uS3/Rps3 controls fidelity of translation termination and programmed stop codon readthrough in co-operation with eIF3

Abstract

Ribosome was long considered as a critical yet passive player in protein synthesis. Only recently the role of its basic components, ribosomal RNAs and proteins, in translational control has begun to emerge. Here we examined function of the small ribosomal protein uS3/Rps3, earlier shown to interact with eukaryotic translation initiation factor eIF3, in termination. We identified two residues in consecutive helices occurring in the mRNA entry pore, whose mutations to the opposite charge either reduced (K108E) or increased (R116D) stop codon readthrough. Whereas the latter increased overall levels of eIF3-containing terminating ribosomes in heavy polysomes in vivo indicating slower termination rates, the former specifically reduced eIF3 amounts in termination complexes. Combining these two mutations with the readthrough-reducing mutations at the extreme C-terminus of the a/Tif32 subunit of eIF3 either suppressed (R116D) or exacerbated (K108E) the readthrough phenotypes, and partially corrected or exacerbated the defects in the composition of termination complexes. In addition, we found that K108 affects efficiency of termination in the termination context-specific manner by promoting incorporation of readthrough-inducing tRNAs. Together with the multiple binding sites that we identified between these two proteins, we suggest that Rps3 and eIF3 closely co-operate to control translation termination and stop codon readthrough.

Structural impact of K63 ubiquitin on yeast translocating ribosomes under oxidative stress

“…3B) and are important for ribosome maturation and assembly (39,40). Although the uS3-h16 latch has been observed in arrested and terminating ribosomes (41), structural changes in uS3 could also impact initiation and binding to mRNA (SI Appendix, Fig. S7A) (31).…”

Two “Edges” in Our Knowledge on the Functions of Ribosomal Proteins: The Revealed Contributions of Their Regions to Translation Mechanisms and the Issues of Their Extracellular Transport by Exosomes

“…The central region of the RP uS3 (RPS3) contains several conserved basic residues (R116, R117, R146, and K148 in the mammalian protein) that are believed to interact with the mRNA stretch between the entry site and the decoding area [28,[58][59][60]. Residues R116/117 are located at the small subunit close to the mRNA entry pore, and mutation studies with yeasts have demonstrated that they are essential for stabilizing interactions between mRNA and the ribosome at the mRNA entry channel at the stages of the formation of pre-initiation and initiation complexes [58].…”

Multifaceted control of mRNA translation machinery in cancer

“…Binding of eIF3 translation factors to the pre-termination 80S ribosome, likely through interaction with the ribosomal protein uS3/Rps3 (Poncová, Wagner et al 2019), was also shown to interfere with the eRF1-mediated decoding of STOP codons that are set in an unfavorable termination context (Beznosková, Wagner et al 2015). This function is important for re-initiation of mRNAs with uORF and translation of important stress-response protein coding mRNAs such as ATF4 under stress conditions (Hronová, Mohammad et al 2017).…”