Two classes of Bacillus subtilis mutants deficient in the adaptive response to simple alkylating agents

Summary Six mutant strains of Bacillus subtilis hypersensitive to N-methyl-N-nitro-N-nitrosoguanidine (MNNG) were shown to be deficient in the adaptive response to MNNG and termed ada mutants (Morohoshi and Munakata 1985). All the mutations mapped between the attSPO2 and lin loci on the chromosome. The mutant and wild-type (ada ⁺) cells contained similar constitutive levels of O⁶-methylguanine-DNA methyltransferase activity. Pretreatment with low concentrations of MNNG increased the activity about nine-fold in the ada ⁺ cells, while it uniformly decreased the activity in the ada cells. The pretreatment of three mutants (ada-3, ada-4, and ada-6) as well as ada ⁺, augumented the activity of methylpurine-DNA glycosylase and rendered the cells resistant to the lethal and mutagenic effects of N-propyl- or N-butyl-N-nitro-N-nitrosoguanidine. With the rest of the mutant strains (ada-1, ada-2, and ada-5), neither of such responses was elicited by the pretreatment. Thus, the former ada strains seem to have a defect in the gene specifically involved in the induction of the methyltransferase, while the latter ada strains have a defect in the gene controlling the adaptive response as a whole.Abbreviations MNNG N-methyl-N-nitro-N-nitrosoguanidine - ENNG N-ethyl-N-nitro-N-nitrosoguanidine - PNNG N-propyl-N-nitro-N-nitrosoguanidine - MNU N-methyl-N-nitrosourea - MMS methyl methanesulphonate     

Exchange of individual ribosomal proteins between ribosomes as studied by heavy isotope-transfer experiments

Summary We have described previously an inducible response in Escherichia coli which occurs during growth on low levels of the methylating agent, N-methyl-N-nitro-N-nitrosoguanidine (MNNG), and which enables cells both to survive better and to be less mutated by a subsequent challenge dose of MNNG than control cultures (Samson and Cairns, 1977). We show here that this response is distinct from previously characterised pathways of DNA repair, and particularly from the SOS response, which is another inducible effect resulting from DNA damage. An examination of the cross-reactivity of this response with other mutagens has shown that it is a generalised mechanism affecting alkylation damage to DNA. It cannot, however, be induced by UV or the UV-mimetic mutagen, 4-nitroquinoline 1-oxide, nor act on lesions put into DNA by those mutagens.     

Lack of correlation between DNA-methylating activity and appearance of the immunogenic phenotype in clones of a murine lymphoma treated with mutagens

Summary The relationship between induction of novel immunogenicity by xenogenizing chemicals and DNA-methylating activity in murine tumors was investigated at the clonal level in L1210Ha cells treated with 5-azacytidine, N-methyl-N-nitro-N-nitrosoguanidine (MNNG) or 1-(p-chlorophenyl)-3,3-dimethyltriazene (DM-Cl). Cells were exposed to the drugs in vitro, cloned by limiting dilution, and assayed for transplantation immunogenicity and 5-methylcytosine content. The results showed that 0% (0/29, 5-azacytidine), 6.8% (2/29, MNNG) and 87.5% (28/32, DM-Cl) of the resulting clones were highly immunogenic, as judged by their tumorigenicity in intact compared to immunodepressed hosts. Frequency distribution analysis of the 5-methylcytosine content of drug-treated and parental clones showed that the methylation pattern was not significantly modified by tumor exposure to either 5-azacytidine or MNNG, and the two immunogenic clones induced by MNNG had methylcytosine levels very close to the 50th percentile value. In contrast, the extent of DNA methylation was increased in the cells treated with DM-Cl, but no obvious association was found between methylation status and immunogenicity of the drug-treated clones. In four 5-azacytidine-treated clones that displayed little or no immunogenicity, additional rounds of drug exposure led to progressive DNA demethylation, but failed, as a rule, to enhance tumor cell immunogenicity. Taken together, the present data indicate that, at least for the examined tumor, immunogenic variants are generated by mutagen treatment at high (MNNG) or very high (DM-Cl) frequencies under conditions in which hypomethylation-induced antigen amplification is unlikely.This work was supported by Progetto Finalizzato Oncologia, C. N. R, Rome-Italy, grant no. 87.01423.44 Abbreviations used: MNNG, N-methyl-N-nitro-N-nitrosoguanidine; DM-Cl, 1-(p-chlorophenyl)-3,3-dimethyltriazene; MST, difference (days) between median survival times of intact and irradiated mice injected with the same cells.     

Influence of benzamide on killing and mutation of density-inhibited V79 cells by MNNG

Density-inhibited V79 cells when held for 24 h in complete medium after exposure to N-methyl-N′-nitroN-nitrosoguanidine (MNNG) show improved survival levels and decreased mutant frequencies at all dose levels, compared to cells not so held. However, when benzamide, an inhibitor of poly(ADP-ribose) synthesis was present during this 24-h holding, the improvement in survival and decrease in mutant frequencies were not observed. Rather, compared to the control, the cells became more sensitive to MNNG and mutant frequency also increased significantly for all doses studied.     

Environmental radiation as the conditioning factor for the survival of yeast Saccharomyces cerevisiae

Whether natural radiation can be a conditioning factor for the growth and survival of a living organism was investigated using diploid yeast S. cerevisiae D7. Yeast cells were conditioned by growing them continuously for at least 100 generation in 3 different radiation background such as i) ambient radiation (1.1 mSv/y), ii) sub-ambient radiation (0.44 mSv/y, within a shielded chamber) and iii) an elevated background radiation (88 and 880 mSv/y in a gamma-field). At the end, the cells were challenged with 60Co gamma, 100 Gy and the viable fractions were determined. Conditioning the cells in 880 mSv/y and in ambient radiation, enabled the cells to reduce the deleterious effect of the challenging dose significantly (P < 0.05) compared to that of sub-ambient radiation. The cellular viability of yeast cultures seems to be influenced by the prevailing radiation background, apart from starvation. Comparatively, a rapid decline in viability was noticed when the cultures were incubated for 60 days in the shielded chamber. The results indicate that some amount of radiation equivalent to background level or little above is needed to confer fitness in biological systems against stress factors, including radiation. The adaptive dose for the diploid yeast was also determined by single exposure. The priming dose ranged from 0.01 to 1.2 Gy. An adaptive dose of 0.25 or 0.4 Gy, almost nullified the deleterious effect of the challenging dose. The adaptive response may have a greater role in the field of cancer therapy and in radiation risk assessment. Understanding the response of an organism at different radiation-background will be helpful for successful space management.     

Antimutagenic activity of Lactobacillus plantarum KLAB21 isolated from kimchi Korean fermented vegetables

Antimutagenic activity of Lactobacillus plantarum KLAB21, isolated from Korean kimchi, was investigated against MNNG (N-methyl-N-nitro-N-nitrosoguanidine), NQO (4-nitroquinoline-1-oxide), NPD (4-nitro-O-phenylenediamine) and aflatoxin B1 using Salmonella typhimurium strains TA100 and TA98. Although all the cell fractions including the culture supernatant, dry cells and cell-free extract exhibited antimutagenic activity against MNNG and NQO, the culture supernatant possessed the highest activity. The antimutagenic ratio of the culture supernatant was 98.4% against MNNG on strain TA100 and 57.3% against NQO on strain TA98. Its antimutagenic activity was reconfirmed by a Bacillus subtilis spore-rec assay. Levels of the antimutagenic ratios of other lactic acid bacteria originating from fermented milk ranged between 26.8 to 53% against MNNG and 28.5 to 43.4% against NQO. The antimutagenic activities of the strain KLAB21 against NPD were 72.6% on TA100 and 62.8% on TA98, and those against aflatoxin B1 were 82.5% on TA100 and 78.2% on TA98.     

Morphological,growth, and chromosomal changes in bovine pancreatic duct epithelial cells exposed toN-methyl-N′-nitro-N-nitrosoguanidine

Summary Epithelial cells cultured from bovine pancreatic ducts were given a single treatment ofN-methyl-N′-nitro-N-nitrosoguanidine (MNNG). Multinucleated cells and giant cells were observed more frequently in carcinogen-treated cultures than in controls. The MNNG-treated cultures also contained a sizeable population of small, dense cells that were not observed in control cultures. At the concentration of 1.0 μg/ml, MNNG caused an initial depression in the growth rate of the cells followed by growth stimulation for several weeks. The MNNG produced chromosomal damage in the cells as indicated by the observation that a substantial proportion of carcinogen-treated cells were heteroploid and contained a high frequency of metacentric and submetacentric chromosomes and a dicentric marker chromosome. The MNNG treated and control cultures did not acquire the ability to grow in soft agar or to produce tumors after transplantation into athymic, nude mice. This work was supported in part by Public Health Service Contract and Interagency Agreement Y01-CP60204 from the Division of Cancer Cause and Prevention, National Cancer Institute.     

Defective excision repair in a mutant of Micrococcus radiodurans hypermutable by some monofunctional alkylating agents

Summary The lethal and mutagenic effects of methyl methanesulphonate (MMS), ethyl methanesulphonate (EMS), and N-methyl-N-nitro-N-nitrosoguanidine (MNNG) can be dissociated in a mitomycin C (MTC)-sensitive mutant, strain 302, of Micrococcus radiodurans.As regards lethality 302 is extremely sensitive, compared with the wild type, to MTC and decarbamoyl MTC (DCMTC), slightly sensitive to EMS, MNNG, nitrous acid, 7-bromomethylbenz {} anthracene (BrMBA), and N-acetoxy-N-2-acetylaminofluorene (AAAF), and resistant to MMS, hydroxylamine, and ICR 191G. As regards mutability it is, compared to the wild type, very sensitive to MMS, EMS, and MNNG, and slightly sensitive to hydroxylamine and nitrous acid but not to any other agent examined.Alkaline sucrose gradient studies indicate that 302 does not incise DNA containing BrMBA adducts, although it does incise DNA damaged by AAAF but probably not to the same extent as wild type.We put forward the hypothesis that the hypermutability of 302 is due to the non-removal of bases or nucleotides, modified in exocyclic positions, which have altered base-pairing capabilities, while lethality results from the non-removal of bases or nucleotides, also modified in exocyclic positions, which no longer form hydrogen-bonded base pairs.     

Involvement of mismatch repair proteins in adaptive responses induced by N-methyl-N'-nitro-N-nitrosoguanidine against γ-induced genotoxicity in human cells

As humans are exposed to a variety of chemical agents as well as radiation, health effects of radiation should be evaluated in combination with chemicals. To explore combined genotoxic effects of radiation and chemicals, we examined modulating effects of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), a direct-acting methylating agent, against genotoxicity of γ-radiation. Human lymphoblastoid TK6 cells and its mismatch-deficient derivative, i.e., MT1 cells, were treated with MNNG for 24h before they were exposed to γ-irradiation at a dose of 1.0 Gy, and the resulting genotoxicity was examined. In TK6 cells, the pretreatments with MNNG at low doses suppressed frequencies of the thymidine kinase (TK) gene mutation and micronucleus (MN) formation induced by γ-irradiation and thus the dose responses of TK and MN assays were U-shaped along with the pretreatment doses of MNNG. In contrast, the genotoxic effects of MNNG and γ-irradiation were additive in MT1 cells and the frequencies of TK mutations and MN induction increased along with the doses of MNNG. Apoptosis induced by γ-radiation was suppressed by the pretreatments in TK6 cells, but not in MT1 cells. The expression of p53 was induced and cell cycle was delayed at G2/M phase in TK6, but not in MT1 cells, by the treatments with MNNG. These results suggest that pretreatments of MNNG at low doses suppress genotoxicity of γ-radiation in human cells and also that mismatch repair proteins are involved in the apparent adaptive responses.     

Differential expression of genes related to levels of mucosal cell proliferation among multiple rat strains by using oligonucleotide microarrays

Induction levels of cell proliferation, in response to gastric mucosal damage by N-methyl-N-nitro-N-nitrosoguanidine (MNNG), are different among rat strains and correlate with susceptibility to MNNG-induced gastric carcinogenesis. Here, we used oligonucleotide microarrays to search for genes that show expression levels accordant with the extents of cell proliferation among six rat strains. Expression levels of 8,800 probe sets were analyzed in the pylorus of ACI, LEW, WKY (strains with strong cell proliferation), F344, (ACI × BUF)F₁, and BUF rats (strains with weak cell proliferation) after 2-week MNNG treatment. No genes showed complete accordance, and 22 genes showed accordance with one or two exceptions. After confirmation by quantitative RT-PCR, four genes—cellular retinoic acid-binding protein II (Crabp2), fatty acid binding protein 1 (Fabp1), progastricsin (pepsinogen C, Pgc), and UDP-glucuronosyltransferase 2 family member 5 (Ugt2b5)—were found to show good accordance with only one exception. Crabp2, Fabp1, and Ugt2b5 were differentially expressed between ACI and BUF rats both before and after MNNG treatment. Although Crabp2 had been identified as one of the 16 genes differentially expressed between ACI and BUF rats with cDNA-RDA, Fabp1 and Ugt2b5 were newly identified in this study. All three genes are known to be involved in retinoic acid-mediated signaling and could be involved in the control of differential induction of cell proliferation.     

Adaptation-like response to the chemical induction of sister chromatid exchanges in human lymphocytes

Summary Experiments have been performed to determine whether human lymphocytes in primary cultures can show an adaptive response to the induction of cellular lesions (manifested as a production of sister chromatid exchanges, SCEs) as previously found in bacteria and established human and mammalian cell lines. Human lymphocytes were pretreated with various subtoxic concentrations (5–50ng/ml) of N-methyl-N-nitro-N-nitrosoguanidine (MNNG) once every 6h for 72h, and subsequently challenged by a high dose (4g/ml) of MNNG. The lymphocytes in MNNG-challenged cultures had the lowest frequency of SCEs when pretreated with 10ng/ml MNNG. Further cross-resistance study revealed that repeaied pretreatments of lymphocytes with 10ng/ml MNNG for 72h can render the cells resistant to the induction of SCEs by the following challenge with a high dose of MNNG, but not of mitomycin C or ethyl nitrosourea. The data also suggest variations in the degree of the adaptation-like response among individuals.     

Random mutagenesis and use of 2-deoxy-D-glucose as an antimetabolite for selection of α-amylase-overproducing mutants of Aspergillus oryzae

By using 2-deoxy-D-glucose, selection of different mutants of Aspergillus oryzae PTCC 5164, which were produced by random mutagenesis by u.v. radiation, nitrous acid and N-methyl-N-nitro-N-nitrosoguanidine (MNNG), was studied. 2-Deoxy-D-glucose, a well-known antimetabolite, was used to isolate derepressed mutants. The mutational and lethal effects of these mutagens on conidia of A. oryzae were compared and the frequency distribution of isolated mutants, in the presence of 2-deoxy-D-glucose, was determined. Potent mutants, which produced higher dextrinizing and saccharogenic activities, were isolated. The best strain was a result of mutagenesis by nitrous acid, which produced 6.73 times more dextrinizing and 5.13 times more saccharogenic activity than the parent strain. In general, the mutants obtained by nitrous acid and u.v. were more potent than those obtained by MNNG.     

Regulation of the two forms of glycogen phosphorylase by cAMP and its analogs in Dictyostelium discoideum

Summary We have recently reported the existence of two forms of glycogen phosphorylase (1,4--D-glucan: orthophosphate--glucosyltransferase; EC 2.4.1.1) in Dictyostelium discoideum. During development the activity of the glycogen phosphorylase b form decreased as the activity of the a form increased. The total phosphorylase activity remained constant. The physical and kinetic properties of the Dictyostelium enzyme were similar to those of the mammalian enzyme. In mammals, cAMP regulates the conversion of the two forms by a cAMP dependent protein kinase (cAMPdPK). We report here that if cAMP is added to a single cell suspension, the Dictyostelium phosphorylase activity becomes independent of 5AMP and a 104kd peptide appears. We also show the effect of several cAMP analogs on the phosphorylase activity in these single-cell suspensions. The cAMP analogs were selected on the basis of their affinities for the membrane-bound cAMP receptor or the cytoplasmic cAMPdPK. We found that relatively low levels, 100 M, of cAMP or 2'd-cAMP added to aggregation-competent cells in shaking culture caused a loss of phosphorylase b activity and the appearance of phosphorylase a activity. The analog, 2'd-cAMP, has a high affinity for the cAMP receptor but a low affinity for the cAMPdPK. Two other analogs, Bt₂-cAMP and 8-Br-cAMP, which have low affinities for the cAMP receptor but high affinities for the cAMPdPK, required high levels (500 M) for b to a conversion. cDNAs to three cAMP-regulated genes-PL3, Dll, and D3-were used as controls in the above experiments. In order to determine if intracellular levels of cAMP were involved in the regulation of phosphorylase activity, both the phosphorylase and the PL3, D11 and D3 mRNA levels were examined in cells suspended in a glucose/albumin mixture - a medium in which adenylate cyclase is inhibited. Under these conditions, neither gene regulation nor a change in the phosphorylase b to a activity occurred in response to added extra cellular cAMP. The results suggest that an intracellular increase in cAMP is involved in the regulation of the two forms of glycogen phosphorylase in Dictyostelium.Abbreviations EGTA Ethyleneglycol-bis-(-aminoethyl ether) - N,N,N N-tetra acetic acid - SDS Sodium Dodecyl Sulfate - PAGE Polyacrylamide Gel Electrophoresis     

Mechanism of potent mutagenic action of N-methyl-N'-nitro-N-nitrosoguanidine on intracellular phage lambda.

N-Methyl-N′-nitro-N-nitrosoguanidine efficiently induces mutations from “clear” to “virulent” in phage λ only during the intracellular growth phase. Lambda DNA extracted from infected bacteria after treatment with MNNG³ produced a mutant yield about 100-fold higher than the spontaneous level upon transfection of MNNG-treated spheroplast cells, whereas the yield diminished an order of magnitude when assayed on untreated spheroplasts. As measured by ¹⁴C incorporation after treatment with [methyl-¹⁴C]MNNG, λ DNA packed in head protein was methylated to about 3% by an MNNG dose of 0.6 mg/ml but was barely mutagenised; whereas intracellular λ DNA was methylated to no more than 0.6% by an MNNG dose of 0.09 mg/ml and was highly mutagenised. Lambda phages treated in vitro with ethyl methanesulfonate produced a rather low mutant yield on untreated cells but the yield increased about tenfold on MNNG-treated cells. Mutability of untreated λ on cells having received an F′ factor was enhanced efficiently by ultraviolet light, but not so by MNNG, previously applied to the F′. Surprisingly similar MNNG dose-effect curves exist for enhancing spontaneous, mispairing (MNNG or EMS induced) and misrepair (ultraviolet light induced) mutagenesis of λ. From these and other data we conclude that MNNG hypermutagenesis results from a synergistic increase in mispairing probability of appropriately methylated bases (by action of MNNG in vivo) in the target gene within an MNNG-induced intracellular environment that has an enhanced mutagenic capacity.     

Cyclin-dependent kinase inhibitor,roscovitine, in combination with exogenous cytokinin,N6-benzyladenine,causes increase of cis-cytokinins in immobilized tobacco cells

Dynamics of the response of tobacco cells (line BY-2) to exogenous cytokinin, N ⁶-benzyladenine, and cyclin-dependent kinase inhibitor, roscovitine, was followed using alginate-immobilized cells packed into a column. N ⁶-Benzyladenine (1.25 M) increased the synthesis of the physiologically-active endogenous cytokinin, isopentenyladenosine, in the effluent up to 0.1 nM. Simultaneously, conversion of the excess of endogenous cytokinins to biologically inactive derivatives of cis-zeatin occurred, up to 0.8 nM. Roscovitine (50 M) further increased cis-cytokinins, up to 2.2 nM.     

Evidence that N-methyl-N′-nitro-N-nitrosoguanidine induces adaptive response in Bacillus thuringiensis

Bacillus thuringiensis is shown to have an inducible error-free repair system for alkylation damage as found in Escherichia coli and Bacillus subtilis. Growth of cells in the presence of low concentrations of N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) induces an adaptive response which is characterized by an increase in resistance to killing and mutagenesis by challenge with higher concentrations of MNNG. In addition, we have noted with interest that adaptive low doses seem to produce lesions at a rate sufficient to induce an increase of mutation frequency, and inhibition of cell division. The possibility of an interaction between SOS and adaptive responses with these low doses of MNNG is discussed.     

The adaptive response of E.coli to low levels of alkylating agent: the role of polA in killing adaptation.

Summary In an attempt to characterise which gene products may be involved in the repair system induced in E. coli by growth on low levels of alkylating agent (the adaptive response) we have analysed mutants deficient in other known pathways of DNA repair for the ability to adapt to MNNG. Adaptive resistance to the killing effects of MNNG seems to require a functional DNA polymerase I whereas resistance to the mutagenic effects can occur in polymerase I deficient strains; similarly killing adaptation could not be observed in a dam3 mutant, which was nonetheless able to show mutational adaptation. These results suggest that these two parts of the adaptive response must, at least to some extent, be separable. Both adaptive responses can be seen in the absence of uvrD ⁺ uvrE ⁺-dependent mismatch repair, DNA polymerase II activity, or recF-mediated recombination and they are not affected by decreased levels of adenyl cyclase. The data presented support our earlier conclusion that adaptive resistance to the killing and mutagenic effect of MNNG is the result of previously uncharacterised repair pathways.     

Inducible DNA-repair systems in yeast: competition for lesions

DNA lesions may be recognized and repaired by more than one DNA-repair process. If two repair systems with different error frequencies have overlapping lesion specificity and one or both is inducible, the resulting variable competition for the lesions can change the biological consequences of these lesions. This concept was demonstrated by observing mutation in yeast cells (Saccharomyces cerevisiae) exposed to combinations of mutagens under conditions which influenced the induction of error-free recombinational repair or error-prone repair. Total mutation frequency was reduced in a manner proportional to the dose of 60Co-gamma- or 254 nm UV radiation delivered prior to or subsequent to an MNNG exposure. Suppression was greater per unit radiation dose in cells gamma-irradiated in O2 as compared to N2. A rad3 (excision-repair) mutant gave results similar to wild-type but mutation in a rad52 (rec-) mutant exposed to MNNG was not suppressed by radiation. Protein-synthesis inhibition with heat shock or cycloheximide indicated that it was the mutation due to MNNG and not that due to radiation which had changed. These results indicate that MNNG lesions are recognized by both the recombinational repair system and the inducible error-prone system, but that gamma-radiation induction of error-free recombinational repair resulted in increased competition for the lesions, thereby reducing mutation. Similarly, gamma-radiation exposure resulted in a radiation dose-dependent reduction in mutation due to MNU, EMS, ENU and 8-MOP + UVA, but no reduction in mutation due to MMS. These results suggest that the number of mutational MMS lesions recognizable by the recombinational repair system must be very small relative to those produced by the other agents. MNNG induction of the inducible error-prone systems however, did not alter mutation frequencies due to ENU or MMS exposure but, in contrast to radiation, increased the mutagenic effectiveness of EMS. These experiments demonstrate that in this lower eukaryote, mutagen exposure does not necessarily result in a fixed risk of mutation, but that the risk can be markedly influenced by a variety of external stimuli including heat shock or exposure to other mutagens.     

Requirement for Cell Dispersion Prior to Selection of Induced Azaguanine-Resistant Colonies of Chinese Hamster Cells

With V79 Chinese hamster cell cultures treated with a mutagen, the maximum frequency of colonies resistant to 8-azaguanine (AZG) was attained when the cells were dispersed after a suitable expression time before adding the selection medium. V79–4 cells were exposed to 500 µM MMS, 7 µM AFAA, or 10 µM MNNG and allowed to multiply before being reseeded at 4 x 10⁴ cells/60 mm dish and selected with 10 µg/ml AZG. Maximum frequencies of 4 x 10^-5, 4 x 10^-4, and 2.4 x 10^-3 were obtained about 100, 130, and 200 hrs after exposure to MMS, AFAA, and MNNG, respectively. The maximum frequencies following MMS or MNNG treatments were about 10-fold greater than those obtained when induction and selection of AZG-resistant colonies were performed in the same culture dish. The reseeding of treated cells eliminated the possibility of metabolic cooperation within mosaic colonies of wild-type and mutant cells and achieved expression of the induced changes before intercolony crossfeeding reduced the frequency of resistant colonies.—AZG-resistant colonies were selected in medium containing dialyzed fetal bovine serum, and the selection medium was replaced at least twice. Both serum dialysis and selection medium replacement were necessary for consistent achievement of background frequencies of resistant colonies near 10^-6. Reconstruction experiments with AZG-resistant V79 lines showed that the efficiency of recovery of resistant cells in the selection medium was constant over a range of 0–20 colonies observed/dish. A mixed population of V79 and AZG-resistant cells was also correctly analyzed by the procedure used in mutagenesis studies.     

N-methyl-N''-nitro-N-nitrosoguanidine-induced resistance to ionizing radiation

Summary N-methyl-N-nitro-N-nitrosoguanidine (MNNG) pretreatments increase the resistance of Escherichia coli to -radiation. The increased resistance is dependent on functional polA, recA, recB, recC, and lexA genes and is partly dependent on recN. The MNNG-induced resistance is additive to resistance induced by pretreatment with -radiation but not by increases induced by hydrogen peroxide. The MNNG-induced resistance occurs in adaptive response mutants and at pretreatment levels of MNNG that do not activate cells to reactivate UV-inactivated lambda phage. The MNNG-induced resistance appears to be distinct from other inductions to -radiation resistance.