Analysis of the role of the pseudoknot component in the SRV-1 gag-pro ribosomal frameshift signal: loop lengths and stability of the stem regions.

The simian retrovirus-1 (SRV-1) gag-pro frameshift signal was identified in previous work, and the overall structure of the pseudoknot involved was confirmed (ten Dam E, Brierley I, Inglis S, Pleij C, 1994, Nucleic Acids Res 22:2304-2310). Here we report on the importance of specific elements within the pseudoknot. Some mutations in stem S1 that maintain base pairing have reduced frameshift efficiencies. This indicates that base pairing in itself is not sufficient. In contrast, frameshifting correlates qualitatively with the calculated stability of mutations in S2. The stems thus play different roles in the frameshift event. The nature of the base in L1 has little influence on frameshift efficiency. It is however required to bridge S2; deleting it lowers frameshifting from 23 to 9%. In L2, frameshift efficiency was not affected in a mutant that changed 10 to 12 bases. This makes it unlikely that the primary sequence of L2 plays a role in -1 frameshifting, in contrast to readthrough in Moloney murine leukemia virus (Wills N, Gesteland R, Atkins J, 1994, EMBO J 13:4137-4144). Deletions of 2 and 3 bases gave more frameshifting than the wild type, probably reflecting the increased stability of the pseudoknot due to a shorter loop L2. Deleting even more bases reduces frameshifting compared to wild-type levels. At this point, stress will build up in L2, and this will reduce overall pseudoknot stability.