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Initiation of protein synthesis from the A site of the ribosome

J E Wilson1, T V Pestova, C U Hellen

  • 1Department of Microbiology and Immunology, Stanford University School of Medicine, California 94305, USA.

Cell
|August 31, 2000
PubMed

Insights

This study reveals that the cricket paralysis virus internal ribosome entry site can initiate translation from the A site, bypassing the need for initiator transfer RNA (tRNA) and initiation factors. This finding expands the understanding of how ribosomes initiate protein synthesis in eukaryotes.

Area of Science:

  • Molecular Biology
  • Virology
  • Genetics

Background:

  • Ribosome binding to messenger RNA (mRNA) typically involves the initiator transfer RNA (tRNA) binding to the start codon in the P site.
  • This process is mediated by eukaryotic initiation factor 2 (eIF2) and requires guanosine triphosphate (GTP) hydrolysis.

Purpose of the Study:

  • To investigate the mechanism of translation initiation at the internal ribosome entry site (IRES) of the cricket paralysis virus.
  • To determine if canonical translation initiation factors and start codon recognition are essential for IRES-mediated translation.

Main Methods:

  • Site-directed mutagenesis of the P-site codon within the viral IRES.
  • Analysis of protein products generated from the modified IRES.
  • Biochemical assays to assess the requirement for initiator tRNA, eIF2, and GTP.

Main Results:

  • The cricket paralysis virus IRES formed 80S ribosomes and initiated translation without initiator tRNA, eIF2, or GTP hydrolysis.
  • Mutagenesis of the P-site codon did not abolish translation, indicating it was not decoded.
  • Protein sequence analysis confirmed that translation initiated from the A-site codon (GCU).

Conclusions:

  • Translation initiation can occur from the A site of the ribosome, challenging the conventional model.
  • Viral IRES elements can employ alternative mechanisms for translation initiation, independent of canonical host factors.
  • This discovery broadens the potential repertoire of translated open reading frames in eukaryotic mRNAs.

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