Allelic amber and ochre suppressors

Ohlsson, Betsy M.; Strigini, Paolo; Beckwith, Jon

doi:10.1016/0022-2836(68)90376-8

Cited by 45 publications

(21 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition to suppressing the gene of interest, the suppressor is also expected to affect genes throughout the genome that end with amber stop codons. Reflecting the relative prevalence of amber and ochre stop codons, amber suppressors do not appear to affect the growth of their hosts, whereas isogenic ochre derivatives cause growth defects proportional to the efficiency of suppression (23,24). It has been shown that amber suppression extends and inactivates the stringent factor RelA (3), but the full extent of extraneous effects has never before been determined.…”

mentioning

confidence: 99%

Global Transcriptional Effects of a Suppressor tRNA and the Inactivation of the Regulator frmR

Herring

Blattner

2004

J Bacteriol

View full text Add to dashboard Cite

Expression of an amber suppressor tRNA should result in read-through of the 326 open reading frames (ORFs) that terminate with amber stop codons in the Escherichia coli genome, including six pseudogenes. Abnormal extension of an ORF might alter the activities of the protein and have effects on cellular physiology, while suppression of a pseudogene could lead to a gain of function. We used oligonucleotide microarrays to determine if any effects were apparent at the level of transcription in glucose minimal medium. Surprisingly, only eight genes had significantly different expression in the presence of the suppressor. Among these were the genes yaiN, adhC, and yaiM, forming a single putative operon whose likely function is the degradation of formaldehyde. Expression of wild-type yaiN was shown to result in repression of the operon, while a suppression-mimicking allele lacking the amber stop codon and extended 7 amino acids did not. The operon was shown to be induced by formaldehyde, and the genes have been renamed frmR, frmA, and frmB, respectively.A suppressor tRNA is a tRNA with a mutation (usually) in the anticodon that allows it to recognize a stop codon and insert an amino acid in its place (reviewed in reference 6). First identified in 1965 (4, 7), they have been widely used in studies of translation, phage biology, and protein engineering. They have been critical to our understanding of the structure and function, processing, and charging of tRNAs, as well as ribosome-tRNA interaction, polarity, codon context effects, and the elucidation of the genetic code (29). Suppressors have also been used to study the impact of single amino acid substitutions and nonstandard amino acids on protein function (15,30). Amber suppressors have been used preferentially because amber (UAG) is the least common of the three stop codons, occurring at the end of 326 out of 4,290 open reading frames (ORFs) in the Escherichia coli genome. In contrast, ochre (UAA) occurs 2,706 times and opal (UGA) occurs 1,258 times. The reasons for this dramatic skew are not known, though it is interesting that 1.5% of fecal coliform isolates (mostly E. coli) were found to contain natural amber suppressors (18). Out of 13 pseudogenes in strain K-12 (identified from the EcoGene website [26] at http://bmb.med.miami.edu/EcoGene/EcoWeb/), 6 are disrupted with amber stop codons (glvC, ybfH, yehQ, yfcU, yjiP, and b2650) and almost all are of unknown function.For the functional characterization of the E. coli genome, we have developed methods to systematically introduce amber mutations into the chromosome and have constructed an arabinose-inducible suppressor tRNA on a high-copy-number plasmid to modulate the effects of the amber mutations (12, 13). To use this system it was important to examine the effects of the suppressor on the cell. In addition to suppressing the gene of interest, the suppressor is also expected to affect genes throughout the genome that end with amber stop codons. Reflecting the relative prevalence of amber and ochre stop codons, ...

show abstract

mentioning

confidence: 99%

Global Transcriptional Effects of a Suppressor tRNA and the Inactivation of the Regulator frmR

Herring

Blattner

2004

J Bacteriol

View full text Add to dashboard Cite

show abstract

“…Such evidence has been obtained for su2 and su3 in mutation studies by Ohlsson, Strigini & Beckwith (1968) and for sul, su2 and su3 by Person & Osborn (1968). Such evidence has been obtained for su2 and su3 in mutation studies by Ohlsson, Strigini & Beckwith (1968) and for sul, su2 and su3 by Person & Osborn (1968).…”

Section: Introductionmentioning

confidence: 58%

“…Su B was isolated by and maps very close to amber su2 (Signer et al 1965) though it has been reported to be distinct from it (Ohlsson et al 1968). Su B was isolated by and maps very close to amber su2 (Signer et al 1965) though it has been reported to be distinct from it (Ohlsson et al 1968).…”

Section: Methodsmentioning

confidence: 98%

Genetic studies of class 2 nonsense suppressors inEscherichia coli

1973

View full text Add to dashboard Cite

Three genetically distinct ochre suppressors have been identified in a strain of Escherichia coli B/r, all of which suppress a tyrosine auxotrophy and classify as class 2 by phage suppression pattern. One ochre suppressor, which was obtained by conversion from a class 2 amber suppressor, and a second ochre suppressor obtained directly from the non-suppressing parent, were found to have separate map locations, though a peculiar phenotype with regard to a leucine auxotrophy is exhibited by strains carrying either suppressor. We suggest that both suppressors correspond to separate genes for glutamine-inserting £RNA. A Leu + mutant of a strain carrying one of these suppressors was studied and was found to contain a further nonsense suppressor having amber-suppressing activity at a reduced level. We suggest that this suppressor might result from a mutation in another part of the translational machinery concerned with glutamine insertion. The third ochre suppressor has no effect on the leucine auxotrophy and mapping data suggest that it may be supL, an ochre suppressor probably inserting a different amino acid from glutamine. 16-2

show abstract

“…The evidence for this mechanism of production of suppressors can be found in references 2, 4, and 5. For UAG suppressors the evidence rests on genetic and chemical evidence (see reference 2), while for UAA suppressors the evidence is entirely genetic [4,5].…”

Section: Method·of Mutation Productionmentioning

confidence: 99%