TAGC notes

PgmNr 714C: Mistranslation elicits different cellular responses based on the amino acid substitution.

Authors:
Matthew Berg ¹; Yanrui Zhu ¹; Bianca Ruiz ²; Joshua Isaacson ¹; Julie Genereaux ¹; Raphael Loll-Krippleber ³; Bryan-Joseph San Luis ³; Charles Boone ³; Grant Brown ³; Judit Villen ²; Christopher Brandl ¹
1) Department of Biochemistry, University of Western Ontario, London, ON, Canada; 2) Department of Genome Sciences, University of Washington, Seattle, WA, USA; 3) Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON, Canada

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Life does not require a perfectly accurate proteome. In fact, errors occur at a rate of one mis-incorporated amino acid in every 10⁴ to 10⁵ codons. Mistranslation, or the mis-incorporation of an amino acid that differs from what is specified by the “standard” genetic code, can also occur due to mutations in the translation machinery. Cells therefore have mechanisms to cope with the resulting errors in protein folding and aggregation. Defects in these pathways may contribute to disease due to a loss of proteostasis. Our goal was to examine how different types of mistranslation affect cells. Using three tRNA variants that mistranslate the genetic code, we investigated genetic interactions and effects of mistranslation on the proteome in Saccharomyces cerevisiae. The tRNA variants mistranslate alanine at proline codons, serine at proline codons or serine at arginine codons with frequencies of 2.9%, 4.7% and 2.8% respectively. The alanine at proline and serine at arginine mistranslating tRNAs cause ~10% increase in doubling time as measured by growth in liquid media, while the more severe serine at proline mistranslating tRNA causes ~20% increase. All mistranslating tRNAs induce a heat shock response. Synthetic genetic array analysis of the tRNAs against the yeast temperature sensitive collection revealed that all the tRNAs had negative genetic interactions with genes involved in protein folding. Interestingly, however, we found distinct differences in the genetic interactions of each tRNA. Similarly, proteome analysis using mass spectrometry identified different subsets of up and down regulated proteins, depending on the type of mistranslation. We conclude that while protein quality control mechanisms are required for all types of mistranslation, the specific amino acid substitutions effect cells in different ways. We previously found variants in human tRNAs that have the potential to mistranslate. Based on the unique genetic and proteomic responses observed for different mistranslating tRNAs, we believe that in addition to exacerbating diseases caused by protein mis-folding, naturally occurring mistranslating tRNAs have the potential to negatively influence a wider range of diseases, depending on the specific amino acid substitution caused by the mistranslation.

The above poster on residue level on protein translation fidelity.