Academic literature on the topic 'Transposable elements; repeats; evolution'

Satellite DNAs are now regarded as powerful and active contributors to genomic and chromosomal evolution. Paired with mobile transposable elements, these repetitive sequences provide a dynamic mechanism through which novel karyotypic modifications and chromosomal rearrangements may occur. In this review, we discuss the regulatory activity of satellite DNA and their neighboring transposable elements in a chromosomal context with a particular emphasis on the integral role of both in centromere function. In addition, we discuss the varied mechanisms by which centromeric repeats have endured evolutionary processes, producing a novel, species-specific centromeric landscape despite sharing a ubiquitously conserved function. Finally, we highlight the role these repetitive elements play in the establishment and functionality of de novo centromeres and chromosomal breakpoints that underpin karyotypic variation. By emphasizing these unique activities of satellite DNAs and transposable elements, we hope to disparage the conventional exemplification of repetitive DNA in the historically-associated context of ‘junk’.

SummaryThe genome of the house fly, Musca domestica, contains an active transposable element system, called Hermes. Using PCR and inverse PCR we amplified and sequenced overlapping segments of several Hermes elements and from these data we have constructed a 2749 bp consensus Hermes DNA sequence. Hermes termini are composed of 17 bp imperfect inverted repeats that are almost identical to the inverted terminal repeats of the hobo element of Drosophila melanogaster. Full length Hermes elements contain a single long ORF capable of encoding a protein of 612 amino acids which is 55% identical to the amino acid sequence of the hobo transposase. Comparison of the ends of the Hermes and hobo elements to those of the Ac element of Zea mays, and the Tam3 element of Antirrhinum majus, as well as several other plant and insect elements, revealed a conserved terminal sequence motif. Thus Hermes is clearly a member of the hobo, Ac and Tam3 (hAT) transposable element family, other members of which include the Tagl element from Arabidopsis thaliana and the Bg element from Zea mays. The evolution of this class of transposable elements and the potential utility of Hermes as a genetic tool in M. domestica and related species are discussed.

Repetitive DNAs are ubiquitous in eukaryotic genomes and, in many species, comprise the bulk of the genome. Repeats include transposable elements that can self-mobilize and disperse around the genome and tandemly-repeated satellite DNAs that increase in copy number due to replication slippage and unequal crossing over. Despite their abundance, repetitive DNAs are often ignored in genomic studies due to technical challenges in identifying, assembling, and quantifying them. New technologies and methods are now allowing unprecedented power to analyze repetitive DNAs across diverse taxa. Repetitive DNAs are of particular interest because they can represent distinct modes of genome evolution. Some repetitive DNAs form essential genome structures, such as telomeres and centromeres, that are required for proper chromosome maintenance and segregation, while others form piRNA clusters that regulate transposable elements; thus, these elements are expected to evolve under purifying selection. In contrast, other repeats evolve selfishly and cause genetic conflicts with their host species that drive adaptive evolution of host defense systems. However, the majority of repeats likely accumulate in eukaryotes in the absence of selection due to mechanisms of transposition and unequal crossing over. However, even these “neutral” repeats may indirectly influence genome evolution as they reach high abundance. In this Special Issue, the contributing authors explore these questions from a range of perspectives.

Fluorescence in situ hybridization (FISH) is widely used in the physical mapping of genes and chromosome landmarks in plants and animals. Bacterial artificial chromosomes (BACs) contain large inserts, making them amenable for FISH mapping. In our BAC-FISH experiments, we selected 56 restriction fragment length polymorphism (RFLP)-locus-specific BAC clones from the libraries of Triticum monococcum and Aegilops tauschii, which are the A- and D-genome donors of wheat (Triticum aestivum, 2n = 6x = 42), respectively. The BAC clone 676D4 from the T. monococcum library contains a dispersed repeat that preferentially hybridizes to A-genome chromosomes, and two BAC clones, 9I10 and 9M13, from the Ae. tauschii library contain a dispersed repeat that preferentially hybridizes to the D-genome chromosomes. These repeats are useful in simultaneously discriminating the three different genomes in hexaploid wheat, and in identifying intergenomic translocations in wheat or between wheat and alien chromosomes. Sequencing results show that both of these repeats are transposable elements, indicating the importance of transposable elements, especially retrotransposons, in the genome evolution of wheat.Key words: bacterial artificial chromosome (BAC), fluorescence in situ hybridization (FISH), transposable elements (TEs), wheat, Triticum aestivum.

The distinctive biology and unique evolutionary features of snakes make them fascinating model systems to elucidate how genomes evolve and how variation at the genomic level is interlinked with phenotypic-level evolution. Similar to other eukaryotic genomes, large proportions of snake genomes contain repetitive DNA, including transposable elements (TEs) and satellite repeats. The importance of repetitive DNA and its structural and functional role in the snake genome, remain unclear. This review highlights the major types of repeats and their proportions in snake genomes, reflecting the high diversity and composition of snake repeats. We present snakes as an emerging and important model system for the study of repetitive DNA under the impact of sex and microchromosome evolution. We assemble evidence to show that certain repetitive elements in snakes are transcriptionally active and demonstrate highly dynamic lineage-specific patterns as repeat sequences. We hypothesize that particular TEs can trigger different genomic mechanisms that might contribute to driving adaptive evolution in snakes. Finally, we review emerging approaches that may be used to study the expression of repetitive elements in complex genomes, such as snakes. The specific aspects presented here will stimulate further discussion on the role of genomic repeats in shaping snake evolution.

While the evolutionary mechanisms driving eukaryote genome size evolution are still debated, repeated element content appears to be crucial. Here, we reconstructed the phylogeny and identified repeats in the genome of 26 Drosophila exhibiting a twofold variation in genome size . The content in transposable elements (TEs) is highly correlated to genome size evolution among these closely related species. We detected a strong phylogenetic signal on the evolution of both genome size and TE content, and a genome contraction in the Drosophila melanogaster subgroup.

Eukaryotic organisms have dynamic genomes, with transposable elements (TEs) as a major contributing factor. Although the large autonomous TEs can significantly shape genomic structures during evolution, genomes often harbor more miniature nonautonomous TEs that can infest genomic niches where large TEs are rare. In spite of their cut-and-paste transposition mechanisms that do not inherently favor copy number increase, miniature inverted-repeat transposable elements (MITEs) are abundant in eukaryotic genomes and exist in high copy numbers. Based on the large number of MITE families revealed in previous studies, accurate annotation of MITEs, particularly in newly sequenced genomes, will identify more genomes highly rich in these elements. Novel families identified from these analyses, together with the currently known families, will further deepen our understanding of the origins, transposase sources, and dramatic amplification of these elements.

Transposable elements (TEs) are ubiquitous in arthropods. However, analyses of large-scale and long-term coevolution between TEs and host genomes remain scarce in arthropods. Here, we choose 14 representative Arthropoda species from eight orders spanning more than 500 million years of evolution. By developing an unbiased TE annotation pipeline, we obtained 87 to 2266 TE reference sequences in a species, which is a considerable improvement compared to the reference TEs previously annotated in Repbase. We find that TE loads are diversified among species and were previously underestimated. The highly species- and time-specific expansions and contractions, and intraspecific sequence diversification are the leading driver of long terminal repeat (LTR) dynamics in Lepidoptera. Terminal inverted repeats (TIRs) proliferated substantially in five species with large genomes. A phylogenetic comparison reveals that the loads of multiple TE subfamilies are positively correlated with genome sizes. We also identified a few horizontally transferred TE candidates across nine species. In addition, we set up the Arthropod Transposable Elements database (ArTEdb) to provide TE references and annotations. Collectively, our results provide high-quality TE references and uncover that TE loads and expansion histories vary greatly among arthropods, which implies that TEs are an important driving force shaping the evolution of genomes through gain and loss.

Abstract While genome-wide surveys of abundance and diversity of mobile elements have been conducted for some class I transposable element families, little is known about the nature of class II transposable elements on this scale. In this report, we present the results from analysis of the sequence and structural diversity of Mutator-like elements (MULEs) in the genome of Arabidopsis thaliana (Columbia). Sequence similarity searches and subsequent characterization suggest that MULEs exhibit extreme structure, sequence, and size heterogeneity. Multiple alignments at the nucleotide and amino acid levels reveal conserved, potentially transposition-related sequence motifs. While many MULEs share common structural features to Mu elements in maize, some groups lack characteristic long terminal inverted repeats. High sequence similarity and phylogenetic analyses based on nucleotide sequence alignments indicate that many of these elements with diverse structural features may remain transpositionally competent and that multiple MULE lineages may have been evolving independently over long time scales. Finally, there is evidence that MULEs are capable of the acquisition of host DNA segments, which may have implications for adaptive evolution, both at the element and host levels.

The research field of comparative genomics is moving from a focus on genes to a more holistic view including the repetitive complement. This study aimed to characterize relative proportions of the repetitive fraction of large, complex genomes in a nonmodel system. The monocotyledonous plant order Asparagales (onion, asparagus, agave) comprises some of the largest angiosperm genomes and represents variation in both genome size and structure (karyotype). Anonymous, low coverage, single-end Illumina data from 11 exemplar Asparagales taxa were assembled using a de novo method. Resulting contigs were annotated using a reference library of available monocot repetitive sequences. Mapping reads to contigs provided rough estimates of relative proportions of each type of transposon in the nuclear genome. The results were parsed into general repeat types and synthesized with genome size estimates and a phylogenetic context to describe the pattern of transposable element evolution among these lineages. The major finding is that although some lineages in Asparagales exhibit conservation in repeat proportions, there is generally wide variation in types and frequency of repeats. This approach is an appropriate first step in characterizing repeats in evolutionary lineages with a paucity of genomic resources.

Transposable elements (TEs) are mobile genetic elements capable of replicating and spreading within, and in some cases, between genomes. I describe a whole-genome analysis of DD34E TEs, which belong to the IS630-Tc1-mariner superfamily of DNA transposable elements, in the African malaria mosquito, Anopheles gambiae. Twenty-six new transposons as well as a new family, gambol, were identified. The gambol family shares the DD34E catalytic motif with Tc1-DD34E transposons, but is distinct from these elements in their phylogenetic relationships. Although gambol appears to be related to a few DD34E transposons from cyanobacteria and fungi, no gambol elements have been reported in any other insects or animals thus far. This discovery expands the already expansive diversity of the IS630-Tc1-mariner TEs, and raises interesting questions as to the origin of gambol elements and their apparent diversity in An. gambiae. Several DD34E transposons discovered in An. gambiae possess characteristics that are associated with recent transposition, such as high sequence identity between copies, and intact terminal-inverted repeats and open reading frames. One such element, AgTango, was also found in a distantly related mosquito species, Aedes aegypti, at high amino acid sequence identity (79.9%). It was discovered that Tango transposons have patchy distribution among twelve mosquito species surveyed using PCR as well as genomic searches, suggesting a possible case for horizontal transfer. Additionally, it was discovered that in some mosquito genomes, there are several Tango transposons. These observations suggest differential evolutionary scenarios and/or TE-host interaction of Tango elements between mosquito species. This strengthened the case that AgTango may be a functional transposase, and I sought to test its potential activity in a cell culture-based inter-plasmid transposition assay using the Herves plasmids as a positive control (Arensburger et al., 2005). AgTango constructs were successfully constructed; however, no transposition events were detected for Tango or Herves. Because the positive control failed to work, no assessment can be made concerning Tango's transposase. Possible causes and solutions for these results, alternative means to detect transposition, as well as future directions with Tango are discussed.
Ph. D.

Cílem práce je seznámení se s problematikou uchovávání informace v DNA, provést rešerši na téma transpozony, bioinformatické nástroje a algoritmy, které jsou používány k jejich detekci v nasekvenovaných genomech a vytvořit tak stručný úvod do obsáhle problematiky, včetně jejího zasazení do kontextu současně probíhajícího výzkumu v dané oblasti. Na základě přehledu stávajících algoritmů a nástrojů pro detekci transpozonů je navržen a implementován nástroj pro hledání tzv. LTR transpozonů.

In this thesis we investigate the mappability of human genome and we look at some reasons that might cause unmappability in a region. We look at transposable elements and genome duplications as the main reasons for unmappability. In this analysis we simulated singled end, paired end and mate paired reads of 6 different lengths and we used BWA to map these simulated reads to the human genome. We assumed that a position in the genome is mappable if there is at least one unique read mapped to that position. We looked at unmappable regions and fraction of transposable elements or genome duplications corresponding to these regions. We also looked at age distribution of transposable elements and genome duplications that are in unmappable regions. Our results shows that regions that are in younger and longer transposable elements are harder to sequence. In order to compare our simulated data with a real sequencing data, we used the output of a sequencing from Illumina to compare coverage of genome in this real data set with our mappability results. We show that 4.1% of genome that is mappable in our simulations result, has low coverage in real sequencing data. We also investigated the reasons behind having low coverage in mappable regions. Our simulation result shows the impact of transposable elements and other repeats on mappability in the human genome and we show that using longer paired end and mate paired reads improves the mappability of the human genome.
Dans cette thèse, nous étudions la "visibilité" du génome humain par des méthodes séquençage modernes et nous regardons quelles sont les raisons qui pourraient causer l'absence de visibilité dans une région donnée. Nous montrons que les éléments transposables et les duplications de génome sont les principaux obstables à la visibilité de régions génomiques. Dans cette analyse, nous avons utilisé des reads simulés, de types individuels ou pairés, de 6 longueurs différentes et nous avons utilisé BWA pour assigner ces reads au génome humain. Nous avons supposé que la position dans le génome est visible s'il y a au moins un read unique assigné à cette position. Nous avons examiné les régions non visibles et la fraction d'éléments transposables ou des duplications de génome correspondant à ces régions. Nous avons également examiné la distribution d'âge des éléments transposables et des duplications de génome qui sont dans les régions non visibles. Nos résultats montrent que les régions qui sont des éléments plus jeunes et plus transposable sont plus difficiles à séquencer. Afin de comparer nos données simulées avec les données réelles de séquençage, nous avons utilisé des données de reséquençage provenant d'un séquençage Illumina pour comparer la couverture observée du génome avec nos résultats provenant de données simulées. Nous montrons que 4,1% du génome qui est visible dans nos simulations a une faible couverture dans les données de séquençage réelles. Nous avons également étudié les raisons pouvant expliquer une faible couverture dans les régions visibles. Les résultats de nos simulations montrent l'impact des éléments transposables et les autres répétitions sur la visibilité dans le génome humain et nous montrent que l'utilisation de long reads pairés améliorent la visibilité du génome humain.

The different genomic environments in which transposable elements reside in the Great Apes and the Drosophila result in substantial differences between the evolution of transposable elements in these two groups of organisms. In the Great Apes, where deletion of transposable elements is relatively rare, elements tend to be retained in the genome to the extent that complete sets of elements belonging to a particular transposable element family can be obtained. In Drosophila, there is a rapid turnover of transposable elements, imposing strong selection pressure on transposable elements to be able to infect new hosts. This study investigates the evolution of transposable elements in these two genomic environments. Complete sets of elements belonging to young Alu subfamilies in humans and closely-related species are used to investigate factors involved in their evolutionary history, such as mutation and gene conversion. The application of the master gene model, and other proposed models of the proliferation of young Alu subfamilies, are considered in light of the results obtained. The evolution of the AluYg, Yh and Yi lineages are investigated using a C++ program to simulate their evolutionary history. The results of the simulations are compared to statistics such as theta and pi, as well as the number of shared mutations and the proliferation time, in order to determine possible, and likely, values for parameters such as the retrotransposition rate and the number of source elements for each subfamily. The results suggest that although the master gene model may apply to some lineages, it is not the best model to explain the evolutionary history of all young Alu subfamilies. The selection pressure on transposable elements in Drosophila results in a high level of horizontal transfer of these elements among species of the Drosophila genus. In this study, the twelve sequenced Drosophila genomes are used to investigate the frequency of horizontal transfer within these twelve species using a large dataset of transposable element sequences from the DNA transposons, as well as LTR, and non-LTR, retrotransposons. Horizontal transfer is inferred where identity between transposable elements of the same family in different species exceeds that between the coding regions of the Adh gene in the relevant species. Cases are further supported by evidence from the distribution of the transposable element family across the Drosophila genus, and phylogenetic incongruence, which in many cases elucidates likely directions of transfer. The results suggest that horizontal transfer may be even more common than previously thought, and appears to be most common for the LTR retrotransposons. The possibility that possession of the env gene may result in higher rates of horizontal transfer of LTR retrotransposons is investigated, and the env open reading frame is found to be under selective constraint.

Los transposones son elementos genéticos que tienen la capacidad de modificar su posición en el genoma. Como consecuencia, tienen un impacto en la evolución de los genomas inactivando o alterando los genes del huésped y proporcionando nuevas funciones génicas. Los transposones ocupan una fracción importante en todos los genomas resecuenciados. El objetivo del trabajo presentado en esta tesis trata en estudiar los distintos impactos de transposones tanto en genes como en la evolución de los genomas de distintas especies de plantas. En esta tesis, se ha analizado la fracción de transposones en melón y pepino, dos especies muy cercanas. Los resultados sugieren que los transposones han proliferado más en melón, causando un aumento del tamaño del genoma. Los transposones no se encuentran distribuidos habitualmente de forma homogénea y tienden a acumularse en las regiones pericentroméricas heterocromáticas, como es el caso de los genomas de melón y pepino. Curiosamente, los resultados presentados muestran que los transposones han expandido las regiones pericentroméricas en melón, demostrando que los transposones pueden modificar la estructura de los genomas. El número de genomas de referencia de plantas disponibles y el número de variedades resecuenciadas ha crecido exponencialmente permitiendo estudiar la correlación entre las variaciones genéticas y fenotípicas. El propósito del trabajo resumido en la segunda parte de esta tesis consiste en analizar el impacto de los transposones en genomas de especies cultivables, detectando los polimorfismos ocasionados por la presencia o ausencia de transposones en un locus concreto, a través de la comparación de una variedad resecuenciada respecto al genoma de referencia. El análisis de inserciones polimórficas de transposones se ha realizado en tres especies distintas: melón, palmera datilera y Physcomitrella patens. Los resultados obtenidos pueden ayudar a identificar familias de transposones activas recientemente y proporcionar información nueva sobre polimorfismos genéticos que pueden estar ligados a caracteres seleccionados durante la evolución reciente de estas tres especies. Para estudiar el impacto de la transposición en la regulación génica, el trabajo presentado en la tercera parte de esta tesis se centra en la capacidad de los transposones en amplificar y redistribuir sitios de unión a factores de transcripción. Los resultados muestran que algunas familias de MITEs se han amplificado y redistribuido los sitios de unión del factor de transcripción E2F durante la evolución de algunas especies del género Brassica. El objetivo de este trabajo ha sido evaluar el impacto de los sitios de unión a E2F localizados dentro de transposones reprogramando la regulación de genes en la red transcripcional de E2F. Los resultados obtenidos han determinado que los sitios de unión a E2F localizados dentro de transposones tienen capacidad de unirse a los factores de transcripción de E2F in vivo, independientemente de las marcas epigenéticas en la región. Además, los transposones se utilizan como herramienta genética útil para generar colecciones de mutantes en animales y plantas debido a su capacidad de integrar copias en el genoma. En plantas, algunos retrotransposones se integran preferentemente cerca de genes siendo particularmente interesantes para la mutagénesis. De entre todos, el retrotransposón de tabaco Tnt1 se ha utilizado para generar mutantes en distintas especies de plantas. La última parte de esta tesis consiste en analizar la capacidad del retrotransposón de tabaco Tnt1 en transponer en el musgo Physcomitrella patens, ya que se demostró que Tnt1 transpone eficientemente en P. patens y se integra preferentemente cerca de genes. Finalmente, este estudio presenta vectores derivados de Tnt1 diseñados para transponer con alta eficiencia y ser utilizados para generar colecciones de mutantes con inserciones estables en esta especie briofita.
Els transposons són elements genètics que tenen la capacitat de modificar la seva posició dins el genoma. Com a conseqüència, tenen un impacte en l’evolució del genomes inactivant o alterant els gens de l’hoste i proporcionant noves funcions gèniques. Els transposons ocupen una fracció important de tots els genomes seqüenciats. L’objectiu del treball presentat en aquesta tesis consisteix en estudiar els diversos impactes de transposons tant en els gens com en l’evolució dels genomes de diferents espècies de plantes. En aquesta tesis, s’ha analitzat la fracció de transposons en meló i cogombre, dues espècies molt properes. Els resultats suggereixen que els transposons han proliferat més en meló, causant un augment de la mida del genoma. Els transposons no es troben distribuïts habitualment de forma homogènia i tendeixen a acumular-se en les regions pericentromèriques heterocromàtiques, com el cas dels genomes de meló i cogombre. Curiosament, els resultats presentats mostren que els transposons han expandit les regions pericentromèriques en meló, demostrant que els transposons poden modificar l’estructura dels genomes. El número de genomes de referència de plantes disponibles i el número de varietats reseqüenciades ha crescut exponencialment permetent estudiar la correlació entre les variacions genètiques i fenotípiques. El propòsit del treball resumit en la segona part d’aquesta tesis consisteix en analitzar l’impacte dels transposons en genomes d’espècies cultivables detectant els polimorfismes deguts a la presència o absència de transposó en un locus concret, comparant una varietat reseqüenciada respecte al seu genoma de referència. L’anàlisi d’insercions polimòrfiques de transposons s’ha realitzat en tres espècies diferents: meló, palmera datilera i Physcomitrella patens. Els resultats obtinguts poden ajudar a identificar famílies de transposons actives recentment i proporcionar informació nova sobre polimorfismes genètics que poden estar lligats a caràcters seleccionats durant l’evolució recent d’aquestes tres espècies. Per tal d’estudiar l’impacte de la transposició en la regulació gènica, el treball presentat en la tercera part d’aquesta tesis se centra en la capacitat dels transposons en amplificar i redistribuir llocs d’unió a factors de transcripció. Els resultats mostren que algunes famílies de MITEs s’han amplificat i han redistribuït els llocs d’unió del factor de transcripció E2F durant l’evolució d’algunes espècies del gènere Brassica. L’objectiu d’aquest treball és avaluar l’impacte dels llocs d’unió a E2F localitzats dins de transposons reprogramant la regulació de gens de la xarxa transcripcional de E2F. Els resultats obtinguts han determinat que els llocs d’unió a E2F localitzats dins de transposons tenen la capacitat d’unir-se als factors de transcripció de E2F in vivo, independentment de les marques epigenètiques de la regió. A més a més, els transposons s’han convertit en eines genètiques útils per generar col·leccions de mutants en animals i plantes degut a la seva capacitat d’integrar còpies en el genoma. En plantes, alguns retrotransposons s’integren preferentment a prop de gens sent particularment interessants per la mutagènesis. Entre tots ells, el retrotransposó de tabac Tnt1 s’ha utilitzat per generar mutants en diferents espècies de plantes. L’última part d’aquesta tesis consisteix en analitzar la capacitat del retrotransposó de tabac Tnt1 en transposar en la molsa Physcomitrella patens. S’ha demostrat que Tnt1 transposa eficientment en P. patens i s’integra preferentment a prop de gens. Aquest estudi presenta vectors derivats de Tnt1 dissenyats per transposar amb alta eficiència i ser utilitzats per generar col·leccions de mutants amb insercions estables en aquest briòfit.
Transposable elements are genetic elements that have the capacity to modify their position within the genome. As a consequence, they impact the evolution of genomes by inactivating or altering host genes and by providing new gene functions. Transposons account for an important fraction of all sequenced genomes. The goal of the work presented in this dissertation is to investigate the diverse impacts of transposons on gene and genome evolution in different plant species. The transposon content has been analyzed in melon and cucumber, two closely related species. The results suggest that transposons have proliferated to a greater extend in melon, causing an increase of its genome size. Transposable elements are usually not homogenously distributed and tend to accumulate in heterochromatic pericentromeric regions. This is also the case of melon and cucumber genomes. Interestingly, the results presented show that transposons have expanded the pericentromeric regions in melon, showing that transposons can modify the structure of genomes. The number of plant reference genomes made available and the number of varieties resequenced is growing exponentially, and this is allowing to study the correlation between genetic and phenotypic variations. The purpose of the work summarized in the second part of this dissertation is to analyze the impact of transposons in crop genomes by detecting polymorphisms due to the presence or absence of transposon at a given locus, comparing one resequenced variety respect to the reference genome. The analysis of transposon-related polymorphism insertions has been performed in three different species: melon, date palm and Physcomitrella patens. The results obtained can help to identify the transposon families recently active and to provide new information on genetic polymorphisms that can be linked to traits selected during the recent evolution of these three species. In order to study the impact of transposition on gene regulation, the work reported in the third part of this dissertation focuses on the capacity of transposons to amplify and redistribute transcription factor binding sites. The results show that some MITE families have amplified and redistributed the binding sites of E2F transcription factor during Brassica evolution. The goal of this study was to assess the impact of E2F binding sites located within a transposon in reprogramming gene regulation on the E2F transcriptional network. The results obtained have determined that E2F binding sites located within transposons have the capacity to bind E2F transcription factor in vivo, regardless the epigenetic mark context. Moreover, transposons have become a useful genetic tool to generate mutant collections in animals and plants due to the capacity to insert copies into the genome. In plants, some retrotransposons have been shown to integrate preferentially near genes making them particularly interesting for mutagenesis. Among them, the tobacco retrotransposon Tnt1 has been used to generate mutants in different plant species. The last part of this dissertation consists in analyzing the capacity of the tobacco retrotransposon Tnt1 to transpose in the moss Physcomitrella patens. It shows that Tnt1 efficiently transposes in P. patens and inserts preferentially in genic regions. This work presents Tnt1-derived vectors designed for high efficiency transposition that could be used to generate stable insertion mutant collections in this bryophyte species.

Thesis (Ph.D.)--Biology, Georgia Institute of Technology, 2008.
Committee Chair: Jordan, I. King; Committee Member: Borodovsky, Mark; Committee Member: Bunimovich, Leonid; Committee Member: Choi, Jung; Committee Member: McDonald, John.

BACKGROUND: Comparative evolutionary analysis of whole genomes requires not only accurate annotation of gene space, but also proper annotation of the repetitive fraction which is often the largest component of most if not all genomes larger than 50 kb in size. RESULTS: Here we present the Rice TE database (RiTE-db) - a genus-wide collection of transposable elements and repeated sequences across 11 diploid species of the genus Oryza and the closely-related out-group Leersia perrieri. The database consists of more than 170,000 entries divided into three main types: (i) a classified and curated set of publicly-available repeated sequences, (ii) a set of consensus assemblies of highly-repetitive sequences obtained from genome sequencing surveys of 12 species; and (iii) a set of full-length TEs, identified and extracted from 12 whole genome assemblies. CONCLUSIONS: This is the first report of a repeat dataset that spans the majority of repeat variability within an entire genus, and one that includes complete elements as well as unassembled repeats. The database allows sequence browsing, downloading, and similarity searches. Because of the strategy adopted, the RiTE-db opens a new path to unprecedented direct comparative studies that span the entire nuclear repeat content of 15 million years of Oryza diversity.

Background: Mutator-like transposable elements, a class of DNA transposons, exist pervasively in both prokaryotic and eukaryotic genomes, with more than 10,000 copies identified in the rice genome. These elements can capture ectopic genomic sequences that lead to the formation of new gene structures. Here, based on whole-genome comparative analyses, we comprehensively investigated processes and mechanisms of the evolution of putative genes derived from Mutator-like transposable elements in ten Oryza species and the outgroup Leersia perieri, bridging similar to 20 million years of evolutionary history. Results: Our analysis identified thousands of putative genes in each of the Oryza species, a large proportion of which have evidence of expression and contain chimeric structures. Consistent with previous reports, we observe that the putative Mutator-like transposable element-derived genes are generally GC-rich and mainly derive from GC-rich parental sequences. Furthermore, we determine that Mutator-like transposable elements capture parental sequences preferentially from genomic regions with low methylation levels and high recombination rates. We explicitly show that methylation levels in the internal and terminated inverted repeat regions of these elements, which might be directed by the 24-nucleotide small RNA-mediated pathway, are different and change dynamically over evolutionary time. Lastly, we demonstrate that putative genes derived from Mutator-like transposable elements tend to be expressed in mature pollen, which have undergone de-methylation programming, thereby providing a permissive expression environment for newly formed/transposable element-derived genes. Conclusions: Our results suggest that DNA methylation may be a primary mechanism to facilitate the origination, survival, and regulation of genes derived from Mutator-like transposable elements, thus contributing to the evolution of gene innovation and novelty in plant genomes.

Ma thèse vise à caractériser l’évolution dynamique et l’organisation des génomes des différentes espèces du blé (genres Triticum et Aegilops) en relation avec la prolifération des éléments transposables (TEs) dans leur génome (>80%), les polyploïdisations récurrentes ainsi que la syntenie avec d’autres espèces de la famille des Poaceae. En constituant des sets de séquences génomiques représentatives et en étudiant la variabilité entre des haplotypes des génomes du blé, j’ai caractérisé la dynamique et la prolifération différentielle des TEs qui est la résultante de l’équilibre entre leurs insertions et aussi leurs éliminations actives. Le taux moyen de remplacement de l’espace TEs, mesurant les différences de séquences dues aux insertions et aux délétions entre deux haplotypes, a été ainsi estimé à 86% par million d’années (Ma) et dépasse celles bien documentées du maïs. Les insertions des TEs mais aussi leurs éliminations par recombinaisons illégitimes de l’ADN (pouvant atteindre plusieurs dizaines de kb) ainsi que les recombinaisons génétiques entre haplotypes divergents représentent les principaux mécanismes à la base des changements rapides de l’espace TEs. Sur une échelle d’évolution plus longue (60 Ma), j’ai analysé la conservation des gènes et l’évolution du locus (Ha) entre différentes espèces des Poaceae. J’ai pu ainsi préciser l’émergence du caractère grain tendre et des gènes Ha, comme nouveaux membres de la famille des gènes de Prolamine, dans l’ancêtre commun des Pooideae (blé et Brachypodium, de la tribu des Triticeae et des Brachypodieae) et des Ehrhartoideae (riz), après leur divergence des Panicoideae (maïs, sorgho)
My PhD aims to characterize dynamic evolution and organization of wheat genomes from différent species (Triticum and Aegilops genera) in relation to transposable element (TE) proliferation in their genomes (>80%), polyploidizations and synteny with other Poaceae species. By constituting and comparing representative genomic sequences and analyzing haplotype variability of the wheat genomes, I have characterized dynamics and differential proliferation of TEs, as resulting from the combinations of their insertions and deletions. Mean replacement rate of the TE space, which measures sequence differences due to insertion and removal of TEs between two haplotypes, was estimated to 86% per one million year (My). This is more important than the well-documented haplotype variability found in maize. It was observed that TE insertions and DNA elimination by illegitimate recombination (implicating several ‘tens’ of kb) as well as homologous recombination between divergent haplotypes represent the main molecular basis for rapid change of the TE space. At a longer evolutionary scale (60 My), I have compared gene conservation at the Ha locus region between different Poaceae species. The comparative genome analysis and evolutionary comparison with genes encoding grain reserve proteins of grasses suggest that an ancestral Ha-like gene emerged, as a new member of the Prolamin gene family, in a common ancestor of the Pooideae (wheat and Brachypodium from the Triticeae and Brachypodieae tribes) and Ehrhartoideae (rice), between 60 and 50 My, after their divergence from Panicoideae (Sorghum)

We have studied the regulation of the maize Ac and MuDR transposable elements activities during plant development. Ac was studied in an heterologous system (transgenic tobacco plants and cell suspensions) while MuDR was studied in the native maize background. The focus of this study was on the transcriptional regulation of Ac and MuDR. For Ac, the major achievements were to show that 1-It is autoregulated in a way that the Ac-encoded transposase can repress the activity of its own promoter; 2-It is expressed at low basal level in all the plant organs that were studied, and its activity is stronger in dividing tissues -- a behaviour reminiscent of housekeeping genes; 3- the activity of Ac promoter is cell cycle regulated -- induced at early S-phase and increasing until mitosis; 4- host factor binding sites were identified at both extremities of Ac and may be important for transposition. For MuDR, It was shown that it encodes two genes, mudrA and mudrB, convergently transcribed from near-identical promoters in the terminal inverted repeats. Distinct 5' start sites, alternative splicing, production of antisense RNA and tissue specificity were all shown to be involved in the regulation of MuDR.

The objectives of the project were to develop expression vectors using the Sleeping Beauty transposon technology and the genetic border elements to provide both enhanced integration rate and stable transgene expression, and to evaluate the application of such vectors in farmed fish such as catfish and carp. The panel recommended adding the objective of evaluating the endogenous transposable elements, particularly in catfish, in order to evaluate the applicability of the expression vectors while reduc1ng efforts in real production of transgenic fish considering the focus of the project was to develop the vector and evaluation of its applicability, not producing transgenic fish. Efficient production of transgenic farmed fish is hindered by two major problems: mosaicism due to delayed integration after single-cell stage, and silencing of transgene expression. In this project, we proposed to combat these problems by coupling the Sleeping Beauty transposon technology that can enhance integration rate and the border elements that can insulate transgene from position effect. Our major objective was to develop a new generation of expression vector that contains both of these elements. We have developed expression vectors containing both the Sleeping Beauty transposon signals, inverted repeats and direct repeats (IR and DR, respectively), and the border elements, scs and scs'. Growth hormone minigene has been cloned into this vector for applications of such vectors in growth enhancement. Luc reporter gene has been also cloned into this vector cascades for relative easy evaluation of transgene expression. Transgenic fish have been produced using these expression vectors in both catfish (US) and carp (Israel). Much effort was also devoted to evaluation of the endogenous transposable elements in catfish as recommended by the BARD grant panel. Multiple families of Tcl-like transposons were identified from catfish. Surprisingly, many Tc I-related transcripts were identified. Among these transcripts, both the sense and antisense transcripts were present. Some of the transcripts may be useful for development of novel transposase-based technology for aquaculture applications in the future. This project has both scientific and aquaculture implications. First, to develop expression vectors containing both IR/DR and scs/scs' repeated elements have been reported being extremely technically difficult due to excision of the repeated sequences by the E. coli host during cloning processes. We have successfully constructed this advanced vector that contained very complex cascades for both gene integration and gene regulation. We have produced transgenic fish using such vectors. This advanced expression vector should be useful for production of transgenic fish. By simply replacing the growth hormone gene, any gene of interest can be readily inserted in this vector. Thus this vector should provide technological possibility for early integration and stable expression of any economically important genes in aquaculture. We have also evaluated the applications of the Sleeping Beauty-based vectors in terms of the impact of gene size and found that the size of trans gene drastically affects transposition. The system will be only useful for transferring genes smaller than 5.6 kb. We have also identified novel transposase-related transcripts that may be useful for the development of novel transposase-based technologies for general scientific research and for aquaculture applications.