Structural and sequence integrity are essential for the replication of the viroid-like satellite RNA of lucerne transient streak virus

Lucerne transient streak virus (LTSV, genus Sobemovirus) supports the replication and encapsidation of a 322 nt untranslated small-circular RNA (scLTSV). Since scLTSV does not code for any proteins or share sequence similarity with its helper virus (LTSV), it is presumed that it uses structural and sequence motifs to signal the helper virus (and host) machinery for its replication and encapsidation. Insertion and deletion mutations were introduced at various locations within the scLTSV molecule. Our results showed that most mutants were not infectious, with only two exceptions, a (-1) nucleotide deletion and a 9 nt, palindromic insertion mutant which preserved the overall rod-like structure of the scLTSV. Sequence analysis of cDNA clones revealed that the palindromic sequence was replicated for up to 12 days of infection, before the sequence reverted back to its wild-type form. Our results indicate that scLTSV has an optimal sequence and secondary structure for replication, movement and/or packaging within the LTSV helper virus.