RNA synthesis by Newcastle disease virus temperature-sensitive mutants in two RNA-negative complementation groups.

The temperature-sensitive RNA-negative mutants of Newcastle disease virus comprise two complementation groups, group A (seven members) and group E (one member). The RNA-synthesizing activities of four representative members of group A and the single member of group E were compared with the activity of the wild type. These mutants were defective to varying extents in primary transcription at the nonpermissive temperature, ranging from mutant A1, which had no activity, to mutant E1, which lost only 50% of its activity. All of the mutants were also defective in a postprimary transcriptive process since after preincubation at the permissive temperature in the presence of cycloheximide, there was no subsequent RNA synthesis at the nonpermissive temperature upon removal of the cycloheximide. Similarly, in experiments in which cycloheximide was not used, shifts from the permissive temperature to the nonpermissive temperature before 3 h postinfection did not result in RNA synthesis. However, later shifts to the nonpermissive temperature did allow RNA synthesis. With the exception of mutant A1, all of the mutants maintained this RNA-synthetic ability for at least 3 h, suggesting that RNA synthesis from progeny genomes was not the major postprimary transcriptive defect in these mutants. In contrast, the RNA-synthetic ability of mutant A1 rapidly decayed at the nonpermissive temperature, suggesting that the A gene product is involved in RNA synthesis from progeny genomes. The postprimary transcriptive defect(s) of the other mutants may be in the processing or stability of a protein, in the processing of mRNA, or in replication. Plaque-forming revertants (ts+) of all of the mutants coreverted for RNA synthesis. This finding strengthens the relationship between temperature sensitivity for plaquing and both the primary and postprimary RNA-negative phenotypes.