The translation machinery recognizes codons that
enter the ribosomal A site with remarkable accuracy to
ensure that polypeptide synthesis proceeds with a minimum
of errors. When a termination codon enters the A site of
a eukaryotic ribosome, it is recognized by the release
factor eRF1. It has been suggested that the recognition
of translation termination signals in these organisms is
not limited to a simple trinucleotide codon, but is instead
recognized by an extended tetranucleotide termination signal
comprised of the stop codon and the first nucleotide that
follows. Interestingly, pharmacological agents such as
aminoglycoside antibiotics can reduce the efficiency of
translation termination by a mechanism that alters this
ribosomal proofreading process. This leads to the misincorporation
of an amino acid through the pairing of a near-cognate
aminoacyl tRNA with the stop codon. To determine whether
the sequence context surrounding a stop codon can influence
aminoglycoside-mediated suppression of translation termination
signals, we developed a series of readthrough constructs
that contained different tetranucleotide termination signals,
as well as differences in the three bases upstream and
downstream of the stop codon. Our results demonstrate that
the sequences surrounding a stop codon can play an important
role in determining its susceptibility to suppression by
aminoglycosides. Furthermore, these distal sequences were
found to influence the level of suppression in remarkably
distinct ways. These results suggest that the mRNA context
influences the suppression of stop codons in response to
subtle differences in the conformation of the ribosomal
decoding site that result from aminoglycoside binding.
