TRANSLATIONAL FRAMESHIFTING BY BARLEY YELLOW DWARF VIRUS-RNA (PAV SEROTYPE) IN ESCHERICHIA-COLI AND IN EUKARYOTIC CELL-FREE-EXTRACTS

The open reading frame (39K ORF) at the 5' end of the genome of barley yellow dwarf virus, PAV serotype (BYDV-PAV), overlaps with a 60K ORF by 13 nucleotides. Several approaches were used to show that the 60K O RF (putative polymerase gene) is translated by a low-frequency framesh ift event in which some ribosomes shift into the 60K ORF rather than t erminate at the 39K ORF stop codon. A sequence encompassing this regio n of overlap induced minus one (-1) translational frameshifting in het erologous and native contexts. In Escherichia coli, with the a subunit of lacZ used as a reporter gene, the rate of frameshifting caused by the BYDV-PAV sequence was approximately 3%. Amino acid sequencing of t he transframe protein confirmed that ribosomes slip into the -1 frame in the overlapping region which includes a consensus shifty heptanucle otide: GGGUUUU. In a wheat germ translation system, BYDV-PAV genomic R NA from virions frameshifted about twice as efficiently as full-length transcripts from a cDNA clone. Frameshifting in rabbit reticulocyte l ysates was much lower for either template. The identity of the 99-kDa wheat germ translation product was verified as the transframe protein by immunoprecipitation with antibody specific for the 60K ORF. These r esults support our previous observations of frameshifting in protoplas ts and illustrate a subtle molecular control mechanism between this pa thogen and its host cells.