Divergent tRNA-like element supports initiation, elongation, and termination of protein biosynthesis

• Published in: Proceedings of the National Academy of Sciences - PNAS 2003-12, Vol.100 (26), p.15410-15415
• Main Authors: Jan, E, Kinzy, T.G, Sarnow, P
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Base Sequence; binding proteins; Biochemistry; Biological Sciences; Codon, Terminator - genetics; Codons; Cricket paralysis virus; elongation factors; eukaryotic elongation factors; Gels; Genes, Reporter; insect viruses; Internal ribosome entry site; Luciferases - genetics; Messenger RNA; Open reading frames; Peptide Chain Elongation, Translational; Peptide Chain Initiation, Translational; Peptide Chain Termination, Translational; Peptide elongation factors; Peptides - chemistry; Peptides - metabolism; Protein Biosynthesis; Proteins; Ribonucleic acid; Ribosomes; Ribosomes - chemistry; Ribosomes - metabolism; RNA; RNA, Transfer - genetics; RNA, Transfer - metabolism; RNA, Transfer, Amino Acyl - genetics; RNA, Transfer, Amino Acyl - metabolism; Stop codon; Terminator codon; Transcription, Genetic; Transfer RNA; translation (genetics)
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.2535183100

The cricket paralysis virus internal ribosome entry site (IRES) can, in the absence of canonical initiation factors and initiator tRNA (Met-tRNAi), occupy the ribosomal P-site and assemble 80S ribosomes. Here we show that the IRES assembles mRNA-80S ribosome complexes by recruitment of 60S subunits to preformed IRES-40S complexes. Addition of eukaryotic elongation factors eEF1A and eEF2 and aminoacylated elongator tRNAs resulted in the synthesis of peptides, implying that the IRES RNA itself mimics the function of Met-tRNAiin the P-site to trigger the first translocation step without peptide bond formation. IRES-80S complexes that contained a stop codon in the A-site recruited eukaryotic release factor eRF1, resulting in ribosome rearrangements in a surprisingly eEF2-dependent manner. Thus, this P-site-occupying IRES directs the assembly of 80S ribosomes, sets the translational reading frame, and mimics the functions of both Met-tRNAiand peptidyl tRNA to support elongation and termination.