tRNA modifications regulate translation during cellular stress

Abstract

The regulation of gene expression in response to stress is an essential cellular protection mechanism. Recent advances in tRNA modification analysis and genome-based codon bias analytics have facilitated studies that lead to a novel model for translational control, with translation elongation dynamically regulated during stress responses. Stress-induced increases in specific anticodon wobble bases are required for the optimal translation of stress response transcripts that are significantly biased in the use of degenerate codons keyed to these modified tRNA bases. These findings led us to introduce the notion of tRNA modification tunable transcripts (MoTTs - transcripts whose translation is regulated by tRNA modifications), which are identifiable using genome-wide codon counting algorithms. In support of this general model of translational control of stress response, studies making use of detailed measures of translation, tRNA methyltransferase mutants, and computational and mass spectrometry approaches reveal that stress reprograms tRNA modifications to translationally regulate MoTTs linked to arginine and leucine codons, which helps cells survive insults by damaging agents. These studies highlight how tRNA methyltransferase activities and MoTTs are key components of the cellular stress response.

Keywords: Codon usage; Modified ribonucleoside; Stress response; Transfer RNA; Translation.

Figure 1. Stress-induced changes in tRNA modification as measured by LC-MS/MS

Budding yeast contain 25 modified ribonucleosides, of which 23 can be measured by chromatography-coupled tandem mass spectrometry (LC-MS/MS). Following tRNA isolation and hydrolysis, individual modified ribonucleosides are resolved by reversed-phase HPLC and quantified by tandem mass spectrometry. The data are used to calculate fold-change values comparing control cells to stressed cells, with the fold-change values analyzed by multivariate statistics to identify patterns of stress-induced changes. The heat map represents fold-change data for four mechanistically distinct toxicants and shows both agent- and dose-specific signatures. The heat map image was reproduced from Chan et al. (2010) PLoS Genetics 6: e1001247.

Figure 2. tRNA modifications and their relation to stress signaling pathways

Description of mutant phenotypes for cells deficient in Trm9 and Trm4, structure of substrates and products for each enzyme catalyzed tRNA modification, the pathways regulated by each tRNA methyltransferases, and pathways whose activation is prevented (underlined) by proper tRNA modification, via the prevention of amino acid misincorporation errors.

Figure 3. Genome-based identification of MoTTs

Iterative analysis of each open reading frame is used to count the number of codons in each gene and determine the frequency of use of synonymous codons for each amino acid (Steps 1 – 3). After analysis of all genes in a genome, the average value for all genes is then used to identify specific genes that are over- (yellow) or under- (purple) using a codon, with groups of genes that have similar codon over- and under-usage patterns identified by clustering and heat map visualization (Step 4).

Figure 4. Changes in tRNA modification regulate the translation of MoTTs

The scheme depicts the concept of stress-induced tRNA reprogramming and selective translation of codon-biased mRNAs (MoTTs) for oxidative stress in budding yeast.