Short Internal Sequences Involved in Replication and Virion Accumulation in a Subviral RNA of Turnip Crinkle Virus

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Abstract
cis -acting sequences and structural elements in untranslated regions of viral genomes allow viral RNA-dependent RNA polymerases to correctly initiate and transcribe asymmetric levels of plus and minus strands during replication of plus-sense RNA viruses. Such elements include promoters, enhancers, and transcriptional repressors that may require interactions with distal RNA sequences for function. We previously determined that a non-sequence-specific hairpin (M1H) in the interior of a subviral RNA (satC) associated with Turnip crinkle virus is required for fitness and that its function might be to bridge flanking sequences (X. Sun and A. E. Simon, J. Virol. 77:7880-7889, 2003). To establish the importance of the flanking sequences in replication and satC-specific virion repression, segments on both sides of M1H were randomized and subjected to in vivo functional selection (in vivo SELEX). Analyses of winning (functional) sequences revealed three different conserved elements within the segments that could be specifically assigned roles in replication, virion repression, or both. One of these elements was also implicated in the molecular switch that releases the 3′ end from its interaction with the repressor hairpin H5, which is possibly involved in controlling the level of minus-strand synthesis.