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Carbon source dependence of transposable element-associated gene activation in Saccharomyces cerevisiae.

Seven cis-dominant mutations leading to the overproduction of the glucose-repressible alcohol dehydrogenase isozyme ADHII (structural gene, ADH2) in Saccharomyces cerevisiae have previously been shown to be due to insertion of a transposable element, Ty, in the 5' regulatory region of the ADH2 gene. We showed that although mating-competent cells (a, alpha, a/a, or alpha/alpha cells) overproduced both ADHII enzyme and ADH2 mRNA, mating-incompetent cells (a/alpha or ste-cells) produced much less ADHII enzyme and ADH2 mRNA. This mating type effect on ADH2 expression was greatest in the presence of a normally derepressing carbon source, glycerol, and much less apparent in the presence of a repressing carbon source, glucose. In addition, Ty insertion led to an aberrant carbon source response in mating-incompetent cells--the normally glucose-repressible ADHII becomes glycerol repressible. The mating type effect and aberrant carbon source response in mating-incompetent cells was specific for Ty-associated mutations in the 5' flanking region of the ADH2 gene in that a non-Ty mutation in the same region did not show these effects. Finally, Ty1 RNA levels also showed a/alpha, suppression, which was apparent only during growth on a nonfermentable carbon source such as glycerol. This suggests that Ty-mediated gene expression is subject to regulation by both mating competence and carbon catabolites.

Alcohol Dehydrogenase↗

Mechanisms regulating the copy numbers of six LTR retrotransposons in the genome of Drosophila melanogaster.

There has been debate over the mechanisms that control the copy number of transposable elements in the genome of Drosophila melanogaster. Target sites in D. melanogaster populations are occupied at low frequencies, suggesting that there is some form of selection acting against transposable elements. Three main theories have been proposed to explain how selection acts against transposable elements: insertions of a copy of a transposable element are selected against; chromosomal rearrangements caused by ectopic exchange between element copies are selected against; or the process of transposition itself is selected against. The three theories give different predictions for the pattern of transposable element insertions in the chromosomes of D. melanogaster. We analysed the abundance of six LTR (long terminal repeat) retrotransposons on the X and fourth chromosomes of multiple strains of D. melanogaster, which we compare with the predictions of each theory. The data suggest that no one theory can account for the insertion patterns of all six retrotransposons. Comparing our results with earlier work using these transposable element families, we find a significant correlation between studies in the particular model of copy number regulation supported by the proportion of elements on the X for the different transposable element families. This suggests that different retrotransposon families are regulated by different mechanisms.

Animals↗

Transposable DNA elements and life history traits: II. Transposition of P DNA elements in somatic cells reduces fitness, mating activity, and locomotion of Drosophila melanogaster.

Some transposable DNA elements in higher organisms are active in somatic cells, as well as in germinal cells. What effect does the movement of DNA elements in somatic cells have on life history traits? It has previously been reported that somatically active P and mariner elements in Drosophila induce genetic damage and significantly reduce lifespan. In this study, we report that the movement of P elements in somatic cells also significantly reduces fitness, mating activity, and locomotion of Drosophila melanogaster. If other elements cause similar changes in life history traits, it is doubtful if transposable DNA elements remain active for long in somatic cells in natural populations.

Animals↗

Complete sequence of the IncP-9 TOL plasmid pWW0 from Pseudomonas putida.

The TOL plasmid pWW0 (117 kb) is the best studied catabolic plasmid and the archetype of the IncP-9 plasmid incompatibility group from Pseudomonas. It carries the degradative (xyl) genes for toluenes and xylenes within catabolic transposons Tn4651 and Tn4653. Analysis of the complete pWW0 nucleotide sequence revealed 148 putative open reading frames. Of these, 77 showed similarity to published sequences in the available databases predicting functions for: plasmid replication, stable maintenance and transfer; phenotypic determinants; gene regulation and expression; and transposition. All identifiable transposition functions lay within the boundaries of the 70 kb transposon Tn4653, leaving a 46 kb sector containing all the IncP-9 core functions. The replicon and stable inheritance region was very similar to the mini-replicon from IncP-9 antibiotic resistance plasmid pM3, with their Rep proteins forming a novel group of initiation proteins. pWW0 transfer functions exist as two blocks encoding putative DNA processing and mating pair formation genes, with organizational and sequence similarity to IncW plasmids. In addition to the known Tn4651 and IS1246 elements, two additional transposable elements were identified as well as several putative transposition functions, which are probably genetic remnants from previous transposition events. Genes likely to be responsible for known resistance to ultraviolet light and free radicals were identified. Other putative phenotypic functions identified included resistance to mercury and other metal ions, as well as to quaternary ammonium compounds. The complexity and size of pWW0 is largely the result of the mosaic organization of the transposable elements that it carries, rather than the backbone functions of IncP-9 plasmids.

Bacterial Proteins↗

Germ line and somatic instability of a white mutation in Drosophila mauritiana due to a transposable genetic element.

A spontaneous white mutation recovered in Drosophila mauritiana is unstable and reverts to normal eye color at a frequency greater than 4 per 1,000 X-chromosomes. Germ line reversion occurs at a high rate in D. mauritiana males and in interspecific hybrid females, while the rate is depressed in D. mauritiana females. These events are not restricted to the germ line, as cases of variegated patterns of eye pigmentation, indicating somatic reversion, are recovered at a frequency comparable to that of the male germ line reversion rate. Germ line reversion events are genetically stable, while the somatic variegation patterns are not heritable. The patterns of eye pigment variegation produced suggests that reversion events are occurring throughout development. Whole genome DNA digests blotted and probed with the cloned D. melanogaster white gene indicate that this unstable white mutation in D. mauritiana is associated with an insertion of DNA that is lost upon reversion to wild type, indicating that this DNA insert is in fact a transposable element.

Animals↗

The role of transposable elements-endogenous retroviruses in embryonic development and regeneration.

Endogenous retroviruses (ERVs) are dynamically regulated across the lifespan and can function as context-dependent components of host gene-regulatory networks. During embryonic development, selected ERV-derived elements are co-opted to support zygotic genome activation, lineage specification, and placental development. In adult tissues, ERV-derived sequences can contribute to tissue and immune homeostasis, whereas potentially disruptive ERV activity is constrained by epigenetic mechanisms. During regeneration and somatic cell reprogramming, ERV and broader transposable-element programs undergo transient, locus-specific remodeling. In aging, the weakening of epigenetic and nuclear restraint can promote aberrant ERV derepression, inflammation, and functional decline. This review summarizes the diverse roles of ERVs across these contexts and discusses the challenges of defining locus-specific functions, resolving repetitive sequences, and developing safe ERV-targeted interventions.

Endogenous Retroviruses↗

A new resource of locally transposed Dissociation elements for screening gene-knockout lines in silico on the Arabidopsis genome.

We transposed Dissociation (Ds) elements from three start loci on chromosome 5 in Arabidopsis (Nossen ecotype) by using a local transposition system. We determined partial genomic sequences flanking the Ds elements and mapped the elements' insertion sites in 1,173 transposed lines by comparison with the published genomic sequence. Most of the lines contained a single copy of the Ds element. One-half of the lines contained Ds on chromosome 5; in particular, insertion "hot spots" near the three start loci were clearly observed. In the other lines, the Ds elements were transposed across chromosomes. We found other insertion hot spots at the tops of chromosomes 2 and 4, near nucleolus organizer regions 2 and 4, respectively. Another characteristic feature was that the Ds elements tended to transpose near the chromosome ends and rarely transposed near centromeres. The distribution patterns differed among the three start loci, even though they possessed the same Ds construct. More than one-half of the Ds elements were inserted irregularly into the genome; that is, they did not retain the perfect inverted repeat sequence of Ds nor leave perfect target site duplications. This precise analysis of distribution patterns will contribute to a comprehensive understanding of the transposing mechanism. From these Ds insertion sites, we have constructed a database for screening gene-knockout mutants in silico. In 583 of the 1,173 lines, the Ds elements were inserted into protein-coding genes, which suggests that these lines are gene-knockout mutants. The database and individual lines will be available freely for academic use from the RIKEN Bio-Resource Center (http://www.brc.riken.go.jp/Eng/index.html).

Arabidopsis↗

Transformation systems in insects.

Genetic transformation is an important technology that provides unique opportunities to find, isolate, and analyze genes, as well as to create organisms with unique functional characteristics. Insect biologists have been developing genetic transformation technologies that rely extensively on transposable elements. A number of class II transposable elements isolated originally from insects have been converted into broad host range insect gene vectors. Class II transposable elements are particularly amenable to gene vector development, although they suffer from some limitations such as low rates of recombination. Use of these gene vectors requires the physical introduction of the vectors into developing insect embryos by microinjection. Microinjection methods vary to accommodate the unique physical and developmental characteristics of the target insects. All methods rely on the use of fine glass needles in conjunction with micromanipulators and a microscope. A serious constraint on the use of existing systems can be the inefficiency of successfully delivering the gene vectors to the germ cells of the developing embryo. The general method for vector delivery to insect germ cells is described, as well as variations that are useful under some conditions.

Animals↗

hAT element population genetics in Anopheles gambiae s.l. in Mozambique.

Herves is a functional Class II transposable element in Anopheles gambiae belonging to the hAT superfamily of elements. Class II transposable elements are used as gene vectors in this species and are also being considered as genetic drive agents for spreading desirable genes through natural populations as part of an effort to control malaria transmission. In this study, Herves was investigated in populations of Anopheles gambiae s.s., Anopheles arabiensis and Anopheles merus in Mozambique over a period of 2 years. The copy number of Herves within these three species was approximately 5 copies per diploid genome and did not differ among species or between years. Based on the insertion-site occupancy-frequency distribution and existing models of transposable element dynamics, Herves appears to be transpositionally active currently or, at least recently, in all species tested. Ninety-five percent of the individuals within the populations of the three species tested contained intact elements with complete Herves transposase genes and this is consistent with the idea that these elements are currently active.

Animals↗

Structural analysis of 5'-flanking regions of rat, mouse and human renin genes reveals the presence of a transposable-like element in the two mouse genes.

The two renin genes of the mouse (Ren1 and Ren2) are expressed at different levels in the submaxillary gland (SMG). In contrast to mice, there is no detectable renin gene expression in the rat SMG. To determine the molecular basis for these different levels of renin expression, we have compared the 5'-flanking regions of the rat and mouse genes. The sequence of mouse, but not rat, genes reveals the presence in Ren1 and Ren2 of a large insertion, probably a new class of transposable elements. A second, apparently unrelated shorter insertion is present only in Ren2. Otherwise, the mouse and rat 5'-flanking sequences are well conserved and resemble the corresponding region of the human Ren gene, indicating that the insertions occurred after the separation of the rat and mouse species but before the duplication of the mouse Ren gene. We suggest that these structural differences may have a role in the differential expression of the Ren genes in mice and other animals.

Animals↗

Sequence similarity of putative transposases links the maize Mutator autonomous element and a group of bacterial insertion sequences.

The Mutator transposable element system of maize is the most active transposable element system characterized in higher plants. While Mutator has been used to generate and tag thousands of new maize mutants, the mechanism and regulation of its transposition are poorly understood. The Mutator autonomous element, MuDR, encodes two proteins: MURA and MURB. We have detected an amino acid sequence motif shared by MURA and the putative transposases of a group of bacterial insertion sequences. Based on this similarity we believe that MURA is the transposase of the Mutator system. In addition we have detected two rice cDNAs in genbank with extensive similarity to MURA. This sequence similarity suggests that a Mutator-like element is present in rice. We believe that Mutator, a group of bacterial insertion sequences, and an uncharacterized rice transposon represent members of a family of transposable elements.

Amino Acid Sequence↗

Interplay between the role of DNA methylation in regulating gene expression and TE-silencing in a reptilian methylome.

DNA methylation is a major component of eukaryotic genomes with an important role in the defence against transposable elements, to transcriptionally silence their activity and prevent transposition. DNA methylation also plays a major role in the regulation of gene expression. This dual role can come into conflict, where DNA methylation in gene regulatory regions becomes perturbed due to transposable element transposition, leading to disruption of gene expression. Here, we describe how this conflict is reflected in DNA methylation patterns in the sand lizard genome where there is recent transposable element activity. Using long-read sequencing technology we show that CpG islands in gene transcriptional start sites are typically hypomethylated and associated with higher gene expression. Outside transcriptional start sites, a majority of CpG islands overlapped transposable elements and were associated with hypermethylation, consistent with a host-defence role in suppressing transposition activity. We identify 605 instances where transcriptional start sites were associated with transposable elements (4.3% of all genes). These instances were far rarer in conjunction with a CpG island, when methylation signatures would be in conflict. Transposable elements were found to be closer to and at higher density the more hypermethylated a transcriptional start site was, suggesting strong selection against selfish genetic elements transposing into hypomethylated transcriptional start sites.

CpG islands↗

The Hermes element from Musca domestica can transpose in four families of cyclorrhaphan flies.

Transgenic insect technology will provide opportunities to explore the basic biology of a broad range of insect species in ways that will prove insightful and important. It is also a technology that will provide opportunities to manipulate the genotypes of insects of practical significance to the health and welfare of humans. The Hermes transposable element from the housefly, Musca domestica, is a short inverted repeat-type element related to hobo from Drosophila melanogaster, Ac from Zea mays, and Tam3 from Antirrhinum majus. It has potential to become a versatile and efficient broad host-range insect transformation vector. The ability of Hermes to transpose when introduced into five species of diptera from four divergent families was tested using an in vivo, interplasmid transpositional recombination assay. Hermes was capable of transposing in all species tested, demonstrating that Hermes has a broad host-range. In addition, the rates of transposition were sufficiently high in all species tested to suggest that Hermes will be an efficient gene transfer vector in a wide range of insect species. The Hermes element also revealed a pattern of integration into the target substrate that permitted factors determining integration site selection to be identified. Primary nucleotide sequence of the integration site played a role as did proximity to preferred integration sites and the nucleosomal organization of the target.

Animals↗

A comparative study of Tam3 and Ac transposition in transgenic tobacco and petunia plants.

Transposition of the Anthirrinum majus Tam3 element and the Zea mays Ac element has been monitored in petunia and tobacco plants. Plant vectors were constructed with the transposable elements cloned into the leader sequence of a marker gene. Agrobacterium tumefaciens-mediated leaf disc transformation was used to introduce the transposable element constructs into plant cells. In transgenic plants, excision of the transposable element restores gene expression and results in a clearly distinguishable phenotype. Based on restored expression of the hygromycin phosphotransferase II (HPTII) gene, we established that Tam3 excises in 30% of the transformed petunia plants and in 60% of the transformed tobacco plants. Ac excises from the HPTII gene with comparable frequencies (30%) in both plant species. When the beta-glucuronidase (GUS) gene was used to detect transposition of Tam3, a significantly lower excision frequency (13%) was found in both plant species. It could be shown that deletion of parts of the transposable elements Tam3 and Ac, removing either one of the terminal inverted repeats (TIR) or part of the presumptive transposase coding region, abolished the excision from the marker genes. This demonstrates that excision of the transposable element Tam3 in heterologous plant species, as documented for the autonomous element Ac, also depends on both properties. Southern blot hybridization shows the expected excision pattern and the reintegration of Tam3 and Ac elements into the genome of tobacco plants.

Agrobacterium tumefaciens↗

FB elements can promote exon shuffling: a promoter-less white allele can be reactivated by FB mediated transposition in Drosophila melanogaster.

Foldback ( FB) elements are transposable elements found in many eukaryotic genomes; they are thought to contribute significantly to genome plasticity. In Drosophila melanogaster, FBs have been shown to be involved in the transposition of large chromosomal regions and in the genetic instability of some alleles of the white gene. In this report we show that FB mediated transposition of w(67C23), a mutation that deletes the promoter of the white gene and its first exon, containing the start codon, can restore expression of the white gene. We have characterized three independent events in which a 14-kb fragment from the w(67C23) locus was transposed into an intron region in three different genes. In each case a local promoter drives the expression of white, producing a chimeric mRNA. These findings suggest that, on an evolutionary timescale, FB elements may contribute to the creation of new genes via exon shuffling.

ATP-Binding Cassette Transporters↗

Biogeographic analysis of the Uhu and LOA elements in the Hawaiian Drosophila.

Two transposable elements have been isolated from the Hawaiian Drosophila, the Uhu and LOA elements. The Uhu element has been shown to be present in a group of closely related species, the planitibia subgroup of the picture-winged Drosophila. This study examines the distribution of the Uhu element in several other subgroups of the picture-winged Drosophila, as well as the modified-mouthparts and antopocerus groups of the Hawaiian Drosophila. The LOA element has a much more limited distribution, having only been found in representatives of the planitibia and grimshawi subgroups of the picture-winged Drosophila. For both the Uhu and LOA elements there is an inverse correlation between the copy number of the element and the age of the island on which the species is endemic, i.e., species endemic to the Island of Hawaii, the youngest island, have the highest copy number, while species endemic to Kauai, the oldest island, have the lowest copy number of the element. The correlation suggests there is a relationship between speciation and the activity of transposable elements.

Animals↗

Gene transfer into corn earworm (Helicoverpa zea) embryos.

Transposable elements with short inverted repeats at their termini have been identified in a number of diverse insect species and have proven to be useful gene delivery vectors for the transformation of Drosophila melanogaster. In this report we examine the ability of the D. melanogaster hobo element to transpose in lepidopteran species. A Trichoplusia ni (cabbage looper) and a Helicoverpa zea (corn earworm) embryonic cell line were found to be capable of supporting productive transposition of the hobo element as measured by a plasmid-based excision assay. Furthermore, hobo transposition was detected in H. zea embryos in a manner consistent with that seen for the cell line. In both cases, transposition/excision was found to be independent of vector-encoded transposase functions, indicating that endogenous genes are involved in hobo mobility. Finally, we demonstrate the stable insertion of the bacterial lacZ gene into the H. zea genome. These data demonstrate that hobo elements are capable of transgressing species boundaries and functioning in non-drosophilid cellular environments. More importantly, this represents the first description of a genetic transformation system for a lepidopteran species.

Animals↗

Gene silencing in plants: relevance for genome evolution and the acquisition of genomic methylation patterns.

Transgenes often become silenced in plants because of repressive influences exerted by flanking plant DNA and/or because of interactions among multiple copies of closely linked transgenes. Repeated transgenes on different chromosomes can also interact in a way that leads to silencing and methylation, suggesting a previously unrecognized ability of unlinked homologous sequences to cross-talk in complex genomes. Non-Mendelian inheritance is a frequent consequence of these interactions because the silenced genes do not fully reactivate or lose methylation after segregating in progeny. Several examples of gene silencing in plants appear to reflect the action of genome defence system that methylates and inactivates foreign or invasive sequences such as transgenes and transposable elements. Because certain types of transposable elements are embedded in regulatory regions of plant genes and have become greatly amplified in plant genomes, they could contribute substantially to normal gene expression and to the generation of genomic methylation patterns. Polyploidy, which has been a major force in plant and vertebrate evolution, might encourage proliferation of transposable elements because genes in polyploids are duplicated and hence less susceptible to the consequences of insertional mutagenesis. Accordingly, the appearance of genome-wide methylation has often coincided with episodes of polyploidization.

DNA Methylation↗