Equine arteritis virus subgenomic mRNA synthesis: analysis of leader-body junctions and replicative-form RNAs

Abstract

In addition to the genomic RNA, a 3' coterminal nested set of six subgenomic mRNAs is produced in equine arteritis virus (EAV)-infected cells. The seven viral RNAs are also 5' coterminal, since they all contain a 206-nucleotide common leader sequence which is identical to the 5' end of the genome. A conserved penta-nucleotide sequence motif, 5' UCAAC 3', was shown to be present at the junctions between the leader and body sequences in each of the mRNAs. In addition, two alternative junction sites were detected for mRNA 3. Seven replicative-form (RF) RNAs (RFs I to VII), corresponding to the genomic RNA and each of the subgenomic EAV mRNAs, could be prepared from lysates of infected cells. The minus-strand RNA contents of these RF RNAs were analyzed by using an RNase protection assay with an RNA probe containing the mRNA 2 leader-body junction. It was established that RF II contained a negative-stranded copy of mRNA 2, including a complementary leader sequence. The presence of subgenomic minus-strand RNA in RFs is indicative of a function as a transcription template during the production of EAV subgenomic mRNAs.