Frequency of non-disjunction and loss of sex chromosomes in oogenesis in the Drosophila melanogaster mutant I(1) ts 403 with a defect in the heat-shock protein system in anoxia and at high temperature]

A unique property of Drosophila melanogaster l(1)ts403 strain with the defect in heat shock protein system (HSP) is high frequency of losses and non-disjunction of sex chromosomes induced by heat shock (HS) (37 degrees C, 1 h). This effect was shown in only 6-14-th stages of oocytes. Anoxia was not effective in induction of these mutations. Successive action of anoxia and HS decreased loss frequency and non-disjunction in comparison with the only action of HS. These findings agree with the data in literature indicating that HSP synthesis was increased in the l(1)ts403 mutant when first anoxia and then HS were administered, in contrast to the action of HS only. The role of HSP in the recovery of HS-induced disruptions (chromosomal proteins and meiotic division apparatus) which can lead to chromosome non-disjunction and losses is discussed.     

Studies on mutagen-sensitive strains of Drosophila melanogaster I. The enhanced X-ray sensitivity of a mutant strain (ebony) which is UV-sensitive and also deficient in photorepair

Yegorova and colleagues (1978) showed that a mutant strain of Drosophila melanogaster (ebony) was more sensitive to UV-induced killing of embryos and also less proficient in photoreactivating (PR) ability than a wild-type (Canton-S) strain and that the genes governing UV sensitivity and PR ability were different and presumably located on the autosomes. The experiments reported in the present paper were designed to compare the patterns of sensitivity of these 2 strains and their hybrids to X-irradiation. The sensitivity of the larvae to the killing effects of X-irradiation, and of male and female germ-cell stages to the X-ray induction of genetic damage was studied.It was found that the larvae of the ebony strain are more sensitive to X-ray-induced killing than those of the Canton-S strain. The frequencies of radiation-induced dominant lethals and sex-linked recessive lethals are higher in spermatozoa sampled from ebony males than in those of Canton-S males. In spermatozoa sampled from hybrid males, the yields of dominant lethals are no higher than in those sampled from Canton-S males and do not seem to depend on the origin of the X-chromosome. There are no statistically significant differences between the ebony and Canton-S strains in the sensitivity of their spermatozoa to the induction of autosomal translocations.Stage-7 oocytes sampled from ebony females are more sensitive to the X-ray induction of dominant lethality than are those from Canton-S females; oocytes sampled from hybrid females manifest a level of sensitivity that is significantly lower than that in either parental strain. The frequencies of X-chromosome losses induced in in this germ-cell stage are significantly lower in ebony than in Canton-S females at least at the exposure level of 3000 R at which 3 experiments were carried out. There are no measurable differences in the amount of dominant lethality induced in stage-14 oocytes of ebony, Canton-S and hybrid females.When X-irradiated Berlin-K males are mated to ebony or Canton-S females, the yields of dominant lethals are higher when ebony females are used, showing that there is a “maternal effect” for this kind of damage. Such a maternal effect is also found for sex-linked recessive lethals (irradiated Muller-5 males mated to ebony or Canton-S females). However, when irradiated ring-X-chromosome-carrying males are mated to ebony or Canton-S females, the frequencies of paternal sex-chromosome losses (scored as XO males) are lower when ebony females are used.These results have been interpreted on the assumption that the ebony strain is homozygous for recessive, autosomal genes that confer increased radiosensitivity and that the Canton-S strain carries the normal, wild-type alleles for these genes. The higher yields of dominant and recessive lethals in mature spermatozoa and of dominant lethals in stage-7 oocytes are a consequence of an enhanced sensitivity to the mutagenic (in particular, to the chromosome-breaking) effects of X-irradiation and/or of defective repair of radiation-induced genetic damage. The lower yield of XO males from irradiated stage-7 oocytes of ebony females is probably a consequence of a defect in the repair of chromosome-breakage effects, resulting in the conversion of potential X losses in females into dominant lethals. The “maternal effects” for dominant lethals, sex-linked recessive lethals and for the loss of ring-X chromosomes are assumed to have a common causal basis, namely, a defective repair of chromosome-breakage events in the females of the ebony strain.     

Effects of a chromosome-3 mutator gene on radiation-induced mutability in Drosophila melanogaster females

A series of X-irradiation experiments was carried out using Drosophila melanogaster females homozygous for a third chromosome mutator gene and females which had a similar genetic background except that the mutator-bearing third chromosomes were substituted by normal wild-type chromosomes. The mutator females had been previously shown by Gold and Green to manifest a higher level of radiation-induced mutability (as measured by the X-ray-induction of sex-linked recessive lethals) in their pre-meiotic germ cells compared to normal females at an exposure of 100 R. In the presence work, the sensitivity of the pre-meiotic germ cells of mutator and normal females to the X-ray induction (2000 R) of sex-linked recessive lethals was studied. In addition, experiments were conducted to examine the sensitivity of the immature (stage 7; prophase I of meiosis) oocytes of both kinds of females to the induction of dominant lethals, X-linked recessive lethals and X-chromosome losses. The result show that in pre-meiotic germ cells, the frequencies of radiation-induced recessive lethals are similar in both kinds of females. However, the proportion of these mutations that occur in clusters of size 3 and higher, is higher in mutator than in normal females. In stage-7 oocytes, the frequencies of radiation-induced dominant lethals and sex-linked recessive lethals were similar in both kinds of females. The X-loss frequencies however, were consistently higher in mutator females although statistical significance was obtained only at higher exposures (3000 and 3750 R) and not at lower ones (750-2250 R). Possible reasons for the discrepancy between the present results and those of Gold and Green with respect to pre-meiotic germ cells are discussed.     

Distribution of MR-induced sex-linked recessive lethal mutations in Drosophila melanogaster

In the ‘doubling-dose’ method currently used in genetic risk evaluation, two principle assumptions are made and these are: (1) there is proportionality between spontaneous and induced mutations and (2) the lesions that lead to spontaneous and induced mutations are essentially similar. The studies reported in this paper were directed at examining the validity of these two assumptions in Drosophila. An analysis was made of the distribution of sex-linked recessive lethals induced by MR, one of the well-studied mutator systems in Drosophila.

Appropriate genetic complementation tests with 15 defined X-chromosome duplications showed that MR-induced lethals occurred at many sites along the X-chromosome (in contrast to the known locus specificity of MR-induced visible-mutations); some, but not all these sites at which recessive lethals arose in the MR-system are the same as those known to be hot-spots for X-ray-induced lethals. With in situ hybridization we were able to demonstrate that a majority of MR-induced lethals is associated with a particular mobile DNA sequence, the P-element, i.e. they arose as a result of transposition.

The differences between the profiles of MR-induced and X-ray-induced recessive lethals, and the nature of MR-induced and X-ray-induced mutations, thus raise questions about the validity of the assumptions involved in the use of the ‘doubling-dose’ method.     

Characteristics of inserted mutations obtained in the P-M hybrid dysgenesis system in the 9F12-10A7 region of the Drosophila melanogaster X-chromosome

19 new mutations in the 9F12-10A7 region of Drosophila melanogaster X chromosome was obtained in the system of P-M hybrid dysgenesis. They appeared to be lethals, as judged from viability of homo- or hemizygous females. In situ hybridization of P DNA with polytene chromosomes revealed P-element insertion in the 10A1-2 band in the majority of the mutants. As a result of complementation analysis, all these mutations were localized at previously known loci: l(1)BP1, l(1)BP5, l(1)BP8, l(1)BP7. No insertion mutations were found at the vermilion locus. This can imply for non-random distribution of insertion mutations in the region studied. Further comparison of these mutations with previously EMS-induced ones revealed that insertion mutations are predominantly hypomorph lethals which do not influence the viability, morphology and fertility of homozygous males and females, but drastically reduce viability of hemizygous females.     

Distribution of MR-induced sex-linked recessove lethal mutations in Drosophila melanogaster

In the ‘doubling-dose’ method currently used in genetic risk evaluation, two principle assumptions are made and these are: (1) there is proportionality between spontaneous and induced mutations and (2) the lesions that lead to spontaneous and induced mutations are essentially similar. The studies reported in this paper were directed at examining the validity of these two assumptions in Drosophila. An analysis was made of the distribution of sex-linked recessive lethals induced by MR, one of the well-studied mutator systems in Drosophila.Appropriate genetic complementation tests with 15 defined X-chromosome duplications showed that MR-induced lethals occurred at many sites along the X-chromosome (in contrast to the known locus specificity of MR-induced visible-mutations); some, but not all these sites at which recessive lethals arose in the MR-system are the same as those known to be hot-spots for X-ray-induced lethals. With in situ hybridization we were able to demonstrate that a majority of MR-induced lethals is associated with a particular mobile DNA sequence, the P-element, i.e. they arose as a result of transposition.The differences between the profiles of MR-induced and X-ray-induced recessive lethals, and the nature of MR-induced and X-ray-induced mutations, thus raise questions about the validity of the assumptions involved in the use of the ‘doubling-dose’ method.     

Radiosensitivity of Drosophila lines. VII. The effect of the maternal genotype on the yield of recessive and dominant lethal mutations induced by the gamma irradiation of males

The effect of material repair on induction of paternal mutations was tested with radiosensitive rad(2)201G1 mutant. Basc males were irradiated at doses from 0 to 60 Gy of gamma-rays and mated to the radiosensitive mutant or control females. Frequencies of sex-linked recessive lethals and dominant lethals (induced in the paternal genome) were determined. With control females, the rate of recessive lethals increased linearly from 0 to 60 Gy. With rad(2)201G1 mutant, an increase in spontaneous and induced rates of paternal dominant lethals was observed; the rate of sex-linked recessive lethals increased non-linearly from 0 to 60 Gy.     

Comparative induction of somatic eye-color mutations and sex-linked recessive lethals in Drosophila melanogaster by tryptophan pyrolysates

The mutagenicities of the products of pyrolysis of tryptophan, Trp-P-1 and Trp-P-2, on Drosophila melanogaster were examined by measuring the effects of these compounds in inducing recessive lethals and somatic eye-color mutations. Since negative results have already been obtained by the standard procedure in males, Trp-P-1 and Trp-P-2 (0.75 to 6 mg/ml) in sucrose solution were given to females for assay of recessive lethal mutations in X-chromosomes. These compounds caused a marginal increase above the control level in the mutation frequency. For the assay of effects on somatic eye-color mutations, Trp-P-1 (200 and 400 ppm) and Trp-P-2 (400 and 800 ppm) were fed to male larvae of a tester strain carrying a genetically unstable marker set of z and w+ on the X-chromosome. These compounds caused dose-dependent increases above the control level in somatic eye-color mutations in adults. It is concluded that, under the conditions used, the somatic eye-color mutation system was more sensitive than the recessive lethal system to the mutagenic effects of tryptophan pyrolysates.     

The effect of high temperature on the frequency of nondisjunction and loss of sex chromosomes in females of Drosophila melanogaster strain I(1)ts403 with a defect in the heat shock protein system

High temperature (37 degrees C) induces sex chromosome non-disjunction and loss in Drosophila melanogaster females l(1)ts403 with the defect in heat-shock protein system. The same temperature has no effect on the females from the T line and other earlier studied lines.     

EMS-induced dominant lethal dose response curve in DBA/1J male mice.

An ethyl methanesulfonate (EMS) induced dominant lethal dose response experiment was conducted in strain DBA/1J male mice. Two methods of scoring for dominant lethals, the classic method (dissecting females at mid-pregnancy) and an alternative method (inspection of uterine scars after litters were weaned) were compared. Results indicate that strain DBA/1J has a similar sensitivity to EMS-induced dominant lethals as has been previously reported for other mouse genotypes. Of the two methods used to score dominant lethals, the classic method is more sensitive.     

Effect of sterol metabolism in the yeast-Drosophila system on the frequency of radiation-induced aneuploidy in Drosophila melanogaster oocytes

The influence of sterol metabolism upon mutagenesis in Drosophila melanogaster was investigated using ecological-genetic yeast - drosophila system. Sterol deficiency in the organism of Drosophila was caused by using the strain of Saccharomyces cerevisiae 9-2P712 with a mutation in the nysr1 locus which blocks synthesis of ergosterol as a nutrition substrate for flies. It was concluded that maintenance of females on the mutant yeast strain causes an increase of radiation-induced X-chromosome loss in mature oocytes. Resistance of oocytes to X-ray irradiation is restored, reaching the control level, when 0,1% cholesterol solution in 10% ethanol is added to the yeast biomass. The possible membrane and hormonal mechanisms of elevation of induced aneuploidy and the role of sterol metabolism in ensuring resistance of insects to damaging factors are discussed.     

Semidominant effect of the l(1)ts403 (sbr10) mutation on nondisjunction of sex chromosomes in meiosis in Drosophila melanogaster females exposed to heat

The sbr gene of Drosophila melanogaster belongs to the NXF (nuclear export factor) family responsible for the mRNA transport from nucleus to cytoplasm. We have shown that in the heat-exposed (37 degrees C, 1 h) females, the l(1)ts403 (sbr10) mutation leads, in particular, to the high-frequency nondisjunction and loss of sex chromosomes in meiosis. For this trait, the incomplete dominance of the sbr10 mutation is observed. At the same time, the sbr10 mutation is recessive for many other traits of the heat-exposed flies: reduced viability, low fertility, impaired synthesis of the heat shock proteins, etc. The females heterozygous for the null allele (Df(1)vL4, a deletion eliminating gene srb) do not differ from females homozygous for the wild-type allele in frequency of the heat shock-induced nondisjunction and loss of sex chromosomes in meiosis. Because of this, the sbr10 mutation can be assigned to the gain-of-function alleles (those gaining the dominance function). Expression of the mutant sbr10 allele against the background of the wild-type allele suggests that in the heat shock-exposed females, the heat-modified product of this ts allele has a strong effect on sex chromosome disjunction in meiosis.     

Temperature-Sensitive Mutations in DROSOPHILA MELANOGASTER. Xvii. Heat- and Cold-Sensitive Lethals on Chromosome 3

Ethyl methanesulfonate-treated third chromosome of Drosophila melanogaster were tested for the presence of dominant and recessive temperature-sensitive lethal mutations at 17 degrees , 22 degrees and 29 degrees C. Out of 1,176 chromosomes tested, no dominant ts lethals, 21 heat-sensitive, 22 cold-sensitive and 10 heat-cold-sensitive lethals were recovered. Heat-cold sensitivity was produced by a single mutation in all cases. Sixty-two percent of the ts lethals were fertile as homozygotes in both sexes. Surprisingly, 88% of the ts lethals mapped between st and Sb, a region straddling the centromere and estimated to comprise 12.9% of the genetic length and 55% of the physical length of chromosome 3. All but one of the heat- and cold-sensitive lethals complemented with each other at their respective restrictive temperatures.     

Radiosensitive Drosophila strains. IX. An analysis of the fertility and frequency of dominant lethal mutations in gamma-irradiated females of the mutant rad(2)201G1 strain

Fertility and frequency of gamma-induced dominant lethals in female oocytes have been studied in a strain of Drosophila melanogaster carrying rad(2)201G1 mutation and in the wild type strain. It was shown that oocytes of the mutant strain exhibited the higher sensitivity during the whole period of oogenesis, as compared to those of the wild type flies. The strongest influence of rad(2)201G1 mutation on the frequency of dominant lethals and fertility was observed.     

Investigations on radiosensitive and radioresistant populations of Drosophila melanogaster. VIII. The system of relative radioresistance in immature oocytes of the irradiated population ROI4

Prophase I oocytes of the irradiated population ROI4 of Drosophila melanogaster are radioresistant relative to those of a control population (+K). The system of relative radioresistance is apparently dose-modifying and can be described by Dose-Reduction Factors (DRFs). At least 3 constituent components of the system can be distinguished, as follows. The genetic factor rar-1 contributes to the system with respect to the induction of dominant (DRF = 1.31) and sex-linked recessive lethals (DRF = 1.31) in a way that is inhibited by caffeine. The factor rar-2, independently reduces both types of lethal to the same amount as does rar-1, but also affects the production of X-chromosome loss (DFR = 1.72). The results of several different approaches allow, as a working hypothesis, the interpretation that rar-2 reduces the association of heterologous, chiasmatic chromosomes in the chromocentre in time and/or space and thus minimizes the preconditions for the production of certain types of interchange and of non-disjunction. A third factor, rar-3, is postulated to contribute, independently from the others, to the system of relative radioresistance with respect to dominant lethals (DRF = 1.58), interchanges and non-disjunction (DRFs = 1.58), and sex-linked recessive lethals (DRF = 1.87).     

Vital Genes That Flank Sex-Lethal, an X-Linked Sex-Determining Gene of DROSOPHILA MELANOGASTER

The X-chromosome:autosome balance in D. melanogaster appears to control both sex determination and dosage compensation through effects on a maternally influenced sex-linked gene called Sex-lethal (Sxl; 1-19.2). To facilitate molecular and genetic analysis of Sxl, we attempted to determine the locations of all ethyl methanesulfonate (EMS)-mutable genes vital to both sexes in the region between 6E1 and 7B1. This area includes approximately 1 cM of the genetic map on each side of Sxl and was reported by C. B. Bridges to contain 26 salivary gland polytene chromosome bands. The region appears rather sparsely populated with genes vital to both sexes, since the 122 recessive lethal mutations we recovered fell into only nine complementation groups. From one to 38 alleles of each gene were recovered. There was a preponderance of embryonic lethals in this area, although the lethal periods of loss-of-function mutations included larval, pupal and adult stages as well. Since the screen required that mutations be recessive and lethal to males, our failure to recover new Sxl alleles was the result expected for a gene with a female-specific function. An attempt was made to identify recessive male-specific lethals in this region, but none were found. Precise map positions were determined for eight of the nine vital genes. An interesting feature of the map is the location of Sxl in the middle of a 0.6- to 0.7-cM interval that appears to be devoid of genes vital to both sexes. The genetic location was determined of breakpoints near Sxl for all available chromosome rearrangements. Sxl is most likely located just to the left of band 7A1. We determined the relationship of our EMS-induced mutations in these nine genes to alleles induced by others. From this we conclude that the various genes appear to differ significantly from each other in their relative sensitivity to mutation by EMS vs. X rays.     

Maternal and paternal effects on the specific mutation l(1)ts403 of thermosensitivity of early Drosophila melanogaster embryos]

High thermosensitivity of early embryos controlled by mutation l(1)ts403 with disturbed heat-shock response was studied. Thermosensitivity was examined in early (0-1 h) and late (3.5-4.5 h) embryos obtained by reciprocal crosses and backcrosses. It was shown that mutation l(1)ts403 lacks maternal effect. In progeny of reciprocal crosses, early embryonic thermosensitivity was intermediate with regard to that of progeny obtained by interlinear crosses. In early embryos of Drosophila, zygotic genes are not expressed and synthesis heat-shock protein synthesis is not induced. Based on this, it was proposed that the product of gene l(1)ts403, which affects early embryonic thermosensitivity, is transmitted both paternally and maternally and shows dosage effect.     

Test of a semiselective screen for induced aneuploidy in germ cells of Drosophila melanogaster females with structurally normal chromosomes

A new semiselective screen (only female progeny survive) for induced aneuploidy in germ cells of Drosophila melanogaster (referred to as 20/Q56 for the X-chromosome mutation markers in the parental females) has been validated by recovering cold, colchicine and N,N-dimethylnitrosamine (DMN) induced chromosome gain and loss events in females that contain structurally normal chromosomes. In addition, the spontaneous and induced results from the 20/Q56 assay, which identifies gain events at division I and loss events at divisions I and II of meiosis, were compared with a nonselective (all progeny survive) modified mating scheme that identifies gains and losses at both divisions of oogenesis. Females with the same genotypes are treated in the two mating schemes and are then mated with males that contain different marked Y chromosomes. The spontaneous rates of chromosome gains and losses were not significantly different in the two mating schemes (these rates ranged from 0.008 to 0.022%), supporting previous reports that spontaneous aneuploidy occurs at a higher frequency at division I of meiosis in females of D. melanogaster than at division II. Both the 20/Q56 and modified screens were able to identify significant increases in aneuploidy after adult treatments with cold shock (10 degrees C and 5 degrees C), colchicine (5 ppm and 10 ppm), and DMN (100 ppm). Brood analysis (five 2-day or five 3-day broods) showed that the largest increases in aneuploidy after cold treatment occurred in the first brood, which contains a high proportion of stage 14 oocytes, whereas colchicine induced the highest frequencies in the latter broods and DMN was effective in all but the last brood. Although the 20/Q56 mating scheme identifies gain events only in division I of meiosis whereas the modified mating scheme identifies gains in both divisions, the 20/Q56 scheme is just as effective in identifying induced aneuploidy as is the modified scheme. There were no significant differences in the frequencies of induced gains or losses in the two schemes. These results also suggest that the 3 treatments induced chromosome gain events mainly at division I of oogenesis. Taken together, the results from this study suggest that the 20/Q56 mating scheme in D. melanogaster, which is semiselective and therefore less expensive and time-consuming to perform, is an appropriate test system to screen for chemical induced aneuploidy in germ cells of a higher organism.