Abstract
Separate conserved copies of suffix, a short interspersed Drosophila retroelement (SINE), and also divergent copies in the 3′ untranslated regions of the three genes, have already been described. Suffix has also been identified on the 3′ end of the Drosophila non-LTR F element, where it forms the last conserved domain of the reverse transcriptase (RT). In our current study, we show that the separate copies of suffix are far more actively transcribed than their counterparts on the F element. Transcripts from both strands of suffix are present in RNA preparations during all stages of Drosophila development, providing the potential for the formation of double-stranded RNA and the initiation of RNA interference (RNAi). Using in situ RNA hybridization analysis, we have detected the expression of both sense and antisense suffix transcripts in germinal cells. These sense and antisense transcripts are colocalized in the primary spermatocytes and in the cytoplasm of the nurse cells, suggesting that they form double-stranded RNA. We performed further analyses of suffix-specific small RNAs using northern blotting and SI nuclease protection assays. Among the total RNA preparations isolated from embryos, larvae, pupae and flies, suffix-specific small interfering RNAs (siRNAs) were detected only in pupae. In wild type ovaries, both the siRNAs and longer suffix-specific Piwi-interacting RNAs (piRNAs) were observed, whereas in ovaries of the Dicer-2 mutant, only piRNAs were detected. We further found by 3′ RACE that in pupae and ovaries, F element transcripts lacking the suffix sequence are also present. Our data provide direct evidence that suffix-specific RNAi leads to the silencing of the relative LINE (long interspersed element), F element, and suggests that SINE-specific RNA interference could potentially downregulate a set of genes possessing SINE stretches in their 5′ or 3′ non-coding regions. These data also suggest that double stranded RNAs possessing suffix are processed by both RNAi and an additional silencing mechanism.
Highlights
Retroelements are ancient components of the genome, and are potential participants in some RNA-related regulatory mechanisms in the cell
The recent discovery of RNA interference (RNAi) has extended our knowledge of such processes by uncovering mechanisms in which short RNA molecules are used by protein complexes for the recognition of specific nucleotide sequences that are important for the regulation of gene expression and the formation of chromosomal structures [1]
Several classes of retroelements have been detected during the last few decades: non-LTR mobile elements, LTR-elements that are closely related to retroviruses, and short retroelements
Summary
Retroelements are ancient components of the genome, and are potential participants in some RNA-related regulatory mechanisms in the cell. A number of mechanisms were subsequently described in which control of mRNA translation, the formation of heterochromatin structures, and the silencing of either mobile elements or unpaired DNA is mediated by RNAs as universal intermediates in homology sensing [3,4,5]. In some of these mechanisms, it has been postulated that ubiquitous retroelements could serve as targets for silencing, and as tools that provide RNA sequences for regulation. The discovery of RNAi mechanisms, which are considered to be an ancient protective mechanism against retroelements, but Academic Editor: Maarten van Lohuizen, Netherlands Cancer Institute, Netherlands
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