Long-distance RNA–RNA interactions between terminal elements and the same subset of internal elements on the potato virus X genome mediate minus- and plus-strand RNA synthesis

Abstract

Potexvirus genomes contain conserved terminal elements that are complementary to multiple internal octanucleotide elements. Both local sequences and structures at the 59 terminus and long-distance interactions between this region and internal elements are important for accumulation of potato virus X (PVX) plus-strand RNA in vivo. In this study, the role of the conserved hexanucleotide motif within SL3 of the 39 NTR and internal conserved octanucleotide elements in minus-strand RNA synthesis was analyzed using both a template-dependent, PVX RNA-dependent RNA polymerase (RdRp) extract and a protoplast replication system. Template analyses in vitro indicated that 39 terminal templates of 850 nucleotides (nt), but not 200 nt, supported efficient, minus-strand RNA synthesis. Mutational analyses of the longer templates indicated that optimal transcription requires the hexanucleotide motif in SL3 within the 39 NTR and the complementary CP octanucleotide element 747 nt upstream. Additional experiments to disrupt interactions between one or more internal conserved elements and the 39 hexanucleotide element showed that long-distance interactions were necessary for minus-strand RNA synthesis both in vitro and in vivo. Additionally, multiple internal octanucleotide elements could serve as pairing partners with the hexanucleotide element in vivo. These cis-acting elements and interactions correlate in several ways to those previously observed for plusstrand RNA accumulation in vivo, suggesting that dynamic interactions between elements at both termini and the same subset of internal octanucleotide elements are required for both minus-and plus-strand RNA synthesis and potentially other aspects of PVX replication.

RNA-RNA and RNA-protein interactions in coronavirus replication and transcription

“…This is the case of high-throughput functional addition, plus-strand genomic and sgRNA synthesis also require interactions between an octa-nucleotide located at the genome 5' end, and the central conserved sequences indicated above. 94 In hepatitis C virus (HCV), genome replication involves RNA-RNA interactions between a cis-acting element located within the coding sequences and other structural motifs inserted up and downstream to this element. 12,19 Genome cyclization is the most distal intramolecular interaction possible within two domains in a viral genome.…”

Dynamic RNA structures in the dengue virus genome

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A Local, Interactive Network of 3′ RNA Elements Supports Translation and Replication of Turnip Crinkle Virus

“…Some, but not all, plus-strand RNA viruses contain a substantial number of genome-wide secondary structure elements (genome-scale ordered RNA structure [GORS]) and adopt a compact spheroid shape that correlates with viral persistence (5,6). While RNA viruses likely make extensive use of local and long-distance RNA interactions between such secondary structure elements to control basic processes, only a limited number of such interactions have been reported, with identified connections forming more readily discernible canonical base pairings (3,4,9,10,12,13,14,15,23,26,28,40,41).…”