Figure 7.

1a-stimulated RNA3 accumulation does not increase expression of a gene translated from RNA3. (A) Schematic of B3–5′CAT, an RNA3 derivative in which the 5′ half of the 3a gene has been replaced by the CAT gene. (B) Representative Northern blot analysis of positive-strand B3–5′CAT RNA accumulation in yeast expressing the indicated BMV components, performed as in Fig. 2A. (C) CAT expression in the yeast of Fig. 7B. Total protein was extracted from a portion of each of the same cultures used for RNA analysis in Fig. 7B. Equal aliquots of each sample were analyzed for CAT activity by using ¹⁴C-chloramphenicol and thin-layer chromatography, and ensuring that all sample activities were within the linear range of the assay (9, 10). The positions of chloramphenicol (Cm) and its acetylated derivatives (1AcCm, 3AcCm) are shown. CAT activity for each strain was normalized to expression in B3–5′CAT-expressing yeast lacking 1a and 2a, averaged over four, independent experiments, and presented with standard deviation below each lane.