Ribosomal protein S28 contributes to translational accuracy in Saccharomyces cerevisiae

Abstract

A method was devised to isolate intron-containing genes, the majority of which encode ribosomal proteins, from the yeast Saccharomyces cerevisiae. A RNA processing mutant was used as a source of spliced intron structures, which were purified and used to probe a yeast genomic library. Fifteen intron-containing genes were obtained in the screen, of which many were identified as ribosomal protein genes. One gene was found to be homologous to the Escherichia coli ribosomal protein gene encoding S12, known to be involved in the accuracy of translation. Genomic Southern analysis revealed that the protein was encoded by two genes in yeast; the second was cloned from a lambda gt11 library. Both genes have been sequenced, and two dimensional gel analysis of the overexpressed protein has established that the genes in question encode ribosomal protein S28.;To determine whether S28 plays a similar role in translational accuracy, mutations in S28, at a site known to be involved in translation in S12, were generated by the polymerase chain reaction. These mutations were tested for their effect on translational suppression and aminoglycoside sensitivity under conditions in which one of the wild type S28 genes had been disrupted, and in conjunction with a previously identified omnipotent suppressor mutation, SUP44. Significant alterations in suppression and antibiotic sensitivity in cells containing background levels of wild type protein have been detected, indicating that the role of this protein in translation has been conserved between prokaryotes and eukaryotes.