The Juan non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity

Abstract

Background: In contrast to DNA-mediated transposable elements (TEs), retrotransposons, particularly non-long terminal repeat retrotransposons (non-LTRs), are generally considered to have a much lower propensity towards horizontal transfer. Detailed studies on site-specific non-LTR families have demonstrated strict vertical transmission. More studies are needed with non-site-specific non-LTR families to determine whether strict vertical transmission is a phenomenon related to site specificity or a more general characteristic of all non-LTRs. Juan is a Jockey clade non-LTR retrotransposon first discovered in mosquitoes that is widely distributed in the mosquito family Culicidae. Being a non-site specific non-LTR, Juan offers an opportunity to further investigate the hypothesis that non-LTRs are genomic elements that are primarily vertically transmitted.

Results: Systematic analysis of the ~1.3 Gbp Aedes aegypti (Ae. aegypti) genome sequence suggests that Juan-A is the only Juan-type non-LTR in Aedes aegypti. Juan-A is highly reiterated and comprises approximately 3% of the genome. Using minimum cutoffs of 90% length and 70% nucleotide (nt) identity, 663 copies were found by BLAST using the published Juan-A sequence as the query. All 663 copies are at least 95% identical to Juan-A, while 378 of these copies are 99% identical to Juan-A, indicating that the Juan-A family has been transposing recently in evolutionary history. Using the 0.34 Kb 5' UTR as the query, over 2000 copies were identified that may contain internal promoters, leading to questions on the genomic impact of Juan-A. Juan sequences were obtained by PCR, library screening, and database searches for 18 mosquito species of six genera including Aedes, Ochlerotatus, Psorophora, Culex, Deinocerites, and Wyeomyia. Comparison of host and Juan phylogenies shows overall congruence with few exceptions.

Conclusion: Juan-A is a major genomic component in Ae. aegypti and it has been retrotransposing recently in evolutionary history. There are also indications that Juan has been recently active in a wide range of mosquito species. Furthermore, our research demonstrates that a Jockey clade non-LTR without target site-specificity has been sustained by vertical transmission in the mosquito family. These results strengthen the argument that non-LTRs tend to be genomic elements capable of persistence by vertical descent over a long evolutionary time.

Figure 1

Structural organization of the Juan-A element of Aedes aegypti. ORF1 encodes a nucleic acid binding protein and ORF2 encodes both an apurinic/apyrimidinic (AP) endonuclease and reverse transcriptase (RT) domain. Arrows indicate the 939 bp region amplified by PCR that was used for phylogenetic inference. A canonical polyadenylation signal sequence is present in the 3' end of Juan-A (not shown) Regions used for copy number determination by database search in Table 1 are shown by horizontal lines.

Figure 2

Phylogenetic comparison of Juan sequences and their hosts. A. Consensus tree of host phylogeny constructed with MrBayes (see methods) using nucleotide sequences of Vg-C, a single copy gene. Vg-C from An. gambiae is used to root the tree. Clade credibility values from 150,000 generations are given at each node. B. Consensus tree constructed with MrBayes using conceptually translated sequences of Juan from PCR and genomic database (Aa, Ae. aegypti, Ag, An. gambiae). Clade credibility values from 200,000 generations are given at each node or beside brackets. Ag-Jen-4 and other An. gambiae sequences correspond to families previously identified (Biedler and Tu 2003). Jockey elements from D. melanogaster (Dm) and C. tentans (Ct) are used to root the tree. Accessions are given beside sequence names. An asterisk indicates that the reading frame was intact. A bold capital "L" indicates that the sequence was obtained from a genomic library. The first three letters of a species name is used to label PCR and library sequences. Genus names are in bold beside brackets. Juan-A is from Ae. aegypti and Juan-C is from C. pipiens. Abbreviations: Ae. aegypti (aeg); Ae. albopictus (alb); Ae. simpsoni (sim); Ae. polynesiensis (pol); Ae. vexans (vex); C. molestus (mol); C. quinquefasciatus (qui); C. restuans (res); C. tarsalis (tar); D. cancer (can); O. atropalpus (atr); O. bahamensis (bah); O. epactius (epa); O. taeniorhinchus (tae); O. triseriatus (tri); P. ciliata (cil); W. michelli (mic). Number indicates clone from PCR. Ae. aegypti and An. gambiae sequences from genomic database (e.g. Ag-Jock-1, AaJockeyEle2) can be found in the TEfam database [57]. See additional files 1 and 2 for alignments used for phylogenetic inference.