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Effect of tandem repeated AUG codons on translation efficiency of eukaryotic mRNA carrying a short leader sequence

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Abstract

The effect on translation of multiple copies of the initiation codon AUG at the initiation site in a eukaryotic mRNA carrying a short leader sequence was tested in translation experiments in vitro. DNA, corresponding to a chimeric mRNA sequence consisting of the 5′ leader region of brome mosaic virus (BMV) RNA4 and the goat pre-α-lactalbumin mRNA sequence, was prepared and transcribed in vitro using SP6 RNA polymerase. Site-directed mutagenesis was carried out to change the sequence around the initiation codon AUG. In a wheat germ translation system, the yield of protein obtained using the mRNA with a duplication of the AUG codons at the initiation site was 1.6 times that achieved when only one AUG was present. The rate of formation of the 80S initiation complex was measured by the ribosome binding assay using cycloheximide. A good correlation was observed between the ability to form the complex and translation efficiency.

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References

  1. Boissel JP, Kasper TJ, Bunn HF (1988) Cotranslational aminoterminal processing of cytosolic proteins. J Biol Chem 263:8443–8449

  2. Jobling SA, Gehrke L (1987) Enhanced translation of chimaeric messenger RNAs containing a plant viral untranslated leader sequence. Nature 325:622–625

  3. Kozak M (1983) Comparison of initiation of protein synthesis in prokaryotes, eukaryotes, and organelles. Microbiol Rev 47:1–45

  4. Kozak M (1991) Structural features in eukaryotic mRNAs that modulate the initiation of translation. J Biol Chem 266:19867–19870

  5. Kramer W, Drutsa V, Jansen HW, Kramer B, Pflugfelder M, Fritz HJ (1984) The gapped duplex DNA approach to oligonucleotidedirected mutation construction. Nucleic Acids Res 12:9441–9456

  6. Kumagai I, Tamaki E, Kakinuma S, Miura K (1987) Molecular cloning and sequencing of cDNA encoding goat pre α-lactalbumin. J Biochem 101:511–517

  7. Melton DA, Krieg PA, Rebagliati MR, Maniatis T, Zinn K, Green MR (1984) Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res 12:7035–7056

  8. Shih DS, Kaesberg P (1973) Translation of brome mosaic viral ribonucleic acid in a cell-free system derived from wheat embryo. Proc Natl Acad Sci USA 7:1799–1803

  9. Shih DS, Lane LC, Kaesberg P (1972) Origin of the small component of brome mosaic virus RNA. J Mol Biol 64:353–362

  10. Steitz JA (1980) RNA-RNA interactions during polypeptide chain initiation. In: Chambliss G, Craven GR, Davies J, Davis K, Kahan L, Nomura M (eds) Ribosomes: structure, function and genetics. University Park Press, Baltimore, pp 479–495

  11. Yamaguchi K, Hidaka S, Miura K (1982) Relationship between structure of the 5′ noncoding region of viral mRNA and efficiency in the initiation step of protein synthesis in a eukaryotic system. Proc Natl Acad Sci USA 79:1012–1016

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Author information

Correspondence to Kin-ichiro Miura.

Additional information

Communicated by M. Sekiguchi

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Wakiyama, M., Hirao, I., Kumagai, I. et al. Effect of tandem repeated AUG codons on translation efficiency of eukaryotic mRNA carrying a short leader sequence. Molec. Gen. Genet. 238, 59–64 (1993). https://doi.org/10.1007/BF00279531

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Key words

  • Eukaryotic mRNA
  • Initiation codon AUG
  • In vitro translation
  • Translation efficiency