Rous sarcoma virus translation revisited: Characterization of an internal ribosome entry segment in the 5 ' leader of the genomic RNA

The 5' leader of Rous sarcoma virus (RSV) genomic RNA and of retroviruses i n general is long and contains stable secondary structures that are critica l in the early and late steps of virus replication such as RNA dimerization and packaging and in the process of reverse transcription. The initiation of RSV Gag translation has been reported to be 5' cap dependent and control led by three short open reading frames located in the 380-nucleotide leader upstream of the Gag start codon. Translation of RSV Gag would thus differ from that prevailing in other retroviruses such as murine leukemia virus, r eticuloendotheliosis virus type A, and simian immunodeficiency virus, in wh ich an internal ribosome entry segment (IRES) in the 5' end of the genomic RNA directs efficient Gag expression despite stable 5' secondary structures . This prompted us to investigate whether RSV Gag translation might be cont rolled by an IRES-dependent mechanism. The results show that the 5' leaders of RSV and v-Src RNA exhibit IRES properties, since these viral elements c an promote efficient translation of monocistronic RNAs in conditions inhibi ting 5' cap-dependent translation. When inserted between two cistrons in a canonical bicistronic construct, both the RSV and v-Src leaders promote exp ression of the 3' cistron. A genetic analysis of the RSV leader allowed the identification of two nonoverlapping 5' and 3' leader domains with IRES ac tivity. In addition, the v-Src leader was found to contain unique 3' sequen ces promoting an efficient reinitiation of translation. Taken together, the se data lead us to propose a new model for RSV translation.