The cytoplasmically-inherited killer virus of yeast will be used as a model to study the expression of the information on a double-stranded RNA genome in a eukaryotic cell. This virus contains a virion-associated RNA polymerase activity which catalyzes the transcription in vitro of a message-polarity single strand of RNA which is released from the virion. In heterologous eukaryotic translation systems, this encodes a 32,000 dalton protein which is apparently identical to the in vivo precurser of the low molecular weight killer toxin protein produced by killer yeast strains which harbor the virus. Transcription and translation in vitro by homologous yeast systems will be used to study this genetically well-characterized virus. Transcription by isolation virions purified from yeast cytoplasm will be studied to elucidate the priming and initiation mechanism of the viral RNA polymerase. Transcripts and isolated separated genomic RNA strands will be tested as translational templates, with attention focused upon structural features required for message activity in the yeast system. Open reading frames elucidated by sequence analysis of RNA will be tested in their function by ribosome binding experiments and sequence determination of the primary translation products. Post-transcriptional modifications of the messenger RNA including cap structures and polyadenylation will be tested for their effects on translation template activity. Thus, the killer virus of yeast will be used to elucidate the mechanisms controlling transcription and translation in eukaryotic cells, processes important in regulation of many cellular functions.
