Elsevier

Gene

Volume 440, Issues 1–2, 1 July 2009, Pages 42-49
Gene

Identification of novel LTR retrotransposons in the genome of Aedes aegypti

https://doi.org/10.1016/j.gene.2009.03.021Get rights and content

Abstract

We have detected seventy-six novel LTR retrotransposons in the genome of the mosquito Aedes aegypti by a genome wide analysis using the LTR_STRUC program. We have performed a phylogenetic classification of these novel elements and a distribution analysis in the genome of A. aegypti. These mobile elements belong either to the Ty3/gypsy or to the Bel family of retrotransposons and were not annotated in the mosquito LTR retrotransposon database (TEfam). We have found that ∼ 1.8% of the genome is occupied by these newly detected retrotransposons that are distributed predominantly in intergenic genomic sequences and introns. The potential role of retrotransposon insertions linked to host genes is described and discussed. We show that a retrotransposon family belonging to the Osvaldo lineage has peculiar structural features, and its presence is likely to be restricted to the A. aegypti and to the Culex pipiens quinquefasciatus genomes. Furthermore we show that the ninja-like group of elements lacks the Primer Binding Site (PBS) sequence necessary for the replication of retrotransposons. These results integrate the knowledge on the complicate genomic structure of an important disease vector.

Introduction

Eukaryotic genomes are largely composed of transposable elements (TE). These elements are classified in two main classes (class I and class II) according their transposition mechanisms (reviewed in Finnegan, 1992). Class II elements are characterized by DNA to DNA transposition using of a self encoded transposase. Class I elements use an RNA intermediate, which is reverse transcribed into cDNA molecules and then inserted in the genome. Class I elements can be further categorized in LTR- and non-LTR retrotransposons depending on the presence or absence of terminal direct repeats. Completely sequenced genomes facilitate the characterization of the full transposon complement in a genome. This is possible both with a sequence similarity search analyses (extrinsic methods) using characterized mobile elements from related model organisms as query and with the development of in silico methods that focus on the structure (intrinsic methods) of TEs rather than the sequence similarity. The latter methods allow a faster identification of mobile elements that have a low sequence similarity with respect to reference elements. This strategy has been successfully applied to the identification of L1 insertions in the human genome (Szak et al., 2002), LTR retrotransposons insertions in A. gambiae (Marsano and Caizzi, 2005) and Mus musculus genomes (McCarty and McDonald, 2004), and MITEs (Miniature Inverted repeat Transposable Elements) in the A. gambiae genome (Tu, 2001).

Aedes aegypti is the primary mosquito vector responsible for the transmission of both the yellow fever and dengue viruses. Recently Nene et al. (2007) have revealed that nearly 50% of its genome consists of transposable elements. LTR retrotransposons built up about 10.5% of the A. aegypti genome. Furthermore an extensive compilation of mobile elements has been reported and a relational database called TEfam (http://tefam.biochem.vt.edu/tefam) was released. Here the sequences of more than one thousand of mosquitoes TE families have been annotated. More than 800 families of the TEs reported in the TEfam database are related to A. aegypti retrotransposons and 642 belong either to the Ty3/gypsy (179 elements), Ty5/copia (233 elements) or Bel/Pao (230 elements) families. In addition, six distinct phylogenetic lineages can be recognized within the Ty3/gypsy family (namely the gypsy lineage (21 elements), Mag lineage (64 elements), CsRn1 lineage (15 elements), mdg1 lineage (26 elements), Osvaldo lineage (30 elements) and mdg3 lineage (23 elements).

The massive presence of transposable elements in the genome of A. aegypti is consistent with two observations. First, A. aegypti genome is 4-fold larger than A. gambiae genome: this must be taken into account when studying repetitive sequences from A. aegypti. Second, A. aegypti's introns are on average longer than introns of related species due to the presence of transposable elements (Nene et al., 2007).

Here we report 76 additional LTR retrotransposon elements in the genome of A. aegypti, identified using the LTR_STRUC program (McCarthy and McDonald, 2003). We have performed classification on the basis of evolutionary relationships with other LTR retrotransposons. We have also analyzed the structure and the genomic distribution of the new elements detected. A novel family belonging to the Osvaldo lineage with unexpected structural features has been identified. Furthermore all members of the ninja group identified in this study lack a discrete PBS sequence (Primer Binding Site). The results of the genomic distribution analysis are consistent with the presence of retrotransposons preferentially in intergenic regions of the genome of A. aegypti or in intron sequences. The possible functional role of some insertions on the host gene organization is also discussed.

Section snippets

LTR_STRUC analysis and classification of LTR retrotransposons

The entire genome of A. aegypti was downloaded from the Broad Institute website (http://www.broad.mit.edu/index.html) and scanned with the LTR_STRUC program (McCarthy and McDonald, 2003) using the default parameters. 4026 putative retrotransposon sequences obtained as output were subjected to an “all against all” BLAST in order to group sequences with % identity greater than 98% over a sequence of at least 1 Kb. Two hundred and seventeen groups (containing at least 2 sequences) and 359 singlets

Results

We screened the A. aegypti genome with the LTR_STRUC program (McCarthy and McDonald, 2003), we have obtained 4026 putative retrotransposon sequences as output. These sequences were arranged in more than 200 groups of sequences sharing 100% identity over at least 99% of the sequence alignment. For each group we chose one representative sequence potentially able to encode protein domains of retrotransposons (GAG PRO RT RH INT). These sequences were used to probe the TEfam and the REPBASE

Discussion

The genome of A. aegypti is particularly rich in transposable elements and this abundance has probably masked the presence of several elements in the initial genomic analyses (Nene et al., 2007). The genomic sequence of A. aegypti is continuously updated which means that other transposable element sequences are probably to be discovered and characterized. Moreover the overall organization of the A. aegypti genome into euchromatin and heterochromatin is poorly understood at the molecular level;

Conclusions

We think that the results obtained integrate the already large amount of data concerning the mobile elements of A. aegypti. At the same time this is an example of how difficult can be the identification of the complete TE repertoire in a eukaryotic genome. The identification of the complete transposon set in a genome is otherwise essential to understand the evolution and the expression of a genome.

Other transposable elements are likely to be identified as the A. aegypti genome will become

Acknowledgments

This work was supported by Istituto Superiore di Sanita` (contract no. OAD/F2) to R. C.

We thank Konstantinos Lefkimmiatis for critical reading and English improvement of the manuscript, Pietro D'Addabbo for helping us with the RepeatMasker program and for computer assistance. We would also like to thank the anonymous reviewers of the previous version of the paper for their useful comments.

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