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PubMed1996

Adjacent sequences influence DNA repair accompanying transposon excision in maize.

Mobile elements transposing via DNA intermediates often leave small rearrangements, or "transposon footprints," at sites where they excise. Each excision event leaves its own footprint and, at any given site, these vary in size and sequence. Footprint formation involves DNA repair of sequences flanking the element. We have analyzed the footprints formed by a 2-kb Ds element excising from six different sites in exons of the maize waxy (Wx) gene. We find that groups of footprints left at individual sites are surprisingly nonrandom; different excision products predominate consistently at each site. Less frequent footprints left by each insertion appear related to the predominant type. The data suggest that flanking sequences affect the DNA repair processes associated with element excision. Two models have been proposed to explain footprint formation, one featuring a 5' exonuclease and the other featuring hairpin loop formation and an endonuclease. Our data have interesting implications for both these models. Evidence is also presented to support the presence of a separate excision mechanism that can remove Ac/Ds elements without leaving any footprint and that operates in parallel with the footprint-forming mechanism.

Alleles↗
PubMed2001

The complex repeats of Dictyostelium discoideum.

In the course of determining the sequence of the Dictyostelium discoideum genome we have characterized in detail the quantity and nature of interspersed repetitive elements present in this species. Several of the most abundant small complex repeats and transposons (DIRS-1; TRE3-A,B; TRE5-A; skipper; Tdd-4; H3R) have been described previously. In our analysis we have identified additional elements. Thus, we can now present a complete list of complex repetitive elements in D. discoideum. All elements add up to 10% of the genome. Some of the newly described elements belong to established classes (TRE3-C, D; TRE5-B,C; DGLT-A,P; Tdd-5). However, we have also defined two new classes of DNA transposable elements (DDT and thug) that have not been described thus far. Based on the nucleotide amount, we calculated the least copy number in each family. These vary between <10 up to >200 copies. Unique sequences adjacent to the element ends and truncation points in elements gave a measure for the fragmentation of the elements. Furthermore, we describe the diversity of single elements with regard to polymorphisms and conserved structures. All elements show insertion preference into loci in which other elements of the same family reside. The analysis of the complex repeats is a valuable data resource for the ongoing assembly of whole D. discoideum chromosomes.

Animals↗
PubMedDate not supplied

Transposable elements controlling genetic instabilities in mammals.

It is proposed that the instabilities in gene action of some alleles at certain loci in the mouse (e.g., a, c, H-2 Mi, p, pe, T, Va, W), which do not seem to conform to traditional hypotheses of gene action, are better interpretable in the light of modern studies of transposable DNA elements (insertion sequences and transposons of prokaryotic organisms; controlling elements of maize; transposable controlling elements of Drosophila). Some phenotypic evidence in the mouse in support of this hypothesis is presented for the a, Mi, p, and W loci, which affect pigmentation.

Animals↗
PubMed1985

Tn4400, a compound transposon isolated from Bacteroides fragilis, functions in Escherichia coli.

Transfer factor pBFTM10, isolated from the obligate anaerobic bacterium Bacteroides fragilis, carries a clindamycin resistance determinant which we have suggested is part of a transposable element. DNA homologous to this determinant is found in many Clnr Bacteroides isolates, either in the chromosome or on plasmids. We have now established that Ccr resides on a transposon, Tn4400. In addition to the Ccr determinant that functions under anaerobic conditions in B. fragilis, Tn4400 also carries a determinant for tetracycline resistance (Tcr) which only functions in Escherichia coli under aerobic conditions. The presence of Tn4400 on pBFTM10 does not confer tetracycline resistance on B. fragilis cells containing it. DNA from pBFTM10 was cloned in E. coli, with pDG5 as the cloning vector, to form pGAT500. Using a mobilization assay involving pGAT500 and an F factor derivative, pOX38, we determined that a 5.6-kilobase region of pBFTM10 DNA was capable of mediating replicon fusion and transposition. Most of the mobilization products resulted from inverse transposition reactions, while some were the result of true cointegrate formation. Analysis of the cointegrate molecules showed that three were formed by the action of one of the ends of Tn4400 (IS4400), and one was formed by the action of the whole element (Tn4400). The cointegrate molecule carrying intact copies of Tn4400 at the junction of the two plasmids could resolve to yield an unaltered donor plasmid (pGAT500) and a conjugal plasmid containing a copy of Tn4400 or a copy of one insertion sequence element (pOX38::Tn4400 or pOX38::IS4400). Thus, Tn4400 is a compound transposon containing active insertion sequence elements as directly repeated sequences at its ends.

Bacteroides fragilis↗
PubMed1987

Rhizobium meliloti insertion element ISRm2 and its use for identification of the fixX gene.

Two of the three plasmids of the wild-type Rhizobium meliloti 41 (pRme41a and pRme41c) carry a copy of ISRm2, a 2.7-kilobase-long transposable element. ISRm2 is terminated by 22-base-pair (bp) inverted repeat sequences, exhibiting some homology to the inverted repeats of elements generating 9-bp target sequence duplication. Transposition of ISRm2 results in a duplication of 8 bp in length, rather rare among transposable elements. DNA sequences homologous to an internal fragment of ISRm2 were found in several Rhizobium species. Transposition of ISRm2 into fixation and nodulation genes located on the symbiotic plasmid pRme41b was detected at a high frequency. Exact locations of two copies of ISRm2 which transposed into the nod-nif region on the megaplasmid were determined. In one case, integration into the protein-coding region of the hsnD gene that determines a host specificity function of nodulation occurred. In the other mutant, ISRm2 was localized upstream of nifA, where a short open reading frame coding for a new fix gene (fixX) was identified. The product of fixX is a ferredoxin carrying a characteristic cluster of cysteine residues. On the basis of the observation that the arrangement of the ISRm2 copies is identical in the free-living wild-type cells and in nitrogen-fixing nodules, we concluded that the involvement of ISRm2 transposition in the development of nitrogen-fixing symbiosis is unlikely.

Cloning, Molecular↗
PubMedDate not supplied

Germinal reversion of the merle allele in Australian shepherd dogs.

A homozygous merle Australian shepherd bitch produced two nonmerle offspring and 64 that carried the merle allele. The two nonmerle offspring produced no merles when bred to nonmerles, indicating that the germinal reversion to nonmerle was stable. This occurrence is further evidence that the merle allele is due to a transposable DNA element.

Alleles↗
PubMed1985

Extrachromosomal DNA forms of copia-like transposable elements, F elements and middle repetitive DNA sequences in Drosophila melanogaster. Variation in cultured cells and embryos.

Drosophila melanogaster embryos and cells in culture were screened for the presence of unintegrated covalently closed circular DNA forms that hybridize to copia-like transposable elements, the F element and uncharacterized dispersed middle repetitive DNA elements. Our results indicate that the majority of copia-like elements (including copia, 297, 412, mdg1, mdg3 and gypsy), the F elements, and 9 of 12 middle repetitive DNA elements are present as free DNA forms in cultured cells and embryos. An 18 base-pair inverted repeat has been reported to flank the long direct repeat of mdg3, implying that mdg3 is not an orthodox copia-like element; however, we have sequenced two independently isolated mdg3 clones and shown that the inverted repeat is not part of the element. The relative abundance with which free DNA forms are found varies between the cultured cells used, and between cultured cells and embryos. This variation, which can be up to 20-fold for some elements, does not correlate well with either the amount of element-specific poly(A)+ RNA present per cell or the number of element-specific sequences integrated in the genome.

Animals↗
PubMed1999

The genetics and evolution of the mariner transposable element in Drosophila simulans: worldwide distribution and experimental population dynamics.

We have studied both the frequency and biogeographical distribution of the transposable DNA element mariner in natural populations of Drosophila simulans and the short-term evolutionary characteristics of mariner in experimental populations. The mariner element has been identified in natural populations of D. simulans from Africa, Europe, the Middle East, Japan, Australia, several Pacific islands, North America, and South America. Only four lines out of 296 were devoid of active mariner elements, as measured by the presence of functional mariner transposase. A slight correlation was found between the latitudinal coordinate of the collection sites and the level of mariner activity in the populations; this correlation became highly significant in Australia where a cline in mariner activity was observed along the eastern coast of the continent. We also observed that wild-type laboratory strains kept for several years as small populations might lose mariner activity over time. Using experimental populations, we modeled what might happen when naturally occurring populations exhibiting high and low levels of mariner activity encounter one another. We found that active mariner elements either will tend to lose their activity over time and gradually become inactive or possibly will be lost from the population; in either case, this will lead to the pattern seen in this experiment of a significant loss of mariner activity over time.

Animals↗
PubMed2002

Presence of a characteristic D-D-E motif in IS1 transposase.

Transposases encoded by various transposable DNA elements and retroviral integrases belong to a family of proteins with three conserved acidic amino acids, D, D, and E, constituting the D-D-E motif that represents the active center of the proteins. IS1, one of the smallest transposable elements in bacteria, encodes a transposase which has been thought not to belong to the family of proteins with the D-D-E motif. In this study, we found several IS1 family elements that were widely distributed not only in eubacteria but also in archaebacteria. The alignment of the transposase amino acid sequences from these IS1 family elements showed that out of 14 acidic amino acids present in IS1 transposase, three (D, D, and E) were conserved in corresponding positions in the transposases encoded by all the elements. Comparison of the IS1 transposase with other proteins with the D-D-E motif revealed that the polypeptide segments surrounding each of the three acidic amino acids were similar. Furthermore, the deduced secondary structures of the transposases encoded by IS1 family elements were similar to one another and to those of proteins with the D-D-E motif. These results strongly suggest that IS1 transposase has the D-D-E motif and thus belongs to the family of proteins with the D-D-E motif. In fact, mutant IS1 transposases with an amino acid substitution for each of the three acidic amino acids possibly constituting the D-D-E motif were not able to promote transposition of IS1, supporting this hypothesis. The D-D-E motif identified in IS1 transposase differs from those in the other proteins in that the polypeptide segment between the second D and third E in IS1 transposase is the shortest, 24 amino acids in length. Because of this difference, the presence of the D-D-E motif in IS1 transposase has not been discovered for some time.

Amino Acid Motifs↗
PubMed1985

Insertion of an unstable element in an intervening sequence of maize Adh1 affects transcription but not processing.

Three independently isolated and unstable mutants of the maize alcohol dehydrogenase 1 gene (Adh1) have arisen by insertion of the Mu transposing element into the first intervening sequence of the progenitor Adh1 allele. The mutants have been selected for their decreased levels of alcohol dehydrogenase 1 activity. The original mutants were unstable, giving rise to both revertant alleles and altered mutant alleles. From one of the original mutants, two derivative mutants have been recovered and described. We analyzed the effect of Mu insertion in all five of these mutants by measuring relative levels of run-off transcripts from the progenitor and mutant alleles and by comparing levels of run-off transcripts corresponding to regions lying 5' and 3' to the insertion sites. In this paper we present evidence that early transcriptional events are affected, but that, in spite of the inclusion of a 1.4-kilobase transposing DNA element, processing of transcripts occurs normally.

Alcohol Oxidoreductases↗
PubMed1997

Transposable element-host interactions: regulation of insertion and excision.

Transposable elements propagate by inserting into new locations in the genomes of the hosts they inhabit. Their transposition might thus negatively affect the fitness of the host, suggesting the requirement for a tight control in the regulation of transposable element mobilization. The nature of this control depends on the structure of the transposable element. DNA elements encode a transposase that is necessary, and in most cases sufficient, for mobilization. In general, regulation of these elements depends on intrinsic factors with little direct input from the host. Retrotransposons require an RNA intermediate for transposition, and their frequency of mobilization is controlled at multiple steps by the host genome by regulating both their expression levels and their insertional specificity. As a result, a symbiotic relationship has developed between transposable elements and their host. Examples are now emerging showing that transposons can contribute significantly to the well being of the organisms they populate.

Animals↗
PubMed1993

The ebb and flow of a fungal genome.

Transposable DNA elements have only recently been described in a few species of filamentous fungi, but may be more abundant than previously believed. Several different elements have been isolated from the rice blast pathogen Magnaporthe grisea. The distribution and amplification of these elements suggest a potential role in the evolution of the fungal genome.

DNA Transposable Elements↗
PubMed2003

Heat shock treatment increases the frequency of loss of an erythromycin resistance-encoding transposable element from the chromosome of Lactobacillus crispatus CHCC3692.

A 3,165-bp chromosomally integrated transposon, designatedTn3692, of the gram-positive strain Lactobacillus crispatus CHCC3692 contains an erm(B) gene conferring resistance to erythromycin at concentrations of up to 250 micrograms/ml. Loss of this resistance can occur spontaneously, but the rate is substantially increased by heat shock treatment. Heat shock treatment at 60 degrees C resulted in an almost 40-fold increase in the frequency of erythromycin-sensitive cells (erythromycin MIC, 0.047 micrograms/ml). The phenotypic change was followed by a dramatic increase in transcription of the transposase gene and the concomitant loss of an approximately 2-kb DNA fragment carrying the erm(B) gene from the 3,165-bp erm transposon. In cells that were not subjected to heat shock, transcription of the transposase gene was not detectable. The upstream sequence of the transposase gene did not show any homology to known heat shock promoters in the gene data bank. Significant homology (>99%) was observed between the erythromycin resistance-encoding gene from L. crispatus CHCC3692 and the erm(B) genes from other gram-positive bacteria, such as Streptococcus agalactiae, Streptococcus pyogenes, Enterococcus faecium, and Lactobacillus reuteri, which strongly indicates a common origin of the erm(B) gene for these species. The transposed DNA element was not translocated to other parts of the genome of CHCC3692, as determining by Southern blotting, PCR analysis, and DNA sequencing. No other major aberrations were observed, as judged by colony morphology, growth performance of the strain, and pulsed-field gel electrophoresis. These observations suggest that heat shock treatment could be used as a tool for the removal of unwanted antibiotic resistance genes harbored in transposons flanked by insertion sequence elements or transposases in lactic acid bacteria used for animal and human food production.

Animals↗
PubMed1997

Analysis of the cis-acting DNA elements required for piggyBac transposable element excision.

The terminal DNA sequence requirements for piggyBac transposable element excision were explored using a plasmid-based assay in transfected, cultured insect cells. A donor plasmid containing duplicate 3' piggyBac terminal inverted repeats was constructed that allowed individual nucleotides or groups of nucleotides within one of the 3' repeats to be mutated. The relative extent of excision using the mutated end versus the wild-type end was then assayed. Removal of even one of the terminal 3' G nucleotides from the piggyBac inverted repeat, or removal of the dinucleotide AA from the flanking TTAA target site prevents excision of piggyBac at the mutated terminus. Incorporation of an asymmetric TTAC target site at the 3' end does not prevent excision from the mutated end. Thus, both piggyBac DNA and flanking host DNA appear to play crucial roles in the excision process.

Animals↗
PubMed1995

Transposon signatures: species-specific molecular markers that utilize a class of multiple-copy nuclear DNA.

Transposable elements are mobile sequences found in nuclear genomes and can potentially serve as molecular markers in various phylogenetic and population genetic investigations. A PCR-based method that utilizes restriction site variation of element copies within a genome is developed. These patterns of site variation, referred to as transposon signatures, are useful in differentiating between closely related groups. Signature data using the magellan retrotransposon, for example, is useful in examining relationships within the genus Zea and Tripsacum. This method allows transposable elements, or even other multiple-copy nuclear DNA sequences, to be generally utilized as molecular markers in discriminating between other closely related species and subspecies.

Base Sequence↗
PubMed2000

Molecular biology. Transposase team puts a headlock on DNA.

Transposable DNA elements jump from one location in the genome to another. But, the cut-and-paste molecular machinations that support this nomadic lifestyle are still being unraveled. In their Perspective, Williams and Baker at the Massachusetts Institute of Technology discuss new details of transposon relocation revealed through resolution of the structure of a transposase enzyme bound to DNA (Davies et al.).

Amino Acid Motifs↗
PubMed2000

Varied expression of a Y-linked P[w+] insert due to imprinting in Drosophila melanogaster.

During gametogenesis, a gene can become imprinted affecting its expression in progeny. We have used the expression of a Y-linked P[w+]YAL transposable DNA element as a reporter system to investigate the effect of parental origination on the expression of the w+ insert. Expression of w+ was greater in male progeny when the Y chromosome, harboring the insert, was inherited from the parental male rather than from the parental female. Imprinting was not due to a genetic background influence in the males, since the only difference among the males was the parental origin of the Y chromosome. It was also observed that the genetic background can affect imprinting, since w+ expression was also higher in males when the Y was derived from C(1)DX attached-X parental females rather than from C(1)RM attached-X parental females. Though the heterochromatic imprinting mechanism is unknown, a mutated Heterochromatin Protein 1 (HP1) gene, which is associated with suppression of position-effect variegation, increases expression of the w+ locus in the P[w+]YAL insert, indicating that HP1 may play a role in Y chromosome packaging.

Animals↗