Abstract
In Saccharomyces cerevisiae, phosphorylation of the alpha subunit of translation initiation factor 2 (eIF-2) by protein kinase GCN2 stimulates translation of GCN4 mRNA. In mammalian cells, phosphorylation of eIF-2 alpha inhibits the activity of eIF-2B, the GDP-GTP exchange factor for eIF-2. We present biochemical evidence that five translational regulators of GCN4 encoded by GCD1, GCD2, GCD6, GCD7, and GCN3 are components of a protein complex that stably interacts with eIF-2 and represents the yeast equivalent of eIF-2B. In vitro, this complex catalyzes guanine nucleotide exchange on eIF-2 and overcomes the inhibitory effect of GDP on formation of eIF-2.GTP.Met-initiator tRNA(Met) ternary complexes. This finding suggests that mutations in GCD-encoded subunits of the complex derepress GCN4 translation because they mimic eIF-2 alpha phosphorylation in decreasing eIF-2B activity. Our results indicate that translational control of GCN4 involves a reduction in eIF-2B function, a mechanism used in mammalian cells to regulate total protein synthesis in response to stress.
MeSH terms
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Amino Acid Sequence
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Chromatography, Affinity
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DNA-Binding Proteins / biosynthesis
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DNA-Binding Proteins / genetics
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Eukaryotic Initiation Factor-2 / isolation & purification
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Eukaryotic Initiation Factor-2 / metabolism*
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Eukaryotic Initiation Factor-2B*
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Fungal Proteins / biosynthesis*
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Fungal Proteins / genetics
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Fungal Proteins / isolation & purification
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Fungal Proteins / metabolism*
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Gene Expression Regulation, Fungal*
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Guanine Nucleotide Exchange Factors*
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Guanosine Triphosphate / metabolism
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Kinetics
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Molecular Sequence Data
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Phosphorylation
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Protein Biosynthesis*
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Protein Kinases / biosynthesis*
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Protein Kinases / genetics
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RNA, Messenger / genetics
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RNA, Messenger / metabolism*
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RNA, Transfer, Amino Acyl / metabolism
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RNA, Transfer, Met*
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Repressor Proteins / genetics
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Repressor Proteins / isolation & purification
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Repressor Proteins / metabolism*
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism*
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Saccharomyces cerevisiae Proteins*
Substances
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DNA-Binding Proteins
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Eukaryotic Initiation Factor-2
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Eukaryotic Initiation Factor-2B
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Fungal Proteins
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GCD2 protein, S cerevisiae
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GCD6 protein, S cerevisiae
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GCD7 protein, S cerevisiae
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GCN3 protein, S cerevisiae
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Guanine Nucleotide Exchange Factors
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RNA, Messenger
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RNA, Transfer, Amino Acyl
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RNA, Transfer, Met
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Repressor Proteins
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Saccharomyces cerevisiae Proteins
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tRNA, formylmethionine-
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Guanosine Triphosphate
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Protein Kinases