Conserved octanucleotide sequences located upstream
of two major potato virus X (PVX) subgenomic RNAs (sgRNAs),
as well as elements in the 5′ end of the genome,
affect accumulation of sgRNA. To determine if complementarity
between these sequences is important for PVX RNA accumulation,
we analyzed the effects of mutations within these elements
and compensatory mutations in a tobacco protoplast system
and in plants. Mutations in the 5′ nontranslated
region (NTR mutants) that reduced complementarity resulted
in lower genomic RNA (gRNA) and sgRNA levels, whereas mutations
to the octanucleotide elements affected only the corresponding
sgRNA levels. However, for both the NTR and octanucleotide
mutants, the extent of reductions in RNA levels did not
directly correlate with the degree of complementarity,
suggesting that the sequences of these elements are also
important. Mutants containing changes in the NTR and compensatory
changes in one of the octanucleotide elements restored
levels of gRNA and the other sgRNA species with an unaltered
octanucleotide element to those of wild-type. Although
compensatory changes significantly increased levels of
the sgRNA species with the modified octanucleotide element,
levels were not restored to those of wild-type. Our data
indicate that long distance RNA–RNA interactions
and the sequences of the interacting elements are required
for PVX plus-strand RNA accumulation.