Stop codons preceded by rare arginine codons are efficient determinants of SsrA tagging in Escherichia coli

Abstract

The SsrA or tmRNA quality control system intervenes when ribosomes stall on mRNAs and directs the addition of a Cterminal peptide tag that targets the modified polypeptide for degradation. Although hundreds of SsrA-tagged proteins can be detected in cells when degradation is prevented, most of these species have not been identified. Consequently, the mRNA sequence determinants that cause ribosome stalling and SsrA tagging are poorly understood. SsrA tagging of Escherichia coli ribokinase occurs at three specific sites at or near the C terminus of this protein. The sites of tagging correspond to ribosome stalling at the termination codon and at rare AGG codons encoding Arg-307 and Arg-309, the antepenultimate and Cterminal residues of E. coli ribokinase. Mutational analyses and studies of the effects of overexpressing the tRNA that decodes AGG reveal that the combination of a rare arginine codon at the C terminus and the adjacent inefficient UGA termination codon act to recruit the SsrA-tagging system, presumably by slowing the rate of translation elongation and termination.

Global analysis of mRNA decay and abundance in Escherichia coli at single-gene resolution using two-color fluorescent DNA microarrays

“…These observations raise the possibility that occurrence in transcripts of codons with corresponding tRNA that is limiting may stimulate mRNA decay by hindering ribosome progression (46)(47)(48)(49). However, we found no association between the frequency per unit length of a particular codon within a message and its measured half-life.…”

SmpB as the handyman of tmRNA during trans-translation

“…These sites can be (1) clusters of rare codons, 27 (2) weak termination codons. [28][29][30] They can also be the result of a flawed co-translational process, such as wrong protein folding or secretion; then, trans-translation is necessary to relieve the subsequent translational arrests, whatever the codon context. 3 The mechanism by which trans-translation is activated on these full-length mRNAs has been elusive until the discovery of the role played by the bacterial toxin RelE in inhibiting protein synthesis under nutrient deprivation conditions.…”

Proline Residues at the C Terminus of Nascent Chains Induce SsrA Tagging during Translation Termination

“…SsrA tagging of otherwise full-length proteins has also been observed, suggesting that ribosomes stall under some circumstances at stop codons (8,10). The identity of the stop codon can affect SsrA tagging of full-length proteins, with less efficient stop codons leading to higher levels of tagging (10). Because termination of translation is a relatively slow process compared with translation elongation (11), SsrA and protein release factors probably compete for binding to the A-site, while the ribosome idles at an inefficient stop codon.…”

“…For example, SsrA tagging has been demonstrated for proteins synthesized from truncated messages lacking in-frame stop codons and at protein positions corresponding to rare codons (4,(7)(8)(9)(10). Ribosomes would be expected to stall at the 3Ј end of the incomplete mRNA or at the rare codon mRNA position when the cognate charged tRNA was scarce.…”

“…Ribosomes would be expected to stall at the 3Ј end of the incomplete mRNA or at the rare codon mRNA position when the cognate charged tRNA was scarce. SsrA tagging of otherwise full-length proteins has also been observed, suggesting that ribosomes stall under some circumstances at stop codons (8,10). The identity of the stop codon can affect SsrA tagging of full-length proteins, with less efficient stop codons leading to higher levels of tagging (10).…”