Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p

John M. Zaborske, Jana Narasimhan, Li Jiang, Sheree A. Wek, Kimberly A. Dittmar, Florien Freimoser, Tao Pan, Ronald Wek

Research output: Contribution to journalArticle

72 Citations (Scopus)

Abstract

When cells are subjected to nutritional stress, uncharged tRNAs accumulate and activate Gcn2p phosphorylation of eukaryotic initiation factor-2 (eIF2) and the general amino acid control pathway. The Gcn2p regulatory domain homologous to histidyl-tRNA synthetases is proposed to bind to uncharged tRNA, directly contributing to activation of Gcn2p. Here we apply a microarray technology to analyze genome-wide changes in tRNA charging in yeast upon activation of Gcn2p in response to amino acid starvation and high salinity, a stress not directly linked to nutritional deficiency. This microarray technology is applicable for all eukaryotic cells. Strains were starved for histidine, leucine, or tryptophan and shown to rapidly induce Gcn2p phosphorylation of eIF2. The relative charging level of all tRNAs was measured before and after starvation, and Gcn2p activation and the intracellular levels of the starved amino acid correlate with the observed decrease in tRNA charging. Interestingly, in some cases, tRNAs not charged with the starved amino acid became deacylated more rapidly than tRNAs charged with the starved amino acid. This increase in uncharged tRNA levels occurred although the intracellular levels for these non-starved amino acids remained unchanged. Additionally, treatment of a wild-type strain with high salinity stress showed transient changes in the charging of several different tRNAs. These results suggest that Gcn2p can be activated by many different tRNA species in the cell. These results also depict a complex cellular relationship between tRNA charging, amino acid availability, and non-nutrient stress. These relationships are best revealed by simultaneous monitoring of the charging level of all tRNAs.

Original languageEnglish
Pages (from-to)25254-25267
Number of pages14
JournalJournal of Biological Chemistry
Volume284
Issue number37
DOIs
StatePublished - Sep 11 2009

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Eukaryotic Initiation Factor-2
Transfer RNA Aminoacylation
Transfer RNA
Phosphotransferases
Genes
Chemical activation
Genome
Amino Acids
Salinity
Starvation
tryptophan-leucine
Phosphorylation
Microarrays
Histidine-tRNA Ligase
Amino Acid-Specific Transfer RNA
Technology
Eukaryotic Cells
Histidine
Malnutrition
Leucine

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Zaborske, J. M., Narasimhan, J., Jiang, L., Wek, S. A., Dittmar, K. A., Freimoser, F., ... Wek, R. (2009). Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p. Journal of Biological Chemistry, 284(37), 25254-25267. https://doi.org/10.1074/jbc.M109.000877

Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p. / Zaborske, John M.; Narasimhan, Jana; Jiang, Li; Wek, Sheree A.; Dittmar, Kimberly A.; Freimoser, Florien; Pan, Tao; Wek, Ronald.

In: Journal of Biological Chemistry, Vol. 284, No. 37, 11.09.2009, p. 25254-25267.

Research output: Contribution to journalArticle

Zaborske, JM, Narasimhan, J, Jiang, L, Wek, SA, Dittmar, KA, Freimoser, F, Pan, T & Wek, R 2009, 'Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p', Journal of Biological Chemistry, vol. 284, no. 37, pp. 25254-25267. https://doi.org/10.1074/jbc.M109.000877
Zaborske JM, Narasimhan J, Jiang L, Wek SA, Dittmar KA, Freimoser F et al. Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p. Journal of Biological Chemistry. 2009 Sep 11;284(37):25254-25267. https://doi.org/10.1074/jbc.M109.000877
Zaborske, John M. ; Narasimhan, Jana ; Jiang, Li ; Wek, Sheree A. ; Dittmar, Kimberly A. ; Freimoser, Florien ; Pan, Tao ; Wek, Ronald. / Genome-wide analysis of tRNA charging and activation of the eIF2 kinase Gcn2p. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 37. pp. 25254-25267.
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