Artificial horizontal transfer of retroposons.
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Many factors may explain why certain transposable elements (TEs) spread in
some species and not others. On the one hand, they include processes that affect the
rate of transposition, such as differences in the regulation of expression; on the other
hand, they include characteristics of a genome that affect the consequences of
transposition. In particular genome size may have an effect: a genome that is large
due to non-essential repetitive DNA may be permissive for TE movement, as insertion
events are less likely to be deleterious. Genome size may also help explain the pattern
of TE distribution between species of mosquitoes, including the important vectors of
arboviruses, Aedes aegypti and Culex pipiens sensu lato. These species have genomes 3-
5 times larger than a third genus, the Anopheles mosquitoes, which includes the
malaria vectors. While all mosquitoes carry a diverse range of TEs, only culicines have
the super abundant retroposon, Juan which can contribute up to 3% of the genome.
The genome sequences of various insect species were compared and the
mosquitoes show a significant trend of increase in genome size, which can be
attributed to the increase in retroposon sequences.
Two variants of Juan are reported, and new information is added regarding
these elements. Previous publication of these elements contained errors in their
sequences. A unique triple repeat of a cysteine rich region with a CCHC motif is
present in the open reading frame. This sequence is a zinc-knuckle domain, important
for the replication mechanism of these elements.
In comparison, a third recently active but very low copy number retroposon,
termed Pip1, is also described. The results show that Pip1 is related to the Juan
elements and also possess the triple CCHC motif. The PCR results also supports
previous findings of polymorphism in insertion sites of this element, suggesting that
Pip1 was active after the establishment of the different strains. Pip1 copies can be
grouped into three distinct groups based on nucleotide differences. Pip1 could also be
using an alternative start codon to initiate transcription.
Full length intact copies of the three TEs in this study were been cloned into a
germline transformation vector based on piggyBac and used for germline
transformation in Drosophila melanogaster. Drosophila melanogaster has no Juan or Pip1
elements and an even smaller genome than anophelines mosquitoes, so insertion
events from unregulated TE movement should be more detectable. We found that the
elements have been successfully introduced into the Drosophila lines. The lines were
inbred to obtain a homozygous population. A range of transformed lines were
monitored. No effects of hybrid dysgenesis was found. Flies with black spotted eyes
were identified in a Pip1 line but this phenotype was not heritable. Whole genome
sequencing was carried out on the flies using next generation sequencing (NGS)
technology. Retroposon sequences was detected at a high frequency. Insertion
junctions were not detected but this result does not eliminate the possibility that a
junction is present but the sequencing was not sensitive enough. A possible
explanation is the retroposon is present as extrachromosomal plasmid DNA
Authors
Yeoh, Joseph Guan ChongCollections
- Theses [4099]