Nucleotide sequence and translation of satellite tobacco mosaic virus RNA

Abstract

Satellite tobacco mosaic virus (STMV) is a plant virus with a 17-nm icosahedral particle encapsidating a 0.3 × 10 6 M r ssRNA genome that depends on tobamoviruses for its replication. The complete nucleotide sequence of STMV RNA deduced in the experiments described here was 1059 nucleotides in length. The efficiency of labeling viral RNA with [γ- 32P]ATP using T4 polynucleotide kinase was not affected by treatment with tobacco acid pyrophosphatase and/or bacterial alkaline phosphatase, indicating that the majority of the 5′ termini of encapsidated STMV RNAs were not phosphorylated. The 240 3′-terminal nucleotides of STMV RNA and either tobacco mosaic virus (TMV) U1 RNA or TMV U2/U5 RNA had greater than 65% overall sequence similarity, with two nearly identical regions of 40 and 50 bases, respectively. There were no other regions of sequence relatedness to TMV RNA. The 19 5′-terminal nucleotides of STMV RNA had greater than 65% sequence similarity with the 16 5′-terminal nucleotides of brome mosaic virus (RNA 3 and 50% sequence similarity with the 12 5′-terminal nucleotides of the Q strain of cucumber mosaic virus RNA 3. The first open reading frame (ORF) beginning at base 53 encoded a 6800 M r protein that corresponded in size to a major in vitro translation product directed by STMV RNA. A second ORF, beginning at nucleotide 163, had the capacity to code for a protein that corresponded in size (17,500 M r) to the other major in vitro translation product. The first 12 codons of this ORF corresponded to the sequence of the N-terminal amino acids of the capsid protein. Western-blot analysis of the in vitro translation products revealed that the 17,500 M r protein had the same electrophoretic mobility as the authentic capsid protein; it was also antigenically related to the capsid protein, but the 6800 M r protein was not. Time course analysis of in vitro translation demonstrated that the 6800 M r protein was synthesized at the same time as the capsid protein and did not arise by the proteolytic cleavage of a larger precursor polypeptide. These results suggest that the genome of STMV functioned as a polycistronic messenger RNA. It has not been determined if the 6800 M r protein is synthesized in vivo. STMV RNA had untranslated regions of 52 and 418 nucleotides at its 5′ and 3′ termini, respectively. Nonphosphorylated 5′ termini, the degree of similarity to the 3′ terminus of two of its helper viruses, the genome organization, and the ability to function as a polycistronic mRNA are unique features for the genome of this satellite virus.