AUG context and mRNA translation in vaccinia virus

Abstract

In this study, we examined the role of the nucleotides flanking AUG initiator codons in modulating translation of vaccinia virus genes. First, the sequence around initiation codons in the genomic sequence of vaccinia was analyzed. A preference for A in position 3 and for A or G in position +4, relative to the A of the initiator AUG codon was noted. These results are in broad agreement with known optimal consensus for the translation of eukaryotic mRNAs. Nucleotides 3 and +4, which are known to affect the efficiency of translation of cellular mRNAs over a 20-fold range, were examined by site-directed mutagenesis and functional assays in the context of vaccinia-infected cells. A collection of virus recombinants expressing beta-Galactosidase (beta-Gal) under the control of a strong promoter and containing all possible nucleotide substitutions in those positions was generated. Quantitation of beta-Gal mRNA and beta-Gal activity showed that the efficiency of translation was not affected significantly by nucleotide substitutions in positions 3 and +4. Therefore, despite the high occurrence of purine nucleotides in those positions in vaccinia mRNAs, no effect of those positions was noted in conditions where â-Gal was overexpressed.