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PubMed1982

Molecular organization of a Drosophila puff site that responds to ecdysone.

The 68C locus in the polytene chromosomes of Drosophila melanogaster salivary glands is puffed during the last half of the third larval instar and harbors the structural gene for sgs3, one of the glue polypeptides synthesized by the glands during this period. This puff regresses in response to the steroid hormone ecdysone. We have isolated a set of overlapping cloned segments that define approximately 50 kb of DNA at the 68C puff locus. Three polysomal poly(A)+ RNAs that are abundant in the salivary glands during the intermolt-puff stage are transcribed from three genes (II, III and IV) that map near the center of the cloned DNA in a 5 kb cluster region. These are the only transcripts from the 50 kb of 68C DNA detectable in these glands, and they are undetectable in other larval tissues at this stage, or in whole animals at other stages of development. Correlative criteria indicate that gene IV, which yields an RNA of 1.1 kb, is the structural gene for sgs3. Genes II and III, which yield RNAs of 0.36 kb and 0.32 kb, respectively, are oppositely oriented so that their promoters are adjacent, suggesting that this pair may form a single regulated unit, a suggestion that is enhanced by the fact that the pair is bounded by an inverted repeat of 0.3 kb elements. The possible, but as yet unidentified, functions of this gene pair are discussed. A 9.2 kb element belonging to a family of transposable elements called roo is inserted adjacent to the 5 kb cluster region in some but not other D. melanogaster strains. This insertion has no obvious effect on the transcription of genes II, III and IV. Although roo elements yield a 9 kb poly(A)+ RNA in embryos, no roo transcripts were detected in intermolt salivary glands, whether they do or do not contain an element at the 68C puff site.

Chromosome Mapping↗
PubMed2006

Characterizing the composition and evolution of homoeologous genomes in hexaploid wheat through BAC-end sequencing on chromosome 3B.

Bread wheat (Triticum aestivum) is one of the most important crops worldwide. However, because of its large, hexaploid, highly repetitive genome it is a challenge to develop efficient means for molecular analysis and genetic improvement in wheat. To better understand the composition and molecular evolution of the hexaploid wheat homoeologous genomes and to evaluate the potential of BAC-end sequences (BES) for marker development, we have followed a chromosome-specific strategy and generated 11 Mb of random BES from chromosome 3B, the largest chromosome of bread wheat. The sequence consisted of about 86% of repetitive elements, 1.2% of coding regions, and 13% remained unknown. With 1.2% of the sequence length corresponding to coding sequences, 6000 genes were estimated for chromosome 3B. New repetitive sequences were identified, including a Triticineae-specific tandem repeat (Fat) that represents 0.6% of the B-genome and has been differentially amplified in the homoeologous genomes before polyploidization. About 10% of the BES contained junctions between nested transposable elements that were used to develop chromosome-specific markers for physical and genetic mapping. Finally, sequence comparison with 2.9 Mb of random sequences from the D-genome of Aegilops tauschii suggested that the larger size of the B-genome is due to a higher content in repetitive elements. It also indicated which families of transposable elements are mostly responsible for differential expansion of the homoeologous wheat genomes during evolution. Our data demonstrate that BAC-end sequencing from flow-sorted chromosomes is a powerful tool for analysing the structure and evolution of polyploid and highly repetitive genomes.

Base Sequence↗
PubMed2002

Conservation of gene order, structure and sequence between three closely linked genes in Drosophila melanogaster and Drosophila virilis.

In Drosophila melanogaster, the apparently unrelated genes anon-66Da, RpL14, and anon-66Db (from telomere to centromere) are located on a 5547 bp genomic fragment on chromosome arm 3L at cytological position 66D8. The three genes are tightly linked, and flanked by two relatively large genes with unknown function. We have taken a comparative genomic approach to investigate the evolutionary history of the three genes. To this end we isolated a Drosophila virilis 7.3 kb genomic fragment which is homologous to a 5.5 kb genomic region of D. melanogaster. Both fragments map to Muller's element D, namely to section 66D in D. melanogaster and to section 32E in D. virilis, and harbor the genes anon-66Da, RpL14, and anon-66Db. We demonstrate that the three genes exhibit a high conservation of gene topography in general and in detail. While most introns and intergenic regions reveal sequence divergences, there are, however, a number of interspersed conserved sequence motifs. In particular, two introns of the RpL14 gene contain a short, highly conserved 60 nt long sequence located at corresponding positions. This sequence represents a novel Drosophila small nucleolar RNA, which is homologous to human U49. Whereas DNA flanking the three genes shows no significant interspecies homologies, the 3'-flanking region in D. virilis contains sequences from the transposable element Penelope. The Penelope family of transposable elements has been shown to promote chromosomal rearrangements in the D. virilis species group. The presence of Penelope sequences in the D. virilis 7.3 kb genomic fragment may be indicative for a transposon-induced event of transposition which did not yet scramble the order of the three genes but led to the breakdown of sequence identity of the flanking DNA.

Amino Acid Sequence↗
PubMedDate not supplied

Independent amplification of two classes of Tourists in some Oryza species.

Transposable elements similar to Tourist elements from maize were isolated from the rice genome. The elements were about 300 bp, exhibited short terminal inverted repeats (TIR), and appeared to show preferential insertion at TAA sites. Some rice Tourist elements seem to have recently transposed. Based on the sequences of cloned elements, two classes of rice Tourist elements have been identified. Members of these two classes apparently amplified independently at different times in the past.

Base Sequence↗
PubMed2001

Gene conversion within regulatory sequences generates maize r alleles with altered gene expression.

The maize r locus encodes a transcription factor that regulates the developmental expression of the plant pigment anthocyanin. In an unusual example of gene regulatory diversity, the R-sc (Sc, strong seed color) and the R-p (P, plant color) alleles of r have nonoverlapping tissue specificity and nonhomologous 5' flanking sequences. Heterozygotes between wild-type P and Sc mutants with Ds6 transposable element inserts (r-sc:m::Ds6 or sc:m) produce colored seed derivatives (Sc+) during meiotic recombination. The sc:m alleles with Ds6 insertion in 3' regions of r produce crossover Sc+ derivatives. sc:m alleles with Ds6 elements inserted in 5' regions produce rare Sc+ derivatives borne on nonrecombinant chromosomes. Among 52 such noncrossover Sc+ derivatives, 18 are indistinguishable from the Sc progenitor in phenotype and DNA sequence [Scp(+) alleles]. The remaining 34 derivatives have strong Sc+ expression, including darkly pigmented aleurone, scutellum, coleoptile, and scutellar node [Scp(e) alleles]. The coleoptile and scutellar node phenotypes are unique from either progenitor but are similar to those of some naturally occurring r alleles. Both classes of Sc+ derivatives are explained by gene conversion between the promoter region of Sc:124 and a homologous region located proximal to P. The recombinational intermediate formed between sc:m alleles and P results in deletion of the Ds6 element alone or both Ds6 and a nearby unrelated transposable element-like sequence.

Alleles↗
PubMed2026

Nuclear DNA of plastid origin (NUPTs), neglected driver of genome variation and evolutionary innovation.

Plant nuclear genomes contain a variable, though typically minor, fraction of DNA sequences of plastid origin known as NUPTs. Unlike the massive transfer of DNA and genes from the proto-organelle genome to the nucleus that occurred during the endosymbiotic event that gave rise to plastids, the formation of NUPTs is an ongoing process that does not imply concomitant DNA loss. Although NUPTs are generally considered to be potentially deleterious insertions that are continuously generated and rapidly eliminated at near-constant turnover rates, accumulating evidence reveals alternative evolutionary trajectories. In this review, we discuss recent findings that highlight the episodic formation of NUPTs, their subsequent proliferation, and their eventual long-term fixation within the nuclear genome. We also explore their non-random spatial association with specific genomic elements. NUPTs show preferential overlap with specific superfamilies of transposable elements, which may facilitate their proliferation and dispersal throughout the nuclear genome. Regarding protein-coding genes, the contribution of NUPTs varies among species. In contrast, NUPTs are found to be consistently enriched among certain classes of non-coding RNA genes, notably rRNA, tRNA, and specific regulatory RNA families, suggesting that they are involved in the evolution of gene regulation and translational machinery. Overall, these findings underscore the unexpected complexity of the mechanisms underlying NUPT formation and support the idea that they are a significant source of genome variation and evolutionary innovation. Further research is necessary to fully elucidate the mechanisms underlying NUPT formation, as well as to determine their potential adaptive significance in plant genome evolution.

Plastids↗
PubMed1982

Transposition of cloned P elements into Drosophila germ line chromosomes.

Recombinant DNA carrying the 3-kilobase transposable element was injected into Drosophila embryos of a strain that lacked such elements. Under optimum conditions, half of the surviving embryos showed evidence of P element-induced mutations in a fraction of their progeny. Direct analysis of the DNA of strains derived from such flies showed them to contain from one to five intact 3-kilobase P elements located at a wide variety of chromosomal sites. DNA sequences located outside the P element on the injected DNA were not transferred. Thus P elements can efficiently and selectively transpose from extrachromosomal DNA to the DNA of germ line chromosomes in Drosophila embryos. These observations provide the basis for efficient DNA-mediated gene transfer in Drosophila.

Animals↗
PubMed1986

Identification and immunochemical analysis of biologically active Drosophila P element transposase.

We have identified proteins encoded by P transposable elements expressed in transformed Drosophila tissue culture cells. Two proteins have been identified by immunochemical techniques. One, an 87,000 dalton polypeptide, is encoded by a P element mRNA lacking the third (ORF2-ORF3) intervening sequence. The other protein, a 66,000 dalton polypeptide, is encoded by an mRNA that retains the third intron and is found in somatic tissues. Furthermore, tissue culture cell lines expressing the 87,000 dalton polypeptide are able to catalyze both the precise and imprecise excision of a nonautonomous P element. The 87,000 dalton protein is encoded by sequences from all four P element open reading frames. Taken together, these data strongly suggest that the 87,000 dalton polypeptide is the P element transposase.

Amino Acid Sequence↗
PubMed2002

The colugo (Cynocephalus variegatus, Dermoptera): the primates' gliding sister?

Although a general agreement on the major groups of eutherian orders and their phylogenetic affiliations is emerging, the evolutionary affiliations among the members constituting these groups are still subject to debate. A prominent example is the recently published molecular evidence that challenges the long assumed monophyly of primates, displaying the colugo or flying lemur (Cynocephalus, Dermoptera) as a sister to anthropoid primates (Arnason et al. 2002 ) and positioning them after the prosimian primates (tarsiers and strepsirhines) split off. The phylogenetic analysis of the complete mitochondrial (mt) genome sequence of Cynocephalus variegatus presented in this study first appears to corroborate interpretations of primates as a paraphyletic group. However, more detailed analyses disclosed that mt nucleotide composition and consequently amino acid (AA) composition varied considerably among the species analyzed. This led us to assume that the flying lemur may be incorrectly grouped with anthropoids on the basis of similar mt nucleotide and AA compositions, rather than reflecting the true evolutionary relationship. To reanalyze the flying lemur's evolutionary association with other eutherian orders from a completely different molecular perspective, a molecular cladistic approach was applied. To this end, we determined the presence/absence pattern of transposable elements that provide a nearly homoplasy-free and copious source of molecular evolutionary markers, with well-defined character polarity. We could identify transposable elements, both on a multilocus and single-locus level, being present in all extant primate infraorders but absent in the flying lemur, thus clearly supporting the monophyly of primates by retropositional evidence.

Animals↗
PubMed1980

Properties of erythromycin-inducible transposon Tn917 in Streptococcus faecalis.

Streptococcus faecalis strain DS16 harbors two plasmids, a conjugative plasmid, pAD1, which encodes hemolysin and bacteriocin activities, and a nonconjugative plasmid, pAD2, encoding resistance to streptomycin, kanamycin, and erythromycin, the latter of which is inducible. The erythromycin resistance determinant is located on a 3.3-megadalton transposable element designated Tn917, which could be transposed to pAD1 as well as to two other plasmids, pAm gamma 1 and pAM alpha 1. When strain DS16 was exposed to low (inducing) concentrations of erythromycin for a few hours, the frequency of Tn917 transposition from pAD2 to pAD1 increased by an order of magnitude. This induction paralleled induction of erythromycin resistance and was prevented by exposing the cells to inhibitors of deoxyribonucleic acid, ribonucleic acid or protein synthesis. The exposure of strain DS16 to inducing concentrations of erythromycin also enhanced the frequency of erythromycin-resistant transconjugants appearing during mating. Initially, cointegrate molecules, whose molecular weights were approximately the sum of pAD1 and pAD2, accounted for these transconjugants; however, as the induction time increased, pAD1::Tn917 became increasingly prominent.

Chloramphenicol↗
PubMed1989

Structural analysis of Tc1 elements in Caenorhabditis elegans var. Bristol (strain N2).

The transposable element Tc1 in the genome of Caenorhabditis elegans var. Bristol strain N2 is very stable. In order to investigate possible causes of Tc1 immobility in this strain 17 individual isolates have been cloned and characterized with regard to their structure and genomic environment. Ten of 16 elements examined had identical restriction maps, and at least 1 of these (#7) showed a high level of somatic excision. Two of the elements had altered restriction sites, 2 had different internal deletions of about 700 bp, 1 had an 89-bp terminal deletion, and 1 a 54-bp insertion. When DNA sequences flanking the N2 Tc1 elements were used as probes in genomic hybridizations, it was found that most N2 elements are located in regions of repetitive DNA. Furthermore when hybridizations to DNA from N2 and var. Bergerac strain B0 were performed, a major band of the same size was observed in both strains. Two flanking sequences identified strain polymorphic sites hP2(IV) and hP3(IV). In at least one of these cases, a rearranged Tc1 was present in the B0 strain at the same location. The fact that all or most of the Tc1 elements are in the same location in N2 and B0 adds support to the hypothesis that the high copy number B0 strain arose from amplification of Tc1 copies in a N2-like strain. The N2 Tc1 elements are highly conserved; however, intact elements had fewer nucleotide changes than the rearranged elements. These results may indicate that the intact Tc1 elements in N2 are functionally active and subject to selective pressure.

Animals↗
PubMed2002

Evolution of a polymorphic regulatory element in interferon-gamma through transposition and mutation.

Mammalian transposable elements have intrinsic regulatory elements that can activate neighboring genes, and it is speculated that they can also carry extrinsic transactivating DNA sequences to new genomic locations. We have identified a polymorphic segment of the human interferon-gamma promoter region where two adjacent binding sites for NF-kappaB and NFAT originated from the insertion of an Alu element approximately 22-34 MYA. Both binding sites lie outside the Alu consensus sequence but within the boundaries of the insertion, suggesting that this segment of DNA was comobilized when the Alu element moved from another part of the genome. Sequence comparisons and examination of DNA-protein interactions across nine different primate species indicate that the inserted sequence contained the intact NFAT binding site, whereas the ability to bind NF-kappaB evolved through a series of mutations after the insertion. These observations are consistent with the notion that retropseudogenes can comobilize intact regulatory sequences to new locations and thereby influence the evolution of gene regulatory networks; however, the extent to which such events have shaped the evolution of gene regulation remains unknown.

Alu Elements↗
PubMed1989

Transpositional behavior of the maize En/Spm element in transgenic tobacco.

The transposable element Enhancer (En/Spm) of Zea mays encodes the functions necessary for transposition of members of the En-I family of elements. En constructs were introduced into tobacco and transposition was demonstrated by cloning and DNA sequence analysis. Transcriptional analysis of En transgenic plants revealed 2.5 and 6 kb En homologous transcripts similar in size to those found in maize, in addition to aberrant splice products. Progeny analysis of a primary transformant indicates En transpositional activity after meiosis. In these progeny somatic variegation, at the NPT II gene, was demonstrated to be caused by En excision. Two re-insertions of En were analyzed and found to reside in unique copy DNA while another was found in middle repetitive DNA.

Journal Article↗
PubMed2006

Comparing the whole-genome-shotgun and map-based sequences of the rice genome.

The rice genome has now been sequenced using whole-genome-shotgun and map-based methods. The relative merits of the two methods are the subject of debate, as they were in the human genome project. In this Opinion article, we will show that the serious discrepancies between the resultant sequences are mostly found in the large transposable elements such as copia and gypsy that populate the intergenic regions of plant genomes. Differences in published gene counts and polymorphism rates are similarly resolved by considering how transposable elements affect the sequence analysis.

Chromosomes, Plant↗
PubMed1997

The enigma of Y chromosome degeneration: TRAM, a novel retrotransposon is preferentially located on the Neo-Y chromosome of Drosophila miranda.

We have cloned a novel transposable element from the neo-Y chromosome of Drosophila miranda. The size of the element, designated as TRAM, is 3.452 bp, including on both sides long terminal direct repeats (LTRs) of 372 bp, respectively. The element is flanked by a 5-bp target site duplication, ATATG. The putative primer binding site (PBS) for minus-strand priming is complementary to 18 nucleotides of the 3'-end of tRNA(Trp). Data base screens for DNA sequence identities were negative, apart from the sequence motif of the PBS. The deduced amino acid sequence from the large ORF does not reveal identities described for other transposons. In situ hybridizations with TRAM subclones show a biased distribution in the genome, with a massive accumulation of TRAM in the neo-Y chromosome, while the former homologue, the X2 chromosome is devoid of TRAM sites. The enriched occurrence of the TRAM element at the evolving neo-Y chromosome of D.miranda adds compelling evidence in favor of the view that Y chromosome degeneration is driven by the accumulation of transposable elements.

Animals↗
PubMed2002

Coevolution of the telomeric retrotransposons across Drosophila species.

As in other eukaryotes, telomeres in Drosophila melanogaster are composed of long arrays of repeated DNA sequences. Remarkably, in D. melanogaster these repeats are produced, not by telomerase, but by successive transpositions of two telomere-specific retrotransposons, HeT-A and TART. These are the only transposable elements known to be completely dedicated to a role in chromosomes, a finding that provides an opportunity for investigating questions about the evolution of telomeres, telomerase, and the transposable elements themselves. Recent studies of D. yakuba revealed the presence of HeT-A elements with precisely the same unusual characteristics as HeT-A(mel) although they had only 55% nucleotide sequence identity. We now report that the second element, TART, is also a telomere component in D. yakuba; thus, these two elements have been evolving together since before the separation of the melanogaster and yakuba species complexes. Like HeT-A(yak), TART(yak) is undergoing concerted sequence evolution, yet they retain the unusual features TART(mel) shares with HeT-A(mel). There are at least two subfamilies of TART(yak) with significantly different sequence and expression. Surprisingly, one subfamily of TART(yak) has >95% sequence identity with a subfamily of TART(mel) and shows similar transcription patterns. As in D. melanogaster, other retrotransposons are excluded from the D. yakuba terminal arrays studied to date.

Animals↗
PubMed1993

Transactivation of Ds elements in plants of lettuce (Lactuca sativa).

The maize transposable element, Activator (Ac), is being used to develop a transposon mutagenesis system in lettuce, Lactuca sativa. In this paper, we describe somatic and germinal transactivation of Ds by chimeric transposase genes in whole plants. Constructs containing either the Ds element or the Ac transposase open reading frame (ORF) were introduced into lettue. The Ds element was located between either the 35S or the Nos promoter and a chimeric spectinomycin resistance gene (which included a transit peptide), preventing expression of spectinomycin resistance. The genomic coding region of the Ac transposase was expressed from the 35S promoter. Crosses were made between 104 independent R1 plants containing Ds and three independent R1 plants expressing transposase. The excision of Ds in F1 progenies was monitored using a phenotypic assay on spectinomycin-containing medium. Green sectors in one-third of the F1 families indicated transactivation of Ds by the transposase at different developmental stages and at different frequencies in lettuce plants. Excision was confirmed using PCR and by Southern analysis. The lack of green sectors in the majority of F1 families suggest that the majority of T-DNA insertion sites are not conducive to excision. In subsequent experiments, the F1 plants containing both Ds and the transposase were grown to maturity and the F2 seeds screened on medium containing spectinomycin. Somatic excision was again observed in several F2 progeny; however, evidence for germinal excision was observed in only one F2 family.

Base Sequence↗
PubMed1997

Abortive gap repair: underlying mechanism for Ds element formation.

The mechanism by which the maize autonomous Ac transposable element gives rise to nonautonomous Ds elements is largely unknown. Sequence analysis of native maize Ds elements indicates a complex chimeric structure formed through deletions of Ac sequences with or without insertions of Ac-unrelated sequence blocks. These blocks are often flanked by short stretches of reshuffled and duplicated Ac sequences. To better understand the mechanism leading to Ds formation, we designed an assay for detecting alterations in Ac using transgenic tobacco plants carrying a single copy of Ac. We found frequent de novo alterations in Ac which were excision rather than sequence dependent, occurring within Ac but not within an almost identical Ds element and not within a stable transposase-producing gene. The de novo DNA rearrangements consisted of internal deletions with breakpoints usually occurring at short repeats and, in some cases, of duplication of Ac sequences or insertion of Ac-unrelated fragments. The ancient maize Ds elements and the young Ds elements in transgenic tobacco showed similar rearrangements, suggesting that Ac-Ds elements evolve rapidly, more so than stable genes, through deletions, duplications, and reshuffling of their own sequences and through capturing of unrelated sequences. The data presented here suggest that abortive Ac-induced gap repair, through the synthesis-dependent strand-annealing pathway, is the underlying mechanism for Ds element formation.

DNA Repair↗