Ribosome rescue by Escherichia coli ArfA (YhdL) in the absence of trans-translation system

Yuhei Chadani, Katsuhiko Ono, Shinichiro Ozawa, Yuichiro Takahashi, Kazuyuki Takai, Hideaki Nanamiya, Yuzuru Tozawa, Kazuhiro Kutsukake, Tatsuhiko Abo

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Abstract

Although SsrA(tmRNA)-mediated trans-translation is thought to maintain the translation capacity of bacterial cells by rescuing ribosomes stalled on messenger RNA lacking an in-frame stop codon, single disruption of ssrA does not crucially hamper growth of Escherichia coli. Here, we identified YhdL (renamed ArfA for alternative ribosome-rescue factor) as a factor essential for the viability of E. coli in the absence of SsrA. The ssrA-arfA synthetic lethality was alleviated by SsrADD, an SsrA variant that adds a proteolysis-refractory tag through trans-translation, indicating that ArfA-deficient cells require continued translation, rather than subsequent proteolysis of the truncated polypeptide. In accordance with this notion, depletion of SsrA in the ΔarfA background led to reduced translation of a model protein without affecting transcription, and puromycin, a codon-independent mimic of aminoacyl-tRNA, rescued the bacterial growth under such conditions. That ArfA takes over the role of SsrA was suggested by the observation that its overexpression enabled detection of the polypeptide encoded by a model non-stop mRNA, which was otherwise SsrA-tagged and degraded. In vitro, purified ArfA acted on a ribosome-nascent chain complex to resolve the peptidyl-tRNA. These results indicate that ArfA rescues the ribosome stalled at the 3' end of a non-stop mRNA without involving trans-translation.

Original languageEnglish
Pages (from-to)796-808
Number of pages13
JournalMolecular Microbiology
Volume78
Issue number4
DOIs
Publication statusPublished - Nov 2010

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Ribosomes
Escherichia coli
Messenger RNA
Proteolysis
Puromycin
Peptides
Terminator Codon
Growth
Transfer RNA
Codon
Proteins

ASJC Scopus subject areas

  • Molecular Biology
  • Microbiology

Cite this

Ribosome rescue by Escherichia coli ArfA (YhdL) in the absence of trans-translation system. / Chadani, Yuhei; Ono, Katsuhiko; Ozawa, Shinichiro; Takahashi, Yuichiro; Takai, Kazuyuki; Nanamiya, Hideaki; Tozawa, Yuzuru; Kutsukake, Kazuhiro; Abo, Tatsuhiko.

In: Molecular Microbiology, Vol. 78, No. 4, 11.2010, p. 796-808.

Research output: Contribution to journalArticle

Chadani, Yuhei ; Ono, Katsuhiko ; Ozawa, Shinichiro ; Takahashi, Yuichiro ; Takai, Kazuyuki ; Nanamiya, Hideaki ; Tozawa, Yuzuru ; Kutsukake, Kazuhiro ; Abo, Tatsuhiko. / Ribosome rescue by Escherichia coli ArfA (YhdL) in the absence of trans-translation system. In: Molecular Microbiology. 2010 ; Vol. 78, No. 4. pp. 796-808.
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