Figure 5.

Protein 1a extends RNA3 half-life. (A) Yeast expressing the indicated BMV components were grown in defined, gal-containing medium to induce RNA3 transcription from the pB3 GAL1 promoter. At time zero, these strains were transferred to defined, glc-containing medium to repress RNA3 transcription. For each strain, equal aliquots of cells were removed at the indicated times after transfer to glc medium and frozen at −70°C to stop further decay. Total RNA was then extracted, and equal amounts of total RNA from each sample were analyzed by Northern blotting, as in Fig. 2A, to follow the decay of positive-strand RNA3. Note the different time intervals used for sampling yeast containing or lacking 1a. For comparison, exposures were adjusted to provide similarly intense starting signals for all strains. (B) For three independent experiments as in A, positive-strand RNA3 levels were measured with a Molecular Dynamics PhosphorImager, expressed for each strain as a percentage of the RNA3 level at time zero, and plotted on a semilogarithmic plot vs. time after transfer to glc medium. Averages and standard deviations are shown. RNA3 half-life corresponds to the time at which each curve intersects 50% (dotted line).