Danielle Nouaud

Combined evidence annotation of transposable elements in genome sequences

Transposable elements (TEs) are mobile, repetitive sequences that make up significant fractions of metazoan genomes. Despite their near ubiquity and importance in genome and chromosome biology, most efforts to annotate TEs in genome sequences rely on the results of a single computational program, RepeatMasker. In contrast, recent advances in gene annotation indicate that high-quality gene models can be produced from combining multiple independent sources of computational evidence. To elevate the quality of TE annotations to a level comparable to that of gene models, we have developed a combined evidence-model TE annotation pipeline, analogous to systems used for gene annotation, by integrating results from multiple homology-based and de novo TE identification methods. As proof of principle, we have annotated “TE models” in Drosophila melanogaster Release 4 genomic sequences using the combined computational evidence derived from RepeatMasker, BLASTER, TBLASTX, all-by-all BLASTN, RECON, TE-HMM and the previous Release 3.1 annotation. Our system is designed for use with the Apollo genome annotation tool, allowing automatic results to be curated manually to produce reliable annotations. The euchromatic TE fraction of D. melanogaster is now estimated at 5.3% (cf. 3.86% in Release 3.1), and we found a substantially higher number of TEs (n = 6,013) than previously identified (n = 1,572). Most of the new TEs derive from small fragments of a few hundred nucleotides long and highly abundant families not previously annotated (e.g., INE-1). We also estimated that 518 TE copies (8.6%) are inserted into at least one other TE, forming a nest of elements. The pipeline allows rapid and thorough annotation of even the most complex TE models, including highly deleted and/or nested elements such as those often found in heterochromatic sequences. Our pipeline can be easily adapted to other genome sequences, such as those of the D. melanogaster heterochromatin or other species in the genus Drosophila.

Molecular domestication — more than a sporadic episode in evolution

Transposable elements are short but complex pieces of DNA or RNA containing a streamlined minimal-genome with the capacity for its selfish replication in a foreign genomic environment. Cis-regulatory sections within the elements orchestrate tempo and mode of TE expression. Proteins encoded by TEs mainly direct their own propagation within the genome by recruitment of host-encoded factors. On the other hand, TE-encoded proteins harbor a very attractive repertoire of functional abilities for a cell. These proteins mediate excision, replication and integration of defined DNA fragments. Furthermore, some of these proteins are able to manipulate important host factors by altering their original function. Thus, if the host genome succeeds in domesticating such TE-encoded proteins by taming their ‘anarchistic behavior,’ such an event can be considered as an important evolutionary innovation for its own benefit. In fact, the domestication of TE-derived cis-regulatory modules and protein coding sections took place repeatedly in the course of genome evolution. We will present prominent cases that impressively demonstrate the beneficial impact of TEs on host biology over evolutionary time. Furthermore, we will propose that molecular domestication might be considered as a resumption of the same evolutionary process that drove the transition from ‘primitive genomes’ to ‘modern’ ones at the early dawn of life, that is, the adaptive integration of a short piece of autonomous DNA into a complex regulatory network.

Detection of new transposable element families in Drosophila melanogaster and Anopheles gambiae genomes.

The techniques that are usually used to detect transposable elements (TEs) in nucleic acid sequences rely on sequence similarity with previously characterized elements. However, these methods are likely to miss many elements in various organisms. We tested two strategies for the detection of unknown elements. The first, which we call “TBLASTX strategy,” searches for TE sequences by comparing the six-frame translations of the nucleic acid sequences of known TEs with the genomic sequence of interest. The second, “repeat-based strategy,” searches genomic sequences for long repeats and clusters them in groups of similar sequences. TE copies from a given family are expected to cluster together. We tested the Drosophila melanogaster genomic sequence and the recently sequenced Anopheles gambiae genome in which most TEs remain unknown. We showed that the “TBLASTX strategy” is very efficient as it detected at least 332 new TE families in D. melanogaster and 400 in A. gambiae. This was unexpected in Drosophila as TEs of this organism have been extensively studied. The “repeat-based strategy” appeared to be very inefficient because of two problems: (i) TE copies are heavily deleted and few copies share homologous regions, and (ii) segmental duplications are frequent and it is not easy to distinguish them from TE copies.

The failure to obtain sexual isolation by artificial selection

When strains of Drosophila melanogaster derived from Japanese and French natural populations were tested for mating preferences, one of three patterns was observed: panmictic, homogamic or selective mating. An attempt was made to select for increased sexual isolation between three pairs of the panmictic strains and four pairs demonstrating isolation or selection. None succeeded. However, an analysis of male and female selection points out no departure from panmixia in the panmictic strains and a selective mating tendency during the first 10 generations of selection in strains demonstrating isolation or selection. We hypothesize that the number of genes controlling sexual selection or isolation is small and that polymorphism for these genes varies from one strain to another.

Éléments transposables et nouveautés génétiques chez les eucaryotes

Les elements transposables sont des sequences d’ADN capables de se deplacer d’un site chromosomique a un autre. Ils se repartissent en deux groupes principaux sur la base de leur mecanisme de transposition. Les elements de classe I, ou retroelements, transposent via la transcription inverse d’un intermediaire ARN, tandis que ceux de classe II transposent directement d’un site chromosomique a un autre grâce a la transposase, une enzyme codee par l’element. Une fraction tres importante des genomes eucaryotes est constituee d’ADN mobile. Par exemple, 10 a 15 % du genome est constitue d’elements transposables chez Drosophila melanogaster, plus de 50 % chez le mais, et on estime que plus de 40 % du genome humain pourrait deriver d’elements transposables [1]. L’interet porte aux elements transposables se focalise sur deux points principaux : l’evolution des elements transposables eux-memes et leur impact sur le genome des hotes. Au cours des dernieres annees, nous avons oriente nos recherches sur ce deuxieme aspect, c’est-a-dire sur le role des elements transposables dans l’evolution de la structure et du fonctionnement du genome de l’hote. Cette demarche est plutot neuve car, il y a peu, les elements transposables etaient consideres comme de l’ADN essentiellement egoiste [2]. Cette derniere vision reste valable si l’on considere une courte fenetre de temps puisque les elements transposables se maintiennent dans une espece grâce a un equilibre entre transposition et elimination, sans apporter aucun benefice adaptatif a leur hote. Cependant, en raison de leur perennite sur de longues periodes evolutives, les elements transposables peuvent etre a l’origine de mutations qui peuvent avoir des consequences majeures sur l’evolution du genome de l’hote [3]. Par exemple, la comparaison des genomes bacteriens met en evidence que de nombreux rearrangements chromosomiques sont associes a des systemes geniques formellement equivalents a des elements genetiques mobiles : les complexes de genes restriction-modifications [4]. Dans ces complexes, le gene d’une enzyme de restriction est souvent physiquement associe a un gene codant une methylase qui modifie les sites cibles reconnus par l’enzyme, les protegeant ainsi de la coupure : le duo endonuclasemethylase est formellement equivalent au duo transposase-regulateur d’un element transposable. Chez les eucaryotes, il existe aujourd’hui de nombreux exemples qui demontrent aussi que des changements de structure ou de fonctionnement du genome sont associes a des elements transposables ou a des sequences derivees d’anciens elements transposables (pour revue, voir [5]). Par ailleurs, de nombreuses observations montrent que les elements transposables peuvent etre inactives par des mecanismes epigenetiques tel que la methylation, l’heterochromatinisation et la co-suppression [68]. Ces decouvertes suggerent que, chez les eucaryotes, les elements transposables pourraient etre consideres comme des agents favorisant la mise en place de certains mecanismes de regulation epigenetique Elements transposables et nouveautes genetiques chez les eucaryotes