TRANSLATION OF SINDBIS VIRUS MESSENGER-RNA - EFFECTS OF SEQUENCES DOWNSTREAM OF THE INITIATING CODON

One incentive for developing the alphavirus Sindbis virus as a vector for the expression of heterologous proteins is the very high level of viral structural proteins that accumulates in infected cells. Although replacement of the structural protein genes by a heterologous gene should lead to an equivalent accumulation of the heterologous protein, the Sindbis virus capsid protein is produced at a level 10- to 20-fold higher than that of any foreign protein. Chimeric mRNAs which contain the first 275 nucleotides of the Sindbis virus 26S mRNA fused to the lacZ gene are also translated at the higher level. The enhancing sequences, located downstream of the AUG codon that initiates translation of the capsid protein, have a predicted hairpin-like structure; deletions in this region destroy the activity. These sequences enhance translation in infected cells but have the opposite effect in uninfected cells. Furthermore, translation of this RNA in infected cells is suppressed by a second viral RNA lacking the hairpin-like structure, but translation of the latter RNA is not affected. We propose that the hairpin-like structure presents a barrier to the movement of the ribosomes during translation of mRNA. In infected cells, under conditions in which this mRNA is essentially the only RNA being translated, a slowdown in the transit of the ribosomes gives factors present at low concentrations a chance to bind to the translation complex and permits a high level of functional complexes to be formed. In uninfected cells and in infected cells translating two different viral subgenomic mRNAs, a pause in the movement of the ribosomes along the RNA is no longer an advantage, because the required factors are now usurped by other translation complexes.