Search PubMed⌕ Search

Biomedical subjects

B A Bridges

Publications and source records attributed to B A Bridges.

At least 37 records · Page 2Linked to original sources
PubMed1997

Mutation in Escherichia coli under starvation conditions: a new pathway leading to small deletions in strains defective in mismatch correction.

Strains of Escherichia coli carrying the mutY mutation lack a mismatch correction glycosylase that removes adenines from various mismatch situations. In growing bacteria, 8-oxoguanine-adenine mispairs persist and can give rise to G-->T transversions during subsequent replication cycles. We now show that when trpA23 mutY bacteria are held under tryptophan starvation conditions the tryptophan-independent mutants that arise include small in-frame deletions in addition to transversions. The trpA23 reversion system appears to be unusual in that small in-frame deletions occurring in a particular region of the gene can lead to the production of a functional protein. We suggest that this is a consequence of the deletion causing the polar group on the arginine at the trpA23 site to be pulled away from the active site of the enzyme. Such deletions are also found with starved bacteria defective in methyl-directed mismatch correction activity (mutH, mutL or mutS), and deletion mutations are also found among the much lower number of mutants that arise in bacteria wild-type for mismatch correction. There is thus a pathway, hitherto undetected, leading to deletions probably from mismatches under conditions of growth restraint. RecA, UmuC, UvrA, MutH,L,S, SbcC and SbcD proteins are not required for the operation of the deletion pathway. A possible explanation is that the deletion pathway is not dependent upon further replication and that it fails to be discernible in growing cells because it is relatively slow acting and mismatches are likely to encounter a DNA replication fork before the initial step of the deletion pathway.

Base Sequence↗
PubMed1997

DNA turnover and mutation in resting cells.

There is growing evidence that mutations can arise in non-dividing cells (both bacterial and mammalian) in the absence of chromosomal replication. The processes that are involved are still largely unknown but may include two separate mechanisms. In the first, DNA lesions resulting from the action of endogenous mutagens may give rise to RNA transcripts with miscoded bases. If these confer the ability to initiate DNA replication, the DNA lesions may have an opportunity to miscode during replication and thus could give rise to apparently 'adaptive' mutations. A second mechanism is suggested by recent work in starved bacteria, showing that there is much more turnover of chromosomal DNA than has been previously thought. This could permit polymerase errors to lead to mutations in non-dividing cells. Such cryptic DNA synthesis, which may essentially replace existing DNA rather than duplicating it, could, in principle, act as an additional source of variability on which selection may act, initially in the absence of cell division. In mammals such processes would undoubtedly have implications for germ cell mutagenesis and carcinogenesis.

Animals↗
PubMed1996

Effect of mutY and mutM/fpg-1 mutations on starvation-associated mutation in Escherichia coli: implications for the role of 7,8-dihydro-8-oxoguanine.

MutY specifies a DNA glycosylase that removes adenines unnaturally paired with various bases including oxidized derivatives of guanine, such as 7,8-dihydro-8-oxoguanine (8-oxoG). The rate of mutation in starved Escherichia coli cells is markedly raised in mutY mutants defective in this glycosylase. As predicted, the mutations produced include G to T transversions. Bacteria carrying mutM or fpg-1 mutations (defective in Fapy glycosylase, which removes oxidized guanine residues such as 8-oxoG) show little or no enhancement of mutation under starvation conditions. When present together with mutY, however, mutM clearly further enhances the rate of mutation in starved cells. Plasmids resulting in overproduction of MutY or Fapy glycosylases reduce the rate of mutation in starved cells. We conclude that, in non-growing bacteria, oxidized guanine residues, including 8-oxoG, constitute an important component of spontaneous mutation. Addition of catalase to the plates did not reduce the mutant yield, indicating that extracellular hydrogen peroxide is not involved in the production of the premutational damage. Singlet oxygen, known to give rise to 8-oxoG, may be the ultimate oxidative species.

Catalase↗
PubMed1996

The tyrT locus of Escherichia coli B.

The tyrT (tRNA(1TYr)) locus of Escherichia coli B differs structurally from that of K-12 strains by the absence of 2 of 3.14 terminal repeat sequences.

Base Sequence↗
PubMed1996

Elevated mutation rate in mutT bacteria during starvation: evidence for DNA turnover?

The rate of appearance of prototrophic revertants when Escherichia coli tyrA14 (ochre) or trpA23 bacteria were incubated on plates lacking the required amino acid was greatly elevated when the organisms also carried a mutT mutation. One possible explanation for this result is that the amount of DNA replication or turnover under these conditions is much greater than has been previously recognized.

Bacterial Proteins↗
PubMed1996

Lack of evidence for an association between the frequency of mutants or translocations in circulating lymphocytes and exposure to radon gas in the home.

Radon measurements in the living room and main bedroom of 41 houses in the town of Street, Somerset, England have been made. Exposure levels, weighted using the formula of the UK National Radiological Protection Board, of 19-484 Bq m-3 (about half > 100 Bq m-3) were found. Blood samples were obtained from a total of 66 occupants in these homes, and the frequency of genetic alterations in lymphocytes was estimated using two different end points. Gene mutations at the hypoxanthine guanine phosphoribosyl transferase locus were determined in T lymphocytes for 65 subjects using a clonal assay, and the frequency of the BCL-2 t(14;18) translocation, a chromosomal event associated with leukemia/lymphoma, was estimated in lymphocytes using a polymerase chain reaction-based technique for 64 subjects. In neither case was a significant correlation with radon levels in the home found, in contrast to our earlier observation with a smaller series.

Adult↗
PubMed1996

Mutation in resting cells: the role of endogenous DNA damage.

In E coli, new spontaneous mutations can arise in bacteria that are non-dividing and in which there is little or no DNA synthesis. These mutations are almost invariably those that enable the cell to resume growth, a phenomenon that has been termed directed or adaptive mutation. Evidence is accumulating from studies with DNA repair deficient strains that damage produced by endogenous mutagens may be an important source of such mutations. A DNA lesion that can miscode can explain the apparent adaptive behaviour since if a "mutant" RNA transcript confers sufficient advantage that the cell is triggered into a cycling state, the ensuing round of DNA replication will be likely to fix the mutation by means of a DNA miscoding event. The most important lesion in this respect appears to be 8-oxoG, which can pair equally well with adenine or cytosine and so give rise to G to T transversions. It is responsible for almost half the G to T transversions arising in non-growing repair proficient bacteria. Alkylations contribute to the production of both transitions and transversions but only those at A:T base pairs are important in repair proficient bacteria. There is also a report of a lesion susceptible to UvrA,B,C dependent excision repair, but whether it is important in bacteria possessing excision repair has not been addressed. Data on mammalian cells are almost non-existent, but there is evidence that point mutations can occur in vivo in postmitotic neurons. The underlying assumption that there is little or no DNA synthesis in non-dividing bacteria has been challenged by recent data suggesting that there may be extensive cryptic DNA turnover.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1995

Are there DNA damage checkpoints in E. coli?

The concept of regulatory 'checkpoints' in the eukaryotic cycle has proved to be a fruitful one. Here, its applicability to the bacterial cell cycle is examined. A primitive DNA damage checkpoint operates in E. coli such that, after exposure to ultraviolet light, while excision repair occurs, chromosome replication continues very slowly with the production of discontinuous daughter strands. The slower the rate of excision of photoproducts, the greater the delay before the normal rate of DNA replication is restored, the additional time for repair ensuring that normal survival is maintained. A model is proposed in which replication rate is controlled by the ratio of RecA-coated to uncoated single stranded regions of DNA in the replication fork. There are also two cell division inhibitors SulA (= SfiA) and SfiC under the control of the SOS system and sensitive to DNA damage, but they are irrelevant to the survival of wild-type bacteria under normal conditions. In strains where SulA and SfiC do not operate, inhibition is not influenced by the rate of excision repair and so fails one of the criteria for a DNA damage checkpoint, namely the monitoring of the DNA for the level of residual damage.

Cell Cycle↗
PubMed1995

Starvation-associated mutation in Escherichia coli strains defective in transcription repair coupling factor.

When E. coli WU3610 (tyrA14 ochre) bacteria are starved of tyrosine on the surface of glucose-salts agar plates, there is a progressive accumulation of slow growing prototrophic mutants that are neither revertants at the ochre codon nor any of the well characterised tRNA ochre suppressors. Isogenic derivatives defective in transcription repair coupling factor (mfd) showed normal starvation-associated mutation (SAM). WU361045, the original mfd strain, showed very much reduced SAM. At 37 degrees C this was associated with progressive loss of viability on plates but the defect in SAM was not due to loss of viability since incubation at 27 degrees C or addition of catalase prevented the loss of viability but did not restore SAM. Furthermore, mutants could not be rescued from starved WU361045 populations by a short period of tyrosine supplementation arguing that WU361045 was defective not in the survival of starvation-associated mutants, but in their formation. The SAM defect in WU361045 was not complemented by the katF gene on a low copy number plasmid. It is concluded that WU361045 carries an unidentified mutation, not under katF control, that greatly reduces SAM. If SAM is attributable to a spontaneously occurring DNA lesion, the latter is unlikely to be formed by hydrogen peroxide or active species derived from it.

Adaptation, Biological↗
PubMed1995

Growth rate effects of mutations conferring streptomycin-dependence and of ancillary mutations in the rpsL gene of Escherichia coli: implications for the clustering (hypermutation) hypothesis for spontaneous mutation.

Colonies of newly arising streptomycin-dependent (SmD) mutants frequently contain a high proportion of cells with additional mutations (ancillary mutations) in the same gene (rpsL). The ancillary mutations appear to have arisen at a rate greatly above expectation. To better estimate this rate it is necessary to allow for any selective advantage conferred by the ancillary mutations. We have previously measured their effect on growth rate of established SmD strains in the presence of streptomycin. In the present work a pair of single and double mutant alleles (rpsL832 and rpsL852 respectively) has been employed together with the wild-type allele to model the situation soon after such mutations first arise, i.e. when the cell still contains wild-type S12 protein (the rpsL gene product). When these alleles, under the control of an IPTG-inducible promoter and carried on a plasmid, were expressed in the presence of a chromosomal wild-type allele, the double mutant allele permitted much faster cell growth than the single mutant allele. In the presence of streptomycin, and with rpsL+ on a plasmid, bacteria with a double mutant chromosomal gene grew faster than those with a single mutant chromosomal gene. If these results can be extrapolated to a bacterial cell in which an SmD mutation has just occurred, the ancillary mutation should be able to confer a selective advantage during a limited period when wild-type S12 protein is still present, both in the absence and in the presence of streptomycin.(ABSTRACT TRUNCATED AT 250 WORDS)

Alleles↗
PubMed1995

Mutant frequencies in workers at the Sellafield installation.

The frequency of hypoxanthine-guanine phosphoribosyltransferase (hprt) mutations has been determined in the peripheral T-lymphocytes of 18 workers at the Sellafield nuclear reprocessing installation with cumulative recorded radiation doses of about 500 mSv or more in comparison with a matched control group of 18 workers with doses less than 50 mSv. The mean dose of the exposed group was 663 mSv (range 449.3-880.8), and of the unexposed group 10.9 mSv (range 0-46.5). During the most recent 6 y the mean doses were 40.1 mSv (range 1.9-93.9) and 2.3 mSv (range 0-8.7), respectively. The logarithmic mean mutant frequency of the exposed group (9.3 x 10(-6)) was lower than that of the unexposed group (12.4 x 10(-6)). The apparent inverse association of mutant frequency and exposure varied in statistical significance tests from borderline to p < 0.01 depending on how the analysis was performed and whether recent or total exposure was considered.

Humans↗
PubMed1994

Starvation-associated mutation in Escherichia coli: a spontaneous lesion hypothesis for "directed" mutation.

When stationary phase E. coli WU3610, carrying an ochre mutation in the tyrA gene, were incubated on plates lacking tyrosine, tyrosine-independent (Tyr+) mutants appeared from day 7 onwards in a time-dependent manner. These starvation-associated mutants did not contain either identifiable tRNA suppressors or reversions at the ochre site and are thus quite distinct from the mutants commonly found to arise during active growth. When an appropriate fluctuation assay protocol was employed slow growing Tyr+ mutants were also found to arise in growing cells, and their distribution was more characteristic of a replication-dependent than a time-dependent process. The rate of appearance of starvation-associated mutants at 37 degrees C was somewhat less than at 27 degrees C and this was attributed to a reduction in viability at the higher temperature. There was no evidence for the accumulation on the plates of mutations in other genes. Tyr+ mutants were, however, shown to arise during incubation of stationary phase cells under conditions where there was no selection for tyrosine independence, provided outgrowth was subsequently permitted on plates lacking tyrosine. This distinguishes the present system from those systems exhibiting genuine "directed" or "adaptive" mutation, should they exist. To explain the specificity which occurs, it is proposed that the appearance of stationary mutants in ochre strains reflects the time-dependent accumulation in the transcribed strand of a DNA lesion that has a high probability of miscoding during transcription and replication. A mutant RNA transcript permits protein synthesis which in turn triggers DNA replication. The mutation is then fixed in the DNA as a permanent heritable change. The apparent "directedness" of the process is, on this model, determined solely by the particular miscoding DNA lesion occurring in a transcribed strand at a site where a change in phenotype permits DNA replication to occur.

Culture Media↗