GCN2-dependent phosphorylation of eukaryotic translation initiation factor-2α in Arabidopsis

Abstract

The yeast regulatory protein kinase, general control non-derepressible-2 (GCN2) plays a key role in general amino acid control. GCN2 phosphorylates the α subunit of the trimeric eukaryotic translation initiation factor-2 (eIF2), bringing about a decrease in the general rate of protein synthesis but an increase in the synthesis of GCN4, a transcription factor that promotes the expression of genes encoding enzymes for amino acid biosynthesis. The present study concerned the phosphorylation of Arabidopsis eIF2α (AteIF2α) by the Arabidopsis homologue of GCN2, AtGCN2, and the role of AtGCN2 in regulating genes encoding enzymes of amino acid biosynthesis and responding to virus infection. A null mutant for AtGCN2 called GT8359 was obtained and western analysis confirmed that it lacked AtGCN2 protein. GT8359 was more sensitive than wild-type Arabidopsis to herbicides that affect amino acid biosynthesis. Phosphorylation of AteIF2α occurred in response to herbicide treatment but only in wild-type Arabidopsis, not GT8359, showing it to be AtGCN2-dependent. Expression analysis of genes encoding key enzymes for amino acid biosynthesis and nitrate assimilation revealed little effect of loss of AtGCN2 function in GT8359 except that expression of a nitrate reductase gene, NIA1, was decreased. Analysis of wild-type and GT8359 plants infected with Turnip yellow mosaic virus or Turnip crinkle virus showed that AteIF2α was not phosphorylated.

Mechanism of Cytoplasmic mRNA Translation

“…Herbicides (i.e., chlorosulfuron, glyphosate) that inhibit amino acid synthesis and thus induce amino acid starvation result in induction of GCN2 and phosphorylation of eIF2α (Zhang et al, 2008b). GCN2 also functions in response to purine starvation, UV irradiation, wounding, hormones, cold shock (Lageix et al, 2008), cadmium stress (Sormani et al, 2011b), amino acid metabolism and sulfur signaling (Byrne et al, 2012), but evidently not virus infection (Zhang et al, 2008b). The phosphorylation of eIF2α in response to purine starvation was correlated with reduced large polysome complexes, suggesting that it can generally inhibit initiation (Lageix et al, 2008;Sormani et al, 2011b).…”

The Circadian Clock Modulates Global Daily Cycles of mRNA Ribosome Loading

“…In contrast, only a few endogenous cues are known to regulate translation, e.g., sucrose and amino acids (Nicolaï et al, 2006;Lageix et al, 2008;Zhang et al, 2008;Roy and von Arnim, 2013). The clock is another endogenous mechanism now known to affect translation.…”

Translational Regulation of Cytoplasmic mRNAs

“…There is evidence for this pathway in plants. First, upon amino acid starvation, which occurs when the synthesis of branched chain or other amino acids in the plastid is disrupted with a herbicide, the Arabidopsis GCN2 kinase is activated and phosphorylates eIF2a (Zhang et al, 2003;Lageix et al, 2008;Zhang et al, 2008;Li et al, 2013a). This is accompanied by a widespread drop in polysome loading of mRNAs.…”

“…One plant kinase targeting eIF2a is GCN2 (Zhang et al, 2003;Immanuel et al, 2012). In response to herbicide (Zhang et al, 2008), amino acid or purine starvation (Lageix et al, 2008), or cadmium stress (Sormani et al, 2011b) the Arabidopsis GCN2 kinase (AtGCN2) is activated and phosphorylates eIF2a followed by a reduction in global protein synthesis (Lageix et al, 2008;Zhang et al, 2008). Vice versa, overexpression of GCN2 in wheat causes numerous changes in gene expression and metabolism, most notably a reduction in free asparagine, a vehicle for nitrogen transport in plants (Byrne et al, 2012).…”