A MUTATION IN TOMATO ASPERMY CUCUMOVIRUS THAT ABOLISHES CELL-TO-CELL MOVEMENT IS MAINTAINED TO HIGH-LEVELS IN THE VIRAL-RNA POPULATION BY COMPLEMENTATION

The nucleotide substitution C-->A at nucleotide 100 of tomato aspermy cucumovirus (TAV) strain V (V-TAV) RNA segment 3 (RNA3) introduces an ocher stop at the fourth codon of the movement protein open reading fr ame. Experiments with RNA transcripts from full-length clones showed t hat this mutation abolished cell-to-cell movement and, thus, infectivi ty in planta. Heterogeneity analyses on stock V-TAV virion RNA showed that an A at position 100 was present in the molecular population of R NA3 at a frequency of 0.76 and that a C at this position was present a t a frequency of 0.24. This result indicates that a fraction of RNA3 m olecules complements cell-to-cell movement of movement-defective molec ules. It was shown that the mutation C-->A conferred enhanced RNA repl ication of the defective mutant in tobacco protoplasts. The effect of the mutation on replication was dependent on sequence context, since t he same mutation did not affect the replication efficiency in the rela ted TAV strain 1 RNA3. Competition experiments in tobacco protoplasts were done to estimate the fitness during a cell invasion cycle of the movement-defective mutant relative to the wild type (wt). From these d ata, a lower limit to the degree of complementation of movement-defect ive molecules by movement-competent ones could be estimated as 0.13. T his estimate shows that complementation may play an important role in the determination of genetic structure in RNA genome populations. A fu rther effect of the enhanced replication of the movement-defective mut ant was the efficient competition with the wt for the initiation of in fection foci in planta.