Abstract
BackgroundThe tmRNA quality control system recognizes stalled translation complexes and facilitates ribosome recycling in a process termed ‘ribosome rescue’. During ribosome rescue, nascent chains are tagged with the tmRNA-encoded SsrA peptide, which targets tagged proteins for degradation. In Escherichia coli, tmRNA rescues ribosomes arrested on truncated messages, as well as ribosomes that are paused during elongation and termination.Methodology/Principal FindingsHere, we describe a new translational pausing determinant that leads to SsrA peptide tagging of the E. coli GalE protein (UDP-galactose 4-epimerase). GalE chains are tagged at more than 150 sites, primarily within distinct clusters throughout the C-terminal domain. These tagging sites do not correspond to rare codon clusters and synonymous recoding of the galE gene had little effect on tagging. Moreover, tagging was largely unaffected by perturbations that either stabilize or destabilize the galE transcript. Examination of GalE-thioredoxin (TrxA) fusion proteins showed that the GalE C-terminal domain is no longer tagged when fused to an N-terminal TrxA domain. Conversely, the N-terminus of GalE induced tagging within the fused C-terminal TrxA domain.Conclusions/SignificanceThese findings suggest that translation of the GalE N-terminus induces subsequent tagging of the C-terminal domain. We propose that co-translational maturation of the GalE N-terminal domain influences ribosome pausing and subsequent tmRNA activity.
Highlights
Bacteria possess several molecular quality control systems to ensure the fidelity of protein synthesis
Truncated mRNA is invariably associated with translational pauses that induce SsrA tagging – including rare codons, inefficient stop codons, and the SecM programmed ribosome arrest [16,23,39,45,46,47]
Perhaps the simplest model to explain GalE tagging is one in which the translation of partially degraded galE transcripts causes translational arrest and subsequent tmRNANSmpB activity. This model accounts for the findings of Yamamoto et al, who reported extensive tagging of protein synthesized from mRNA lacking a stable 39-structure [5]
Summary
Bacteria possess several molecular quality control systems to ensure the fidelity of protein synthesis. The tmRNANSmpB quality control system functions in all eubacteria to recycle stalled translational complexes in a reaction termed ‘ribosome rescue’. The SsrA peptide is a degradation signal that targets tagged polypeptides to a number of proteases [1,6,7,8]. These activities ensure that defective mRNAs and proteins are identified and destroyed, thereby reducing the burden of non-productive protein synthesis. The tmRNA quality control system recognizes stalled translation complexes and facilitates ribosome recycling in a process termed ‘ribosome rescue’. Nascent chains are tagged with the tmRNA-encoded SsrA peptide, which targets tagged proteins for degradation. In Escherichia coli, tmRNA rescues ribosomes arrested on truncated messages, as well as ribosomes that are paused during elongation and termination
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