Saccharomyces cerevisiae protein phosphorylation and translation accuracy

Protein-protein and protein-rRNA interactions involving ribosomal proteins uS4 and uS5 are thought to maintain the accuracy of protein synthesis by increasing selection of cognate aminoacyl-tRNAs. Selectivity involves a major conformational change—domain closure—that stabilizes aminoacyl-tRNA in the ribosomal acceptor (A) site. This has been thought a constitutive function of the ribosome ensuring consistent accuracy. Recently, the Saccharomyces cerevisiae Ctk1 cyclin-dependent kinase was demonstrated to ensure translational accuracy and Ser238 of uS5 proposed as its target. Surprisingly, Ser238 is outside the uS4-uS5 interface and no obvious mechanism has been proposed to explain its role. We show that the true target of Ctk1 regulation is another uS5 residue, Ser176, which lies in the interface opposite to Arg57 of uS4. Based on site-specific mutagenesis, we propose that phospho-Ser176 forms a salt bridge with Arg57, which should increase selectivity by strengthening the interface. Genetic data show that Ctk1 regulates accuracy indirectly by stimulating phosphorylation of Ser176 by the kinase Ypk2. A second kinase pathway involving TORC1 and Pkc1 can inhibit this effect. The level of accuracy appears to depend on competitive action of these two pathways to regulate the level of Ser176 phosphorylation.

Methods
This dataset is the result of in vivo analysis of misreading error frequency in the yeast Saccharomyces cerevisiae using a set of misreading error reporter systems developed in our laboratory.