HIV-1 frameshift efficiency is primarily determined by the stability of base pairs positioned at the mRNA entrance channel of the ribosome

Mouzakis, Kathryn D.; Lang, Andrew L.; Vander Meulen, Kirk A.; Easterday, Preston D.; Butcher, Samuel E.

Our data support an 8-nt spacer length between the slippery site and the stem-loop, as the effect of deletions in the spacer correlates with corresponding changes in stem-loop local stability 8-nt downstream of the slippery site ( Figure 4 ). Consistent with this idea, deletion of 1 nt in the spacer region of the Beet western yellow virus (BWYV) À1 PRF site promotes the melting of the first base pair in the downstream structure (81) . If the mRNA channel length is maintained, it follows that spacer lengths in all À1 PRF sites should be !7 nt in length. Yet, some frameshift sites have been drawn with 5-to 6-nt spacers [reviewed in (4, 5) ], including that of BWYV (81) . Our data suggest that these frameshift site structures may be partially unwound at the time of frameshifting, in order to accommodate the requisite spacer length and positioning of the slippery sequence in the ribosomal decoding center. Unfortunately, there are currently no high-resolution structural views of ribosomes engaged with frameshift site structures. In conjunction with functional studies such as the one presented here, high-resolution structural views will ultimately be required to define the frameshifting mechanism. Prior studies have observed relationships between spacer length and À1 PRF efficiency in various systems (22, 39, 82, 83) and are consistent with a spacer length of 7-8 nt and local stability being the primary determinant in frameshift efficiency. Spacer lengths of 6-8 nt produced the highest level of À1 PRF for the antisense oligonucleotides, stem-loop and pseudoknot stimulatory structures (82) . In agreement with our conclusions, these spacers position base pairs with strong local stabilities at the entrance to the mRNA channel.