Ribosome rescue by Escherichia coli ArfA (YhdL) in the absence of trans-translation system.
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 SsrA(DD) , 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.
Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan.
Yuhei
Y
Chadani
Katsuhiko
K
Ono
Shin-Ichiro
S
Ozawa
Yuichiro
Y
Takahashi
Kazuyuki
K
Takai
Hideaki
H
Nanamiya
Yuzuru
Y
Tozawa
Kazuhiro
K
Kutsukake
Tatsuhiko
T
Abo
D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D005798:Genes, Bacterial; D020043:Genes, Essential; D014176:Protein Biosynthesis; D012329:RNA, Bacterial / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
C556014:ArfA protein, E coli; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; C096222:tmRNA