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.     

DNA sequence requirements for hobo transposable element transposition in Drosophila melanogaster

We have conducted a structure and functional analysis of the hobo transposable element of Drosophila melanogaster. A minimum of 141 bp of the left (L) end and 65 bp of the right (R) end of the hobo were shown to contain sequences sufficient for transposition. Both ends of hobo contain multiple copies of the motifs GGGTG and GTGGC and we show that the frequency of hobo transposition increases as a function of the copy number of these motifs. The R end of hobo contains a unique 12 bp internal inverted repeat that is identical to the hobo terminal inverted repeats. We show that this internal inverted repeat suppresses transposition activity in a hobo element containing an intact L end and only 475 bp of the R end. In addition to establishing cis-sequences requirements for transposition, we analyzed trans-sequence effects of the hobo transposase. We show a hobo transposase lacking the first 49 amino acids catalyzed hobo transposition at a higher frequency than the full-length transposase suggesting that, similar to the related Ac transposase, residues at the amino end of the transposase reduce transposition. Finally, we compared target site sequences of hobo with those of the related Hermes element and found both transposons have strong preferences for the same insertion sites.     

Sequences homologous to the hobo transposable element in E strains of Drosophila melanogaster

Hobo is one of the three Drosophila melanogaster transposable elements, together with the P and I elements, that seem to have recently invaded the genome of this species. Surveys of the presence of hobo in strains from different geographical and temporal origins have shown that recently collected strains contain complete and deleted elements with high sequence similarity (H strains), but old strains lack hobo elements (E strains). Besides the canonical hobo sequences, both H and E strains show other poorly known hobo-related sequences. In the present work, we analyze the presence, cytogenetic location, and structure of some of these sequences in E strains of D. melanogaster. By in situ hybridization, we found that euchromatic hobo-related sequences were in fixed positions in all six E strains analyzed: 38C in the 2L arm; 42B and 55A in the 2R arm; 79E and 80B in the 3L arm; and 82C, 84C, and 84D in the 3R arm. Sequence comparison shows that some of the hobo-related sequences from Oregon-R and iso-1 strains are similar to the canonical hobo element, but their analysis reveals that they are substantially diverged and rearranged and cannot code for a functional transposase. Our results suggest that these ubiquitous hobo-homologous sequences are immobile and are distantly related to the modern hobo elements from D. melanogaster.     

hobo transposable elements in Drosophila melanogaster and D. simulans.

Genomic patterns of occurrence of the transposable element hobo are polymorphic in the sibling species Drosophila melanogaster and D. simulans. Most tested strains of both species have apparently complete (3.0 kb) and smaller hobo elements (H lines), but in both species some strains completely lack such canonical hobo elements (E lines). The occurrence of H and E lines in D. simulans as well as in D. melanogaster implies that an hypothesis of recent introduction in the latter species is inadequate to explain the phylogenetic occurrence of hobo. Particular internally deleted elements, the approximately 1.5 kb Th1 and Th2 elements, are abundant in many lines of D. melanogaster, and an analogous 1.1 kb internally deleted element, h del sim, is abundant in most lines of D. simulans. Besides the canonical hobo sequences, both species (and their sibling species D. sechellia and D. mauritiana) have many hobo-hybridizing sequences per genome that do not appear to be closely related to the canonical hobo sequence.     

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 the hobo transposable element with regard to TPE repeats in transgenic lines of Drosophila melanogaster

The hobo transposable element of Drosophila melanogaster is known to induce a hybrid dysgenesis syndrome. Moreover it displays a polymorphism of a microsatellite in its coding region: TPE repeats. In European populations, surveys of the distribution of hobo elements with regard to TPE repeats revealed that the 5TPE element is distributed along a frequency gradient, and it is even more frequent than the 3TPE element in Western populations. This suggests that the invasive ability of the hobo elements could be related to the number of TPE repeats they contain. To test this hypothesis we monitored the evolution of 16 lines derived from five initial independent transgenic lines bearing the 3TPE element and/or the 5TPE element. Four lines bearing 5TPE elements and four bearing 3TPE elements were used as a noncompetitive genetic background to compare the evolution of the 5TPE element to that of the 3TPE element. Eight lines bearing both elements provided a competitive genetic context to study potential interactions between these two elements. We studied genetic and molecular aspects of the first 20 generations. At the molecular level, we showed that the 5TPE element is able to spread within the genome at least as efficiently as the 3TPE element. Surprisingly, at the genetic level we found that the 5TPE element is less active than the 3TPE element, and moreover may be able to regulate the activity of the 3TPE element. Our findings suggest that the invasive potential of the 5TPE element could be due not only to its intrinsic transposition capacity but also to a regulatory potential.     

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 × 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.     

The role of the transposable element hobo in the origin of endemic inversions in wild populations of Drosophila melanogaster

Evidence from in situ hybridizations of DNA from the transposable element hobo to polytene salivary gland chromosome squashes reveals that hobo occupies both cytological breakpoints of three of four endemic inversions sampled from natural populations of Drosophila melanogaster in the Hawaiian islands. The fourth endemic inversion has a single hobo insert at one breakpoint. Cosmopolitan inversions on the same chromosomes do not show this association. Frequencies of both endemic and cosmopolitan inversions in Hawaiian populations fall in ranges typical for natural populations of D. melanogaster sampled worldwide, suggesting that these results may be typical of other regions besides Hawaii. This appears to be the first direct demonstration that transposable elements are responsible for causing specific rearrangements found in nature; consequently, it is also the first direct demonstration that chromosome rearrangements can arise in nature in a manner predicted by results of hybrid dysgenic crosses in the laboratory. Possible population genetic and evolutionary consequences are discussed.     

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.     

Fixation of transposable elements in the Drosophila melanogaster genome

We have investigated at the molecular level four cases in which D. melanogaster middle repetitive DNA probes consistently hybridized to a particular band on chromosomes sampled from a D. melanogaster natural population. Two corresponded to true fixations of a roo and a Stalker element, and the others were artefacts of the in situ hybridization technique caused by the presence of genomic DNA flanking the transposable elements (TEs) in the probes. The two fixed elements are located in the beta-heterochromatin (20A and 80B, respectively) and are embedded in large clusters of other elements, many of which may also be fixed. We also found evidence that this accumulation is an ongoing process. These results support the hypothesis that TEs accumulate in the non-recombining part of the genome. Their implications for the effects of TEs on determining the chromatin structure of the host genomes are discussed in the light of recent evidence for the role of TE-derived small interfering-RNAs as cis -acting determinants of heterochromatin formation.     

Maintenance of a large pericentric inversion generated by the hobo transposable element in a transgenic line of Drosophila melanogaster

The impact of the hobo transposable element in the global reorganization of the Drosophila melanogaster genome has been investigated in transgenic lines generated by the injection of hobo elements into the Hikone strain, which lacked them previously. Extensive surveys of transgenic lines followed for 250 generations have identified 13 inversions with hobo inserts at most breakpoints. One of these inversions is pericentric on chromosome 2. It has been maintained in the line where it was discovered and in several sublines at frequencies from 0.19 to 0.45, generating stable chromosomal polymorphisms, similar to cosmopolitan paracentric inversions in natural populations. Individuals homozygous for this inversion were viable and fertile, allowing the creation of a new homozygous strain.     

Distribution of hobo transposable elements in the genus Drosophila

This study describes the distribution of hobo-hybridizing sequences in the genus Drosophila. Southern blot analysis of 134 species revealed that hobo sequences are limited to the melanogaster and montium subgroups of the melanogaster-species group. Of the hobo-bearing species, only D. melanogaster and two of its sibling species, D. simulans and D. mauritiana, were found to contain potentially complete hobo elements. The distribution of hobo sequences is one of the narrowest distributions thus far described for any Drosophila transposable element.     

Recurrent insertion and duplication generate networks of transposable element sequences in the Drosophila melanogaster genome

Background  

The recent availability of genome sequences has provided unparalleled insights into the broad-scale patterns of transposable element (TE) sequences in eukaryotic genomes. Nevertheless, the difficulties that TEs pose for genome assembly and annotation have prevented detailed, quantitative inferences about the contribution of TEs to genomes sequences.     

Transposition of the vertebrate Tol2 transposable element in Drosophila melanogaster

The Tol2 element is a transposon found from a genome of a vertebrate, a small teleost medaka fish. Tol2 encodes a gene for a transposase which is active in vertebrate animals so far tested; for instance, in fish, frog, chicken and mammals, and transgenesis methods using Tol2 have been developed in these model vertebrates. However, it has not been known whether Tol2 can transpose in animals other than vertebrates. Here we report transposition of Tol2 in an invertebrate Drosophila melanogaster. First, we injected a transposon donor plasmid containing a Tol2 construct and mRNA encoding the Tol2 transposase into Drosophila eggs, and found that the Tol2 construct could be excised from the plasmid. Second, we crossed the injected flies, raised the offspring, and found that the Tol2 construct was integrated into the genome of germ cells and transmitted to the next generation. Finally, we constructed a Tol2 construct containing the white gene and injected the transposon donor plasmid and the transposase mRNA into fertilized eggs from the white mutant. We analyzed their offspring, and found that G1 flies with wild type red eyes could be obtained from 35% of the injected fly. We cloned and sequenced 34 integration loci from these lines and showed that these insertions were indeed created through transposition and distributed throughout the genome. Our present study demonstrates that the medaka fish Tol2 transposable element does not require vertebrate-specific host factors for its transposition, and also provides a possibility that Tol2 may be used as a new genetic tool for transgenesis and genome analysis in Drosophila.     

The pogo transposable element family of Drosophila melanogaster.

Summary A 190 by insertion is associated with the white-eosin mutation in Drosophila melanogaster. This insertion is a member of a family of transposable elements, pogo elements, which is of the same class as the P and hobo elements of D. melanogaster. Strains typically have many copies of a 190 by element, 10–15 elements 1.1–1.5 kb in size and several copies of a 2.1 kb element. The smaller elements all appear to be derived from the largest by single internal deletions so that all elements share terminal sequences. They either always insert at the dinucleotide TA and have perfect 21 bp terminal inverse repeats, or have 22 by inverse repeats and produce no duplication upon insertion. Analysis by DNA blotting of their distribution and occupancy of insertion sites in different strains suggests that they may be less mobile than P or hobo. The DNA sequence of the largest element has two long open reading frames on one strand which are joined by splicing as indicated by cDNA analysis. RNAs of this strand are made, whose sizes are similar to the major size classes of elements. A protein predicted by the DNA sequence has significant homology with a human centrosomal-associated protein, CENP-B. Homologous sequences were not detected in other Drosophila species, suggesting that this transposable element family may be restricted to D. melanogaster.     

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.     

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%.