Spacer-length dependence of programmed -1 or -2 ribosomal frameshifting on a U(6)A heptamer supports a role for messenger RNA (mRNA) tension in frameshifting

Lin, Zhaoru; Gilbert, Robert J. C.; Brierley, Ian

It is clear that 25OMe has a significant effect on ribosome progression, as evidenced by the appearance of a polypeptide corresponding to ribosomes stalled at the bound AON. This d.o. product accounted for up to 10% of the overall synthesis at high 25OMe or 15OMe concentration (10 mM). It has been speculated recently that ribosomal frameshifting frequencies have been generally underestimated as a failure to take into account ribosomes that have frameshifted yet failed to progress on the mRNA, which are often scored as non-frameshifted products (43) . While the observation here of what appears to be AON-mediated drop-off of ribosomes lends support to the idea that mRNA structures can act as roadblocks to the elongating ribosome, such drop-off is far less apparent when ribosomes are challenged with natural frameshift-stimulatory RNAs (37) . Indeed, in this study, the abundance of the d.o. product was greatly reduced when a 15mer unmodified RNA AON replaced 25OMe (Supplementary Figure S1) . The relatively high proportion of ribosomes that appear to be stalled for an extended period probably reflects the very stable association of the OMe AONs with the mRNA template.