Serine 577 Is Phosphorylated and Negatively Affects the tRNA Binding and eIF2α Kinase Activities of GCN2

Garcia-Barrio, Minerva T.; Dong, Jinsheng; Cherkasova, Vera A.; Zhang, Xiaolong; Zhang, Fan; Ufano, Sandra; Lai, Ruby; Qin, Jun; Hinnebusch, Alan G.

doi:10.1074/jbc.m203187200

Cited by 42 publications

(49 citation statements)

References 41 publications

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“…TORC1 deploys a complex phosphatase-signaling network involving Tap42 and Sit4 (Loewith et al 2002) and indirectly mediates the inhibitory phosphorylation of Gcn2 on Ser577, providing a link between nitrogen and amino acid signaling (Cherkasova and Hinnebusch 2003). Inhibition of TORC1 activates Sit4 leading to dephosphorylation of Ser577 and constitutive activation of Gcn2 via enhanced binding of uncharged tRNAs to the Gcn2 HisRS domain (Garcia-Barrio et al 2002;Kubota et al 2003). Genome-wide studies confirm that Gcn4 targets are activated by rapamycin treatment, an allosteric TORC1 inhibitor (Staschke et al 2010).…”

Section: Gcn4mentioning

confidence: 85%

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

2016

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In this review, we provide an overview of protein synthesis in the yeast Saccharomyces cerevisiae. The mechanism of protein synthesis is well conserved between yeast and other eukaryotes, and molecular genetic studies in budding yeast have provided critical insights into the fundamental process of translation as well as its regulation. The review focuses on the initiation and elongation phases of protein synthesis with descriptions of the roles of translation initiation and elongation factors that assist the ribosome in binding the messenger RNA (mRNA), selecting the start codon, and synthesizing the polypeptide. We also examine mechanisms of translational control highlighting the mRNA cap-binding proteins and the regulation of GCN4 and CPA1 mRNAs.

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Section: Gcn4mentioning

confidence: 85%

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

2016

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“…Strains used in this study are listed in Table 1. Plasmids encoding the FLAG epitope-tagged Gcn2(pDH101) and FLAG epitope-tagged Gcn2-S577A(pCB149) as well as the GCN4-lacZ reporter in plasmid p180 were generously provided by Alan Hinnebusch and were described previously (9,15,19). Yeast was grown on yeast extract-peptonedextrose (YEPD) or synthetic complete medium as described elsewhere (42).…”

Section: Methodsmentioning

confidence: 99%

TOR Controls Transcriptional and Translational Programs via Sap-Sit4 Protein Phosphatase Signaling Effectors

Rohde

Campbell

Zurita-Martinez

et al. 2004

Molecular and Cellular Biology

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The Tor kinases are the targets of the immunosuppressive drug rapamycin and couple nutrient availability to cell growth. In the budding yeast Saccharomyces cerevisiae, the PP2A-related phosphatase Sit4 together with its regulatory subunit Tap42 mediates several Tor signaling events. Sit4 interacts with other potential regulatory proteins known as the Saps. Deletion of the SAP or SIT4 genes confers increased sensitivity to rapamycin and defects in expression of subsets of Tor-regulated genes. Sap155, Sap185, or Sap190 can restore these responses. Strains lacking Sap185 and Sap190 are hypersensitive to rapamycin, and this sensitivity is Gcn2 dependent and correlated with a defect in translation, constitutive eukaryotic initiation factor 2␣ hyperphosphorylation, induction of GCN4 translation, and hypersensitivity to amino acid starvation. We conclude that Tor signals via Sap-Sit4 complexes to control both transcriptional and translational programs that couple cell growth to amino acid availability.

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“…Recently, we and others showed that a direct interaction between GCN1 and GCN2 is necessary for general control response, thereby providing the first insight into the underlying mechanism (15,16).…”

mentioning

confidence: 99%

Budding Yeast GCN1 Binds the GI Domain to Activate the eIF2α Kinase GCN2

Konishi

Ota

Sakaki

et al. 2001

Journal of Biological Chemistry

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When starved for a single amino acid, the budding yeast Saccharomyces cerevisiae activates the eukaryotic initiation factor 2␣ (eIF2␣) kinase GCN2 in a GCN1-dependent manner. Phosphorylated eIF2␣ inhibits general translation but selectively derepresses the synthesis of the transcription factor GCN4, which leads to coordinated induction of genes involved in biosynthesis of various amino acids, a phenomenon called general control response. We recently demonstrated that this response requires binding of GCN1 to the GI domain occurring at the N terminus of GCN2 (Kubota, H., Sakaki, Y., and Ito, T. (2000) J. Biol. Chem. 275, 20243-20246). Here we provide the first evidence for the involvement of GCN1-GCN2 interaction in activation of GCN2 per se. We identified a C-terminal segment of GCN1 sufficient to bind the GI domain and used a novel dual bait two-hybrid method to identify mutations rendering GCN1 incapable of interacting with GCN2. The yeast bearing such an allele, gcn1-F2291L, fails to display derepression of GCN4 translation and hence general control response, as does a GI domain mutant, gcn2-Y74A, defective in association with GCN1. Furthermore, we demonstrated that phosphorylation of eIF2␣ is impaired in both mutants. Since GCN2 is the sole eIF2␣ kinase in yeast, these findings indicate a critical role of GCN1-GCN2 interaction in activation of the kinase in vivo.Protein synthesis in eukaryotic cells is suppressed by stressinduced phosphorylation of eukaryotic initiation factor 2␣ (eIF2␣) 1 on a serine residue at position 51 (1). The phosphorylation converts eIF2␣ from the substrate to an inhibitor of eIF2B, the guanine nucleotide exchange factor of eIF2; phosphorylated eIF2-GDP forms a stable complex with eIF2B to hamper recycling of eIF2-GDP to eIF2-GTP (2). Scarcity of eIF2-GTP accordingly decreases the level of the ternary complex composed of eIF2, GTP, and the charged initiator tRNA, a prerequisite for translational initiation, and hence leads to general suppression of protein synthesis. Thus, eIF2␣ kinases play pivotal roles in this famous translational control.

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Serine 577 Is Phosphorylated and Negatively Affects the tRNA Binding and eIF2α Kinase Activities of GCN2

Cited by 42 publications

References 41 publications

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

TOR Controls Transcriptional and Translational Programs via Sap-Sit4 Protein Phosphatase Signaling Effectors

Budding Yeast GCN1 Binds the GI Domain to Activate the eIF2α Kinase GCN2

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