Coevolution of ribosomes and the translational apparatus: The structure and function of eukaryotic ribosomal protein uS7 from yeast, Saccharomyces cerevisiae.

摘要

High-resolution structures of yeast ribosomes revealed that eukaryotic and bacterial ribosomes share a common ribonucleoprotein core with majority of the changes (addition of rRNA expansion segments and proteins) occurring on the outer shell of the ribosome. The 80S yeast ribosome contains 79 proteins, of which 46 are eukaryote-specific and 34 proteins (15 and 19 in the small and large subunit respectively) are universally conserved. Despite general similarity in sequence and structure, many conserved ribosomal proteins have evolved eukaryote-specific extensions, whose functional significance is largely unknown and/or just beginning to emerge. It has been hypothesized that eukaryote-specific extensions of the conserved ribosomal proteins have evolved to accommodate eukaryote-specific features of the eukaryotic translational apparatus. Yeast, uS7 protein (known as rpS5 in yeast) from the small (40S) ribosomal subunit has evolved an N-terminal extension of ∼60 amino acid residues in length. The eukaryote-specific N-terminal extension of rpS5 also forms contacts with N-terminal extension of uS9 (known as rpS16 in yeast).;To understand the evolutionary complexity of the eukaryotic (yeast) ribosome and the function of yeast ribosomal protein rpS5, we have obtained and characterized yeast strains in which the wild-type yeast rpS5 was replaced by its truncated (from the N-terminal end) and/or mutant variants. Mutations of rpS5 led to the disruption of the eukaryotic-specific rpS5-rpS16 interaction and impair translation initiation, cell growth and induction of GCN4 mRNA translation in a manner suggesting incomplete assembly of 48S preinitiation complexes (PICs) at upstream AUG codons in GCN4 mRNA. We have hypothesized that rpS5-rpS16 interaction modulates the correct placement of the eIF2•GTP•Met-tRNAiMet ternary complex (TC) in the P-site of the 40S ribosomal subunit and eIF5-stimulated GTP-hydrolysis/Pi release, in a manner involving an altered location of the rpS16 C-terminal tail in the 40S decoding center. The rpS5-rpS16 interaction was also suggested to influence the placement of eIF1, TC and eIF5 following start codon recognition.;Our study provided the first experimental evidence supporting the functional significance of eukaryote-specific extensions and protein-protein interactions within the ribosome that are absent in prokaryotes but represent a defining feature of eukaryotic (yeast) ribosome.