Abstract
Helitrons are eukaryotic DNA transposons that have profoundly affected genome variability via capture and mobilization of host genomic sequences. Defining their mode of action is therefore important for understanding how genome landscapes evolve. Sequence similarities with certain prokaryotic mobile elements suggest a “rolling circle” mode of transposition, involving only a single transposon strand. Using the reconstituted Helraiser transposon to study Helitron transposition in cells and in vitro, we show that the donor site must be double-stranded and that single-stranded donors will not suffice. Nevertheless, replication and integration assays demonstrate the use of only one of the transposon donor strands. Furthermore, repeated reuse of Helraiser donor sites occurs following DNA synthesis. In cells, circular double-stranded intermediates that serve as transposon donors are generated and replicated by Helraiser transposase. Cell-free experiments demonstrate strand-specific cleavage and strand transfer, supporting observations made in cells.
Highlights
Helitrons are eukaryotic DNA transposons that have profoundly affected genome variability via capture and mobilization of host genomic sequences
It has been experimentally established that double-stranded DNA (dsDNA) Helraiser circles propagated in E. coli can be used as transposon donors in HeLa cells[21], it remained unclear whether the original transposon circles generated during transposition in HeLa cells were in single- or double-stranded form and whether those transposon circles were substrates for the Helraiser transposase
Helitrons are a unique group of DNA transposons that have profoundly impacted eukaryotic genomes and that transpose using a mechanism that differs from all other characterized eukaryotic DNA transposons
Summary
Helitrons are eukaryotic DNA transposons that have profoundly affected genome variability via capture and mobilization of host genomic sequences. Initially deemed “junk DNA,” they have since been found to be integral components of genome evolution, organization, and stability Major evolutionary innovations, such as the vertebrate adaptive immune system[1,2] and placenta[3,4], as well as novel transcription factors and regulatory networks 5,6), have their origin in transposition, and active mobile elements continue to reshape genomes of prokaryotes and eukaryotes alike This genometransforming property can impact both the evolutionary trajectory of a transposon host Most eukaryotic DNA transposons and all those that have found applications to date are the so-called “cut-and-paste” type DNA transposons These precisely cut themselves out of the donor DNA site by introducing double-strand breaks, and integrate the mobilized DNA segment to a target site, often with little or no sequence preference. Helitrons integrate precisely between 5′-A and T-3′ nucleotides into host AT target sites, but do not generate target site duplications
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