Pattern of transposable elements distribution on <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> polytene chromosomes before and after quantitative trait selection

The effect of selecting the length of a radial vein on the distribution of hybridization sites for P and hobo transposons and mdg1 and mdg2 retrotransposons on polytene chromosomes of Drosophila melanogaster salivary glands was studied. The pattern of the transposable element (TEs) distribution was polymorphic in both parental and selected strains. The similarity in mdg1 and mdg2 distribution between strains selected in one direction was closer than between strains selected in the opposite direction, but the selected strains were closer to each other than to the parental strain, regardless of direction of selection. No new mdg2 hybridization sites that would be absent from the control were found in the selected strains compared to the control. The number of mdg1 and hobo hybridization sites was more selected in strains in the (+) direction than in the (−) direction. The mobility of hobo copies in the strains correlated with the presence of its full-sized copy in the genome. The polymorphism of all TEs studied except for mdg1 was higher in strains selected in the (+) direction than in the (−) direction. These results suggest that some TEs migrate over the genome independently of selection, whereas the other are markers of evolutionary events rather than their causes.     

Fluorescence in situ hybridization analysis of hobo, mdg1 and Dm412 transposable elements reveals genomic instability following the Drosophila melanogaster genome sequencing

The genome of Drosophila melanogaster strain y cn bw sp has been sequenced and the transposable elements insertion sites have been determined. We hybridized fluorescence-labeled probes directed to the hobo transposon, Dm412 and mdg1 retrotransposons to polytene chromosomes and compared the observed sites to those published in the annotated genome sequence. We observed an almost twofold increase in the number of hobo hybridization sites (46 found as compared to 24 annotated sites). There was no evidence that the hobo transposition rate is slowing over the 10-year period. The patterns of Dm412 and mdg1 sites have changed less dramatically since the time of genome sequencing. Three novel Dm412 hybridization sites were detected while 4 out of 30 annotated sites were missing. Only one additional mdg1 site was found, while 1 out of 29 annotated sites has been lost.     

Change in distribution of mobile genetic elements in genome of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>: Cause or consequence of selection for quantitative traits?

The distribution pattern of the hobo transposon and Dm412 retrotransposon hybridization sites on the salivary gland polytene chromosomes from larvae of the Drosophila melanogaster isogenic strain 51 used to analyze the effect of the transposition of transposable elements (TEs) on selection for quantitative traits was studied. It was shown that no more than half of the Dm412 hybridization sites were retained 15 years after isogenization; the frequency of the Dm412 transposition varied from 2.0 × 10⁻⁴ to 8.8 × 10⁻⁵ sites per genome for generation depending on whether the appearance of the same hybridization sites in a part of individuals was considered as independent events or as the manifestation of the appearing sample heterogeneity. The distribution patterns of hobo hybridization sites in two isofemale strains derived from the isogenic strain 51 differed much more markedly; the number of the hobo sites in one of the derivative strains was threefold smaller than in the other one and only some of the sites were common. Within each derivative strain, the TE distribution was uniform, which suggests that inbreeding had no effect on Dm412 activity in this strain. The rates of change in the distribution patterns of various TE in the strain 51 corresponded to their spontaneous transposition rates. Since the isogenic strain accumulates polymorphism in the TE distribution without selection, the TEs are more likely to be the markers of selection events rather than their inducers. Thus, when studying the effects of various environmental factors on TE transposition even in isogenic strains, it is necessary to perform additional close inbreeding to reduce the potential polymorphism.     

Molecular genetic analysis of hobo mobile genetic element polymorphism in the genome of Drosophila melanogaster line subjected to long-term selection

The distribution of mobile genetic element hobo was examined in Drosophila melanogaster lines HA (high male mating activity) and LA (low male mating activity) before and after their isogenization using Southern blot hybridization. The probe containing a full-size hobo copy was shown to produce polymorphic multilocus hybridization with chromosomal DNA. The polymorphism was line-specific. A comparison of hybridization patterns in isogenic and original lines showed that isogenization in dysgenic crosses resulted in the appearance of additional hobo localization sites in LA but not in HA. The hobo destabilization in the LA genome correlated with genetic instability and the ability to induce H-E hybrid dysgenesis. The results obtained are discussed in relation to the possible role of hobo in inducing genetic variability in lines with low male mating activity, which may counteract deleterious consequences of inbreeding and selection in the negative direction.     

Chromosomal Distribution of Transposable Elements in Drosophila Melanogaster: Test of the Ectopic Recombination Model for Maintenance of Insertion Site Number

Data of insertion site localization and site occupancy frequency of P, hobo, I, copia, mdg1, mdg3, 412, 297, and roo transposable elements (TEs) on the polytene chromosomes of Drosophila melanogaster were extracted from the literature. We show that TE insertion site number per chromosomal division was significantly correlated with the amount of DNA. The insertion site number weighted by DNA content was not correlated with recombination rate for all TEs except hobo, for which a positive correlation was detected. No global tendency emerged in the relationship between TE site occupancy frequency, weighted by DNA content, and recombination rate; a strong negative correlation was, however, found for the 3L arm. A possible dominant deleterious effect of chromosomal rearrangements due to recombination between TE insertions is thus not the main factor explaining the dynamics of TEs, since this hypothesis implies a negative relationship between recombination rate and both TE insertion site number and site occupancy frequency. The alternative hypothesis of selection against deleterious effects of insertional mutations is discussed.     

Transposable elements and the penetrance of quantitative characters in Drosophila melanogaster

We wanted to determine whether there is a correlation between the quantitative character, the penetrance of the loss of humeral bristles in scute lines, and the distribution of transposable genetic elements in their genomes. We derived 18 isogenic lines with penetrance ranging between 2.8% and 92.0% from six mutant lines. The localization of the transposable elements (TEs) P, mdg1, Dm412, copia, gypsy and B104 was determined in all isogenic derivatives by in situ hybridization. The total number of the TE sites over all lines was 180. A comparison of the distribution of the TEs in the isogenic lines revealed the location of sites typical of lines with similar penetrance, no matter which parental line was involved. The results obtained suggest that such typical sites appear to tag the genome regions where the polygenes affecting the character in question are most likely to be found.     

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.     

Behavior of hobo and P transposons in yellow2-717 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 y2-717, 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 x 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 insertions was higher than that of excisions. One of the derivative strains, y1t-717alk3-2, was characterized by high intra-strain instability of hobo element localization. In the y2-717a1k3 and y1t-717alk3-2 strains a large inversion, In(1)1B; 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 x 10(-2) per site per X chromosome per generation.     

Wake up of transposable elements following Drosophila simulans worldwide colonization.

Transposable elements (TEs) make up around 10%-15% of the Drosophila melanogaster genome, but its sibling species Drosophila simulans carries only one third as many such repeat sequences. We do not, however, have an overall view of copy numbers of the various classes of TEs (long terminal repeat [LTR] retrotransposons, non-LTR retrotransposons, and transposons) in genomes of natural populations of both species. We analyzed 34 elements in individuals from various natural populations of these species. We show that D. melanogaster has higher average chromosomal insertion site numbers per genome than D. simulans for all TEs except five. The LTR retrotransposons gypsy, ZAM, and 1731 and the transposon bari-1 present similar low copy numbers in both species. The transposon hobo has a large number of insertion sites, with significantly more sites in D. simulans. High variation between populations in number of insertion sites of some elements of D. simulans suggests that these elements can invade the genome of the entire species starting from a local population. We propose that TEs in the D. simulans genome are being awakened and amplified as they had been a long time ago in D. melanogaster.     

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.     

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.     

Changes in the chromosomal insertion pattern of the copia element during the process of making chromosomes homozygous in Drosophila melanogaster

In situ hybridization on polytene chromosomes of Drosophila melanogaster was used to compare the insertion patterns of copia and mdgl transposable elements on chromosome 2 in male gametes sampled by two different methods: (i) by crossing the males tested with females from a highly inbred line with known copia and mdgl insertion profiles; (ii) by crossing the same males with females from a marked strain, and analysing the resulting homozygous chromosomes. Crossing of the males with the inbred line led to homogeneous insertion profiles for both the copia and mdgl elements in larvae, thus giving an accurate estimation of the patterns in the two gamete classes of each male. Crossing with the marked strain led, however, to heterogeneity in insertion patterns of the copia transposable element, while no significant polymorphism was observed for mdgl. The use of balancer chromosomes is thus not an adequate way of inferring transposable element insertion patterns of Drosophila males, at least for the copia element. This technique could, however, be powerful for investigating the control of movements of this element.     

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.     

Structural organization of transposable element mdg4 from Drosophila melanogaster and a nucleotide sequence of its long terminal repeats

A mobile dispersed genetic element, mdg4 , approximately 7.5 kilobases (kb) long has been cloned from D. melanogaster genome. Chromosomal bands have only few sites of mdg4 , but it always hybridizes to the chromocenter. The location of mdg4 varies among D. melanogaster strains. Blot hybridization shows that, in contrast to other mdg elements, mdg4 sequences are rather heterogeneous. Only few copies are full-length. A strong amplification of mdg4 has occurred during the in vitro cultivation of cells involving only one mdg4 variant. Long terminal repeats (LTRs) and flanking sequences have been sequenced in two cloned copies of transposable element mdg4 . In both cloned copies of mdg4 , LTRs have an identical nucleotide sequence 479 bp long. The mdg4 is flanked by four-base-pair direct repeats, short mismatched palindromes being present at the ends of each LTR. The termini of the mdg4 body contain an oligopurine stretch and a region partially complementary to D. melanogaster tRNA-Lys. Thus, structural organization of mdg4 LTRs is similar to that of several other mdg elements and retroviral proviruses.     

Structural heterogeneity and genomic distribution of Drosophila melanogaster LTR-retrotransposons

Structural heterogeneity of five long terminal repeat (LTR) retrotransposon families (297, mdg 1, 412, copia, and 1731) was investigated in Drosophila melanogaster. The genomic distribution of canonical and rearranged elements was studied by comparing hybridization patterns of Southern blots on salivary glands from adult females and males with in situ hybridization on polytene chromosomes. The proportion and genomic distribution of noncanonical copies is distinctive to each family and presents constant features in the four different D. melanogaster strains studied. Most elements of families 297 and mdg 1 were noncanonical and presented large interstock and intrastock polymorphism. Noncanonical elements of these two families were mostly located in euchromatin, although not restricted to it. The elements of families 412 and copia were better conserved. The proportion of noncanonical elements was lower. The 1731 family is mainly composed of noncanonical, beta-heterochromatic elements that are highly conserved among stocks. The relation of structural polymorphism to phylogeny, transpositional activity and the role of natural selection in the maintenance of transposable elements are discussed.     

Changes in the chromosomal insertion pattern of the copia element during the process of making chromosomes homozygous in Drosophila melanogaster

In situ hybridization on polytene chromosomes of Drosophila melanogaster was used to compare the insertion patterns of copia and mdgl transposable elements on chromosome 2 in male gametes sampled by two different methods: (i) by crossing the males tested with females from a highly inbred line with known copia and mdgl insertion profiles; (ii) by crossing the same males with females from a marked strain, and analysing the resulting homozygous chromosomes. Crossing of the males with the inbred line led to homogeneous insertion profiles for both the copia and mdgl elements in larvae, thus giving an accurate estimation of the patterns in the two gamete classes of each male. Crossing with the marked strain led, however, to heterogeneity in insertion patterns of the copia transposable element, while no significant polymorphism was observed for mdgl. The use of balancer chromosomes is thus not an adequate way of inferring transposable element insertion patterns of Drosophila males, at least for the copia element. This technique could, however, be powerful for investigating the control of movements of this element.     

Distribution of hobo transposable elements in natural populations of Drosophila melanogaster.

Forty-six strains derived from American and French natural populations of Drosophila melanogaster were tested for the presence and activity of hobo elements by using Southern blotting and a gonadal dysgenesis assay. The oldest available strains exhibited weak detectable hybridization to the hobo-element probe and revealed neither hobo-activity potential nor hobo-repression potential. In contrast, all recently collected strains harbored hobo sequences and revealed a strong hobo-repression potential but no strong hobo-activity potential. On the basis of restriction-enzyme analysis, old strains appear to have numerous fragments hybridizable to hobo sequences, several probably conserved at the same locations in the genome of the tested strain and others dispersed. In recently isolated strains, and unlike the situation in the published sequence of the cloned hobo108 element, a PvuII site is present in the great majority of full-sized hobo elements and their deletion derivatives. When the genetic and molecular characteristics are considered together, the available evidence is consistent with the hypothesis of a worldwide hobo-element invasion of D. melanogaster during the past 50 years. Comparison of data from the I-R and P-M systems suggests that the putative invasion followed the introduction of the I element but preceded that of the P element. This hypothesis poses the problem of the plausibility of three virtually simultaneous element invasions in this species. Such a possibility might be due to a modification of the genetic structure of American populations of D. melanogaster during the first part of the 20th century.     

Transposable Element-Induced Response to Artificial Selection in Drosophila Melanogaster: Molecular Analysis of Selection Lines

Artificial selection lines for abdominal bristle score of Drosophila melanogaster established from P-M hybrid dysgenic crosses showed increases in selection response, heritability and phenotypic variance compared to similar lines started from nondysgenic crosses. To determine whether this increased genetic variance could be due to enhanced transposition of P elements following the dysgenic cross, the cytological locations (sites) of P elements were determined by in situ hybridization for the whole genome of samples of 20 individuals from the parental P strain, 20 individuals from each of the eight dysgenic selection lines, and ten individuals from each of the eight nondysgenic selection lines. Variation among and within the selection lines and the parental P strain in P element insertion sites was exceptionally high. A total of 601 sites were identified, but there was no difference in total number of sites per line, mean number of sites per individual, mean copy number per individual, or site frequency between dysgenic and nondysgenic selection lines, or between lines selected for high and low bristle score. Transposition following nondysgenic crosses may explain additional observations of accelerated selection responses in nondysgenic selection lines. It was not possible to deduce which, if any, of the several hundred insertions in the dysgenic selection lines were responsible for their extreme bristle phenotypes.     

Autonomous transposition of gypsy mobile elements and genetic instability in Drosophila melanogaster

Summary The laboratory imitator strain (MS) of Drosophila melanogaster is characterized by an elevated frequency of spontaneous mutation (10^–3–10^–4). Mutations occur in both sexes at premeiotic stages of germ cell development. The increased mutability is a characteristic feature of MS itself, since it appears in the absence of outcrossing. Most of the mutations arising in this strain are unstable: reversions to wild type, high frequency mutation to new mutant states and replicating instability were observed. We have investigated the localization of the transposable genetic elements mdg1, 412, mdg3, gypsy (mdg4), copia and P in the X chromosomes of the MS and in the mutant lines y, ct, sbt derived from it by in situ hybridization. The P element was not found in any of these strains. The distributions of mdg1, 412, mdg3 and copia were identical in the X chromosomes of the MS and its derivatives. However, the sites of hybridization with gypsy differ in the various lines tested. In the polytene chromosomes of MS animals significant variation in location and number of copies of the gypsy element was demonstrated between different larvae; copy numbers as high as 30–40 were observed. These results suggest autonomous transposition of gypsy in the MS genome while several other mobile elements remain stable.     

On the abundance and distribution of transposable elements in the genome of Drosophila melanogaster

The abundance and distribution of transposable elements (TEs) in a representative part of the euchromatic genome of Drosophila melanogaster were studied by analyzing the sizes and locations of TEs of all known families in the genomic sequences of chromosomes 2R, X, and 4. TEs contribute to up to 2% of the sequenced DNA, which corresponds roughly to the euchromatin of these chromosomes. This estimate is lower than that previously available from in situ data and suggests that TEs accumulate in the heterochromatin more intensively than was previously thought. We have also found that TEs are not distributed at random in the chromosomes and that their abundance is more strongly associated with local recombination rates, rather than with gene density. The results are compatible with the ectopic exchange model, which proposes that selection against deleterious effects of chromosomal rearrangements is a major force opposing element spread in the genome of this species. Selection against insertional mutations also influences the observed patterns, such as an absence of insertions in coding regions. The results of the analyses are discussed in the light of recent findings on the distribution of TEs in other species.