Specific mutations in a viral RNA pseudoknot drastically change ribosomal frameshifting efficiency

Many viruses regulate protein synthesis by -1 ribosomal frame-shifting usin g an RNA pseudoknot. Frameshifting is vital for viral reproduction. Using t he information gained from the recent high-resolution crystal structure of the beet western yellow virus pseudoknot, a systematic mutational analysis has been carried out in vitro and in vivo. We find that specific nucleotide tertiary interactions at the junction between the two stems of the pseudok not are crucial. A tripler is found between stem 1 and loop 2, and tripler interactions are required for frameshifting function. For some mutations, l oss of one hydrogen bond is sufficient to abolish frameshifting. Furthermor e, mutations near the 5' end of the pseudoknot can increase frameshifting b y nearly 300%, possibly by modifying ribosomal contacts. It is likely that the selection of suitable mutations can thus allow viruses to adjust frames hifting efficiencies and thereby regulate protein synthesis in response to environmental change.