A Comprehensive tRNA Deletion Library Unravels the Genetic Architecture of the tRNA Pool

Abstract

Deciphering the architecture of the tRNA pool is a prime challenge in translation research, as tRNAs govern the efficiency and accuracy of the process. Towards this challenge, we created a systematic tRNA deletion library in Saccharomyces cerevisiae, aimed at dissecting the specific contribution of each tRNA gene to the tRNA pool and to the cell's fitness. By harnessing this resource, we observed that the majority of tRNA deletions show no appreciable phenotype in rich medium, yet under more challenging conditions, additional phenotypes were observed. Robustness to tRNA gene deletion was often facilitated through extensive backup compensation within and between tRNA families. Interestingly, we found that within tRNA families, genes carrying identical anti-codons can contribute differently to the cellular fitness, suggesting the importance of the genomic surrounding to tRNA expression. Characterization of the transcriptome response to deletions of tRNA genes exposed two disparate patterns: in single-copy families, deletions elicited a stress response; in deletions of genes from multi-copy families, expression of the translation machinery increased. Our results uncover the complex architecture of the tRNA pool and pave the way towards complete understanding of their role in cell physiology.

High-resolution mapping of transcriptional dynamics across tissue development reveals a stable mRNA–tRNA interface

“…Further, the expression of the anticodon isoacceptor families, relative to each other, was stable across mammalian development. In yeast, tRNA genes can compensate for changing levels of expression of another member of their isoacceptor family, an effect demonstrated by systematic deletion of tRNA genes (Yona et al 2013;Bloom-Ackermann et al 2014). Our data are a first direct indication that a compensatory mechanism must also operate dynamically in mammals to stabilize the collective expression of a given isoacceptor family.…”

“…We did this since prior studies have shown that sequence variation in internal regulatory sequences of tRNA genes have no clear relationship with their expression levels (Oler et al 2010;Canella et al 2012). We searched for regulatory sequence elements acting in cis that could direct the Pol III recruitment to tRNA genes as described previously (Giuliodori et al 2003;Bloom-Ackermann et al 2014). To investigate the differences in Pol III binding for every pair of timepoints, we used MEME (Bailey and Elkan 1994) and TOMTOM (Gupta et al 2007) to search for enrichment of specific motifs in the 500-bp upstream regions of differentially expressed tRNAs, using stably expressed tRNA genes as the background (Supplemental Table 10; Methods).…”

A resource for functional profiling of noncoding RNA in the yeast Saccharomyces cerevisiae

“…Although a tRNA deletion resource has already been developed and has revealed interesting and distinct cellular responses to the deletion of tRNA families (Bloom-Ackermann et al 2014), it cannot be used for large-scale studies with pooled strains, since the deletion strains are not barcoded. In this respect our collection is a step forward, and will allow comprehensive genome profiling of cellular fitness in different environments (Delneri et al 2008).…”

Modify or die? - RNA modification defects in metazoans

“…Furthermore, the ratio of cognate versus near-cognate tRNA genes can potentially affect the interference by unmodified tRNAs. 148 Finally, tissue specific RNA expression or different genetic mouse backgrounds can affect the outcome of a phenotype significantly (please refer also to the checklist in Table 4). 149,150 All these factors need to be kept in mind, when modeling RNA modification pathologies, and it is possible that certain defects in humans can be best mimicked by studying inducible pluripotent stem cells (iPSC) and differentiated cells derived from them.…”