The fail-safe system to rescue the stalled ribosomes in Escherichia coli

Abstract

Translation terminates at stop codon. Without stop codon, ribosome cannot terminate translation properly and reaches and stalls at the 3'-end of the mRNA lacking stop codon. Bacterial tmRNA-mediated trans-translation releases such stalled ribosome and targets the protein product to degradation by adding specific "degradation tag." Recently two alternative ribosome rescue factors, ArfA (YhdL) and ArfB (YaeJ), have been found in Escherichia coli. These three ribosome rescue systems are different each other in terms of molecular mechanism of ribosome rescue and their activity, but they are mutually related and co-operate to maintain the translation system in shape. This suggests the biological significance of ribosome rescue.

Keywords: ArfA; ArfB; Escherichia coli; ribosome rescue; trans-translation.

FIGURE 1

Schematic representation of ribosome rescue in E. coli. (A) tmRNA (brown line)- and SmpB (blue circle)-mediated trans-translation. In trans-translation, ribosome (orange ovals) switches its translating template from non-stop mRNA (black line) to tmRNA. Then it terminates translation at the stop codon (purple box) exist on the tmRNA. Protein product will be degraded due to the SsrA-tag (shown in brown) attached to the C-terminus. (B) ArfA-mediated ribosome rescue. RF2 is indispensable in this pathway. tRNA enhances this reaction by unknown mechanism. (C) ArfB-mediated ribosome rescue. As a stop codon-independent RF homolog, ArfB binds to A-site of the ribosome within NTC and hydrolyzes peptidyl-tRNA. Only the relevant factors are shown.

FIGURE 2

Model for trans-translation-mediated regulation of ArfA expression. arfA is transcribed as mRNA with (A) or without (B) stop codon. In both cases, mRNA is cleaved by RNase III at the site indicated by triangle. This results in the non-stop mRNA formation. ArfA translated from arfA mRNA with stop codon (A, bottom) will be degraded due to its hydrophobic C-terminus (red box). Translation of arfA non-stop mRNAs (A, top; B) results in NTC formation. The NTC is normally resolved by trans-translation and translated ArfA will be degraded due to the SsrA-tag (closed box) attached to its C-terminus. Once the level of NTC exceeds the capacity of trans-translation, NTC will be resolved first by SsrA-tagged ArfA escaped from proteolysis or ArfB, then by truncated ArfA which is produced from non-stop mRNA (C). ArfA thus produced functions as a back-up ribosome rescue factor for trans-translation system. Amino acid sequence of ArfA is shown at the bottom. Positions corresponding to the inverted repeat (arrows) and RNase III cleavage site (triangle) are shown below the sequence. ArfA produced from the RNase III-processed arfA mRNA lacks its C-terminus portion which shown in gray characters.

FIGURE 3

Biological significance of ribosome rescue. Non-stop mRNA is produced either by immature transcription or degradation of mature mRNA and causes NTC formation when translated. E. coli cell is equipped with at least three ribosome rescue systems mediated by tmRNA–SmpB complex, ArfA, or ArfB. In either case, NTC is resolved and rescued ribosome can be utilized in another round of translation.