Antizyme frameshifting as a functional probe of eukaryotic translational termination

Zemfira N. Karamysheva, Andrey L. Karamyshev, Koichi Ito, Takashi Yokogawa, Kazuya Nishikawa, Yoshikazu Nakamura, Senya Matsufuji

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Translation termination in eukaryotes is mediated by the release factors eRF1 and eRF3, but mechanisms of the interplay between these factors are not fully understood, due partly to the difficulty of measuring termination on eukaryotic mRNAs. Here, we describe an in vitro system for the assay of termination using competition with programmed frameshifting at the recoding signal of mammalian antizyme. The efficiency of antizyme frameshifting in rabbit reticulocyte lysates was reduced by addition of recombinant rabbit eRF1 and eRF3 in a synergistic manner. Addition of suppressor tRNA to this assay system revealed competition with a third event, stop codon readthrough. Using these assays, we demonstrated that an eRF3 mutation at the GTPase domain repressed termination in a dominant negative fashion probably by binding to eRF1. The effect of the release factors and the suppressor tRNA showed that the stop codon at the antizyme frameshift site is relatively inefficient compared to either the natural termination signals at the end of protein coding sequences or the readthrough signal from a plant virus. The system affords a convenient assay for release factor activity and has provided some novel views of the mechanism of antizyme frameshifting.

Original languageEnglish (US)
Pages (from-to)5949-5956
Number of pages8
JournalNucleic Acids Research
Volume31
Issue number20
DOIs
StatePublished - Oct 15 2003

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Terminator Codon
Transfer RNA
Rabbits
Plant Viruses
GTP Phosphohydrolases
Reticulocytes
Eukaryota
Messenger RNA
Mutation
Proteins
In Vitro Techniques

ASJC Scopus subject areas

  • Genetics

Cite this

Karamysheva, Z. N., Karamyshev, A. L., Ito, K., Yokogawa, T., Nishikawa, K., Nakamura, Y., & Matsufuji, S. (2003). Antizyme frameshifting as a functional probe of eukaryotic translational termination. Nucleic Acids Research, 31(20), 5949-5956. https://doi.org/10.1093/nar/gkg789

Antizyme frameshifting as a functional probe of eukaryotic translational termination. / Karamysheva, Zemfira N.; Karamyshev, Andrey L.; Ito, Koichi; Yokogawa, Takashi; Nishikawa, Kazuya; Nakamura, Yoshikazu; Matsufuji, Senya.

In: Nucleic Acids Research, Vol. 31, No. 20, 15.10.2003, p. 5949-5956.

Research output: Contribution to journalArticle

Karamysheva, ZN, Karamyshev, AL, Ito, K, Yokogawa, T, Nishikawa, K, Nakamura, Y & Matsufuji, S 2003, 'Antizyme frameshifting as a functional probe of eukaryotic translational termination', Nucleic Acids Research, vol. 31, no. 20, pp. 5949-5956. https://doi.org/10.1093/nar/gkg789
Karamysheva ZN, Karamyshev AL, Ito K, Yokogawa T, Nishikawa K, Nakamura Y et al. Antizyme frameshifting as a functional probe of eukaryotic translational termination. Nucleic Acids Research. 2003 Oct 15;31(20):5949-5956. https://doi.org/10.1093/nar/gkg789
Karamysheva, Zemfira N. ; Karamyshev, Andrey L. ; Ito, Koichi ; Yokogawa, Takashi ; Nishikawa, Kazuya ; Nakamura, Yoshikazu ; Matsufuji, Senya. / Antizyme frameshifting as a functional probe of eukaryotic translational termination. In: Nucleic Acids Research. 2003 ; Vol. 31, No. 20. pp. 5949-5956.
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