Structure, self-cleavage, and replication of two viroid-like satellite RNAs (virusoids) of subterranean clover mottle virus.

Both the genomic and viroid-like satellite RNAs (virusoids) from four subterranean clover mottle virus isolates described by Francki et al. were analyzed in detail. Restriction endonuclease mapping of cDNAs prepared from the genomic RNAs from all isolates showed that these RNAs are closely related if not identical. The two virusoids, which can occur together in the same isolate or individually, were sequenced and shown to be able to form highly base-paired viroid-like secondary structures. The left-hand portions of these structures are almost entirely homologous but the right-hand portions show little similarity. The plus, but not the minus, virusoid RNAs contain sequences that can form the hammerhead self-cleavage structure of certain other self-cleaving viroid, virusoid, and satellite RNAs. Plus, but not minus, RNA transcripts from cDNA clones self-cleaved essentially to completion at the predicted site during transcription in vitro. Northern blot analysis of infected leaf tissue extracts revealed the presence of an oligomeric series of plus RNAs (of monomer size and greater) but minus RNAs were present only as high molecular weight species of heterogeneous size. These findings are in agreement with the lack of minus RNA self-cleavage in vitro. Hence, these virusoid RNAs appear to replicate by a rolling-circle mechanism in which only the plus RNAs self-cleave to form monomeric RNAs.