The soybean retroelement SIRE1 uses stop codon suppression to express its envelope-like protein

Abstract

The soybean SIRE1 family of Ty1/copia retrotransposons encodes an envelope-like gene (env-like). We analysed the DNA sequences of nine SIRE1 insertions and observed that the gag/pol and env-like genes are in the same reading frame and separated by a single UAG stop codon. The six nucleotides immediately downstream of the stop codon conform to a degenerate nucleotide motif, CARYYA, which is sufficient to facilitate stop codon suppression in tobacco mosaic virus. In vivo stop codon suppression assays indicate that SIRE1 sequences confer leakiness to the UAG stop codon at an efficiency of 5%. These data suggest that SIRE1 retro-elements use translational suppression to express their envelope-like protein; this is in contrast with all characterized retroviruses, which express the envelope protein from a spliced genomic messenger RNA.

Figure 1

Organization of the SIRE1 open reading frames. (A) gag and pol are in one open reading frame (ORF) separated from the envelope-like (env-like) gene by a single stop codon. Boxed arrows represent the long terminal repeats. (B) The three SIRE1 sequence variations after the UAG stop codon conform to the CARYYA consensus of tobacco mosaic virus (TMV; Skuzeski et al., 1991) and the CARNBA consensus sufficient for stop codon suppression in yeast (Namy et al., 2001) (R, purines; Y, pyrimidines; N, any nucleotide; B represents U, C or G). The numbers after SIRE1 define distinct insertions. PR, protease; IN, integrase; RT, reverse transcriptase.

Figure 2

Translational readthrough of SIRE1. For these experiments, levels of readthrough are relative to the initial readthrough level of the CAAUUA SIRE1 variant. UAG stop codons are in bold. Mutations in the SIRE1 sequences are underlined. Error bars represent one standard deviation above and below the average. (A) The three SIRE1 sequence variations result in the same level of readthrough, averaging 5% of the normal activity of a β-glucuronidase reporter gene (data not shown). (B) Readthrough levels of clones with mutations in sequences upstream of the stop codon. (C) Readthrough levels of clones with mutations in the CARYYA consensus. R, any purine. Y, any pyrimidine.