The roles of the red clover necrotic mosaic virus capsid and cell-to-cell movement proteins in systemic infection

The red clover necrotic mosaic dianthovirus (RCNMV) genome is split between two single-stranded RNA species termed RNA-1 and RNA-2. RNA-2 is required for infection of whole plants but is dispensable for infection and virion formation in protoplasts. We have used full-length cDNA clones of RNA-1 and -2 from which infectious in vitro transcripts can be derived to construct a number of mutations in the RNA-1 encoded capcid protein and the RNA-2 encoded cell-to-cell movement protein genes. The capsid protein and the RNA sequence encoding the capsid protein were dispensable for infection of the inoculated leaves of Nicotiana benthamiana and N. clevelandii at both 15 and 25°. In addition, capsid protein was not necessary for systemic infection of N. benthamiana at 15°. As many as 39 amino acid residues could be deleted from the carboxyl-terminus of the RNA-2 encoded 35-kDa cell-to-cell movement protein without lose of or reduction in the rate of cell-to-cell movement or systemic infection. However, larger deletions within the cell-to-cell movement protein gene prevented cell-to-cell movement and syetemic infection of N. benthamiana. These data suggest that the spread of RCNMV in a systemic host is a combination of two distinct events: cell-to-cell movement and long distance transport. We conclude that the RCNMV 35-kDa movement protein is required for cell-to-cell movement, whereas the capsid protein is not necessary for cell-to-cell movement and, depending on host genotype and environmental factors, may or may not be required for long distance transport.