Ambushing the ambush hypothesis: predicting and evaluating off-frame codon frequencies in Prokaryotic Genomes

Abstract

BackgroundIn this paper, we address the evidence for the Ambush Hypothesis. Proposed by Seligmann and Pollock, this hypothesis posits that there exists a selection for off-frame stop codons (OSCs) to counteract the possible deleterious effects of translational frameshifts, including the waste of resources and potential cytotoxicity. Two main types of study have been used to support the hypothesis. Some studies analyzed codon usage and showed that codons with more potential to create OSCs seem to be favored over codons with lower potential; they used this finding to support the Ambush Hypothesis. Another study used 342 bacterial genomes to evaluate the hypothesis directly, finding significant excesses of OSCs in these genomes.ResultsWe repeated both analyses with newer datasets and searched for other factors that could explain the observed trends. In the first case, the relative frequency of codons with the potential to create OSCs is directly correlated with the GC content of organisms, as stop codons are GC-poor. When evaluating the frequency of OSCs directly in 1,976 bacterial genomes we also detected a significant excess. However, when comparing the excess of OSCs with similarly obtained results for the frequency of out-of-frame sense codons, some sense codons have a more significant excess than stop codons.ConclusionsTwo avenues of study have been used to support the Ambush Hypothesis. Using the same methods as these previous studies, we demonstrate that the evidence in support of the Ambush Hypothesis does not hold up against more rigorous testing.

Adenine Enrichment at the Fourth CDS Residue in Bacterial Genes Is Consistent with Error Proofing for +1 Frameshifts

“…Furthermore, incorrectly folded mistranslated proteins may have an adverse effect on cellular interactions or form toxic aggregates ( Tank and True 2009 ). The possible evolutionary advantage of capturing these frameshifts is conjectured to be reflected by an overrepresentation of out of frame stop codons, termed the “ambush hypothesis” ( Seligmann and Pollock 2004 ; Singh and Pardasani 2009 ; Tse et al 2010 ), although the frequency with which codons that form out of frame stops are used is largely predictable from the underlying GC pressure ( Morgens et al 2013 ). Alternatively, selection to reduce costs in genomes where frameshifting is most deleterious (notably GC rich ones) can explain the richer tRNA repertoire found in such genomes ( Warnecke et al 2010 ).…”

Protein Mis-Termination Initiates Genetic Diseases, Cancers, and Restricts Bacterial Genome Expansion

“…When a larger data set was analyzed, Morgens et al found that only a slight overexpression of PSCs in close to 2000 bacterial genomes. They also found that some out-of-frame sense codons are even more overexpressed than the PSCs[155]. Bacteria with high GC content or metabolically versatile bacteria were found to contain very few PSCs in their genome raising questions regarding the validity of the ambush hypothesis[31,155].…”

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“…For example, the polyketide synthase (PKS) genes presented with significantly lower level of hidden stop codons than expected, suggesting both non-adherence to the ambush hypothesis and a suppression of hidden stop codon evolution [8]. In addition, it was reported that some sense codons have a more significant excess than stop codons [4]. These controversial results can be well explained if the emerging of the hidden stops after the occurrence of a frameshift is considered to be a signal to trigger the cell machine for readthrough and then for reading frame recovery, rather than a signal for translational termination.…”

“…The "Ambush Hypothesis" [3] presumed that most frameshifts would yield non-functional proteins, lead to waste of energy, resources and activity of the biosynthetic machinery, and some peptides synthesized after frameshifts were thought to be cytotoxic [3][4][5][6][7][8][9][10][11].…”