The molecular aetiology of tRNA synthetase depletion: induction of a GCN4 amino acid starvation response despite homeostatic maintenance of charged tRNA levels

Matthew R. McFarland, Corina D. Keller, Brandon M. Childers, Stephen A. Adeniyi, Holly Corrigall, Adélaïde Raguin, M. Carmen Romano, Ian Stansfield*

*Corresponding author for this work

Research output: Contribution to journalArticle

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Abstract

During protein synthesis, charged tRNAs deliver amino acids to translating ribosomes, and are then re-charged by tRNA synthetases (aaRS). In humans, mutant aaRS cause a diversity of neurological disorders, but their molecular aetiologies are incompletely characterised. To understand system responses to aaRS depletion, the yeast glutamine aaRS gene (GLN4) was transcriptionally regulated using doxycycline by tet-off control. Depletion of GIn4p inhibited growth, and induced a GCN4 amino acid starvation response, indicative of uncharged tRNA accumulation and Gcn2 kinase activation. Using a global model of translation that included aaRS recharging, Gln4p depletion was simulated, confirming slowed translation. Modelling also revealed that Gln4p depletion causes negative feedback that matches translational demand for Gln-tRNA(Gln) to aaRS recharging capacity. This maintains normal charged tRNA(Gln) levels despite Gln4p depletion, confirmed experimentally using tRNA Northern blotting. Model analysis resolves the paradox that GIn4p depletion triggers a GCN4 response, despite maintenance of tRNA(Gln )charging levels, revealing that normally, the aaRS population can sequester free, uncharged tRNAs during aminoacylation. Gln4p depletion reduces this sequestration capacity, allowing uncharged tRNA(Gln) to interact with Gcn2 kinase. The study sheds new light on mutant aaRS disease aetiologies, and explains how aaRS sequestration of uncharged tRNAs can prevent GCN4 activation under non-starvation conditions.

Original languageEnglish
Pages (from-to)3071-3088
Number of pages18
JournalNucleic Acids Research
Volume48
Issue number6
Early online date4 Feb 2020
DOIs
Publication statusPublished - 6 Apr 2020

Keywords

  • translation
  • tRNA synthetase
  • Saccharomyces cerevisiae
  • GCN4
  • Totally Asymmetric Simple Exclusion Process
  • FACTOR 2B COMPLEX
  • TRANSLATIONAL REGULATION
  • ESCHERICHIA-COLI
  • YEAST
  • MESSENGER-RNAS
  • IN-VIVO
  • GROWTH
  • GENOME-WIDE ANALYSIS
  • GENE-EXPRESSION
  • MUTATIONS

ASJC Scopus subject areas

  • Genetics

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