Structural studies of plant
viral RNA molecules have been based on in vitro chemical and enzymatic modification. That approach, along with mutational analysis, has proven valuable in predicting structural models for some plant viruses such as tobacco mosaic tobamovirus and brome mosaic bromovirus. However, in planta conditions may be dramatically different from those found in vitro. In this study we analyzed the structure of cucumber mosaic cucumovirus
satellite RNA (sat
RNA) strain D4 in vivo and compared it to the structures found in vitro and in purified virions. Following a methodology developed to determine the structure of
18S rRNA within intact plant tissues, different patterns of
adenosine and
cytosine modification were found for D4-sat
RNA molecules in vivo, in vitro, and in virions. This chemical probing procedure identifies
adenosine and
cytosine residues located in unpaired regions of the
RNA molecules. Methylation data, a genetic algorithm in the STAR RNA folding program, and sequence alignment comparisons of 78 satellite CMV RNA sequences were used to identify several helical regions located at the 5' and 3' ends of the
RNA molecule. Data from previous mutational and sequence comparison studies between
satellite RNA strains inducing
necrosis in tomato plants and those strains not inducing
necrosis allowed us to identify one helix and two tetraloop regions correlating with the necrogenicity syndrome.