The terminal loop of a 3′ proximal hairpin plays a critical role in replication and the structure of the 3′ region of Turnip crinkle virus

Plus-strand RNA viruses serve as templates for translation and then transcription by newly synthesized RdRp. A ribosome-binding tRNA-shaped structure (TSS) and upstream hairpin H4 in the 3′ UTR of Turnip crinkle virus (TCV) play key roles in translation and transcription. Second-site mutations generated to compensate for altering the critical asymmetric internal loop of H4 included a three- to two-base alteration in the terminal loop of a 3′ proximal hairpin (Pr) located downstream of the TSS. Unlike the non-deleterious three-base alteration, single mutations in Pr loop were detrimental for RdRp transcription while enhancing translation and RdRp binding. One deleterious mutation in the Pr loop altered the structures of both the TSS and H4. These complex interactions in the 3′ UTR support a compact structural arrangement likely permitting RdRp access to a number of residues within a 195-base region including the 3′ end that are necessary for efficient transcription initiation.