Project description:The increased selection pressure of the herbicide glyphosate has played a role in the evolution of glyphosate-resistance in weedy species, an issue that is becoming a threat to global agriculture. The molecular components involved in the cellular toxicity response to this herbicide at the expression level are still unidentified.In this study, we identify the protein kinase GCN2 as a cellular component that fosters the action of glyphosate in the model plant Arabidopsis thaliana. Comparative studies using wild-type and gcn2 knock-out mutant seedlings show that the molecular programme that the plant deploys after the treatment with the herbicide, is compromised in gcn2. Moreover, gcn2 adult plants show a lower inhibition of photosynthesis, and both seedlings and adult gcn2 plants accumulate less shikimic acid than wild-type after treatment with glyphosate.These results points to an unknown GCN2-dependent factor involved in the cascade of events triggered by glyphosate in plants. Data suggest either that the herbicide does not equally reach the target-enzyme in a gcn2 background, or that a decreased flux in the shikimate pathway in a gcn2 plants minimize the impact of enzyme inhibition.
Project description:The yeast regulatory protein kinase, general control non-derepressible-2 (GCN2) plays a key role in general amino acid control. GCN2 phosphorylates the alpha 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 eIF2alpha (AteIF2alpha) 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 AteIF2alpha 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 AteIF2alpha was not phosphorylated.
Project description:In yeast, the interaction of General Control Non-derepressible 1 (GCN1) with GCN2 enables GCN2 to phosphorylate eIF2? (the alpha subunit of eukaryotic translation initiation factor 2) under a variety of stresses. Here, we cloned AtGCN1, an Arabidopsis homologue of GCN1. We show that AtGCN1 directly interacts with GCN2 and is essential for the phosphorylation of eIF2? under salicylic acid (SA), ultraviolet (UV), cold stress and amino acid deprivation conditions. Two mutant alleles, atgcn1-1 and atgcn1-2, which are defective in the phosphorylation of eIF2?, showed increased sensitivity to cold stress, compared with the wild type. Ribosome-bound RNA profiles showed that the translational state of mRNA was higher in atgcn1-1 than in the wild type. Our result also showed that cold treatment reduced the tendency of the tor mutant seedlings to produce purple hypocotyls. In addition, the kinase activity of TOR was transiently inhibited when plants were exposed to cold stress, suggesting that the inhibition of TOR is another pathway important for plants to respond to cold stress. In conclusion, our results indicate that the AtGCN1-mediated phosphorylation of eIF2?, which is required for inhibiting the initiation of protein translation, is essential for cold tolerance in Arabidopsis.
Project description:To understand the role of GCN2 in regulating translation, we compared the polysome loading state and overall transcript level between Arabidopsis thaliana wild type (ecotype Landsberg erecta) and gcn2 (Genetrap line GT8359, Cold Spring Harbor Laboratory) seedlings with or without herbicide chlorosufuron treatment RNA was fractionated using sucrose gradients into polysomal and nonpolysomal RNAs. We also determined overall total transcript levels. We used Affymetrix ATH1 microarrays. Overall design: wild type and gcn2 mutant were compared in their total transcript and translation state. RNA was fractionated using sucrose gradients into nonpolysomal and polysomal RNAs. The sucrose gradient fractions and the total RNA were analyzed using Affymetrix ATH1 microarrays (platform GPL198).
Project description:Comparison of Arabidopsis thaliana (Arabidopsis) gene expression induced by Myzus persicae (green peach aphid) feeding, aphid saliva infiltration and abscisic acid (ABA) treatment showed a significant positive correlation. In particular, ABA-regulated genes are over-represented among genes that are induced by M. persicae saliva infiltration into Arabidopsis leaves. This suggests that the induction of ABA-related gene expression could be an important component of the Arabidopsis-aphid interaction. Consistent with this hypothesis, M. persicae populations induced ABA production in wild-type plants. Furthermore, aphid populations were smaller on Arabidopsis aba1-1 mutants, which cannot synthesize ABA, and showed a significant preference for wild-type plants compared with the mutant. Total free amino acids, which play an important role in aphid nutrition, were not altered in the aba1-1 mutant line, but the levels of isoleucine (Ile) and tryptophan (Trp) were differentially affected by aphids in wild-type and mutant plants. Recently, indole glucosinolates have been shown to promote aphid resistance in Arabidopsis. In this study, 4-methoxyindol-3-ylmethylglucosinolate was more abundant in the aba1-1 mutant than in wild-type Arabidopsis, suggesting that the induction of ABA signals that decrease the accumulation of defence compounds may be beneficial for aphids.