonymous site conservation analysis failed to reveal extended conservation in the vicinity of the RT site ( Figure 4 ). Thus the furovirus context and predicted structure is alphavirus-like while the pomovirus context and lack of predicted structure is tobamovirus-like (see below). The animal-infecting coltiviruses also have an alphavirus-like RT site (UGA-CGG) in the VP9/VP9 0 -coding sequence and, again, there is potential to form a 3 0 -adjacent RNA stem-loop structure (Figure 3 ; as noted previously in ref. 65) , which is tentatively supported by our conservation analysis ( Figure 4) .