FIG. 4.

Neither sgRNA transcription nor sgRNA itself inhibits replication of exogenous RNA3 replicons. (A) Effect of trans-expressed sgRNA on the replication of an RNA3-derived replicon. The upper diagram represents expression cassettes for B3ΔSG and DNA-derived BMV sgRNAs with wt [B4(WT-sgRNA)] and frameshifted CP ORF [B4(CPfs)]. The lower panel shows a Northern blot analysis of total RNA from yeast expressing B3ΔSG and sgRNA (with or without translationally active CP ORF) alone or in combination with BMV 1a and 2a replication factors. The asterisk indicates a positive-strandRNA whose appearance is associated with the inactivation of sgRNA transcription. (B) Effect of sgRNA transcription on the replication of an exogenous RNA3-derived replicon. The upper diagram depicts the RNA3 derivatives used in this experiment. The lower panel shows Northern blot analyses of total RNA from yeast expressing 1a, 2a, and the indicated RNA3 cassettes to compare the replication level of B3ΔSGΔCP to that of B3ΔCP. Ethidium bromide-stained rRNA is shown below the Northern blots as a loading control.