Behavior of hobo and P transposons in yellow 2-717 unstable line of Drosophila melanogaster and its derivatives after crossing with a laboratory strain

Using fluorescent in situ hybridization technique (FISH), the frequency of hobo and P mobile elements transpositions on X chromosomes from the y ^2-717 , isolated from the Uman’ population of Drosophila melanogaster, as well as from its phenotypically normal and mutant derivatives, obtained as a result of crosses the males examined with the C(I)DX,ywf/Y females, was evaluated. It was demonstrated that the maximum frequency of hobo transpositions on X chromosomes of the males from derivative strains, subjected to repeated hobo-dysgenic crosses reached a value of 1.2 × 10^?2 per site per X chromosome per generation. The number of hobo copies in male X chromosomes from derivative strains was 3 times higher than in the original initial strain. Furthermore, the “old” hobo sites remained unchanged. In derivative strains, the frequency of hobo insertion was higher than that of excisions. One of the derivative strains, y ^1t-717a1k3-2 , was characterized by high intrastrain instability of hobo element localization. In the y ^2-717a1k3 and y ^1t-717a1k3-2 strains a large inversion, In(1)IB; 13CD, was described. At the absence of the full-sized P element in the strains involved in crosses, maximum frequency of P element transpositions in the derivative strains reached a value of 1.2 × 10^?2 per site per X chromosome per generation.     

The effect of gamma-radiation on induction of the hobo element transposition in Drosophila melanogaster

The transposition frequency of the hobo mobile element in four successive generations of Drosophila melanogaster strain y2-717 after an acute gamma-irradiation with a dose of 30 Gr amounted to 7.5 x 10(-4) per site per genome per generation. Under the same conditions, PCR analysis of the genomic DNA of y2-717 flies detected new variants of defective hobo sequence. No changes in the hobo localization and PCR products compared with the control were detected in the case of single irradiation with doses of 3 and 30 Gr. The localizations of hobo element on polytene chromosomes of y2-717 strain did not change during 11 generations after five exposures of flies to 30 Gr. Irradiation of a highly unstable D. melanogaster strain y+743 did not increase the number of families with mutant progeny, yet increased the total number of mutant descendants almost twofold, from 5 to 9%.     

Three-dimensional organization of telomeres in the Drosophila melanogaster salivary glands nuclei

The 3D-FISH was employed to investigate the telomere topology in polytene nuclei of salivary glands of Drosophila melanogaster. The majorities of telomeres in polytene nuclei of salivary glands in Drosophila strain y(2-717) are localized in the nuclear central area and have no contacts with nuclear membrane. In females of this strain, ectopic contacts between telomeres occur at 25 % higher frequency than in males. HeT-A DNA in y(2-717alk3-2) strain, which is a derivative of y(2-717) carrying an inversion between 1D and 13C bands, is found in region 13 of X chromosome. The frequency of ectopic contacts of telomeres in y(2-717alk3-2) males is 10 % higher than that in y(2-717) strain. The number of ectopic contacts can be significantly different in independent experiments, possibly indicating the role of random factors in the contact formation.     

Transposition of the hobo element in Drosophila melanogaster somatic cells

Somatic mutation and recombination test on wing cells of Drosophila melanogaster showed that the recombination frequency in the somatic tissues of strains studied correlated with the presence of a full-length copy of the hobo transposable element in the genome. Transposition of hobo in somatic tissue cells at a frequency 3.5 x 10-2 per site per X chromosome was shown by fluorescence in situ hybridization with salivary gland polytene chromosomes of larvae of one of the D. melanogaster strains having a full-length hobo copy.     

Persistent locus-specific instability of yellow(27-717) and Noth(Uc-1) in Drosophila melanogaster coincides with hobo multiplication

According to FISH data the presence of multiple hobo element copies in the unstable yellow and Notch loci in y(2-717) and Uc-1 Drosophila melanogaster stocks, respectively, was found. Locus-specific instability in these strains is caused by hobo multiplication in the respective loci and its subsequent recombination with neighboring hobo copies rather than its insertion-excision.     

Genetic instability and transposition of mobile element mdg4 in a mutator strain of Drosophila melanogaster

Laboratory mutator strain of Drosophila melanogaster is characterized by increased (up to 10(-3)-10(-4) frequency of spontaneous mutability. Mutations appear in premeiotic stages of gametes development. The majority of mutations were unstable (high frequencies of reversions, appearance of new mutations at the same and other loci, replicating instability). Localization of mobile elements mdg1, mdg2, mdg3, mdg4, copia and P element in X chromosomes of mutator individuals and its mutations y, ct, sbt was studied by hybridization in situ. In all strains P element was absent. The distribution of mdg1, mdg2, mdg3 and copia was identical in mutator strains and its derivatives, but distribution of mdg4 was different. The essential heterogeneity in localization of mdg4 and increased (up to 30-40) copy number in the mutator strain individuals was observed. The ability of single element mdg4 to autonomous transpositions was thus shown.     

Mobilization of hobo elements residing within the decapentaplegic gene complex: suggestion of a new hybrid dysgenesis system in Drosophila melanogaster

We present results demonstrating that the hobo family of transposable elements can promote high rates of chromosomal instability. Using strains with a hobo element inserted within the decapentaplegic gene complex (DPP-C), we have recovered numerous DPP-C mutations involving chromosomal rearrangements and deletions with one endpoint in the vicinity of the pre-existing hobo element. This hypermutability occurred in the germ lines of hybrid progeny from crosses involving strains containing hobo elements to strains lacking them. In some crosses, the offspring had rudimentary gonads, reminiscent of GD sterility. The germline hypermutability and infertility are similar to those produced by P-element-mediated hybrid dysgenesis. Given the many genetic and molecular similarities of the P and hobo systems, we propose that a system analogous to P-M hybrid dysgenesis has been activated in the hobo+ X hobo- crosses.     

The effect of genetic environment on locus-specific instability of the yellow gene in Uman' population of Drosophila melanogaster

The effects of genotype of the laboratory strains, C(1)DX, ywf/Y, 23.5 MRF/CyL4, and C(1)DX,yf; pi2, on locus-specific instability in the yellow gene of the strains y(2-717, y(2-715), and y(2-700 ) from Uman' population of Drosophila melanogaster was studied. Crosses of the males from Uman'-derived lines with the C(1)DX, ywf/Y females yielded a cascade of derivatives, mostly consisting of y+ and y2 alleles, while their crosses with the 23.5 MRF/CyL4 and C(1)DX,yf; pi2 females mostly resulted in the appearance of y+ and y(1) derivatives. The genomes of laboratory strains used in the study contained the full-sized hobo elements, which could differ from one another relative to the structure of variable region and affinity to different DNA sequences.     

Participation of mobile element hobo in transposition events in the long-term instability system of Drosophila melanogaster]

The lines with an active hobo elements as well as those without any hobo fragments were hybridized with the y2sc1waG line. This resulted in the appearance of a number of mutations at the white, miniature, and some other loci. The authors analysed, in which way the hobo transposable elements take part in mutagenesis in these crosses. Most of the white mutants obtained were analysed and transpositions of hobo and Stalker elements were demonstrated. Both independent and simultaneous transpositions were found. It was shown by means of the Southern blot analysis that additional hobo or Stalker insertion into or close to the parental unknown waG insertion resulted in mutant white phenotype's shift toward both extreme and partial reversion. Possible participation in mutagenesis of other mobile elements is also under debate.     

Three-dimensional organization of telomeres in nuclei of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> salivary glands

3D-FISH method was employed to investigate the telomere topology in polytene nuclei of salivary glands in Drosophila melanogaster. The majority of telomeres in polytene nuclei of salivary glands in Drosophila strain y^2-717 is localized in the nuclear central area and have no contacts with nuclear membrane. In the females ectopic contacts between telomeres occur in frequency 25% higher than in males. HeT-A DNA in y^2-717alk3-2 strain, which is a derivative of the y^2-717 stain carrying an inversion between 1B and 13C bands, is found in the region 13 of X chromosome. The frequency of ectopic contacts of telomeres in y^2-717alk3-2 males is 10% higher than in the y^2-717 strain. The number of ectopic contacts can be differ significantly in independent experiments, possibly indicating the role of random factors in the contact formation.     

Polymorphism of yellow locus in Drosophila melanogaster from natural populations

We studied molecular characteristics of yellow (y; 1-0.0) locus, which determines the body coloration of phenotypically wild-type and mutant alleles isolated from geographically distant populations of Drosophila melanogaster in different years. According to Southern data, restrictions map of yellow locus of all studied strains differ from each other as well as from that of Oregon stock. FISH analysis shows that in the neighborhood of yellow locus in X chromosome neither P nor hobo elements are found in y1-775 stock, while only hobo is found there in y1-859 and y1-866 stocks, only P element in y+sn849 stock, and both elements in y1-719 stock. Thus, all studied mutant variants of yellow are of independent origin. Yellow locus residing at the very end of X chromosome (region 1A5-8 of cytologic map) carries significantly more transposon than retrotransposon-induced mutations, as compared to white locus (regions 3C2). It is possible that transposons are more active than retrotransposons at the chromosomal ends of D. melanogaster.     

Ectopic interactions of Drosophila melanogaster polytene chromosomes in the H-E system of hybrid dysgenesis]

Analysis of salivary gland polytene chromosomes of Drosophila melanogaster age III larvae has demonstrated that inversions make up high percentage of aberrations induced by transposition of the hobo element. The frequency of inversions increases upon irradiation. In view of the fact that transpositions of mobile elements and irradiation can considerably affect the homology of associating regions, ectopic chromosome contacts were analysed. On the one hand, this analysis has revealed general features for all types of crosses, such as the prevalence of intrachromosomal contacts over interchromosomal ones, predominant localization of both types of contacts in telomeric regions of chromosomes and lower frequency of their occurrence on X chromosome in males as compared with females. On the other hand, the specificity of ectopic conjugation in different types of crosses has been determined. Given a constant average frequency of ectopic contacts in the D. melanogaster genome, differences in their distribution in chromosomes have been detected. Chromosomes of pure lines differ from their homologues in dysgenic hybrids by distribution of ectopic conjugation peaks and by localization of a number of unique events. On the basis of our own experiments and literature data, the role of ectopic contacts as structural formations connecting chromosome regions similar in their functional activity, rather than in nucleic-protein characteristics, is discussed.     

Transpositions of mobile elements mdg4 (gypsy) and hobo in somatic and germ cells of a genetically unstable mutator strain of Drosophila melanogaster]

Analysis of distribution of the several families of mobile genetic elements has been performed. The analysis dealt with the X chromosomes of male progeny from the crosses of individual males of Mutator strain (MS) with attached-X females. The experimental results demonstrated different localization of the elements gypsy and hobo in the salivary gland squashes of different males-brothers. Location of other elements under study--mdg1, 412, mdg3, copia, 297, 17.6, Beagle, BS, Doc, FB, Springer--was invariant in all larvae. The analysis is equal to the study of transposition events at the level of gametes. Thus, doubtless, the capability of gypsy and hobo to transpose in germ cells of the MS individuals has been detected. Mobilization of the elements occurs at premiotic stages of gametes' development, as indicated by appearance of the clusters of transpositions. In the process of studies on coincidence of gypsy and hobo transposition acts, independent character of the elements' movement has been revealed. It has been detected in the same experiment that the distribution of the gypsy copies in different cells of the same salivary gland varies strongly. All hybridization sites were divided into two groups: "constant" sites common for all cells and "additional" ones, whose locations did not coincide in neighbouring cells of salivary gland. The existence of additional sites is major evidence of gypsy transpositions in somatic cells of MS. Transposition events have been as well discovered for hobo in somatic cells.     

hobo is responsible for the induction of hybrid dysgenesis by strains of Drosophila melanogaster bearing the male recombination factor 23.5MRF

The male recombination factor 23.5MRF, isolated ten years ago from a natural Greek population of Drosophila melanogaster, has been shown to induce hybrid dysgenesis when crossed to some M strains, in a fashion slightly different from that of most P strains. Furthermore, it was recently shown that 23.5MRF can also induce GD sterility when crossed to specific P strain females (e.g., Harwich, pi 2 and T-007). In these experiments, the P strains mentioned behaved like M strains in that they did not induce sterility in the reciprocal crosses involving 23.5MRF. We extended the analysis to show that 23.5MRF does not destabilize snW(M) and that a derivative with fewer full-length P elements behaves like an M strain toward the same P strains and still retains its dysgenic properties in the reciprocal crosses. We show that there is a strong correlation between the site of dysgenic chromosomal breakpoints induced by 23.5MRF and the localization of hobo elements on the second chromosome, and also that hobo elements are found associated with several 23.5MRF induced mutations. These results suggest that hobo elements are responsible for the aberrant dysgenic properties of this strain, and that they may express their dysgenic properties independent of the presence of P elements.     

Efficient and Dispersed Local P Element Transposition from Drosophila Females

We have investigated how Drosophila P element insertions are distributed in the chromosomal region near their starting site. A single P element residing in the euchromatin of minichromosome Dp1187 was mobilized following a cross to the Δ2-3 (99B) strain, and progeny bearing transpositions were identified with a minimum of bias by performing Southern blots on progeny. Approximately 1-2% of all progeny minichromosomes contained new insertions. Many of these ``local transpositions' landed very close to or within the starting P element; however, nearly 1% of all progeny chromosomes contained new insertions 1-180 kb from the donor element. More local insertions were observed in the progeny of females than from male parents, and most occurred in a preferred orientation relative to the starting element. These observations suggested that donor elements are frequently excised and reinserted locally without ever dissociating from a transposition complex. The high frequency and diverse distribution of local transpositions recovered from females suggested that the efficiency of insertional mutagenesis can be significantly enhanced by using a starting P element(s) located near the target of interest.     

Analysis of Dysgenesis-Induced Lethal Mutations on the X Chromosome of a Q Strain of DROSOPHILA MELANOGASTER

The Q strain known as v6 was tested for its ability to induce X-linked lethal mutations in male and female hybrids from crosses with M strains in the P-M system of hybrid dysgenesis. All measurements of the mutation rate were made on the X chromosome derived from the v6 strain. The lethal rate for young hybrid males from the cross M female X v6 male was 1.11% per chromosome. For older males, it was only 0.44%, suggesting that there is less mutational or more repair activity in the germ cells of the older males or that mutant cells are selectively eliminated as the hybrid males age. The lethal rate for hybrid females from comparable crosses was approximately the same for both ages that were tested. However, it was substantially less than the rate for the hybrid males--only 0.26% per chromosome. Genetically identical hybrid females from reciprocal crosses also showed a low mutation rate, 0.13% per chromosome. Again, there was no difference between young and old flies. Mapping experiments established that most of the lethal mutations that were recovered from the male and female hybrids were located in two regions on the X chromosome, one between bands 14B13 and 15A9 , the other between bands 19A1 and 20A , which encompasses the maroonlike locus. More refined mapping of the lethals in the maroonlike region demonstrated that the vast majority of these affected a single gene located in band 19C4 . Cytological analysis of the lethal chromosomes revealed that several carried rearrangements, including inversions, duplications and deficiencies. Chromosome breakage occurred primarily in bands 14D1 -3 and 18F- 20A , and most of the breaks in the latter segment were located in 19C . However, rearrangements involving 19C and mutations of the gene in 19C4 were mutually exclusive events. In situ hybridization of a P element probe to the chromosomes of v6 demonstrated that P elements reside at a minimum of five sites on the X chromosome. These P element sites correspond to the mutational and breakage hot spots on that chromosome. The combined genetic and cytological data imply that most of the X-linked lethal mutations that occur in M X v6 hybrids are due to local P element action. Consideration of these and other data suggest that v6 is a weak P strain in the P-M system of hybrid dysgenesis and that other Q strains might also be regarded in this way.     

Transposable Element-Induced Response to Artificial Selection in DROSOPHILA MELANOGASTER

The P family of transposable elements in Drosophila melanogaster transpose with exceptionally high frequency when males from P strains carrying multiple copies of these elements are crossed to females from M strains that lack P elements, but with substantially lower frequency in the reciprocal cross. Transposition is associated with enhanced mutation rates, caused by insertion and deletion of P elements, and chromosome rearrangements. If P element mutagenesis creates additional variation for quantitative traits, accelerated response to artificial selection of progeny of M female female X P male male strain crosses is expected, compared with that from progeny of P female female X M male male strain crosses.--Divergent artificial selection for number of bristles on the last abdominal tergite was carried out for 16 generations among the progeny of P-strain males (Harwich) and M-strain females (Canton-S) and also of M-strain males (Canton-S) and P-strain females (Harwich). Each cross was replicated four times. Average realized heritability of abdominal bristle score for the crosses in which P transposition was expected was 0.244 +/- 0.017, 1.5 times greater than average heritability estimated from crosses in which transposition was expected to be rare (0.163 +/- 0.010). Phenotypic variance of abdominal bristle score increased by a factor of four in lines selected from M female female X P male male crosses when compared with those selected from P female female X M male male hybrids. Not all quantitative genetic variation induced by P elements is additive. A substantial fraction of nonadditive genetic variation is implicated by chromosomal analysis, which demonstrates deleterious fitness effects of the mutations when homozygous.--Several putative "quantitative" mutations were identified from chromosomes extracted from the selected lines; these will form the basis for further investigation at the molecular level of the genes controlling quantitative inheritance.     

Transposition of mobile dispersed genes (MDG) after substitution of various chromosome pairs in inbred Drosophila melanogaster strains

Transpositions of MDG-1, MDG-3 and copia were detected as a result of crosses of the inbred maladaptive LA stock with laboratory stocks, in order to construct the genomes carrying different combinations of the LA or non-La chromosomal pairs. Changes of the mobile gene distributions were revealed in chromosomes of hybrid genotypes, as compared to parental chromosomal pairs. A trivial source of variability of chromosomal molecular structure ensured by crossing over was excluded by inversions which serve as suppressors of crossing over in corresponding crosses. Multiple transpositions of mobile genes in definite chromosomal sites were detected in genotypes carrying chromosomal pair 2 originated from the LA stock. No such transpositions were observed, when the pair 2 was substituted by the chromosome 2 originated from the Swedish-b line or in control crosses, where the LA stock was not involved. Both LA chromosomes 2 and 3 were shown to be the targets of transpositions. Comparison of hot spot transposition sites of MDG-1, as a result of crosses, with the earlier described rare events of spontaneous transpositions in the LA stock, coupled with its fitness increase, revealed that the hot spot sites were shared in both series of experiments. The data obtained show that transpositions of mobile genetic elements may change the genetic and molecular structure of the chromosome involved in crosses, in spite of suppression of crossing over by inversions usually suggested as a tool for keeping chromosomal genetic structure intact.     

Dynamics of the hobo transposable element in transgenic lines of Drosophila melanogaster

The impact of the hobo transposable element in global reorganization of the Drosophila melanogaster genome has been investigated in transgenic lines generated by injection of hobo elements into the Hikone strain, which lacked them. In the present extensive survey, the chromosomal distribution of hobo insertion sites in the line 28 was found to be homogeneous and similar for all chromosomal arms, except 3L, when compared with other transgenic lines. However, some original features were observed in this line at the genetic and chromosomal levels. Several hotspots of insertion sites were observed on the X, second and third chromosomes. Five sites with a high frequency of hobo insertions were present on the 3L arm in most individuals tested, suggesting the action of selection for hobo element in some sites. The presence of doublets or triplet was also observed, implying that hobo inserts can show local jumps or insertions in preferred regions. This local transposition occurred independently in 11 specific genomic regions in many individuals and generations. The dynamics of this phenomenon were analysed across generations. These results support the use of the hobo system as an important tool in fundamental and applied Drosophila genetics.