The accuracy of translation in Escherichia coli is profoundly influenced by three interacting ribosomal proteins, S12, S4, and S5. Mutations at lysine-42 of S12, originally isolated as causing resistance to streptomycin, increase accuracy. Countervailing "ribosomal ambiguity mutations" (ram) in S4 or S5 decrease accuracy. In the eukaryotic ribosome of Saccharomyces cerevusiae, mutations in SUP46 and SUP44, encoding the proteins equivalent to S4 and S5, lead to omnipotent suppression-i.e., to less accurate translation. The evolution of ribosomal protein S12 can be traced, by comparison with archaebacteria and Tetrahymena, to S28 of S. cerevisiae, even though the two proteins share only very limited regions of homology. However, one region that has been conserved contains a lysine residue whose mutation leads to increased accuracy in E. coil. We have introduced into S28 of yeast the same amino acid substitutions that led to the original streptomycin-resistant mutations in E. coil. We rind that they have a profound effect on the accuracy of translation and interact with SUP44 and SUP46, just as predicted from the E. coi model. Thus, the interplay of these three proteins to provide ths optimal level of accuracy of translation has been conserved during the 2 billion years of evolution that separate E. coil from S. cerevisiae.In the selection of amino acyl-tRNAs during translation, there is a balance between rate and accuracy (reviewed in ref. 1). That this balance can be influenced by ribosomal proteins was clearly demonstrated by the work ofGorini, who showed that ribosomal protein mutations conferring resistance in Escherichia coli to the aminoglycoside streptomycin caused hyperaccurate translation (2). These mutations were found frequently in ribosomal protein S12 (3) and occurred at specific residues (4). Later work established that mutations in ribosomal proteins S4 and S5 (i.e., ram mutations, for ribosomal ambiguity) could cause a loss of translational accuracy and increased sensitivity to streptomycin (5-8).More recently, Noller's group (9, 10) established that mutations in S12, S4, and S5 can result in alterations in rRNA structure. They hypothesize that these proteins affect a structural equilibrium in the rRNA that influences the accuracy of translation.Many mutations affecting translational accuracy in the eukaryote Saccharomyces cerevisiae have been described (11). SUP44 and SUP46, two "omnipotent suppressor" mutations that suppress all three classes of nonsense mutations and that cause increased sensitivity to the aminoglycoside paromomycin, have phenotypes similar to the ram mutations of E. coli (12)(13)(14). SUP44 and SUP46 encode ribosomal proteins S4 (15) and S13 (16-18), the evolutionary homologs of the E. coli S5 and S4, respectively. While many hyperaccurate "antisuppressor" mutations have been described in yeast, none have been shown to occur in ribosomal protein genes (11). Alterations in seven ribosomal proteins have been linked to increases or decreases of translational accuracy in...

Three small-subunit ribosomal proteins shown to influence translational accuracy in Saccharomyces cerevisiae are conserved in structure and function with their procaryotic counterparts. One of these, encoded by RPS28A and RPS28B (RPS28), is comparable to bacterial S12. The others, encoded by sup44 (RPS4) or, sup46 and YS11A (RPS13), are homologues of procaryotic S5 and S4, respectively. In Escherichia coli, certain alterations in S12 cause hyperaccurate translation or antibiotic resistance that can be counteracted by other changes in S5 or S4 that reduce translational accuracy. Using site-directed and random mutagenesis, we show that different changes in RPS28 can have diametrical influences on translational accuracy or antibiotic sensitivity in yeast. Certain substitutions in the amino-terminal portion of the protein, which is diverged from the procaryotic homologues, cause varying levels of nonsense suppression or antibiotic sensitivity. Other alterations, found in the more conserved carboxyl-terminal portion, counteract SUP44- or SUP46-associated antibiotic sensitivity, mimicking E. coli results. Although mutations in these different parts of RPS28 have opposite affects on translational accuracy or antibiotic sensitivity, additive phenotypes can be observed when opposing mutations are combined in the same protein.