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At least 109 records · Page 6Linked to original sources
PubMed1988

The DNA intermediate in yeast Ty1 element transposition copurifies with virus-like particles: cell-free Ty1 transposition.

Yeast Ty1 elements are retrotransposons that transpose via an RNA intermediate found in a virus-like particle (Ty-VLP). A Ty-encoded reverse transcriptase activity found inside the particles is capable of giving rise to full-length reverse transcripts. The predominant form of these reverse transcripts is a full-length linear duplex DNA. We have developed a cell-free system for transposition of Ty1 DNA molecules into a bacteriophage lambda target. Purified Ty-VLPs and target DNA are the only macromolecular components required for the transposition reaction. A TYB-encoded protein, p90-TYB, contains amino acid sequences that are similar to those of retroviral integrase proteins. Mutations in the integrase coding region abolish transposition both in vivo and in vitro.

Antibodies, Monoclonal↗
PubMed1988

The Drosophila su(Hw) gene, which controls the phenotypic effect of the gypsy transposable element, encodes a putative DNA-binding protein.

Homozygous mutations at the suppressor of Hairy-wing [su(Hw)] locus reverse the phenotype of gypsy-induced alleles in a number of genes located throughout the Drosophila genome. To understand the molecular basis of this phenomenon, the su(Hw) locus was isolated by chromosomal walking from a cloned homeo-box-containing sequence. The exact location of the gene was determined by Southern analysis of the DNA alterations associated with several su(Hw) alleles. A 9.5-kb KpnI-SalI fragment, where all the DNA changes associated with su(Hw) mutations were mapped, was able to rescue the su(Hw) mutant phenotype after P-element-mediated germ-line transformation. This DNA fragment encodes a 3.3-kb RNA that is expressed in all stages of Drosophila development; the size or abundance of this RNA is affected in several su(Hw) alleles tested. This transcript encodes a protein that contains a highly acidic region and 12 repeats of the 'Zn finger' domain characteristic of some DNA-binding and transcription-activating proteins, supporting the hypothesis that the su(Hw) locus might encode a transcription factor that plays a role in the expression of the gypsy element.

Alleles↗
PubMed2001

The Tol2 transposable element of the medaka fish: an active DNA-based element naturally occurring in a vertebrate genome.

Several DNA-based transposable elements are known to be present in vertebrate genomes, but few of them have been demonstrated to be active. The Tol2 element of the medaka fish is one such element and, therefore, is potentially useful for developing a gene tagging system and other molecular biological tools applicable to vertebrates. Towards this goal, analyses of the element at the molecular, cellular and population levels are in progress. Results so far obtained are described here.

Animals↗
PubMed1993

SPT13 (GAL11) of Saccharomyces cerevisiae negatively regulates activity of the MCM1 transcription factor in Ty1 elements.

The Ty transposable elements of Saccharomyces cerevisiae consist of a single large transcription unit whose expression is controlled by a combination of upstream and downstream regulatory sequences. Errede (B. Errede, Mol. Cell. Biol. 13:57-62, 1993) has shown that among the downstream control sequences is a binding site for the transcription factor, MCM1. A small restriction fragment containing the Ty1 MCM1-binding site exhibits very weak activation of heterologous gene expression. The absence of SPT13 (GAL11) causes a dramatic increase in activity directed by these sequences. This effect is mediated through the MCM1-binding site itself. MCM1 mRNA and protein levels, as well as its affinity for its binding site, are unchanged in the absence of SPT13. Our results suggest that SPT13 has a role in the negative control of MCM1 activity that is likely to be posttranslational. A role for SPT13 in the negative regulation of the activity of the Ty1 MCM1-binding site is consistent with our previous proposal that spt13-mediated suppression of Ty insertion mutations could be attributed to the loss of negative regulation of genes adjacent to Ty elements.

Base Sequence↗
PubMed1999

Identification of mutable slender glume gene in rice (Oryza sativa L.).

The segregation pattern and chromosomal location of a slender glume mutation, induced by gamma-ray irradiation, was investigated. The mutation is genetically unstable: in the selfed progenies of slender glumed plants, not only plants with normal glumes but also plants that are chimeric for glume shape almost always appear at low frequency. The results showed that the mutation is controlled by a single recessive, mutable mutant gene slg. The frequency of reversion of slg to its wild-type state was little affected by crossing, back-crossing, genetic background or cytoplasmic factors. Conventional trisomic and linkage analyses revealed that the slg locus was located close to the rfs (rolled fine stripe leaf) locus on chromosome 7. In a subsequent RFLP analysis, slg was found to be located between the two RFLP loci XNpb20 and XNpb33, with recombination values of 3.0 and 3.2%, respectively. Southern analysis indicated that the mutability of slg is caused by none of the known transposable elements in rice. From these results, we infer that slg has a novel transposable DNA insert in its vicinity, which was possibly activated by gamma-ray irradiation.

Chimera↗
PubMed1997

Genetic analyses of the interactions of the IS1-encoded proteins with the left end of IS1 and its insertion hotspot.

Insertion sequence IS1 specifies the InsA, delta InsA-B'-InsB and InsA-B'-InsB protein species. These three proteins have the identical alpha-helix-turn-alpha-helix motif that is likely to be responsible for DNA binding. In fact, InsA binds to the ends of IS1, and regulates gene expression and transposition of IS1. delta InsA-B'-InsB and/or InsA-B'-InsB has been thought to possess a transposase-like activity. Here, I examined the actions of these proteins in vivo on the promoter (pinsL) in the left end of IS1. InsA repressed pinsL-driven gene expression, both in cis and in trans. delta InsA-B'-InsB inhibited it efficiently only when pinsL was located near the construct where delta InsA-B'-InsB is expressed. Furthermore, it has been shown that the possible -10 sequence of pinsL is required for delta InsA-B'-InsB to act on, but the -35 sequence where InsA binds specifically, is not. InsA-B'-InsB appeared not to work on a nearby pinsL. The cis-action of delta InsA-B'-InsB is consistent with the previous observation that the IS1 transposase acts preferentially in cis. Interestingly, delta InsA-B'-InsB acted on a nearby P3 promoter in the IS1 insertion hotspot, and on another promoter outside the hotspot. delta InsA-B'-InsB may generally interact with the regions in or around promoters owing to their low DNA helix stability. Note that IS1 transposes preferentially into A + T-rich DNA segments, and that DNA is unwound from the -10 region of a promoter in transcription. The cis-preference of delta InsA-B'-InsB would result in an overall reduction of transposition of IS1 and its defective copy in a cell, allowing stable existence of the element in its bacterial host.

Base Sequence↗
PubMed2005

Genomic deletions and precise removal of transposable elements mediated by short identical DNA segments in primates.

Insertion of transposable elements is a major cause of genomic expansion in eukaryotes. Less is understood, however, about mechanisms underlying contraction of genomes. In this study, we show that retroelements can, in rare cases, be precisely deleted from primate genomes, most likely via recombination between 10- to 20-bp target site duplications (TSDs) flanking the retroelement. The deleted loci are indistinguishable from pre-integration sites, effectively reversing the insertion. Through human-chimpanzee-Rhesus monkey genomic comparisons, we estimate that 0.5%-1% of apparent retroelement "insertions" distinguishing humans and chimpanzees actually represent deletions. Furthermore, we demonstrate that 19% of genomic deletions of 200-500 bp that have occurred since the human-chimpanzee divergence are associated with flanking identical repeats of at least 10 bp. A large number of deletions internal to Alu elements were also found flanked by homologies. These results suggest that illegitimate recombination between short direct repeats has played a significant role in human genome evolution. Moreover, this study lends perspective to the view that insertions of retroelements represent unidirectional genetic events.

Alu Elements↗
PubMed1982

The physiological genetics of denitrifying bacteria.

The genetics of denitrification is a relatively unexplored area that has great promise. Species of Pseudomonas are probably best suited for study because they are widely found among natural denitrifying populations and are quite readily amenable to genetic analysis. The techniques for mutagenesis and for the exchange of chromosomal genes to characterize mutant strains have been well-developed in P. aeruginosa and are being developed in P. stutzeri. Mutants defective in the denitrification of nitrate, nitrite, and nitrous oxide are now available and will aid in describing the catalytic and regulatory elements of the denitrification pathway.

DNA Transposable Elements↗
PubMed1995

Mu transposase-stimulated illegitimate recombination of Tn3kan- and IS101-containing plasmids.

The transposable bacteriophage Mu and the mobile genetic elements Tn3 and IS101 replicatively transpose to random target sites, produce 5 bp target site duplications, and contain the sequence 5'-PuCGAAAPu-3' starting at bp 21 from their ends. The presence of these shared characteristics, plus the fact that Mu transposase can specifically bind to the termini of Tn3 and IS101 in vitro, suggests that the elements may be evolutionarily conserved and retain some functional capacity to transpose each other's DNA. To examine this proposition, in vivo transposition-mating assays were performed and demonstrated that Mu transposase stimulated the formation of recA-independent recombination products between Tn3kan- or IS101-containing plasmids and a target plasmid (pOX38cam) up to 200-fold. However, when transferred to recA+ hosts, these recA-independent products yielded resolution products suggestive of illegitimate recombination, as similar recombination and resolution products were generated, at reduced frequencies, in the absence of Mu transposase. Thus, Mu transposase may stimulate a host-mediated, recA-independent illegitimate recombination reaction. As adjacent pSC101 sequences, including a formerly unknown but functional IHF site (bp 2238-2251), were required for Mu transposase-stimulated IS101 illegitimate recombination, IHF may be one of the putative host factors involved in these recombination reactions.

Bacteriophage mu↗
PubMed1985

[Different effect of mutations in Escherichia coli K12 dna-genes on the transposition of Tn5- and Tn10-elements].

The effect of mutations in dnaA(dnaA46), dnaG(dnaG3), dnaC (dnaC1 and dnaC2) and dnaB genes on transposition of two transposons, Tn5 and Tn10, from bacteriophage lambda genome into the chromosome of host cells has been studied. Transposition was performed at permissive temperatures for the mutant recipients. The mutations in dnaA, dnaC, dnaG genes were shown to decrease the transposition of Tn10 for some orders of magnitude as compared with transposition registered in wild type cells. Independence of Tn5 transposition of the above mentioned genes was demonstrated, providing evidence on the different modes of transposition of these two Tn-elements.

Bacteriophage lambda↗
PubMed2003

Excision of the Drosophila mariner transposon Mos1. Comparison with bacterial transposition and V(D)J recombination.

It has been proposed that the modern immune system has evolved from a transposon in an ancient vertebrate. While much is known about the mechanism by which bacterial transposable elements catalyze double-strand breaks at their ends, less is known about how eukaryotic transposable elements carry out these reactions. We have examined the mechanism by which mariner, a eukaryotic transposable element, performs DNA cleavage. We show that the nontransferred strand is cleaved initially, unlike prokaryotic transposons which cleave the transferred strand first. First strand cleavage is not tightly coupled to second strand cleavage and can occur independently of synapsis, as happens in V(D)J recombination but not in transposition of prokaryotic transposons. Unlike V(D)J recombination, however, second strand cleavage of mariner does not occur via a hairpin intermediate.

Animals↗
PubMed1999

Organization and dynamics of the Mu transpososome: recombination by communication between two active sites.

Movement of transposable genetic elements requires the cleavage of each end of the element genome and the subsequent joining of these cleaved ends to a new target DNA site. During Mu transposition, these reactions are catalyzed by a tetramer of four identical transposase subunits bound to the paired Mu DNA ends. To elucidate the organization of active sites within this tetramer, the subunit providing the essential active site DDE residues for each cleavage and joining reaction was determined. We demonstrate that recombination of the two Mu DNA ends is catalyzed by two active sites, where one active site promotes both cleavage and joining of one Mu DNA end. This active site uses all three DDE residues from the subunit bound to the transposase binding site proximal to the cleavage site on the other Mu DNA end (catalysis in trans). In addition, we uncover evidence that the catalytic activity of these two active sites is coupled such that the coordinated joining of both Mu DNA ends is favored during recombination. On the basis of these results, we propose that the DNA joining stage requires a cooperative transition within the transposase-DNA complex. The cooperative utilization of active sites supplied in trans by Mu transposase provides an example of how mobile elements can ensure concomitant recombination of distant DNA sites.

Bacteriophage mu↗
PubMed2006

AT-rich sequences from the arbuscular mycorrhizal fungus Gigaspora rosea exhibit ARS function in the yeast Saccharomyces cerevisiae.

Autonomous replicating sequences are DNA elements that trigger DNA replication and are widely used in the development of episomal transformation vectors for fungi. In this paper, a genomic library from the mycorrhizal fungus Gigaspora rosea was constructed in the integrative plasmid YIp5 and screened in the budding yeast Saccharomyces cerevisiae for sequences that act as ARS and trigger plasmid replication. Two genetic elements (GrARS2, GrARS6) promoted high-rates of yeast transformation. Sequence analysis of these elements shows them to be AT-rich (72-80%) and to contain multiple near-matches to the yeast autonomous consensus sequences ACS and EACS. GrARS2 contained a putative miniature inverted-repeat transposable element (MITE) delimited by 28-bp terminal inverted repeats (TIRs). Disruption of this element and removal of one TIR increased plasmid stability several fold. The potential for palindromes to affect DNA replication is discussed.

AT Rich Sequence↗