Distribution, evolution, and diversity of retrotransposons at the flamenco locus reflect the regulatory properties of piRNA clusters

Abstract

Most of our understanding of Drosophila heterochromatin structure and evolution has come from the annotation of heterochromatin from the isogenic y; cn bw sp strain. However, almost nothing is known about the heterochromatin's structural dynamics and evolution. Here, we focus on a 180-kb heterochromatic locus producing Piwi-interacting RNAs (piRNA cluster), the flamenco (flam) locus, known to be responsible for the control of at least three transposable elements (TEs). We report its detailed structure in three different Drosophila lines chosen according to their capacity to repress or not to repress the expression of two retrotransposons named ZAM and Idefix, and we show that they display high structural diversity. Numerous rearrangements due to homologous and nonhomologous recombination, deletions and segmental duplications, and loss and gain of TEs are diverse sources of active genomic variation at this locus. Notably, we evidence a correlation between the presence of ZAM and Idefix in this piRNA cluster and their silencing. They are absent from flam in the strain where they are derepressed. We show that, unexpectedly, more than half of the flam locus results from recent TE insertions and that most of the elements concerned are prone to horizontal transfer between species of the melanogaster subgroup. We build a model showing how such high and constant dynamics of a piRNA master locus open the way to continual emergence of new patterns of piRNA biogenesis leading to changes in the level of transposition control.RNAi | gene silencing | epigenetics O ver the course of evolution, transposable elements (TEs) have accumulated in the genomes of eukaryotes, where they can account for up to 85% of the DNA (1). Most of these sequences have lost their ability to transpose. They are now stable components of the genomes. Their conservation throughout evolution suggests that they may confer advantageous effects to their hosts. However, transposition of the copies that remain functional could generate deleterious mutations if they were not severely repressed by their host. RNAi, which is a gene-silencing mechanism triggered by small RNAs (reviewed in ref.2), has been identified as being the main cellular machinery involved in the "taming" of TEs (reviewed in refs. 3-5). RNAi pathways involve small RNAs of diverse families. Among them, Piwiinteracting RNAs (piRNAs) have been shown to be involved in TE silencing in the Drosophila ovary. These piRNAs, 23-29 nt long, are bound by the Argonaute proteins Piwi, Argonaute 3, or Aubergine. They are produced by discrete genomic loci named piRNA clusters, which have been described as containing vestiges of TEs (6). One of these loci, the flamenco (flam) locus, extends over 180 kilobases (kb) on the Drosophila X chromosome. It is proximal to the DISCO interacting protein 1 gene (DIP1) and close to pericentromeric heterochromatin. Before the identification of piRNAs, this locus had been shown to regulate the Gypsy retrotransposon (7, 8 (6) showed the potential for the flam cluster to pr...

The Release 6 reference sequence of the Drosophila melanogaster genome

“…This region is a complex nest of fragmented TE sequences and is associated with the flam locus and production of piRNAs. Zanni et al (2013) reported that the Release 5 unmapped scaffold CP000194 (Release 5 U: 964336-1041768) containing piRNA cluster 17, includes a 30-kb segmental duplication of a portion of the flam locus. This scaffold is incorporated in Release 6 in the newly mapped scaffold between the two gaps in the X: 20B region.…”

The peculiarities of piRNA expression upon heat shock exposure inDrosophila melanogaster

“…It is well-known that piRNA-clusters exhibit structural polymorphism in different D. melanogaster strains. 52 In order to directly demonstrate the role of Hsp70 in the response of piRNA system to HS it is necessary to introduce one or several hsp70 genes into the hsp70-strain or alternatively introduce interference constructs inhibiting hsp70 genes into w 1118 strain (work in progress).…”

piRNA Biogenesis in Drosophila melanogaster

“…The differences between the two cluster types extend far beyond the strand of transcription (Figure 1A). The transcription of uni-strand clusters seems to be similar to canonical mRNA transcription as these clusters have unique promoters and produce 5’ capped and polyadenylated RNA that are sometimes spliced [29–31]. Dual-strand clusters produce non-polyadenylated RNA and the majority of dual-strand clusters does not have clear signatures of Pol II promoters, such as peaks of Pol II and H3K4me2 [31–33].…”