Genetic interactions between a null allele of the RIT1 gene encoding an initiator tRNA-specific modification enzyme and genes encoding translation factors in Saccharomyces cerevisiae

The Saccharomyces cerevisiae gene RIT1 encodes a phospho-ribosyl transferas e that exclusively modifies the initiator tRNA (tRNA(i)(Met)) by the additi on of a 2'-O-ribosyl phosphate group to Adenosine 64. As a result, tRNA(i)( Met) is prevented from participating in the elongation steps of protein syn thesis. We previously showed that the modification is not essential for the function of tRNA(i)(Met) in the initiation of translation, since rit1 null strains are viable and show no obvious growth defects. Here, we demonstrat e that yeast strains in which a rit1 null allele is combined with mutations in any of the genes for the three subunits of eukaryotic initiation factor -2 (eIF-2), or with disruption alleles of two of the four initiator methion ine tRNA (IMT) genes, show synergistic growth defects. A multicopy plasmid carrying an IMT gene can alleviate these defects. On the other hand, introd uction of a high-copy-number plasmid carrying the TEF2 gene, which encodes the eukaryotic elongation factor 1 alpha (eEF-1 alpha), into rit1 null stra ins with two intact IMT genes had the opposite effect, indicating that incr eased levels of eEF-1 alpha are deleterious to these strains, presumably du e to sequestration of the unmodified met-tRNA(i)(Met) for elongation. Thus, under conditions in which the components of the ternary met-tRNA(i)(Met):G TP:eIF-2 complex become limiting or are functionally impaired, the presence of the 2'-O-ribosyl phosphate modification in tRNA(i)(Met) is important fo r the provision of adequate amounts of tRNA(i)(Met) for formation of this t ernary complex.