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.
Highlights
Messenger RNA translation is a central molecular process in any living cell and is among the most complicated and highly regulated of cellular processes [1,2]
The tRNA pool is composed of various tRNA isoacceptor families, each family carries a different anti-codon sequence that decodes the relevant codon by Watson-Crick base pairing, or codons with non-perfect base pairing of the third nucleotide by the wobble interaction. tRNA families are further classified to isotypes if they carry the same amino acid
When a deleted tRNA belongs to a family which contains multiple genes with the same anti-codon, the affected cells responded by up-regulating the translation machinery, but upon deletion of singleton tRNAs, the cellular response resembled that of proteotoxic stress
Summary
Messenger RNA translation is a central molecular process in any living cell and is among the most complicated and highly regulated of cellular processes [1,2]. Each tRNA family can be encoded by a single or multiple gene copies [9,10] It was previously shown for several organisms that the concentrations of various tRNA isoacceptors positively correlates with the tRNA family’s gene copy-number [11,12]. These observations along with detailed analysis of the relationship between gene copy-number of tRNA families and codon-usage, established the notion that the multiplicity of tRNA genes in yeast is not functionally redundant. Such multiplicity might establish the correct balance between tRNA concentrations and the codon usage in protein-coding genes [13], justifying the use of the tRNA gene copy-number as a proxy for actual tRNA amounts [12,14,15]
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