Nuage, a well-conserved perinuclear organelle found in germline cells, is thought to mediate retroelement repression in Drosophila melanogaster by regulating the production of Piwi-interacting RNAs (piRNAs). In this study, we present evidence that the nuage–piRNA pathway components can be found in cytoplasmic foci that also contain retroelement transcripts, antisense piRNAs, and proteins involved in messenger RNA (mRNA) degradation. These mRNA degradation proteins, decapping protein 1/2 (DCP1/2), Me31B (maternal expression at 31B), and pacman (PCM), are normally thought of as components of processing bodies. In spindle-E (spn-E) and aubergine (aub) mutants that lack piRNA production, piRNA pathway proteins no longer overlap the mRNA degradation proteins. Concomitantly, spn-E and aub mutant ovaries show an accumulation of full-length retroelement transcripts and prolonged stabilization of HeT-A mRNA, supporting the role of piRNAs in mediating posttranscriptional retroelement silencing. HeT-A mRNA is derepressed in mRNA degradation mutants twin, dcp1, and ski3, indicating that these enzymes also aid in removing full-length transcripts and/or decay intermediates.
Nuage, a well conserved perinuclear organelle found in germline cells, is thought to mediate the repression of retroelements in Drosophila melanogaster by regulating the production of Piwiinteracting RNAs (piRNAs). Here, we present evidence that the nuage/piRNA pathway components can be found in cytoplasmic foci that also contain retroelement transcripts, anti-sense piRNAs and proteins involved in mRNA degradation. These mRNA degradation proteins, Decapping Proteins 1/2, Me31B and Pacman, are normally thought of as components of the Processing bodies. In spn-E and aub mutants where the production of piRNAs is compromised, piRNA pathway proteins no longer overlap the mRNA degradation proteins. Concomitantly, spn-E and aub mutant ovaries show a substantial accumulation of the full-length retroelement transcripts and prolonged stabilization of HeT-A mRNA, supporting a role of piRNAs in mediating post-transcriptional retroelement silencing. HeT-A mRNA is de-repressed in the canonical mRNA degradation mutants, indicating that these enzymes aid in removal of the full-length transcripts and/or decay intermediates. We have characterized small RNA expression profiles in differentiating mouse Embryonic Stem (ES) cell as a model for early mammalian development. High-throughput 454 pyrosequencing was performed on 19-30 nt RNAs isolated from undifferentiated male and female ES cells, as well as day 2 and 5 differentiating derivatives. A discrete subset of microRNAs largely dominated the small RNA repertoire and the dynamics of their accumulation could be readily used to discriminate pluripotency from early differentiation events. Unsupervised Partitioning Around Meloids (PAM) analysis revealed that differentiating ES cell miRNAs fall into three statistically significant expression clusters with highly contrasted patterns. PAM analysis afforded an unprecedented level of definition in the temporal fluctuations of individual members of several microRNA genomic clusters. Notably this unravelled highly complex post-transcriptional regulations of the key pluripotency miR-290 locus, and helped identify miR-293 as a clear outlier within this cluster, suggesting that previous conclusions drawn from whole miR-290 over-expression need to be reconsidered. Our analysis also uncovered a striking male-specific enrichment of the miR-302 family, which share the same seed sequence with most miR-290 family members. Accordingly, a miR-302 representative was strongly enriched in embryonic germ cells derived from primordial germ cells of male, but not female mouse embryos. Identifying the chromatin remodelling and E2F-dependent transcription repressors Ari4a and Arid4b as additional targets of miR-302 and miR-290 supports and expands a model integrating possible overlapping functions of the two miRNA families in mouse cell totipotency during early development. MicroRNAs (miRNAs) are a class of small RNAs that bind to specific mRNAs targets, directing their degradation and/or repressing their translation, which have been implicated in the control o...
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