Search PubMed⌕ Search

Biomedical subjects

P Quillardet

Publications and source records attributed to P Quillardet.

31 records · Page 2Linked to original sources
PubMed1987

Induction of the SOS system in a dam-3 mutant: a diagnostic strain for chemicals causing DNA mismatches.

We have developed a strain of E. coli in which expression of the SOS function sfiA, monitored by means of a sfiA::lacZ operon fusion, is efficiently triggered by the two base analogues 2-aminopurine and 5-bromo-2'-deoxyuridine. This strain resulted from introduction of a dam-3 mutation into a Uvr+, Rfa+ derivative of strain PQ37 used in the SOS chromotest, a bacterial colorimetric assay for genotoxins (Quillardet et al., 1982). The dam-3 mutation affects the mismatch correction system in E. coli. We show that the SOS-inducing capacity of a weak SOS inducer such as the alkylating agent ethyl methanesulfonate was also increased in the dam-3 strain. We provide evidence that the increase in SOS inducibility due to the dam-3 mutation is specific for compounds causing DNA mismatches and propose the use of the dam-3 derivative of PQ37 as a diagnostic strain for such agents. This diagnostic strain can be a useful addition to the SOS chromotest.

2-Aminopurine↗
PubMed1986

Genotoxic activity of two furan analogues of benzo[a]pyrene and their 2-nitro derivatives.

We measured the genotoxic activities in two bacterial tests, the Salmonella/histidine assay (a reverse mutation assay) and the SOS Chromotest (an assay for SOS induction in E. coli), of two pairs of isomeric furan analogues of benzo[a]pyrene: pyreno[1,2-b]furan (R7490) and pyreno[2,1-b]furan (R7692) and their 2-nitro derivatives, 8-nitro-pyreno[1,2-b]furan (R7489) and 8-nitro-pyreno[2,1-b]furan (R7691). We found that: For all 4 compounds, the responses were correlated in the two tests. For the 2-nitro derivatives, R7489 and R7691, the responses were extremely high, reaching SOS-inducing potencies of 5.2 X 10(3) and 10(5)/nmole in the SOS Chromotest and mutagenic potencies of 6.3 X 10(4) and 3.7 X 10(7) revertants/nmole in the Salmonella/histidine assay (strain TA98), respectively; the responses were only slightly decreased in nitroreductase-deficient strains. The responses to the two pyrenofurans were increased in the presence of an "activating mixture" but were still lower than that to benzo[a]pyrene. In contrast to benzo[a]pyrene and pyreno[2,1-b]furan (R7692), pyreno[1,2-b]furan (R7490) also gave a response in the absence of an "activating mixture". (5) Compounds with the oxygen heteroatom within the "bay region" gave lower responses than their isomers with the oxygen heteroatom outside the "bay region".

Animals↗
PubMed1985

Effect of an umuC mutation on phage lambda induction.

A possible role of the umuC gene product in the induction of the SOS responses was examined. We compared the expression of a genetic fusion, in which gene lacZ, encoding beta-galactosidase in Escherichia coli is under the direct control of the cI repressor from prophage lambda, in a umuC+ strain and in an otherwise isogenic umuC- mutant. We found that two times higher UV doses were required to obtain a similar induction in the umuC+ strain as in the umuC mutant. In addition we showed that, at the same UV dose after a lag period, the specific activity of beta-galactosidase increased more rapidly in the umuC mutant. We suggest that the wild-type umuC gene product participates in the processing of the SOS inducing structures caused by UV irradiation or prevents formation of some of them. This is compatible with a role of the UmuC protein in DNA synthesis past a replication block.

Bacteriophage lambda↗
PubMed1985

The SOS Chromotest, a colorimetric bacterial assay for genotoxins: procedures.

The SOS Chromotest is a quantitative bacterial colorimetric assay for genotoxins. Substantial validation is now available (Quillardet et al., 1985). We describe here in detail the tester strain as well as the effects of the variation of some parameters on the assay. We report a simple spot-test procedure as well as a new standard procedure which incorporate recent technical improvements aimed at simplifying the assay further.

Alkaline Phosphatase↗
PubMed1985

The SOS Chromotest, a colorimetric bacterial assay for genotoxins: validation study with 83 compounds.

The SOS Chromotest is a simple bacterial colorimetric assay for genotoxicity. It is based on the measure of the induction of sfiA, a gene controlled by the general repressor of the SOS system in E. coli. Expression of sfiA is monitored by means of a gene fusion with lacZ, the structural gene for beta-galactosidase. We have examined 83 compounds of various chemical classes with the SOS Chromotest using a standard procedure. Comparison of the results with those obtained in the Mutatest (the Ames test) showed that most (90%) of the mutagenic compounds were also SOS inducers. For these compounds a quantitative correlation was observed between the mutagenic potency and the SOS-inducing potency (SOSIP). The case of the 10% remaining compounds giving conflicting results in the two tests is discussed. Sensitivity, specificity and accuracy for carcinogenicity prediction have been evaluated for the SOS Chromotest and the Mutatest using 73 chemicals for which carcinogenicity data were available. In spite of some differences, similar results were obtained in the two tests. The present data indicate that the SOS Chromotest has many practical advantages and may be used as a primary screening tool or as part of a battery of short-term tests for carcinogens.

Biotransformation↗
PubMed1984

Induction by UV light of the SOS function sfiA in Escherichia coli strains deficient or proficient in excision repair.

The influence of the nucleotide excision repair system on the induction by UV irradiation of the SOS function sfiA has been investigated. The level of sfiA expression was monitored by means of a sfiA::lacZ operon fusion in both the wild-type strain and a uvrA mutant. We found that the initial steady rate of sfiA expression was proportional to the UV dose and was identical in uvr+ and uvrA backgrounds. This suggests that the initial steady rate of sfiA expression is determined by the initial number of lesions and before any effect of excision repair. We confirmed that after 2 h of expression the net synthesis of sfiA product is, for the same UV dose, about five times lower in uvr+ than in uvrA strains. We show that this is due to earlier repression of the SOS system in uvr+ than in uvrA strains and not to different initial rates.

DNA Repair↗
PubMed1982

Cell survival, UV-reactivation and induction of prophage lambda in Escherichia coli K12 overproducing RecA protein.

The effect of the cellular level of RecA protein on the ability of E. coli K12 bacteria to (i) survive UV-irradiation (ii) promote UV-reactivation of UV-damaged phage lambda (iii) induce prophage lambda was determined in bacterial mutants with discrete increasing levels of RecA protein. The various levels of RecA protein were obtained by combining lexA and recA alleles. Except for the double mutant lexA3 recAo98, whose repair ability was 25% less than that observed in wild type bacteria, bacterial survival was proportional to the level of RecA protein measured after 90 min of incubation. In lexA3 recAo98 bacteria, RecA protein, at a constitutive high basal level, failed to compensate totally for the lack of LexA repressor cleavage; UV-reactivation of UV-damaged phage lambda was not restored; yet, prophage lambda was induced with 35% efficiency. Efficient UV-induction of prophage lambda is linked to the induction of lexA-controlled host processes that repair the UV-damaged prophage.

Bacterial Proteins↗
PubMed1982

SOS chromotest, a direct assay of induction of an SOS function in Escherichia coli K-12 to measure genotoxicity.

We present and evaluate the SOS chromotest, a bacterial test for detecting DNA-damaging agents. It is a colorimetric assay based on the induction by these agents of the SOS function sfiA, whose level of expression is monitored by means of a sfiA::lacZ operon fusion. The response is rapid (a few hours), and does not require survival of the tester strain. Dose-response curves for various chemicals include a linear region. The slope of this region is taken as a measure of the SOS inducing potency. Comparison for a number of substances of known genotoxicity of the SOS inducing potency determined in the SOS chromotest with the mutagenic potency determined in the Salmonella assay (mutatest) revealed a striking quantitative correlation over more than 7 orders of magnitude. The sensitivity of the SOS chromotest (lowest amount detected) is equal to that of the mutatest and generally 4-40 times higher than that of a phage induction assay (inductest). From a practical point of view our observations contribute to the validation of the SOS chromotest as a test for detecting genotoxins and in particular genotoxic carcinogens. From a theoretical standpoint the results suggest that mutagenic potency measured in the mutatest reflects the level of induction of an SOS function and that most genotoxins are inducers of the SOS response in bacteria.

Alkaline Phosphatase↗
PubMedDate not supplied

The screening, diagnosis and evaluation of genotoxic agents with batteries of bacterial tests.

Bacterial tests can be used for several related purposes: the screening for genotoxic agents, genetic analysis of the mode of action of genotoxins and attempts to predict their effects in mammals. We examine various aspects of the assembly of tests into batteries with emphasis on the genetic properties of target bacterial cells. We discuss the problems of carcinogenicity prediction, the identification of particular types of DNA lesions, the study of mutagenic specificity and the elucidation of metabolic steps towards the ultimate genotoxin.

Bacteria↗
PubMedDate not supplied

Damaged-site independent mutagenesis of phage lambda produced by inducible error-prone repair.

The existence of damaged-site independent mutagenesis is confirmed here by scoring the appearance of clear-plaque (c-) or virulent (vir) forward mutations on intact (non-irradiated) phage lambda grown on UV-irradiated E. coli K12 hosts. The mutation frequency was measured as a function of the incubation time between the occurrence of host DNA lesions and phage infection. The time course of mutagenesis of intact phage followed the induction pattern observed upon UV-reactivation of UV-damaged phage by Defais et al. (1976). Intact phage did not mutate in UV-irradiated hosts carrying the uvm-25 mutation known to prevent the occurrence of UV-reactivation. These findings suggest that damaged-site independent mutagenesis results from inducible error-prone repair. Clear-plaque mutations arising on intact phage were mostly found in phage bursts consisting of clear and turbid plaque formers whereas UV-damaged phage gave rise to mostly clear-plaque formers. Contrarily to damaged-site dependent mutagenesis, damaged-site independent mutagenesis can arise even at late times during the phage replication cycle. Our data indicate that about half of the phage mutations that arise upon UV-reactivation are damaged-site independent mutations. Replication of intact phage DNA in a host during induction of SOS functions provides a sensitive assay for the detection of damaged-site independent mutagenesis.

Bacteriophage lambda↗
PubMedDate not supplied

The SOS chromotest: direct assay of the expression of gene sfiA as a measure of genotoxicity of chemicals.

We used a gene fusion, placing the lacZ gene encoding beta-galactosidase under the control of the sfiA promoter, to construct a new tester strain for genotoxic agents. The assay is performed in a few hours and involves simple enzymatic assays. The dose response curves contain a linear portion which enables to define the SOS Inducing Potency (SOSIP) of compounds. For the compounds tested SOSIPs extend over 7 decades and correlate generally well with the mutagenic potency assayed in the Salmonella/microsome assay (Mutatest) and in a phage induction assay (Inductest). Sensitivities (lowest amount detected) are comparable in the SOS Chromotest and Mutatest but lower in the Inductest. Our results suggest that at least part of the response in the Mutatest depends on the induction of an SOS function, and that most of the genotoxins are inducer of the SOS system -i.e. can lead to activation of the RecA protease.

4-Nitroquinoline-1-oxide↗
PubMedDate not supplied

The 'carry-over' of aflatoxin into milk of cows fed ammoniated rations: use of an HPLC method and a genotoxicity test for determining milk safety.

Aflatoxin B1 (Af.B1) contaminated and detoxified commodities were used to feed lactating cows. The detoxification treatment was performed using an autoclaving process. Af.B1 contents in untreated groundnut meal, treated groundnut meal, and a mixture of treated groundnut meal and soyameal were 475.0, 6.0 and 3.0 micrograms/kg, respectively. Af.M1 determination in milk was performed using an HPLC method with a detection limit of 0.025 micrograms/l. Also, an attempt was made to apply a genotoxicity test, the SOS Chromotest, to check for the presence of genotoxic residues in the various milk samples. The total excreted Af.M1 was 1.08%, 14.7% and 6.35% of the total ingested Af.B1 from untreated, treated and mixed treated feed, respectively. An unusual Af.M1 excretion peak was observed during the first four days of feeding the treated diet. Approximately 30% of the unreacted ingested Af.B1 appeared in the milk as the metabolite Af.M1. On the following days, the amount of Af.M1 in the milk samples decreased quickly and became almost undetectable. However, the genotoxicity test assays did not show any difference between the various experimental milk samples.

Aflatoxin B1↗