Premature translation termination mutations are efficiently suppressed in a highly conserved region of yeast Ste6p, a member of the ATP-binding cassette (ABC) transporter family.
The requirements for efficient translation termination are incompletely understood. Since the local context surrounding stop codons can influence the efficiency of translation termination, premature termination codons introduced by random mutation may not always terminate at the optimal efficiencies expected of naturally occurring stop codons. To investigate whether this could result in physiologically significant levels of read through, we examined the suppression of premature translation termination mutations within a sequence motif of the yeast Ste6 protein (Ste6p) that is highly conserved among members of the ATP-binding cassette (ABC) transporter family. The human cystic fibrosis transmembrane conductance regulator (CFTR), which is defective in individuals with the disease cystic fibrosis, is also a member of this protein family. The mutations examined in Ste6p were chosen because a premature termination codon at the corresponding residue of CFTR has previously been reported to cause less severe pulmonary involvement than some missense mutations, suggesting that low level suppression of this stop codon could be occurring. Our results indicate that these premature stop codons in Ste6p can be suppressed at frequencies as high as 10%. Characterization of this phenomenon using a beta-galactosidase read through assay system showed that a limited sequence context surrounding this site contained information that was sufficient to cause suppression of translation termination. Amino acid sequence analysis of the full-length translation products produced by read through of an amber codon demonstrated that termination suppression was mediated by near-cognate tRNA mispairing that resulted in the insertion of tyrosine, lysine, or tryptophan.
Fearon
K
K
Department of Microbiology, University of Alabama at Birmingham 35294.
McClendon
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V
Bonetti
B
B
Bedwell
D M
DM
D018528:ATP-Binding Cassette Transporters; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000145:classification; D001483:Base Sequence; D002352:Carrier Proteins / Q000145:classification; D017124:Conserved Sequence; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004271:DNA, Fungal; D005656:Fungal Proteins / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D006023:Glycoproteins; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013728:Terminator Regions, Genetic
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; C505032:CFTR protein, human; D002352:Carrier Proteins; D004271:DNA, Fungal; D005656:Fungal Proteins; D006023:Glycoproteins; D008565:Membrane Proteins; C489277:STE6 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate