The eIF1A C-terminal domain promotes initiation complex assembly, scanning and AUG selection in vivo

Christie A. Fekete, Drew J. Applefield, Stephen A. Blakely, Nikolay Shirokikh, Tatyana Pestova, Jon R. Lorsch, Alan G. Hinnebusch

Research output: Contribution to journalArticle

Abstract

Translation initiation factor 1A stimulates 40S-binding of the eukaryotic initiation factor 2 (eIF2)/GTP/Met-tRNAiMet ternary complex (TC) and promotes scanning in vitro. eIF1A contains an OB-fold present in bacterial IF1 plus N- and C-terminal extensions. Truncating the C-terminus (ΔC) or mutating OB-fold residues (66-70) of eIF1A reduced general translation in vivo but increased GCN4 translation (Gcd- phenotype) in a manner suppressed by overexpressing TC. Consistent with this, both mutations diminished 40S-bound TC, eIF5 and eIF3 in vivo, and ΔC impaired TC recruitment in vitro. The assembly defects of the OB-fold mutation can be attributed to reduced 40S-binding of eIF1A, whereas ΔC impairs eIF1A function on the ribosome. A substitution in the C-terminal helix (98-101) also reduced 43S assembly in vivo. Rather than producing a Gcd- phenotype, however, 98-101 impairs GCN4 derepression in a manner consistent with defective scanning by reinitiating ribosomes. Indeed, 98-101 allows formation of aberrant 48S complexes in vitro and increases utilization of non-AUG codons in vivo. Thus, the OB-fold is crucial for ribosome-binding and the C-terminal domain of eIF1A has eukaryotic-specific functions in TC recruitment and scanning.

Original languageEnglish (US)
Pages (from-to)3588-3601
Number of pages14
JournalEMBO Journal
Volume24
Issue number20
DOIs
StatePublished - Oct 19 2005

Keywords

  • GCN4
  • Scanning
  • Translation
  • eIF1A
  • eIF2

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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  • Cite this

    Fekete, C. A., Applefield, D. J., Blakely, S. A., Shirokikh, N., Pestova, T., Lorsch, J. R., & Hinnebusch, A. G. (2005). The eIF1A C-terminal domain promotes initiation complex assembly, scanning and AUG selection in vivo. EMBO Journal, 24(20), 3588-3601. https://doi.org/10.1038/sj.emboj.7600821