title: Association of GCN1-GCN20 regulatory complex with the N-terminus of eIF2alpha kinase GCN2 is required for GCN2 activation.
abstract: Stimulation of GCN4 mRNA translation due to phosphorylation of the alpha-subunit of initiation factor 2 (eIF2) by its specific kinase, GCN2, requires binding of uncharged tRNA to a histidyl-tRNA synthetase (HisRS)-like domain in GCN2. GCN2 function in vivo also requires GCN1 and GCN20, but it was unknown whether these latter proteins act directly to promote the stimulation of GCN2 by uncharged tRNA. We found that the GCN1-GCN20 complex physically interacts with GCN2, binding to the N-terminus of the protein. Overexpression of N-terminal GCN2 segments had a dominant-negative phenotype that correlated with their ability to interact with GCN1-GCN20 and impede association between GCN1 and native GCN2. Consistently, this Gcn(-) phenotype was suppressed by overexpressing GCN2, GCN1-GCN20 or tRNA(His). The requirement for GCN1 was also reduced by overexpressing tRNA(His) in a gcn1Delta strain. We conclude that binding of GCN1-GCN20 to GCN2 is required for its activation by uncharged tRNA. The homologous N-terminus of Drosophila GCN2 interacted with yeast GCN1-GCN20 and had a dominant Gcn(-) phenotype, suggesting evolutionary conservation of this interaction.
authors.affiliation: Laboratory of Eukaryotic Gene Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: M
authors.initials: M
authors.lastname: Garcia-Barrio
authors.forename: J
authors.initials: J
authors.lastname: Dong
authors.forename: S
authors.initials: S
authors.lastname: Ufano
authors.forename: A G
authors.initials: AG
authors.lastname: Hinnebusch
mesh_terms: D018528:ATP-Binding Cassette Transporters; D000483:Alleles; D000596:Amino Acids / Q000378:metabolism; D015153:Blotting, Western; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins; D004789:Enzyme Activation; D015852:Eukaryotic Initiation Factor-2 / Q000737:chemistry / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D010445:Peptide Elongation Factors; D010641:Phenotype; D011233:Precipitin Tests; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015003:Yeasts / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D000596:Amino Acids; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D015852:Eukaryotic Initiation Factor-2; D005656:Fungal Proteins; C501389:GCN1 protein, S cerevisiae; C094764:GCN20 protein, C elegans; D010445:Peptide Elongation Factors; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D005982:Glutathione Transferase; D011494:Protein Kinases; C512912:GCN2 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases