The ribosomal fraction mediates the translational enhancement associated with the 5′-leader of tobacco mosaic virus

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Abstract
The Ω sequence at the 5′-terminus of tobacco mosaic virus (TMV) RNA acts as a translational enhancer. The differential in Ω-associated translational enhancement between the in vitro translation system derived from wheat germ (WG) and that from rabbit reticulocytes (MDL) was exploited to identify that lysate component which was responsible for a lysate's characteristic reponse to Ω. Using fractionated MDL and WG lysates, which were reconstituted in various combinations, the high salt-washed ribosomal fraction was determined to be the responsive element in a lysate. Analysis of Ω's ability to enhance translation was greatest at low mRNA and high ribosomal concentrations and to occur in the early phase of an in vitro translation assay. Translation of Ω-containing CAT mRNA was more sensitive to the presence of micrococcal nuclease than CAT mRNA without an Ω. In substitution experiments, WG ribosomes functioned at much reduced efficiency in MDL as did MDL ribosomes in WG lysate. The initiation factor-containing fraction of one system could not, as a whole, functionally replace that of the other and actually acted to inhibit translation in the heterologous system.