Deletion analysis of a translational enhancer upstream from the coat protein open reading frame of potato virus S

Summary

We have previously demonstrated that the 101 nucleotides upstream from the ATG of the potato virus S (PVS) coat protein gene has the ability to act as a translational enhancer in vitro and in vivo when fused to a range of marker genes. These 101 nucleotides, which have been designated VTE (viral translational enhancer), contain a block of nucleotide homology that is conserved between members of the carlavirus group, centered around a core sequence of CCTTTAGGTT. When deletions were generated that lacked 47 of the 5′ nucleotides, but still retained the conserved block, and analysed in vitro and in vivo, it was observed that these leaders had lost their ability to act as translational enhancers. These results suggest that the sequences 5′ to the conserved block may be acting as a translational enhancer or may be important in placing the conserved block in optimal context. This is confirmed to some extent by hybrid arrest translation results in which the effects on translation of oligonucleotides, complementary to regions within the VTE leader, were investigated. It was observed that oligonucleotides complementary to the nucleotides 5′ to the conserved block had a dramatic effect on the translational competence of transcripts derived from VTE-luciferase constructs, decreasing levels by 53%, whereas oligonucleotides complementary to sequences 3′ to the conserved block reduced levels of translation by only 15.6%.