Translational quality control mechanisms that mitigate stop codon readthrough and ensure protein homeostasis

Müller, Martin (2023): Translational quality control mechanisms that mitigate stop codon readthrough and ensure protein homeostasis. Dissertation, LMU München: Fakultät für Chemie und Pharmazie

Abstract

Cells invest tremendously to maintain the fidelity for transcription and translation to ensure accurate transmission of the genetic code into proteins. Yet, errors may occur at each stage. During transcription, errors arise at a rate of ~10-5-10-4 per base, whereas the error rate during translation is around a magnitude higher with ~10-4-10-3 amino acid misincorporations per codon. Such errors become increasingly frequent with ageing, posing a sizable risk for the organism. In some instances, this leads to missing or misread stop codons, allowing translation to continue into the 3’UTRs of transcripts. Such C-terminal extensions may interfere with the folding of proteins, or worse, promote promiscuous interactions with other proteins, which in turn may disturb cellular processes and reduce overall fitness. Translation into the polyA-tail of transcripts leads to the activation of the ribosome quality control (RQC) complex, which clears both aberrant protein and mRNA. However, in most cases, translation would be terminated at stop codons within the 3’UTR before the ribosome reaches the polyA-tail. Such readthrough events would therefore not be recognized by the RQC. While previous studies suggested that such readthrough products are recognized and efficiently cleared by cells, the underlying mechanism remained unclear. Given the decline in translational fidelity during ageing, this clearance pathway is expected to become increasingly important to release the burden on the proteostasis network. Using the nematode C. elegans as a model for ageing, we aimed to identify the quality control mechanisms mitigating translational readthrough and investigated the consequences of their failure during ageing. Using this approach, we identified in C. elegans and human cells that readthrough proteins are cleared through a coupled, two-level quality control pathway involving the BAG6 chaperone complex and the ribosome collision-sensing protein GCN1. Readthrough proteins with hydrophobic C-terminal extensions are recognized by SGTA-BAG6 and ubiquitylated by RNF126 for proteasomal degradation. Additionally, cotranslational mRNA decay mediated by GCN1 and CCR4/NOT limits the accumulation of readthrough proteins. Selective ribosome profiling uncovered a general role of GCN1 in regulating translation dynamics when ribosomes encounter non-optimal codons, a feature of 3′UTR sequences. Dysfunction of GCN1 results in mRNA and proteome imbalance, increasingly affecting transmembrane proteins and collagens during ageing. These results define GCN1 as a key factor acting during translation in maintaining protein homeostasis.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 540 Chemie
• Fakultäten: Fakultät für Chemie und Pharmazie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 28. September 2023
• 1. Bericht­erstatter:in: Hartl, F. Ulrich