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Abstract
BackgroundR2 is a non-long terminal repeat (non-LTR) retrotransposable element that inserts site specifically into the 28S genes of the ribosomal (r)RNA gene loci. Encoded at the 5' end is a ribozyme that generates the precise 5' end by self-cleavage of a 28S gene cotranscript. Sequences at the 3' end are necessary for the R2 protein to bind RNA and initiate the target primed reverse transcription (TPRT) reaction. These minimal RNA requirements suggested that if recombination/DNA repair conjoined the 5' and 3' ends of R2, the result would be a non-autonomous element that could survive as long as autonomous R2 elements supplied the TPRT activity.ResultsA PCR-based survey of 39 Drosophila species aided by genomic sequences from 12 of these species revealed two types of non-autonomous elements. We call these elements SIDEs (for ‘Short Internally Deleted Elements’). The first consisted of a 5' ribozyme and a 3' end of an R2 element as predicted. Variation at the 5' junctions of the R2 SIDE copies was typical for R2 insertions suggesting their propagation by TPRT. The second class of SIDE contained sequences from R1 elements, another non-LTR retrotransposon that inserts into rRNA gene loci. These insertions had an R2 ribozyme immediately upstream of R1 3' end sequences. These hybrid SIDEs were inserted at the R1 site with 14 bp target site duplications typical of R1 insertions suggesting they used the R1 machinery for retrotransposition. Finally, the survey revealed examples of U12 small nuclear (sn)RNA and tRNA sequences at the 5' end of R2 elements suggesting the R2 reverse transcriptase can template jump from the R2 transcript to a second RNA during TPRT.ConclusionsThe R2 SIDE and R2/R1 hybrid SIDEs are rare examples of non-autonomous retrotransposons in the Drosophila genome. Associated non-autonomous elements and in vivo template jumps are two additional characteristics R2 shares with other non-LTR retrotransposons such as mammalian L1s. Analysis of the hybrid SIDEs provides supporting evidence that R1 elements, like R2 elements, recognize their 3' untranslated region (UTR) sequences and, thus, belong to the stringent class of non-LTR elements.
Highlights
R2 is a non-long terminal repeat retrotransposable element that inserts site into the 28S genes of the ribosomal (r)RNA gene loci
R2 SIDE While analyzing R2 ribozyme sequences from Drosophila willistoni, a sequence located in the R2 insertion site was identified which showed only 64% sequence identity to the 5' untranslated region (UTR) of the R2 elements in this species [22]
PCR amplification using a degenerate primer to conserved sequences in the ribozyme paired with a reverse primer to 28S sequences 30 to 50 bp downstream of the R2 site (Figure 1B, primers 1 and 2) generated the expected 3.5 kb R2 element product as well as a much shorter product
Summary
R2 is a non-long terminal repeat (non-LTR) retrotransposable element that inserts site into the 28S genes of the ribosomal (r)RNA gene loci. The genomes of all eukaryotes contain examples of transposable elements, sequences that generally appear to be genomic parasites such sequences are occasionally co-opted for the host's benefit [1,2] These mobile elements fall into families that differ in basic structure and method of transposition [3,4]. Non-long terminal repeat (non-LTR) retrotransposable elements comprise one of the two major families of mobile elements whose movement requires reverse transcriptase. The elements that parasitize the retrotransposition machinery of autonomous LINEs (for ‘Long INterspersed Elements’) have been called SINEs (for ‘Short INterspersed Elements’) They are represented by Alu elements in primates dozens of SINE families have been found in other eukaryotic genomes [6,7,8]. This is accomplished either by sequence identity at the 3' end between the LINE and its associated SINE (stringent elements) or a less strict recognition of a simple sequence, frequently a poly(A) tail, (relaxed elements) [11,12,13,14]
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