The genome of pea enation mosaic virus (PEMV) consists of two plus-sense RNAs, both of which are required for mechanical transmission. RNA 1 (5706 nucleotides) has strong sequence similarity with members of the luteovirus group, a similarity that is also manifested in the symptomatology, cytopathology and vector transmission of this virus. RNA 2 (4253 nucleotides) is hypothesized to facilitate systemic invasion and mechanical transmission, attributes that distinguish PEMV from the phloem-limited luteoviruses. Sequence analysis of RNA 2 has demonstrated that PEMV is unique among multicomponent viruses in that it lacks Tand 5'-terminal homology between its genomic RNAs. Sequence analysis of RNA 2 has identified an open reading frame encoding a putative product of 65K that contains a series of polymerase-like motifs typical of viral RNA-dependent RNA polymerases. This protein sequence lacks homology with the polymerase encoded on RNA 1 of PEMV, instead being more closely affiliated with the polymerases of viruses related to the carmoand tombusvirus groups. Inoculation of pea protoplasts with RNA transcripts derived from a full-length cDNA clone ofRNA 2 has demonstrated that RNA 2 replicates autonomously in the absence of RNA 1, although comparable inoculation of whole plants failed to establish a systemic infection. There is no evidence that RNA 2 encodes structural proteins, suggesting that encapsidation functions are supplied in trans by RNA 1, comparable to the helper-dependent complexes occurring within the luteovirus group. These data suggest that the PEMV genome can be characterized as a symbiotic association of two taxonomically distinct viral RNAs cooperatively interacting in the establishment of a systemic virus infection.

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An examination of the genomic strategy of pea enation mosaic virus (PEMV) RNA 1 has verified strong organizational and sequence relationships between PEMV and the beet western yellows-potato leafroll luteovirus subgroup. Sequence analysis of RNA 1 demonstrated five predominant open reading frames (ORFs). The extreme 5' ORF encodes a 34K product of unknown function. The second ORF encodes an 84K product which overlaps 90% of ORF l (in a unique reading frame) and is expressed by internal initiation beginning at the second start codon from the 5' terminus. This protein contains a protease-like motif characteristic of serine-and cysteine-based proteases, suggesting involvement in post-translational processing of viral translation products. The third ORF is characterized by a number of RNA polymerase motifs and a helicase-like motif typical of RNA-dependent RNA polymerases. It overlaps (out of frame) the ORF 2 product and is proposed to be expressed by a frameshift fusion of the ORF 2 and ORF 3 products. The fourth ORF encodes the viral coat protein, and is immediately followed in frame by a 33K ORF thought to represent the aphid transmission subunit of the PEMV virion. Northern blot analysis of polysomeassociated RNA suggests that both products are expressed from an 1800 nucleotide subgenomic mRNA, with the 33K product expressed as a readthrough fusion with the coat protein monomer.

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