RNA helicase Belle/DDX3 regulates transgene expression in Drosophila

Abstract

Belle (Bel), the Drosophila homolog of the yeast DEAD-box RNA helicase DED1 and human DDX3, has been shown to be required for oogenesis and female fertility. Here we report a novel role of Bel in regulating the expression of transgenes. Abrogation of Bel by mutations or RNAi induces silencing of a variety of P-element-derived transgenes. This silencing effect depends on downregulation of their RNA levels. Our genetic studies have revealed that the RNA helicase Spindle-E (Spn-E), a nuage RNA helicase that plays a crucial role in regulating RNA processing and PIWI-interacting RNA (piRNA) biogenesis in germline cells, is required for loss-of-bel-induced transgene silencing. Conversely, Bel abrogation alleviates the nuage-protein mislocalization phenotype in spn-E mutants, suggesting a competitive relationship between these two RNA helicases. Additionally, disruption of the chromatin remodeling factor Mod(mdg4) or the microRNA biogenesis enzyme Dcr-1 also rescued the transgene-silencing phenotypes in bel mutants, suggesting the involvement of chromatin remodeling and microRNA biogenesis in loss-of-bel-induced transgene silencing. Finally we showed that genetic inhibition of Bel function led to the de novo generation of piRNAs from the transgene region inserted in the genome, suggesting a potential piRNA-dependent mechanism that may mediate transgene silencing as Bel function is inhibited. Our findings have demonstrated a novel involvement of Bel in regulating transgene expression and its loss triggers a transgene silencing mechanism mediated by protein regulators implicated in RNA processing, piRNA biogenesis, chromatin remodeling and the microRNA pathway.

The RNA Helicase BELLE Is Involved in Circadian Rhythmicity and in Transposons Regulation in Drosophila melanogaster

“…We have observed a reduction of the transposons’ transcription in belle mutant, in comparison to wild-type; notably, other piRNA mutants normally exhibit an increased TEs’ expression (Aravin et al, 2007; Specchia et al, 2008, 2017; Specchia and Bozzetti, 2009; Senti and Brennecke, 2010). This observation well-agrees with the recently reported data on the effect of belle mutants in the silencing of P-transposon-derived constructs in ovaries, depending on de novo piRNAs generated from the transgenes in the region of their insertion (Lo et al, 2016). Our data led us to infer that belle regulates the transposons probably via the piRNA-mediated pathway.…”

“…belle emerged in a screen for genes involved in the RNAi pathway: it binds to Ago1 and Ago2, involved in the small RNA-mediated regulation (Kim et al, 2005; Ulvila et al, 2006; Zhou et al, 2008). Very recently, a role of belle in the regulation on specific P-derived constructs in the ovary has been described (Lo et al, 2016), and therefore it is now considered a “bona fide” piRNA gene. belle encodes a DEAD-box RNA helicase: it is the closest paralog of vasa and plays roles in the reproductive system allowing fertility in Drosophila melanogaster.…”

Systematic analysis reveals tumor-enhancing and -suppressing microRNAs inDrosophilaepithelial tumors

“…Approximately 200 wing imaginal discs from wild-type ( w1118 ) and tumorous environments were dissected. Small RNA libraries were prepared per instructions for Illumina TrueSeq Small RNA sample prep kit and some modifications, as describe in [ 51 ]. For sequencing, Illumina HiSeq 2500 system was used, and samples were under single end, 50-base pair conditions.…”

Germline silencing of UASt depends on the piRNA pathway

“…In Drosophila , piRNA biogenesis employs a unique ‘ping-pong cycle’ mechanism for piRNA processing and amplification in the nuage , a perinuclear structure surrounding the nurse cells in developing egg chambers (Saito et al, 2006; Brennecke et al 2007; Malone et al 2009; Siomi et al, 2011). The localization of nuage related proteins such as Vasa, Aub, and AGO3 can be used as an indicator for the piRNA biogenesis pathway (Findley et al, 2003; Lim and Kai, 2007; Malone et al, 2009; Handler et al, 2013; Lo et al, 2016).…”