Non-canonical translation mechanisms in plants: efficient in vitro and in planta initiation at AUU codons of the tobacco mosaic virus enhancer sequence

Abstract

The 5' untranslated leader (Omega sequence) of tobacco mosaic virus (TMV) genomic RNA was utilized as a translational enhancer sequence in expression of the 17 kDa putative movement protein (pr17) of potato leaf roll luteovirus (PLRV). In vitro translation of RNAs transcribed from appropriate chimeric constructs, as well as their expression in transgenic potato plants, resulted in the expected wild-type pr17 protein, as well as in larger translational products recognized by pr17-specific antisera. Mutational analyses revealed that the extra proteins were translated by non-canonical initiation at AUU codons present in the wild-type Omega sequence. In the plant system translation initiated predominantly at the AUU codon at positions 63-65 of the Omega sequence. Additional AUU codons in a different reading frame of the Omega sequence also showed the capacity for efficient translation initiation in vitro. These results extend the previously noted activity of the TMV 5' leader sequence in ribosome binding and translation enhancement in that the TMV translation enhancer can mediate non-canonical translation initiation in vitro and in vivo.