[A dual system of superinstability at the yellow and scute loci in Drosophila melanogaster]

We have characterized the phenomenon of super-unstability in yellow and scute loci of Drosophila melanogaster. A few derivatives with different combinations of yellow and scute phenotypes appeared after dysgenic cross between potentially super-unstable stock y2nsscme and P[ry+, (delta 2-3)] (99B). Essentially, the double alterations of yellow and scute phenotypes constitute more than 40% of all derivatives. Most frequently the mutations in both yellow and scute loci change coordinately giving rise to the y+nssc+ allele. Lethal derivatives were not observed. The spectrum and the frequency of the y2nsscme mutagenesis in females differ considerably from the analogous in males. Possibly, the neutral homologous chromosome in females changes contacts between two insertions, and because of that mutagenesis changes too. Thus, all alterations in the double super-unstable system seem to be connected with the recombination between two unstable insertions A genetic exchange may by initiated by a double-strand breaks induced by the transposase of the ends of insertions.     

Involvement of eddy currents in the mutagenicity of ELF magnetic fields

Possible carcinogenic and/or mutagenic activity of extremely low frequency magnetic fields was examined using somatic mutation and recombination test system of Drosophila melanogaster. An X-linked semi-dominant DNA repair defective mutation mei-41(D5) was introduced into the conventional mwh/flr test system to enhance mutant spot frequency. Virgin females of w mei-41(D5)/FM6; flr/TM6 were crossed with w mei-41(D5)/Y; mwh jv; spa(pol) males. The F(1) third instar larvae were exposed to a 50Hz, 20mT sinusoidal AC magnetic field for 24h. After moulting from pupal cases, their wings were examined under a bright field microscope to detect hair spots with mwh or flr mutant morphology. The exposure caused a statistically significant enhancement in somatic recombination spot frequency. Mutant spots arising due to chromosomal non-disjunction or terminal deletion also increased but the frequency of spots resulting from point mutation was not altered. The enhancement in the recombination spot frequency was suppressed to the control level when a culture medium without electrolytes was used during exposure. When larvae were exposed to a magnetic field in an annular dish, flies from the outer ring showed more mutant spots compared to those from the inner ring. These results suggest that the detected mutagenic activity was that of the induced eddy current, rather than that of the magnetic field itself.     

The interaction of the rad201 gene with mei-9 and mei-41 genes in germline cells of Drosophila female

The effect of Drosophila mutation rad201G1 together with mutations mei-41D5 and mei-9a on the sensitivity of oocytes to induction of dominant lethals (DLs) was studied. To this end, the frequencies of spontaneous and gamma-radiation-induced DLs in consecutive egg batches of females carrying double or single mutations were estimated. Since the effects of the mutations examined are expressed only at the previtellogenetic stages of oogenesis, only newly hatched (0-5-hour-old) females, whose oocytes did not develop farther than stage 7, were irradiated. The results obtained indicated that in intact and irradiated oocytes of double mutants mei-9a rad201G1 and mei-41D5 rad201G1, mutation rad201G1 epistatically suppresses the mutations of the both mei genes.     

The Effect of mei-41 on Rdna Redundancy in DROSOPHILA MELANOGASTER

The recombination and repair defective mutant, mei-41, exhibits three rather striking effects on the genetic properties and chromosomal stability of rDNA in Drosophila. First, mei-41 inhibits rDNA magnification. However, mei-9, another recombination and repair defective mutation has no similar effect. This indicates that magnification requires some, but not all, of the gene products necessary for meiotic exchange. Second, under magnifying conditions, mei-41 induces interchanges between the X rDNA and either arm of the Ybb- chromosome. These interchanges occur at high frequency and are independent of rDNA orientation. Third, in mei-41 bb+/Ybb+ males, bobbed mutants in the X, but not the Y, also arise at high frequency. Evidence suggests that these events involve the rDNA type I insertion. The recombination and repair defective properties of mei-41 together with our results regarding its unusual and specific effects involving rDNA are explained in a simple model that has general implications for chromosome structure.     

The Drosophila Meiotic Recombination Gene Mei-9 Encodes a Homologue of the Yeast Excision Repair Protein Rad1

Meiotic recombination and DNA repair are mediated by overlapping sets of genes. In the yeast Saccharomyces cerevisiae, many genes required to repair DNA double-strand breaks are also required for meiotic recombination. In contrast, mutations in genes required for nucleotide excision repair (NER) have no detectable effects on meiotic recombination in S. cerevisiae. The Drosophila melanogaster mei-9 gene is unique among known recombination genes in that it is required for both meiotic recombination and NER. We have analyzed the mei-9 gene at the molecular level and found that it encodes a homologue of the S. cerevisiae excision repair protein Rad1, the probable homologue of mammalian XPF/ERCC4. Hence, the predominant process of meiotic recombination in Drosophila proceeds through a pathway that is at least partially distinct from that of S. cerevisiae, in that it requires an NER protein. The biochemical properties of the Rad1 protein allow us to explain the observation that mei-9 mutants suppress reciprocal exchange without suppressing the frequency of gene conversion.     

Hypersensitivity of Drosophila mei-41 mutants to hydroxyurea is associated with reduced mitotic chromosome stability

6 mutant alleles of the mei-41 locus in Drosophila melanogaster are shown to cause hypersensitivity to hydroxyurea in larvae. The strength of that sensitivity is directly correlated with the influence of the mutant alleles on meiosis in that: alleles exhibiting a strong meiotic effect (mei-41D2, mei-41D5, mei-41D7) are highly sensitive; alleles with negligible meiotic effects (mei-41(104)D1, mei-41(104)D2) are moderately sensitive and an allele which expresses meiotic effects only under restricted conditions (mei-41D9) has an intermediate sensitivity. This sensitivity is not a general feature of strong postreplication repair-deficient mutants, because mutants with that phenotype from other loci do not exhibit sensitivity (mus(2)205A1, mus(3)302D1, mus(3)310D1). The observed lethality is not due to hypersensitivity of DNA synthesis in mei-41 larvae to hydroxyurea as assayed by tritiated thymidine incorporation. Lethality is, however, potentially attributable to an abnormal enhancement of chromosomal aberrations by hydroxyurea in mutant mei-41 larvae. Both in vivo and in vitro exposure of neuroblast cells to hydroxyurea results in an increase in 3 types of aberrations which is several fold higher in mei-41 tissue. Since hydroxyurea disrupts DNA synthesis, these results further implicate the mei-41 locus in DNA metabolism and provide an additional tool for an elucidation of its function. The possible existence of additional genes of this nature is suggested by a more modest sensitivity to hydroxyurea which has been detected in two stocks carrying mutagen-sensitive alleles of alternate genes.     

Effects of DNA-repair-deficient mutants on somatic and germ line mutagenesis in the UZ system in Drosophila melanogaster

To analyze the underlying mechanisms of the UZ system in Drosophila melanogaster, i.e., the unstable white locus with an IS element included originally described by Rasmuson and Green (1974), the repair deficient mutants mei-9b and mei-4lD5 and the meiotic mutant mei-2l8 were introduced into X-chromosomes containing the UZ system. These strains were submitted to larval feeding of MMS to analyze differences occurring in mutation rate and survival. The mei-9b and mei-4l strains were markedly sensitive to MMS treatment, while mei-2l8 did not affect survival. Only the mutant mei-4l, which is lacking in post-replication repair, affected the somatic mutation rate of the UZ system, decreasing it by 50%. The spontaneous germ line mutation rate of UZ, on the other hand, was increased by introducing mei-4l or mei-9b, possibly as a result of the high frequency of unrepaired spontaneous chromosome breaks that occur in these mutants.     

Analysis of the visible mutations induced by x-rays and ethyl methanesulfonate in the mature spermatozoa of Drosophila melanogaster

The recessive visible mutations spectrum of chromosome II induced by X-rays and ethylmethanesulfonata (EMS) in mature Drosophila melanogaster spermatozoa has been studied. Treatment of both mutagens resulted in mutations in all 5 genes in stock mei-9LI and only 4--in D-32. The comparison of mutation frequencies of the same genes in two stocks under EMS-treatment demonstrated the statistical difference of mutation frequencies j, pr, cn of two stocks, genes b and vg did not differ. Under the influence of X-rays the differencies have been observed only for gene b. In stock D-32 the mutation frequency differes from the control for b and vg (EMS treatment) and j, pr, vg (under the action of X-rays), in mei-9LI--the mutation frequency of all 5 genes (under the X-rays) and 4 of 5 genes (EMS treatment).     

Effect of gamma-irradiation on oogenesis of Drosophila mutants defective for reparation and meiotic recombination

Effects of mutations rad201, mei-9, and mei-41 on cell sensitivity to gamma-radiation in Drosophila oogenesis were studied. Females of the control (Oregon R) and mutant strains were irradiated at a dose of 15 Gy. For 9 days after the irradiation, the number of eggs in consecutive day batches, the frequency of dominant lethals (DLs) among the eggs, and the cytologically recorded distribution of oocytes for stages of their development, and the frequency of egg chamber degeneration in female ovaries were estimated. As a result of joint analysis of the data, different oogenesis stages were characterized with regard to the frequency of two radiation-induced events: appearance of DLs in oocytes and degeneration of egg chambers due to apoptosis of nurse cells. It was shown that the mutations affect these parameters only at particular stages of early oogenesis, at which previtellogenetic growth of egg follicles and meiotic recombination in oocytes occur. Mutation rad201G1 increased the frequency of DLs and egg chamber degeneration, mei-41D5 affected only the DL frequency, and mei-9a, in addition to enhancing the chamber degeneration frequency, promoted radiation "rescue" of some oocytes from the DL induction.     

Genetic effects of cosmic radiation in Drosophila melanogaster]

To examine possible effects of space radiation on living organism, we have analyzedtwo types of mutations, sex-linked recessive lethal mutations and somatic mutations, in fruit fly of the species Drosophila melanogaster. Drosophila strains used were wild type strains and a radiation-sensitive strain mei-41. Two different developmental stages of samples were sent into space; young adult males to analyze sex-linked recessive lethal mutations and about 30hr-old larvae to detect somatic mutations in wing epidermal cells. For wild type and mei-41 strains each, about 200 adult male flies and about 6,000 larvae were loaded on space shuttle Endeavour. The male flies returned from space were mated to virgin female flies of a tester strain, and the presence of the lethal mutations was analyzed at F2 generation. The frequencies of sex-linked recessive lethal mutations in flight groups were 2 and 3 times higher for wild type Canton-S and mei-4 1, respectively, than those in ground control groups. Most larvae sent to space emerged as adult flies within about 10 days after the landing. The presence of wing-hair somatic mutations, which give morphological change in hairs growing on the surface of wing epidermal cells, was analyzed under microscope. In wild type strain Muller-5, the frequency of wing hair mutant spots in flight group was about 1.5-fold higher than that in ground control, and in Canton-S-derived wild type strain the frequencies were similar between the two groups. By contrast, for mei-41 strain the mutation frequency was lower in flight group than in control group. The observed higher frequency of lethal mutations in the flight group might be due to a possibility that radiation effects on reproductive cells could be greatly enhanced under micro gravity. However, if this would be the case, we do not have appropriate explanation for the apparent absence of such synergistic effects on somatic wing-hair mutation system.     

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.     

The influence of gamma-irradiation in low doses on the rate of dominant lethal mutations in drosophila melanogaster

The dose-rate effect of acute and chronic irradiation in the dose of 0.2 Gy in Drosophila melanogaster repair (mei-41, mus209 [Russian character: see text] mus309) and free radicals detoxication (sod) mutant strains was investigated. Was shown the lack of dose rate effect on the rate of dominant lethal mutations in mei-41, mus209 and sod. However in mus309, that has defect in the main Drosophila pathway of the DNA double strand breack repair, the increase of the mutation rate after chronic irradiation was observed (inverse dose-rate effect). The obtained results suggest the main role of DNA double strand breack repair in dose-rate effect formation in Drosophila.     

Meiosis as a source of spontaneous mutations in Schizophyllum commune

Spontaneous mutation frequencies were determined for two loci in the fungus Schizophyllum commune, at meiosis and at mitosis. For both loci the meiotic frequency is significantly higher than the mitotic frequency. No correlation was found between meiotic mutagenesis and recombination of markers bracketing the mutant site. The meiotic temperature affected the spontaneous mutation frequency but not the recombination frequency in the cross examined.A number of suppressor mutations were detected for both loci examined. Almost all the suppressors are closely linked to the site they suppress. The distribution of mutations among the suppressor sites was different at meiosis and at mitosis.     

The Interaction of the rad201Gene with Genes mei-9and mei-41in Germline Cells of DrosophilaFemales

The effect of Drosophilamutation rad201 ^G1together with mutations mei-41 ^D5and mei-9 ^aon the sensitivity of oocytes to induction of dominant lethals (DLs) was studied. To this end, the frequencies of spontaneous and gamma-radiation-induced DLs in consecutive egg batches of females carrying double or single mutations were estimated. Since the effects of the mutations examined are expressed only at the previtellogenetic stages of oogenesis, only newly hatched (0–5-hour-old) females, whose oocytes did not develop farther than stage 7, were irradiated. The results obtained indicated that in intact and irradiated oocytes of double mutants mei-9 ^a rad201 ^G1and mei-41 ^D5 rad201 ^G1, mutation rad201 ^G1epistatically suppresses the mutations of the both meigenes.     

mei-41 is required for precocious anaphase in Drosophila females

This paper reports on a new role for mei-41 in cell cycle control during meiosis. This function is revealed by the requirement of mei-41 for the precocious anaphase observed in crossover-defective mutants. Normally in Drosophila oocytes, tension on the meiotic spindle causes a metaphase I arrest. This tension results because crossovers, and the resulting chiasmata, hold homologs together that are being pulled by kinetochore microtobules toward opposite spindle poles. In the absence of tension, such as in a recombination-defective mutant, metaphase arrest is not observed and meiosis proceeds through the two divisions. Here we show that in some recombination-defective mutants, the precocious anaphase requires the mei-41 gene product. For example, metaphase arrest is not observed in mei-218 mutants because of the severe reduction in crossing over. In mei-41 mei-218 double mutants, however, metaphase arrest was restored. The effect of mei-41 is dependent on double-strand break formation. Thus, in mutants that fail to initiate meiotic recombination the absence of mei-41 has no effect. Received: 15 October 1999; in revised form: 9 December 1999 / Accepted: 13 December 1999     

Recombination nodules and synaptonemal complex in recombination-defective females of Drosophila melanogaster

The cytological effects of mutant alleles of the mei-9, mei-218, and mei-41 loci during prophase I have been examined by electron microscopy. None of these mutants affect synaptonemal complex structure, continuity, or temporal behavior. Both the precondition-defective mutants mei-218 and mei-41 affect both number and morphology of spherical recombination nodules and apparently affect at least the numbers of ellipsoidal recombination nodules, whereas in the exchange-defective mutant mei-9 the numbers and morphologies of both ellipsoidal and spherical recombination nodules are normal. The parallel effects of mei-218 and mei-41 on meiotic recombination and on recombination nodules indicate that spherical recombination nodules at least mark the site of exchange events; the effects of these mutants on nodule morphology suggest that the nodule performs an active role in the recombination process. The nodule phenotype of mei-9 indicates that spherical nodules are present, and presumably functioning, well before the concluding stages of the recombination event. The parallel effects of all 3 mutants on ellipsoidal and spherical nodules indicate that these are indeed related structures but does not ellucidate the nature of the relationship. It is suggested that all aspects of meiotic recombination are under the aegis of recombination nodules.     

Mutational analysis of the Drosophila DNA repair and recombination gene mei-9

Drosophila mei-9 is essential for several DNA repair and recombination pathways, including nucleotide excision repair (NER), interstrand crosslink repair, and meiotic recombination. To better understand the role of MEI-9 in these processes, we characterized 10 unique mutant alleles of mei-9. These include a P-element insertion that disrupts repair functions but not the meiotic function; three nonsense mutations, one of which has nearly wild-type levels of protein; three missense mutations, one of which disrupts the meiotic function but not repair functions; two small in-frame deletions; and one frameshift.     

Heteroduplex DNA in meiotic recombination in Drosophila mei-9 mutants

Meiotic recombination gives rise to crossovers, which are required in most organisms for the faithful segregation of homologous chromosomes during meiotic cell division. Characterization of crossover-defective mutants has contributed much to our understanding of the molecular mechanism of crossover formation. We report here a molecular analysis of recombination in a Drosophila melanogaster crossover-defective mutant, mei-9. In the absence of mei-9 activity, postmeiotic segregation associated with noncrossovers occurs at the expense of crossover products, suggesting that the underlying meiotic function for MEI-9 is in crossover formation rather than mismatch repair. In support of this, analysis of the arrangement of heteroduplex DNA in the postmeiotic segregation products reveals different patterns from those observed in Drosophila Msh6 mutants, which are mismatch-repair defective. This analysis also provides evidence that the double-strand break repair model applies to meiotic recombination in Drosophila. Our results support a model in which MEI-9 nicks Holliday junctions to generate crossovers during meiotic recombination, and, in the absence of MEI-9 activity, the double Holliday junction intermediate instead undergoes dissolution to generate noncrossover products in which heteroduplex is unrepaired.