Interaction between the tRNA-binding and C-terminal domains of Yeast Gcn2 regulates kinase activity in vivo

Sebastien Lageix, Jinwei Zhang, Stefan Rothenburg, Alan G. Hinnebusch

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

The stress-activated protein kinase Gcn2 regulates protein synthesis by phosphorylation of translation initiation factor eIF2α. Gcn2 is activated in amino acid-deprived cells by binding of uncharged tRNA to the regulatory domain related to histidyl-tRNA synthetase, but the molecular mechanism of activation is unclear. We used a genetic approach to identify a key regulatory surface in Gcn2 that is proximal to the predicted active site of the HisRS domain and likely remodeled by tRNA binding. Mutations leading to amino acid substitutions on this surface were identified that activate Gcn2 at low levels of tRNA binding (Gcd- phenotype), while other substitutions block kinase activation (Gcn- phenotype), in some cases without altering tRNA binding by Gcn2 in vitro. Remarkably, the Gcn- substitutions increase affinity of the HisRS domain for the C-terminal domain (CTD), previously implicated as a kinase autoinhibitory segment, in a manner dampened by HisRS domain Gcd- substitutions and by amino acid starvation in vivo. Moreover, tRNA specifically antagonizes HisRS/CTD association in vitro. These findings support a model wherein HisRS-CTD interaction facilitates the autoinhibitory function of the CTD in nonstarvation conditions, with tRNA binding eliciting kinase activation by weakening HisRS-CTD association with attendant disruption of the autoinhibitory KD-CTD interaction.

Original languageEnglish (US)
Pages (from-to)e1004991
JournalPLoS Genetics
Volume11
Issue number2
DOIs
StatePublished - Feb 1 2015
Externally publishedYes

Fingerprint

Transfer RNA
yeast
phosphotransferases (kinases)
substitution
Phosphotransferases
Yeasts
yeasts
amino acid
phenotype
amino acids
amino acid substitution
Amino Acid Substitution
mitogen-activated protein kinase
active sites
translation (genetics)
protein
starvation
Histidine-tRNA Ligase
phosphorylation
protein synthesis

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Molecular Biology
  • Genetics
  • Genetics(clinical)
  • Cancer Research

Cite this

Interaction between the tRNA-binding and C-terminal domains of Yeast Gcn2 regulates kinase activity in vivo. / Lageix, Sebastien; Zhang, Jinwei; Rothenburg, Stefan; Hinnebusch, Alan G.

In: PLoS Genetics, Vol. 11, No. 2, 01.02.2015, p. e1004991.

Research output: Contribution to journalArticle

Lageix, Sebastien ; Zhang, Jinwei ; Rothenburg, Stefan ; Hinnebusch, Alan G. / Interaction between the tRNA-binding and C-terminal domains of Yeast Gcn2 regulates kinase activity in vivo. In: PLoS Genetics. 2015 ; Vol. 11, No. 2. pp. e1004991.
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