Mutations induced in Drosophila during space flight.

To examine the possible effects of space radiation on living organisms, fruit flies Drosophila melanogaster were loaded on the US Space Shuttle Endeavour, and after the flight we have analyzed two types of mutations, sex-linked recessive lethal mutations induced in male reproductive cells and somatic mutations which give rise to morphological changes in hairs growing on the surface of wing epidermal cells. Wild type strains and a radiation-sensitive strain mei-41 were used. The frequencies of sex-linked recessive lethal mutations in flight groups were 2 and 3 times higher for wild type Canton-S and mei-41 strains, respectively, than those in ground control groups. By contrast, the frequencies of wing-hair somatic mutations differed little between flight and control groups. The possibility that the space environment causes mutations in certain types of cells such as male reproductive cells, is discussed.     

Induction of mutations in unirradiated X-chromosome of Drosophila melanogaster hybrids of repair deficient females mei-41 [D5] and mus209 [B1] and irradiated males M5

It has been analyzed the frequency of the recessive lethal mutations in the unirradiated X-chromosome of Drosophila. Females of wild type (CS) as well as of error-prone (mei-41) and error-free (mus209) mutant strains were used. In CS hybrids the increasing of the mutation rate (p < 0.05) was found. In muc209 hybrids the mutation rate was not affected. In mei-41 hybrids the tendency to decreasing of the mutation rate was found. The obtained results demonstrate the possible role of error-prone repair in the inducing of mutations in the unirradiated X-chromosome in the presence of irradiated homologue.     

Genotoxicity of Rogor studied in the sex-linked recessive lethal test and wing, eye and female germ-line mosaic assays in Drosophila melanogaster

The genotoxic potential of Rogor (dimethoate), an anticholinesterase organophosphate insecticide, has been studied in the sex-linked recessive lethal test and the wing, eye and female germ-line mosaic assays in Drosophila melanogaster. Larvae of different instars carrying suitable recessive genetic markers on their first and third chromosomes were exposed to the LD50 or half of this dose for the entire larval life. The Basc technique was followed for the detection of the induction of sex-linked recessive lethals. The wings and eyes of the adult flies and the eggs laid by the heterozygous females were checked for the induction of mosaicism. It is concluded that Rogor induces sex-linked recessive lethals in immature male germ cells and is recombinogenic and/or mutagenic in both the somatic and the germ-line cells of Drosophila.     

Genotoxicity of ziram established through wing, eye and female germ-line mosaic assays and the sex-linked recessive lethal test in Drosophila melanogaster

The genotoxicity of ziram (zinc-dimethyl dithiocarbamate, CAS No. 137-30-4), a carbamate fungicide, is studied in the wing, eye and female germ-line mosaic assays and the sex-linked recessive lethal test in Drosophila melanogaster. First-, second- and third-instar larvae, carrying suitable recessive genetic markers on their first and third chromosomes, were exposed to ziram. Wings and eyes of adults were screened for the induction of mosaic spots and the eggs laid by adult females for germ-line mosaicism. The Basc method was used to detect sex-linked recessive lethals. Ziram is genotoxic to the somatic and germ cells of Drosophila melanogaster.     

The genotoxicity of the anti-cancer drug mitoxantrone in somatic and germ cells of Drosophila melanogaster.

The novel antineoplastic drug mitoxantrone was studied for its genotoxic effects in Drosophila melanogaster. In male germ cells, the clinical preparation Novantrone, the dihydrochloride salt of mitoxantrone, did not induce sex-linked recessive lethal mutations in feeding and injection experiments with adult flies, although statistically the results were inconclusive rather than truly negative. However, the free base mitoxantrone was weakly, but significantly genotoxic in this test (0.14% lethals/mM exposure concentration); this is most probably the result of prolonged exposure. On the other hand, both forms of mitoxantrone assayed were clearly genotoxic in the somatic mutation and recombination test of the wing. This test assays the cells of the proliferating imaginal wing discs of larvae. Depending on the feeding method used, the overall clone induction frequency was in the range of about 2-6 x 10(-5) per cell and cell generation and per mM exposure dose. Correction of these frequencies according to mean clone size led to slightly higher estimates (by about 5-25% higher). Although the majority of the clone induction events are due to mitotic recombination, a significant proportion can be attributed to mutational events (gene and chromosome mutations). The genotoxicity of mitoxantrone seems to depend mainly on impaired DNA synthesis in cycling cells owing to the compound's ability to inhibit topoisomerase II by intercalation into DNA.     

Mutagenic activity of selected aromatic amines and polycyclic hydrocarbons in Drosophila melanogaster

With the use of a series of wild-type and repair-deficient strains and appropriate application procedures, it is possible to demonstrate that carcinogenic aromatic amines and polycyclic hydrocarbons are mutagens in Drosophila. We have shown evidence that AAF, N-OH-AAF, AcO-AAF, BP, DAS and DMBA produce recessive lethals when fed to or injected into adult males. Mutagenic activity was also observed when male larvae were exposed to AAF, BP, DMBA, 3-MC or NA. DA was not mutagenic in the recessive lethal assay under the conditions of the test. DMBA can now be considered as a potent mutagen for Drosophila, although demonstration of its activity depends upon the choice of the treatment procedure and the strain selected. One of the questions concerning the action of aromatic amines and polycyclic hydrocarbons is how their genetic effectiveness in Drosophila can be enhanced. The observation that none of several enzyme inducers (PB, BF, AC, 3-MC) increased their mutagenicity may be interpreted in terms of a more efficient metabolic activation or deactivation. This assumes that active metabolite(s) did not reach the testis in doses sufficient for mutation induction. It also appears that, since the problems pertaining to mutagenicity in Drosophila of aromatic hydrocarbons are obviously a matter of metabolism, the use of repair-deficient strains is no longer an attractive proposal for their elucidation. The present investigation shows that, with weak mutagens, usage of strains mei-9Li or y mei-9a mei-4lD5 does not improve the sensitivity of the recessive lethal method or the test for chromosomal loss. As an alternative, in our opinion more attention should be devoted to possible differences in metabolism between somatic and gonadal tissue. We feel strongly that somatic assay systems might be particularly valuable as a complement to recessive lethal tests on the germ line.     

Mutagenicity of sumithion tested in Drosophila somatic and germ cells

Sumithion, a broad-spectrum insecticide, was tested for its mutagenicity in the Drosophila wing-spot test and sex-linked recessive lethal test. Strains carrying the recessive mutant markers mwh and flr3 in their third chromosomes, expressed phenotypically as multiple trichomes or thickened and misshapen wing hairs in the adult wings, were used in the wing-spot test. Larvae transheterozygous for these markers were exposed to the insecticide in instant food and the sex-linked recessive lethal test was performed by the standard technique using the Basc strain. The compound is mutagenic in the wing primordial cells and induces recombination at high doses. Further, the frequency of induction of sex-linked recessive lethals is significant only at high treatment doses.     

Studies on mutagen-sensitive strains of Drosophila melanogaster. II. Detection of qualitative differences between genetic damage induced by X-irradiation of mature spermatozoa in oxygenated and anoxic atmospheres through the use of the repair-deficient mutant mei-9a

Muller-5 males were irradiated with X-rays in nitrogen, in air or in oxygen (followed by nitrogen or oxygen post-treatments in the nitrogen and oxygen series) and were mated to females of a repair-proficient strain (mei+) or to those of a strain known to be deficient in excision repair of UV damage (in somatic cells). The latter strain, designated as mei-9a, is also known to be sensitive, in the larval stages, to the killing effects of UV, X-rays and to a number of chemical mutagens. The frequencies of sex-linked recessive lethals and autosomal translocations induced in the spermatozoa of males were determined and compared. The frequencies of sex-linked recessive lethals in the mei-9 control groups were consistently higher than in the mei+ groups. Irradiation in air or in nitrogen led to significantly higher yields of recessive lethals when the irradiated males were mated to mei-9 females, whereas, after irradiation in oxygen, the yields were similar with both kinds of female. No significant differences in the frequencies of reciprocal translocations were observed between the mei+ and mei-9 groups after irradiation of the males in nitrogen, in air or in oxygen. Likewise, no differential effects of the contrasting post-treatments (nitrogen versus oxygen), either for recessive lethals or for translocations, could be discerned. These results are considered to support the notion that the kinds of genetic damage induced in mature spermatozoa in air or in nitrogen are qualitatively similar (at least with respect to the component(s) that lead to the production of recessive lethal mutations), but clearly different when induced in an oxygen atmosphere. The enhanced yields of recessive lethals with mei-9 females (after irradiation of the males either in air or in nitrogen) has been interpreted on the assumption that the mei-9 mutant is also deficient for the repair of X-ray-induced, recessive lethal-generating premutational lesions. Possible reasons for the lack of differences between the mei+ and mei-9 groups with respect to translocation yields and for the absence of measurable differences in response between the contrasting post-treatments (after irradiation of the males in nitrogen) are discussed.     

Mutagenicity of dimecron studied in 3 mosaic assays and the sex-linked recessive lethal test in Drosophila melanogaster

The genotoxicity of dimecron, a systemic organophosphate pesticide, has been tested in the wing, eye and germ line mosaic assays and the sex-linked recessive lethal test in Drosophila melanogaster. Larvae heterozygous for recessive marker mutations were fed the compound for various periods of time. On emergence, the wings and eyes of the adults were screened for mosaic spots and the eggs laid by the females were checked for induction of female germ line mosaicism. Dimecron is mutagenic to the somatic and germ line cells of Drosophila and induces a high frequency of sex-linked recessive lethals.     

Mutagen sensitivity of Drosophila melanogaster. VI. Evidence from the excision-defective mei-9AT1 mutant for the timing of DNA-repair activity during spermatogenesis

The sex-linked recessive lethal test has been used to compare mutation induction by ethyl methanesulfonate and methyl methanesulfonate in spermatogenic stages of the DNA repair-deficient mei-9AT1 mutant and a repair-proficient control strain. For both agents, the data demonstrate that induced mutation rates are similar in both strains for the meiotic and post-meiotic broods. Conversely, for spermatogonial broods, the data indicate that the excision-deficient strain exhibits a 4-8 fold increase in induced mutation rate in comparison to the excision-proficient control strain. These experiments suggest that the low mutability of gonial cells normally observed for these agents is due to effective excision-repair processes which function until the commencement of meiosis. From alkylation mutagenesis experiments with repair-deficient E. coli strains, we note that the mei-9 strain exhibits pleiotropic mutant phenotypes very similar to those displayed by the uvr D mutant. By analogy with these studies, we speculate that mei-9, like uvr D, is deficient in a DNA unwinding protein.     

Germinal and somatic mutation induction in Drosophila after treatment of larvae with tritiated water.

The present study was carried out to evaluate the mutagenicity of tritium, administered as tritiated water, in Drosophila melanogaster. Larvae were fed on tritium-treated medium during their development. Germinal and somatic mutation induction was detected by means of the sex-linked recessive lethal and the wing spot tests, respectively. Our results show that beta-radiation from tritium is able to induce significant increases in the frequency of both germinal and somatic mutations.     

Acrylamide is genotoxic to the somatic and germ cells of Drosophila melanogaster

The genotoxic effects of acrylamide, a recently detected carcinogen, have been studied in the somatic (wing primordia) and germ cells of Drosophila melanogaster by the wing mosaic assay and the sex-linked recessive lethal test respectively. Larvae, 72 +/- 4 h old, were exposed to 6 different concentrations of acrylamide ranging between 0.25 mM and 5.0 mM in instant medium for 48 h. It is observed that acrylamide is both mutagenic and recombinogenic in the wing disc cells and induces sex-linked recessive lethals.     

Farm-grade chlorpyrifos (Durmet) is genotoxic in somatic and germ-line cells of Drosophila.

The mutagenic potential of Durmet, a farm-grade formulation of chlorpyrifos, was studied in the Drosophila wing mosaic and sex-linked recessive lethal tests. Larvae of the 2nd or 3rd instar carrying suitable recessive genetic markers on chromosome 3 were exposed to different concentrations of the insecticide and the frequency of induction of mutant mosaic spots on the wings was noted. The Basc technique was followed to study the induction of sex-linked recessive lethals. On the basis of the frequency of induction of mosaic wing spots and sex-linked recessive lethals, it is concluded that Durmet is genotoxic in somatic cells as well as germ cells of Drosophila.     

DNA Repair Dependence of Somatic Mutagenesis of Transposon-Caused WHITE Alleles in DROSOPHILA MELANOGASTER after Treatment with Alkylating Agents

DNA repair-defective alleles of the mei-9, mei-41, mus-104 and mus-101 loci of Drosophila melanogaster were introduced into stocks bearing the UZ and SZ marker sets. Males with the UZ marker set, z1 (zeste allele) and w+(TE) (genetically unstable white allele presumably caused by a transposable element), or the SZ marker set, z1 and w+R (semistable white allele caused by partial duplication of the w+ locus plus transposon insert), were exposed to EMS at the first instar. After emergence, adult males bearing red spots on lemon-yellow eyes were scored as flies with somatic reversions of w+(TE) or w+R. The relative mutabilities (relative values of reversion frequency at an equal EMS dose) of either w+(TE) or w+R in a repair-proficient strain and in mei-9, mei-41, mus-104 and mus-101 strains were 1: approximately 1.2:0.3:0.3:0.7, despite the fact that w+(TE) reverted two to three times as frequently as w+R under both the repair-proficient and repair-deficient genetic conditions. Similarly, after treatment with MMS, MNNG and ENNG, w+(TE) was somatically more mutable in the mei-9 strain and less mutable in the mei-41 and mus-104 strains than in the repair-proficient strain. From these results, we propose that mutagenic lesions produced in DNA by treatment with these chemicals are converted to mutant DNA sequences via the error-prone repair mechanisms dependent on the products of the genes mei-41+ (mei-41 and mus-104 being alleles of the same locus) and mus-101+, whereas they are eliminated by mei-9+-dependent excision repair. In contrast to the approximately linear responses of induced reversions of w+(TE) with ENNG in the repair-proficient, mei-9, and mei-41 strains, seemingly there were dosage insensitive ranges for induced reversion with MNNG in the repair-proficient and mei-41 strains, but not for reversion in the mei-9 strain; w+(TE) in the mus-104 strain was virtually nonmutable with MNNG and ENNG. These results suggest that O6-methylguanine (O6MeG) produced in DNA with MNNG, but not O6-ethylguanine produced with ENNG, is almost completely repaired in a low dose range by constitutive activity of DNA O6MeG transmethylase. From the distribution of clone sizes of spontaneous revertant spots and other data, we propose that both w+(TE) and w+R have a similar tendency to spontaneously revert more frequently at early rather than at later developmental stages probably reflecting a common property of their inserted transposons.     

Genotoxicity of 5-azacytidine in somatic cells of Drosophila

The newly developed somatic mutation and recombination test, utilizing the wing-hair mutations mwh and flr3, was used to evaluate the genotoxicity of the base analog 5-azacytidine in larvae of Drosophila melanogaster. Third instar larvae were fed media wetted with various concentrations of the compound, and wings of surviving adults were removed and scored for the presence of clones of cells possessing malformed hairs. Wings of exposed flies trans-dihybrid for mwh and flr3 had significantly increased frequencies of twin spots, small single spots and large single spots. Significant linear regression of twin-spot frequencies upon concentration was also obtained. Induction of twin spots by 5-azacytidine unambiguously demonstrates its recombinogenic activity in somatic cells of Drosophila. Significantly increased frequencies of large single spots on wings of inversion-heterozygous flies were also observed and suggest that 5-azacytidine may also be inducing somatic gene mutations (or deletions).     

Testing the mutagenicity of malondialdehyde and formaldehyde by the Drosophila mosaic and the sex-linked recessive lethal tests

The mutagenicities of malondialdehyde and formaldehyde were tested by screening each for genetic mosaics of Drosophila melanogaster and by the Muller-5 test for sex-linked recessive lethal mutations. For comparison, the effects of X-rays were also assayed by the above technique. Malondialdehyde, a degradation product of polyunsaturated fatty acids, was found to be a weak mutagen by the above criteria; it induced point mutations and chromosome exchanges at low frequency, as proved by the mosaic test, but failed to induce detectable sex-linked lethality. Formaldehyde was more mutagenic than malondialdehyde; beside induction of mosaic spots it induced sex-linked recessive lethal mutations, but only in the larval testes of Drosophila. Formaldehyde also induced disintegration of the clones. Formaldehyde treatment (feeding larvae with formaldehyde-containing food for about 4 days) was 5 times more mutagenic than malondialdehyde treatment and 5 times less effective than irradiation by 1000 R of X-rays. Wing mosaicism offers a more sensitive way to detect mutagenesis as compared with eye mosaicism. It is suggested that aldehyde-induced mosaic spots derive from mitotic recombination and point mutations.     

Radiation-induced lethal mutations appearing in cells of somatic tissue in Drosophila

Exposure of 3-hour drosophila male embryos to gamma-radiation during the topographic segregation of the germ anlage nuclei caused recessive sex-linked lethals in somatic cells only. The selectivity of the screening was determined by the ratio of mutation frequencies induced in embryos and adult males. Analysis of lethal mutations shows that a minimal rate of the divergence between germinal and somatic patterns of the cell development is observed in the embryogenesis, the 3d instar larva and prepupa, and maximal in the 1st and 2nd larva and pupa.     

Evaluation in Drosophila melanogaster of the mutagenic potential of furfural in the mei-9a test for chromosome loss in germ-line cells and the wing spot test for mutational activity in somatic cells.

The mutagenic potential of furfural was evaluated by means of the chromosome loss test in germ cells and the wing spot test in somatic cells of Drosophila melanogaster. The chromosome loss test was carried out employing repair-proficient as well as repair-deficient females. Males carried the compound Y chromosome, BSYy+. Two routes of administration were used: injection and feeding of adult males. Genetic damage was demonstrable after matings of treated males with females carrying the excision repair-deficient mutant mei-9a. The somatic mutation and recombination test was carried out treating 72-h transheterozygous mwh+/+flr3 larvae. Acute treatment of larvae was chosen as the method of exposure. Evidence indicates that furfural induces somatic damage as measured in the wing spot test.     

Detection of lethal mutations appearing in the somatic tissue cells of Drosophila

A method of selective screening of recessive sex-linked lethal mutations, that are manifested only in somatic cells of D. melanogaster, has been developed. Lethals were induced with X-irradiation of X-chromosomes at the moment of their topographic segregation from embryonic somatic cells. Analysis of mutation expression confirmed the method selectivity.     

Mutagen-sensitive mutants in Drosophila melanogaster: effects on premutational damage.

Drosophila melanogaster males from a Basc stock were mutagenized with either X-rays, ethyl methanesulfonate (EMS), or nitrogen mustard (HN2). Groups of identically treated males were crossed to different types of female. Sex-linked recessive lethals were scored as a genetic end point. The females used were homozygous for X-chromosomal mutations (mus(1)101D1, mus(1)104D1, mei-9 or mei-41D5) which lead to defective DNA repair and which increase the mutagen sensitivity of larvae. Females from a white stock with normal DNA repair capacities served as controls. The premutational lesions induced in mature sperm are only processed after insemination by the maternal enzyme systems present in the oocytes. Differences in the efficiency of the processing of lesions can lead to maternal effects on the frequency of mutations recovered from mutagenized sperm. It was found that, with the exception of mus(1)104D1, all mutants analysed significantly modify the mutation fixation of one or more types of premutational lesions. The most drastic effect is found with the mus(1)101D1 stock in which HN2-induced DNA cross-links do not lead to sex-linked recessive lethals. It is assumed that mus(1)101D1 is defective in an early step of DNA cross-link repair. Our first set of data clearly demonstrates that the study of maternal effects in Drosophila is an efficient tool to analyse the in vivo function of repair mutations on chemically induced mutagenesis.