Abstract
Translation initiation factor 1A (eIF1A) is predicted to bind in the decoding site of the 40S ribosome and has been implicated in recruitment of the eIF2-GTP-Met-tRNA i Met ternary complex (TC) and ribosomal scanning. We show that the unstructured C-terminus of eIF1A interacts with the C-terminus of eIF5B, a factor that stimulates 40S-60S subunit joining, and removal of this domain of eIF1A diminishes translation initiation in vivo. These findings support the idea that eIF1A-eIF5B association is instrumental in releasing eIF1A from the ribosome after subunit joining. A larger C-terminal truncation that removes a 3(10) helix in eIF1A deregulates GCN4 translation in a manner suppressed by overexpressing TC, implicating eIF1A in TC binding to 40S ribosomes in vivo. The unstructured N-terminus of eIF1A interacts with eIF2 and eIF3 and is required at low temperatures for a step following TC recruitment. We propose a modular organization for eIF1A wherein a core ribosome-binding domain is flanked by flexible segments that mediate interactions with other factors involved in recruitment of TC and release of eIF1A at subunit joining.
MeSH terms
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Binding Sites
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Eukaryotic Initiation Factor-1*
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Eukaryotic Initiation Factor-2 / chemistry
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Eukaryotic Initiation Factor-2 / genetics
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Eukaryotic Initiation Factor-2 / metabolism*
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Eukaryotic Initiation Factor-3 / chemistry
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Eukaryotic Initiation Factor-3 / genetics
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Eukaryotic Initiation Factor-3 / metabolism*
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Eukaryotic Initiation Factor-5 / chemistry
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Eukaryotic Initiation Factor-5 / genetics
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Eukaryotic Initiation Factor-5 / metabolism*
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Macromolecular Substances
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Peptide Chain Initiation, Translational
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Peptide Initiation Factors / chemistry
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Peptide Initiation Factors / genetics
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Peptide Initiation Factors / metabolism*
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Phenotype
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Protein Binding
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Protein Biosynthesis
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Protein Conformation*
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Ribosomes / metabolism
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Saccharomyces cerevisiae / genetics
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Saccharomyces cerevisiae / metabolism
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Saccharomyces cerevisiae Proteins / chemistry
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Two-Hybrid System Techniques
Substances
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Eukaryotic Initiation Factor-1
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Eukaryotic Initiation Factor-2
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Eukaryotic Initiation Factor-3
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Eukaryotic Initiation Factor-5
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Macromolecular Substances
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Peptide Initiation Factors
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Recombinant Fusion Proteins
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Saccharomyces cerevisiae Proteins
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eukaryotic peptide initiation factor-1A