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

<p>Abstract</p> <p>Background</p> <p>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. <it>Juan </it>is a Jockey clade non-LTR retrotransposon first discovered in mosquitoes that is widely distributed in the mosquito family <it>Culicidae</it>. Being a non-site specific non-LTR, <it>Juan </it>offers an opportunity to further investigate the hypothesis that non-LTRs are genomic elements that are primarily vertically transmitted.</p> <p>Results</p> <p>Systematic analysis of the ~1.3 Gbp <it>Aedes aegypti </it>(<it>Ae. aegypti</it>) genome sequence suggests that <it>Juan-A </it>is the only <it>Juan</it>-type non-LTR in <it>Aedes aegypti</it>. <it>Juan-A </it>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 <it>Juan-A </it>sequence as the query. All 663 copies are at least 95% identical to <it>Juan-A</it>, while 378 of these copies are 99% identical to <it>Juan-A</it>, indicating that the <it>Juan-A </it>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 <it>Juan-A</it>. <it>Juan </it>sequences were obtained by PCR, library screening, and database searches for 18 mosquito species of six genera including <it>Aedes</it>, <it>Ochlerotatus</it>, <it>Psorophora</it>, <it>Culex</it>, <it>Deinocerites</it>, and <it>Wyeomyia</it>. Comparison of host and <it>Juan </it>phylogenies shows overall congruence with few exceptions.</p> <p>Conclusion</p> <p><it>Juan-A </it>is a major genomic component in <it>Ae. aegypti </it>and it has been retrotransposing recently in evolutionary history. There are also indications that <it>Juan </it>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.</p

Evolutionary analysis of the kinesin light chain genes in the yellow fever mosquito Aedes aegypti: gene duplication as a source for novel early zygotic genes

<p>Abstract</p> <p>Background</p> <p>The maternal zygotic transition marks the time at which transcription from the zygotic genome is initiated and a subset of maternal RNAs are progressively degraded in the developing embryo. A number of early zygotic genes have been identified in <it>Drosophila melanogaster </it>and comparisons to sequenced mosquito genomes suggest that some of these early zygotic genes such as <it>bottleneck </it>are fast-evolving or subject to turnover in dipteran insects. One objective of this study is to identify early zygotic genes from the yellow fever mosquito <it>Aedes aegypti </it>to study their evolution. We are also interested in obtaining early zygotic promoters that will direct transgene expression in the early embryo as part of a <it>Medea </it>gene drive system.</p> <p>Results</p> <p>Two novel early zygotic kinesin light chain genes we call <it>AaKLC2.1 </it>and <it>AaKLC2.2 </it>were identified by transcriptome sequencing of <it>Aedes aegypti </it>embryos at various time points. These two genes have 98% nucleotide and amino acid identity in their coding regions and show transcription confined to the early zygotic stage according to gene-specific RT-PCR analysis. These <it>AaKLC2 </it>genes have a paralogous gene (<it>AaKLC1</it>) in <it>Ae. aegypti</it>. Phylogenetic inference shows that an ortholog to the <it>AaKLC2 </it>genes is only found in the sequenced genome of <it>Culex quinquefasciatus</it>. In contrast, <it>AaKLC1 </it>gene orthologs are found in all three sequenced mosquito species including <it>Anopheles gambiae</it>. There is only one KLC gene in <it>D. melanogaster </it>and other sequenced holometabolous insects that appears to be similar to <it>AaKLC1</it>. Unlike <it>AaKLC2</it>, <it>AaKLC1 </it>is expressed in all life stages and tissues tested, which is consistent with the expression pattern of the <it>An. gambiae </it>and <it>D. melanogaster </it>KLC genes. Phylogenetic inference also suggests that <it>AaKLC2 </it>genes and their likely <it>C. quinquefasciatus </it>ortholog are fast-evolving genes relative to the highly conserved <it>AaKLC1-like </it>paralogs. Embryonic injection of a luciferase reporter under the control of a 1 kb fragment upstream of the <it>AaKLC2.1 </it>start codon shows promoter activity at least as early as 3 hours in the developing <it>Ae. aegypti </it>embryo. The <it>AaKLC2.1 </it>promoter activity reached ~1600 fold over the negative control at 5 hr after egg deposition.</p> <p>Conclusions</p> <p>Transcriptome profiling by use of high throughput sequencing technologies has proven to be a valuable method for the identification and discovery of early and transient zygotic genes. The evolutionary investigation of the KLC gene family reveals that duplication is a source for the evolution of new genes that play a role in the dynamic process of early embryonic development. <it>AaKLC2.1 </it>may provide a promoter for early zygotic-specific transgene expression, which is a key component of the <it>Medea </it>gene drive system.</p

Next-Generation Sequencing Reveals Recent Horizontal Transfer of a DNA Transposon between Divergent Mosquitoes

Horizontal transfer of genetic material between complex organisms often involves transposable elements (TEs). For example, a DNA transposon mariner has been shown to undergo horizontal transfer between different orders of insects and between different phyla of animals. Here we report the discovery and characterization of an ITmD37D transposon, MJ1, in Anopheles sinensis. We show that some MJ1 elements in Aedes aegypti and An. sinensis contain intact open reading frames and share nearly 99% nucleotide identity over the entire transposon, which is unexpectedly high given that these two genera had diverged 145–200 million years ago. Chromosomal hybridization and TE-display showed that MJ1 copy number is low in An. sinensis. Among 24 mosquito species surveyed, MJ1 is only found in Ae. aegypti and the hyrcanus group of anopheline mosquitoes to which An. sinensis belongs. Phylogenetic analysis is consistent with horizontal transfer and provides the basis for inference of its timing and direction. Although report of horizontal transfer of DNA transposons between higher eukaryotes is accumulating, our analysis is one of a small number of cases in which horizontal transfer of nearly identical TEs among highly divergent species has been thoroughly investigated and strongly supported. Horizontal transfer involving mosquitoes is of particular interest because there are ongoing investigations of the possibility of spreading pathogen-resistant genes into mosquito populations to control malaria and other infectious diseases. The initial indication of horizontal transfer of MJ1 came from comparisons between a 0.4x coverage An. sinensis 454 sequence database and available TEs in mosquito genomes. Therefore we have shown that it is feasible to use low coverage sequencing to systematically uncover horizontal transfer events. Expanding such efforts across a wide range of species will generate novel insights into the relative frequency of horizontal transfer of different TEs and provide the evolutionary context of these lateral transfer events

Improved reference genome of the arboviral vector Aedes albopictus

Background: The Asian tiger mosquito Aedes albopictus is globally expanding and has become the main vector for human arboviruses in Europe. With limited antiviral drugs and vaccines available, vector control is the primary approach to prevent mosquito-borne diseases. A reliable and accurate DNA sequence of the Ae. albopictus genome is essential to develop new approaches that involve genetic manipulation of mosquitoes. Results: We use long-read sequencing methods and modern scaffolding techniques (PacBio, 10X, and Hi-C) to produce AalbF2, a dramatically improved assembly of the Ae. albopictus genome. AalbF2 reveals widespread viral insertions, novel microRNAs and piRNA clusters, the sex-determining locus, and new immunity genes, and enables genome-wide studies of geographically diverse Ae. albopictus populations and analyses of the developmental and stage-dependent network of expression data. Additionally, we build the first physical map for this species with 75% of the assembled genome anchored to the chromosomes. Conclusion: The AalbF2 genome assembly represents the most up-to-date collective knowledge of the Ae. albopictus genome. These resources represent a foundation to improve understanding of the adaptation potential and the epidemiological relevance of this species and foster the development of innovative control measures

The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity-1

<p><b>Copyright information:</b></p><p>Taken from "The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity"</p><p>http://www.biomedcentral.com/1471-2148/7/112</p><p>BMC Evolutionary Biology 2007;7():112-112.</p><p>Published online 9 Jul 2007</p><p>PMCID:PMC1947958.</p><p></p>Vg-C from 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 from PCR and genomic database (Aa, , Ag, ). Clade credibility values from 200,000 generations are given at each node or beside brackets. Ag-Jen-4 and other sequences correspond to families previously identified (Biedler and Tu 2003). Jockey elements from (Dm) and (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. is from and is from . Abbreviations: (); (); (); (); (); (); (); (); (); (); (); (); (); (); (); (); (). Number indicates clone from PCR. and sequences from genomic database (e.g. Ag-Jock-1, AaJockeyEle2) can be found in the TEfam database [57]. See additional files and for alignments used for phylogenetic inference

The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity-3

<p><b>Copyright information:</b></p><p>Taken from "The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity"</p><p>http://www.biomedcentral.com/1471-2148/7/112</p><p>BMC Evolutionary Biology 2007;7():112-112.</p><p>Published online 9 Jul 2007</p><p>PMCID:PMC1947958.</p><p></p>Vg-C from 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 from PCR and genomic database (Aa, , Ag, ). Clade credibility values from 200,000 generations are given at each node or beside brackets. Ag-Jen-4 and other sequences correspond to families previously identified (Biedler and Tu 2003). Jockey elements from (Dm) and (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. is from and is from . Abbreviations: (); (); (); (); (); (); (); (); (); (); (); (); (); (); (); (); (). Number indicates clone from PCR. and sequences from genomic database (e.g. Ag-Jock-1, AaJockeyEle2) can be found in the TEfam database [57]. See additional files and for alignments used for phylogenetic inference

Evolution and Horizontal Transfer of a DD37E DNA Transposon in Mosquitoes

ITmD37E, a unique class II transposable element (TE) with an ancient origin, appears to have been involved in multiple horizontal transfers in mosquitoes as ITmD37E sequences from 10 mosquito species of five genera share high nucleotide (nt) identities. For example, ITmD37E sequences from Aedes aegypti and Anopheles gambiae, which have an estimated common ancestor of 145–200 million years ago, display 92% nt identity. The comparison of ITmD37E and host mosquito phylogenies shows a lack of congruence. The wide distribution of conserved ITmD37Es in mosquitoes and the presence of intact copies suggest that this element may have been recently active

Identification of early zygotic genes in the yellow fever mosquito Aedes aegypti and discovery of a motif involved in early zygotic genome activation.

During early embryogenesis the zygotic genome is transcriptionally silent and all mRNAs present are of maternal origin. The maternal-zygotic transition marks the time over which embryogenesis changes its dependence from maternal RNAs to zygotically transcribed RNAs. Here we present the first systematic investigation of early zygotic genes (EZGs) in a mosquito species and focus on genes involved in the onset of transcription during 2-4 hr. We used transcriptome sequencing to identify the "pure" (without maternal expression) EZGs by analyzing transcripts from four embryonic time ranges of 0-2, 2-4, 4-8, and 8-12 hr, which includes the time of cellular blastoderm formation and up to the start of gastrulation. Blast of 16,789 annotated transcripts vs. the transcriptome reads revealed evidence for 63 (P<0.001) and 143 (P<0.05) nonmaternally derived transcripts having a significant increase in expression at 2-4 hr. One third of the 63 EZG transcripts do not have predicted introns compared to 10% of all Ae. aegypti genes. We have confirmed by RT-PCR that zygotic transcription starts as early as 2-3 hours. A degenerate motif VBRGGTA was found to be overrepresented in the upstream sequences of the identified EZGs using a motif identification software called SCOPE. We find evidence for homology between this motif and the TAGteam motif found in Drosophila that has been implicated in EZG activation. A 38 bp sequence in the proximal upstream sequence of a kinesin light chain EZG (KLC2.1) contains two copies of the mosquito motif. This sequence was shown to support EZG transcription by luciferase reporter assays performed on injected early embryos, and confers early zygotic activity to a heterologous promoter from a divergent mosquito species. The results of these studies are consistent with the model of early zygotic genome activation via transcriptional activators, similar to what has been found recently in Drosophila

The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity-0

<p><b>Copyright information:</b></p><p>Taken from "The non-LTR retrotransposon in mosquitoes: genomic impact, vertical transmission and indications of recent and widespread activity"</p><p>http://www.biomedcentral.com/1471-2148/7/112</p><p>BMC Evolutionary Biology 2007;7():112-112.</p><p>Published online 9 Jul 2007</p><p>PMCID:PMC1947958.</p><p></p>ase (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 (not shown) Regions used for copy number determination by database search in Table 1 are shown by horizontal lines