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B A Bridges

Publications and source records attributed to B A Bridges.

At least 55 records · Page 3Linked to original sources
PubMed1994

The performance of short-term tests in identifying potential germ cell mutagens: a qualitative and quantitative analysis.

A retrospective analysis was undertaken to assess the performance of selected short-term tests in the discrimination of mammalian germ cell mutagens and nonmutagens using data derived from the U.S. Environmental Protection Agency/International Agency for Research on Cancer Genetic Activity Profile (EPA/IARC GAP) and EPA GENE-TOX databases. The short-term tests selected were gene mutation in Salmonella (S. typhimurium), cultured mammalian cell gene mutation and chromosomal aberrations, and mammalian bone marrow cytogenetics (micronucleus and chromosomal aberrations). These are the first level tests used in the EPA mutagenicity testing guidelines. The results of this analysis showed good sensitivity of short-term in vitro tests for mammalian cell gene mutation (96%) or chromosomal aberrations (92%) in identifying germ cell mutagens, while the sensitivity of tests for gene mutation in S. typhimurium was lower (79%). Bone marrow micronucleus or chromosomal aberration assays in vivo each displayed a sensitivity of 96%. Thus, both the in vitro and in vivo tests may be used effectively to screen chemicals for potential germ cell mutagenicity. In contrast, the in vitro tests mentioned above performed poorly in discriminating putative germ cell nonmutagens, giving results for specificity at or below what is expected due to chance alone (50-11%). The bone marrow assays were more efficient in this regard, the micronucleus test yielding a specificity of 63% and the chromosomal aberrations assay 64%. The mouse bone marrow micronucleus test also performed well on a quantitative basis, responding at or below the lowest effective doses tested in the mouse dominant lethal assay. Regression analysis of the mean lowest effective doses of chemicals evaluated in vivo showed approximately 1:1 linear correlations for mouse germ cell assays (heritable translocation vs dominant lethal or specific locus tests) as well as for mouse bone marrow assays (micronucleus vs chromosomal aberration). The results suggest the value of the bone marrow micronucleus test as an assay for potential germ cell mutagenicity and the dominant lethal test as a relatively inexpensive choice for confirmation of germ cell damage. The sensitivity of the in vitro assays investigated and the discriminatory capability of the in vivo bone marrow assay affirmed the utility of these tests within the framework of the EPA mutagenicity testing guidelines.

Animals↗
PubMed1993

Spontaneous mutation in stationary-phase Escherichia coli WP2 carrying various DNA repair alleles.

Escherichia coli WP2 trpE65(ochre), when grown to stationary phase and plated on glucose salts medium, gave rise to slow growing tryptophan-independent mutants which formed increasing numbers of visible colonies from about day 6 onwards. These mutants were neither revertants at the ochre codon nor mutants at the tRNA suppressor loci normally found in this strain with logarithmic phase mutagenesis. The yield of mutants was not affected by the presence of the following alleles, umuC122, lexA102, polA1, recA1, recA56 or del(srlR-recA)306, except that in the three recA-defective strains, mutant colonies were initially slower to appear, possibly reflecting a lower viability in the inoculum. Stationary-phase spontaneous mutation in bacteria carrying on ochre mutation is thus a distinct and specific process that does not require the SOS system, or UvrA protein or DNA polymerase I. It may reflect the occurrence of a type of non-bulky DNA damage with altered base pairing specificity. In 3 out of 4 experiments with a strain carrying recA441 plus lexA51(Def) the rate of stationary-phase mutagenesis was elevated suggesting that there may be an additional component requiring an activated SOS system.

Adenosine Triphosphatases↗
PubMed1993

Double, independent mutational events in the rpsL gene of Escherichia coli: an example of hypermutability?

A proportion (up to 20%) of newly arising streptomycin-dependent (SmD) colonies of Escherichia coli WP2 contain bacteria where, in addition to a known SmD-determining (primary) mutation in the rpsL gene, there is a further ancillary mutation in the same gene. These ancillary mutations occur at 10 sites between 11 and 201bp away from the primary mutation. Ancillary mutations have been found in mutant colonies arising both spontaneously and after treatment with ultraviolet light and some have been found repeatedly. Ancillary mutations were frequently found to occur in mixed clones with an excess of bacteria carrying only the primary SmD mutation. No ancillary mutations were found in an adjacent non-coding region and there were no coding sequence changes that did not alter the amino acid specified. Although a selective advantage for bacteria containing ancillary mutations could not be demonstrated directly in every case, some small advantage must be presumed to have occurred to explain the absence of mutations at the other sites and particularly at third (wobble) codon positions. Ancillary mutations appear to occur fairly early in the life of a newly arisen SmD mutant clone in some sort of hypermutable process. Whether they are noticed appears to depend on their conferment of some selective advantage on the bacteria carrying them. While the ancillary mutations within rpsL lie close to, and may be consequent upon the formation of a primary SmD mutation, their mechanism of formation appears to be at least to some extent independent and does not involve the recA or umuC genes.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

Hazard identification: efficiency of short-term tests in identifying germ cell mutagens and putative nongenotoxic carcinogens.

For more than a decade, mutagenicity tests have had a clearly defined role in the identification of potential human mutagens and an ancillary role in the identification of potential human carcinogens. The efficiency of short-term tests in identifying germ cell mutagens has been examined using a combined data set derived from the U.S. Environmental Protection Agency/International Agency for Research on Cancer Genetic Activity Profile (EPA/IARC GAP) and EPA Gene-Tox databases. Our review of these data indicates adequate sensitivity of batteries of in vitro short-term mutagenicity tests in identifying germ cell mutagens. The analysis also supports the inclusion of an in vivo assay as suggested in proposed regulatory testing guidelines. In the context of carcinogenicity testing, the ability of short-term bioassays to detect genotoxic or mutagenic carcinogens is well established. Such tests are not considered to be as sensitive to nongenotoxic or nonmutagenic carcinogens. However, analyses presented in this report using the EPA/IARC GAP database demonstrate that many putative nongenotoxic carcinogens that have been adequately tested in short-term genetic bioassays induce gene or chromosomal mutation or aneuploidy. Further investigation should reveal whether the mutagenicity of these agents plays an important mechanistic role in their carcinogenicity.

Animals↗
PubMed1992

Mutagenic DNA repair in Escherichia coli. XXI. A stable SOS-inducing signal persisting after excision repair of ultraviolet damage.

Mutations to streptomycin resistance induced by ultraviolet light in Escherichia coli can lose their susceptibility to photoreversing light during excision repair and in the absence of chromosomal replication and protein synthesis, i.e., under conditions where SOS induction cannot occur. Using fusions of lac with sulA and umuC we have shown that after excision of UV damage in the presence of chloramphenicol there is a persisting, relatively stable signal capable of inducing SOS genes when protein synthesis is subsequently permitted. The persisting signal is formed roughly in proportion to the square of the UV dose and is about 30% photoreversible. It is suggested that the persisting SOS-inducing signal comprises a UV photoproduct (the target lesion) opposite a gap in the opposing DNA strand, and is formed by excision of one (the ancillary lesion) of a pair of closely opposed photoproducts. Calculations suggest that as few as two or three such configurations in a cell can lead to induction of sulA when protein synthesis is permitted. It is not clear whether these configurations can directly induce the SOS system because of their region of single-stranded DNA or whether the ultimate SOS-inducing signal is a more extensive single-stranded region formed when such configurations encounter a replication fork. Photoproduct/gap configurations have been previously suggested to be potentially mutagenic. UV-induced mutations to streptomycin resistance are mostly at A:T sites and are not photoreversible in fully SOS-induced bacteria in the absence of excision repair, indicating that they are not targeted at cyclobutane-type pyrimidine dimers. In SOS-induced excision-proficient bacteria there is about 39% photoreversibility which is rapidly lost after UV. This photoreversibility is attributed to many ancillary lesions being cyclobutane-type pyrimidine dimers which are excised leading to the exposure of target lesions on the opposing strand which, at these particular sites, are mostly non-photoreversible photoproducts.

Bacterial Proteins↗
PubMed1992

Mutagenesis after exposure of bacteria to ultraviolet light and delayed photoreversal.

The induction of mutations by ultraviolet light and delayed photoreversal in bacteria defective for SOS mutagenesis is discussed in terms of two models: the two-step misincorporation and bypass model, and the model involving simple deamination of cytosine-containing dimers. In phage S13 the latter appears to be the predominant mechanism. In Escherichia coli there is little evidence that the simple deamination mechanism is of any significance except in ung strains lacking uracil glycosylase where uracils left after photoreversal are not removed. Deamination might, however, occur during the operation of translesion synthesis via the two-step model and if it did, subsequent photoreversal would lead to the "mutation" being extended from one to both strands by uracil glycosylase repair rather than being removed.

Coliphages↗
PubMed1992

Mutant sequences in the rpsL gene of Escherichia coli B/r: mechanistic implications for spontaneous and ultraviolet light mutagenesis.

Mutants able to grow in the presence of 1.2 mg/ml streptomycin were isolated from Escherichia coli WP2 after exposure to ultraviolet light (UV) or in the absence of any treatment (spontaneous), and from a umuC derivative after exposure to UV and delayed photoreversal. These mutants, characterized as streptomycin resistant (Smr) or dependent (Smd), carry mutations in the rpsL gene. This gene was amplified using the polymerase chain reaction and sequenced. Mutations induced by UV were largely (76%) of the Smr phenotype, all of which were changes at an A:T base pair at codons 42 or 87. Mutations induced by UV plus delayed photoreversal in the non-UV-mutable umuC122 derivative of WP2 were exclusively of the Smd phenotype and all occurred at G:C base pairs at codons 41, 90 or 91. These results are consistent with current understanding of the mechanism of mutagenesis by UV and delayed photoreversal. A broader spectrum of mutations was seen in the spontaneous series including three-base deletions leading to amino acid loss (2 of codon 93, 1 of codon 87). Of particular note was the number of intragenic second site mutations in the spontaneous series, most if not all of which appeared to be silent with respect to streptomycin phenotype. It is necessary to postulate a high rate of formation of such mutations at some stage during the experiment. One possibility is that spontaneous mutation may often occur in bursts when an error correction mechanism (eg., proofreading, mismatch correction) is temporarily inactive.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1992

Workshop on DNA repair.

A workshop on DNA repair with emphasis on eukaryotic systems was held, under the auspices of the EC Concerted Action on DNA Repair and Cancer, at Noordwijkerhout (The Netherlands) 14-19 April 1991. The local organization of the meeting was done under the auspices of the Medical Genetic Centre South-West, The Netherlands (MGC), c/o Department of Radiation Genetics and Chemical Mutagenesis, University of Leiden (The Netherlands). Local organizers were: D. Bootsma (chairman), W. Ferro, J.H.J. Hoeijmakers, A.R. Lehmann, P.H.M. Lohman, L. Mullenders, and A.A. van Zeeland (secretarial assistance: Mrs. C. Escher-van Heerden and Mrs. R. Bontre). Over 190 scientists participated, and the format of the meeting followed that of the 1987 workshop on the 'Molecular Aspects of DNA Repair' (Friedberg et al., 1987). Plenary review talks in the mornings were followed, in the afternoon, by poster viewing in three or four parallel sessions. Groups of 15-20 posters were discussed in detail, and later on, in plenary sessions, chairpersons of the poster discussions reviewed the afternoons' posters. The principal themes of the meeting were the isolation and characterisation of repair genes and proteins, repair in specific sequences, consequences of defective DNA repair, and new methods for detecting DNA damage and repair. Remarkable progress has been made recently in all of these areas, and many exciting new results were presented. It is impossible to summarize all contributions to this (intensive) one-week meeting. Therefore, and for the sake of coherence, presentations that did not fit easily into any of the general themes of the meetings have not been included.

Animals↗
PubMed1991

Possible association between mutant frequency in peripheral lymphocytes and domestic radon concentrations.

To investigate whether previously found geographical correlations between leukaemia incidence and exposure to radon are reflected in a detectable mutagenic effect on individuals, the frequency of mutations in the hypoxanthine guanine phosphoribosyl transferase gene (hprt) in peripheral blood T lymphocytes was measured in subjects with known domestic radon concentrations. These concentrations were measured in December, 1989, in houses in Street, Somerset, UK, by passive alpha-track radon detectors. 20 non-smoking subjects aged 36-55 years were selected from the patient list at the local health centre on the basis of the radon concentrations in their homes--the range selected varied by a factor of ten. Blood samples for preparation of T lymphocytes were taken in July, 1990. There was a significant association between the log mutant frequency and radon concentration (t = 3.47, p less than 0.01). A second analysis of a further set of radon measurements (October, 1990, to January, 1991), in both living rooms and bedrooms, and repeated mutant frequency determinations also showed a significant relation, which remained significant even after exclusion of the highest frequency and adjustment for subject's age and cloning efficiency. These data must be regarded as preliminary and further more extensive studies should be done to determine whether the observed association is causal.

Acute Disease↗
PubMed1991

The mutagenicity of 2-amino-N6-hydroxyadenine to L5178Y tk +/- 3.7.2C mouse lymphoma cells: measurement of mutations to ouabain, 6-thioguanine and trifluorothymidine resistance, and the induction of micronuclei.

2-Amino-N6-hydroxyadenine (AHA) was tested in the mouse lymphoma L5178Y tk +/- assay using the microtitre cloning technique over concentrations from 0.005 micrograms/ml-1 (100% viability) to 6 micrograms/ml (10% viability) as measured by cloning efficiency immediately after treatment. At low, non-toxic concentrations (0.005-0.25 micrograms/ml) a dose-related linear increase in the frequency of ouabain-resistant mutants was seen, in addition to an increase in 6-thioguanine- and trifluorothymidine-resistant mutants. No consistent induction of micronucleated cells was observed in this concentration range. Toxic concentrations (20-90% kill) induced a dose-related increase in micronuclei, while the frequency of ouabain-resistant mutants fell (although it was still highly significantly above the control value). These results suggest that the mechanism of action of AHA depends on the concentration, with point mutations being induced at low, non-toxic doses and detectable chromosome breakage occurring only at higher doses. Both large-colony and small-colony trifluorothymidine-resistant mutants were induced at all concentrations. The utility of using multiple genetic end-points in one cell line and the importance of dose range selection for risk assessment and an understanding of the mode of action of test substances is underlined.

Adenine↗
PubMed1991

Mutagenic DNA repair in Escherichia coli. XIX. On the roles of RecA protein in ultraviolet light mutagenesis.

An experimental system was used in which His+ mutations induced by ultraviolet light (UV) arise from non-photo-reversible photoproducts whereas lethality is largely determined by photoreversible photoproducts. By exposing a strain with a deletion through recA to light immediately after UV, it was possible to examine mutagenesis under conditions where survival was not significantly different from 100%. No UV mutagenesis was seen in the absence of RecA protein even though the rest of the SOS system was fully expressed due to the presence of a defective LexA repressor and the active carboxy-terminal fragment of UmuD was present as a result of an engineered plasmid-borne gene. We conclude that RecA protein has a third essential function if UV mutagenesis is to be detected in excision-deficient-bacteria. Another experiment showed that in exerting this function RecA protein does not need activation by pyrimidine dimers elsewhere on the genome, in contrast to its protein-cleavage mediation functions with LexA and UmuD proteins. RecA1730 protein blocked UV mutagenesis unless delayed photoreversal was given showing that the third function of RecA protein is not in the misincorporation step. It is therefore most likely to be in the bypass step where UmuD' and UmuC are postulated to act, although the possibility cannot be excluded that RecA protein is required for some other survival function distinct from translesion synthesis.

Alleles↗
PubMed1991

Interpretation of results with the 8-azaguanine resistance system in Salmonella typhimurium: no evidence for direct acting mutagenesis by 15-oxosteviol, a possible metabolite of steviol.

15-Oxosteviol, postulated to be the mutagenic metabolite of steviol, was observed to be non-mutagenic in preliminary experiments using a number of different systems. Repetition of the original experiment in Salmonella TM677 failed to show any significant induction of 8-azaguanine resistant mutants by 15-oxosteviol even when the number of bacteria tested was greatly increased. Examination of the earlier positive result showed that it could not be justified from the data and revealed a commonly applied way of mishandling data obtained with the TM677 system.

Azaguanine↗
PubMed1990

Comparative induction of gene mutations and chromosome damage by 1-methoxy-1,3,5-cycloheptatriene (MCHT), 2. Results using L5178Y mouse lymphoma cells to detect both gene and chromosome damage; validation with ionizing radiation, methyl methanesulphonate, ethyl methanesulphonate and benzo[a]pyrene.

A variation of the mouse lymphoma (L5178Y TK+/(-)-3.7.2c) assay has been developed using a microtiter cloning technique instead of the standard agar method. The cell line has been used to detect both gene mutations (at the Na+/K+ ATPase and thymidine kinase loci) and chromosome damage (micronucleus induction) in the same experiment. The system was validated using gamma-irradiation (a known clastogen), 2 direct-acting mutagens, ethyl and methyl methanesulphonate and an indirect-acting mutagen, benzo[a]pyrene. Using the assay, 1-methoxy-1,3,5-cycloheptatriene was shown to be a clastogenic mutagen in the presence of S9, since a clear dose-dependent increase in micronuclei was observed, mainly small colony thymidine kinase mutants were observed, and no ouabain-resistant mutants were induced, a profile very similar to gamma-irradiation. The results suggest that metabolic activation potential explains the results in the accompanying paper (Asquith et al., 1990). The implications for mutagenicity testing are discussed.

Animals↗
PubMed1990

Mutagenic DNA repair in Escherichia coli. XVII. Effect of temperature-sensitive DnaE proteins on the induction of streptomycin-resistant mutations by UV light.

In contrast to the dnaE486 mutation, which is nearly 'dead-stop', dnaE1026 allows DNA synthesis for some time at restrictive temperatures. When bacteria carrying the dnaE486 or dnaE1026 temperature-sensitive mutations were incubated at restrictive temperature after exposure to UV light they showed little or no fixation of mutations as determined by loss of photoreversibility and the mutation frequency fell progressively. These results confirm a role for DnaE protein (the alpha-subunit of DNA polymerase III holoenzyme) in UV mutagenesis. A derivative of dnaE1026 carrying the umuC122 allele which blocks normal UV mutagenesis showed induction of mutations when photoreversing light was given to UV-irradiated bacteria after a period of incubation at either permissive or restrictive temperature. The defective DnaE1026 protein is therefore able to carry out the misincorporations which have been postulated to be the first step in the mutagenic process. Its inability to give rise to mutations in umu+ bacteria may therefore be attributed to its inability to participate in a later step, perhaps because it is unable to interact with the UmuD, C gene products. In contrast, UV-irradiated dnaE486 umuC122 bacteria did not show mutagenesis when incubated at restrictive temperature before photoreversal, suggesting that the altered DnaE486 protein was not able to carry out the postulated misincorporation step at 43 degrees C. DNA polymerase III alpha-subunit therefore appears to be required for both the misincorporation and bypass steps in the two-step model for UV mutagenesis.

DNA Repair↗
PubMed1990

Mutagenic DNA repair in Escherichia coli. XVIII. Involvement of DNA polymerase III alpha-subunit (DnaE protein) in mutagenesis after exposure to UV light.

UV light was unable to induce rifampicin-resistant mutations at 43 degrees C in Escherichia coli ER11 dnaE486. Although DnaE486 gene product is inactive at 43 degrees C, these bacteria contain the pcbA1 mutation which allows DNA replication provided DNA polymerase I is functional. The experiments were carried out under conditions where full expression of rifampicin-resistant mutations could occur so that the lack of induced mutations cannot be ascribed to an effect of incubation at 43 degrees C on mutation expression. UV-mutability at 43 degrees C was restored by the presence of the dnaE+ allele on a plasmid. It is concluded that functional DnaE protein is essential for UV mutagenesis. The dnaE486 mutation also blocked the induction at 43 degrees C of mutations induced by UV plus delayed photoreversal, a procedure that has been postulated to reflect an early misincorporation step in the UV mutagenic process.

DNA Polymerase III↗
PubMed1990

Sunlight-induced cancer: some new aspects and implications of the xeroderma pigmentosum model.

Symptoms of xeroderma pigmentosum, a hereditary disease characterized by accelerated light-induced ageing of the skin, are due to deficiencies in the repair of damaged DNA. Following UV irradiation a high incidence of thioguanine-resistant mutations have been observed, which may be a model for the abnormally high incidence of freckling and benign and malignant transformation observed in these patients. Cells from patients with Cockayne's syndrome and trichothiodystrophy have also been shown to be hypermutable by UV radiation with a similar DNA repair defect, but unlike xeroderma pigmentosum patients they do not experience a higher incidence of skin cancer or freckling. An immunological defect may be a further crucial factor determining the dermatological symptoms of xeroderma pigmentosum patients. Much can be learned about the way normal individuals function from a study of aberrant functioning of individuals with defined genetic deficiencies. In the field of senescence of the skin, the disease xeroderma pigmentosum (XP) has been of particular value, as the exposed skin of these sun-sensitive individuals shows at an early age many of the features of aged sun-exposed skin of normal individuals. These features of XP skin include hyperkeratosis, freckling, benign lesions and malignant tumours, including melanomas. A full discussion of this syndrome has been given by Kraemer et al.

DNA Damage↗