Drosophila INterspersed Elements (DINEs) constitute an abundant but poorly understood group of Helitrons present in several Drosophila species. The general structure of DINEs includes two conserved blocks that may or not contain a region with tandem repeats in between. These central tandem repeats (CTRs) are similar within species but highly divergent between species. It has been assumed that CTRs have independent origins. Herein, we identify a subset of DINEs, termed DINE-TR1, which contain homologous CTRs of approximately 150 bp. We found DINE-TR1 in the sequenced genomes of several Drosophila species and in Bactrocera tryoni (Acalyptratae, Diptera). However, interspecific high sequence identity (∼ 88 %) is limited to the first ∼ 30 bp of each tandem repeat, implying that evolutionary constraints operate differently over the monomer length. DINE-TR1 is unevenly distributed across the Drosophila phylogeny. Nevertheless, sequence analysis suggests vertical transmission. We found that CTRs within DINE-TR1 have independently expanded into satellite DNA-like arrays at least twice within Drosophila. By analyzing the genome of Drosophila virilis and Drosophila americana, we show that DINE-TR1 is highly abundant in pericentromeric heterochromatin boundaries, some telomeric regions and in the Y chromosome. It is also present in the centromeric region of one autosome from D. virilis and dispersed throughout several euchromatic sites in both species. We further found that DINE-TR1 is abundant at piRNA clusters, and small DINE-TR1-derived RNA transcripts (∼25 nt) are predominantly expressed in the testes and the ovaries, suggesting active targeting by the piRNA machinery. These features suggest potential piRNA-mediated regulatory roles for DINEs at local and genome-wide scales in Drosophila.

Rolling-circle replication (RCR) elements constitute a diverse group that includes viruses, plasmids, and transposons, present in hosts from all domains of life. Eukaryotic RCR transposons, also known as Helitrons, are found in species from all eukaryotic kingdoms, sometimes representing a large portion of their genomes. Despite the impact of Helitrons on their hosts, knowledge about their relationship with other RCR elements is still elusive. Here, we compared the endonuclease domain sequence of Helitron transposases with the corresponding region from RCR proteins found in a wide variety of mobile genetic elements. To do that, we used a stepwise alignment approach followed by phylogenetic and multidimensional scaling analyses. Although it has been suggested that Helitrons might have originated from prokaryotic transposons or eukaryotic viruses, our results indicate that Helitron transposases share more similarities with proteins from prokaryotic viruses and plasmids instead. We also provide evidence for the division of RCR endonucleases into three groups (Y1, Y2, and Yx), covering the whole diversity of this protein family. Together, these results point to prokaryotic elements as the likely closest ancestors of eukaryotic RCR transposons, and further demonstrate the fluidity that characterizes the boundaries separating viruses, plasmids, and transposons.

Bracoviruses associate symbiotically with thousands of parasitoid wasp species in the family Braconidae, working as virulence gene vectors, and allowing the development of wasp larvae within hosts. These viruses are composed of multiple DNA circles that are packaged into infective particles, and injected together with wasp’s eggs during parasitization. One of the viral segments of Cotesia vestalis bracovirus contains a gene that has been previously described as a helicase of unknown origin. Here, we demonstrate that this gene is a Rep/Helicase from an intact Helitron transposable element that covers the viral segment almost entirely. We also provide evidence that this element underwent at least two horizontal transfers, which appear to have occurred consecutively: first from a Drosophila host ancestor to the genome of the parasitoid wasp C. vestalis and its bracovirus, and then from C. vestalis to a lepidopteran host (Bombyx mori). Our results reinforce the idea of parasitoid wasps as frequent agents of horizontal transfers in eukaryotes. Additionally, this Helitron-bracovirus segment is the first example of a transposable element that effectively became a whole viral circle.

Helitrons are the only group of rolling-circle transposons that encode a transposase with a helicase domain (Hel), which belongs to the Pif1 family. Because Pif1 helicases are important components of eukaryotic genomes, it has been suggested that Hel domains probably originated after a host eukaryotic Pif1 gene was captured by a Helitron ancestor. However, the few analyses exploring the evolution of Helitron transposases (RepHel) have focused on its Rep domain, which is also present in other mobile genetic elements. Here, we used phylogenetic and non-metric multidimensional scaling analyses to investigate the relationship between Hel domains and Pif1-like helicases from a variety of organisms. Our results reveal that Hel domains are only distantly related to genomic helicases from eukaryotes and prokaryotes, and thus are unlikely to have originated from a captured Pif1 gene. Based on this evidence, and on recent studies indicating that Rep domains are more closely related to rolling-circle plasmids and phages, we suggest that Helitrons are descendants of a RepHel-encoding prokaryotic plasmid element that invaded eukaryotic genomes before the radiation of its major groups. We discuss how a Pif1-like helicase domain might have favored the transposition of Helitrons in eukaryotes beyond simply unwinding DNA intermediates. Finally, we demonstrate that some examples in the literature describing genomic helicases from eukaryotes actually consist of Hel domains from Helitrons, a finding that underscores how transposons can hamper the analysis of eukaryotic genes. This investigation also revealed that two groups of land plants appear to have lost genomic Pif1 helicases independently.

Satellite DNAs are among the most abundant repetitive DNAs found in eukaryote genomes, where they participate in a variety of biological roles, from being components of important chromosome structures to gene regulation. Experimental methodologies used before the genomic era were insufficient, too laborious and time-consuming to recover the collection of all satDNAs from a genome. Today, the availability of whole sequenced genomes combined with the development of specific bioinformatic tools are expected to foster the identification of virtually all the “satellitome” of a particular species. While whole genome assemblies are important to obtain a global view of genome organization, most of them are incomplete and lack repetitive regions. We applied short-read sequencing and similarity clustering in order to perform a de novo identification of the most abundant satellite families in two Drosophila species from the virilis group: Drosophila virilis and D. americana, using the Tandem Repeat Analyzer (TAREAN) and RepeatExplorer pipelines. These species were chosen because they have been used as models to understand satDNA biology since the early 70’s. We combined the computational approach with data from the literature and chromosome mapping to obtain an overview of the major tandem repeat sequences of these species. The fact that all of the abundant tandem repeats (TRs) we detected were previously identified in the literature allowed us to evaluate the efficiency of TAREAN in correctly identifying true satDNAs. Our results indicate that raw sequencing reads can be efficiently used to detect satDNAs, but that abundant tandem repeats present in dispersed arrays or associated with transposable elements are frequent false positives. We demonstrate that TAREAN with its parent method RepeatExplorer may be used as resources to detect tandem repeats associated with transposable elements and also to reveal families of dispersed tandem repeats.