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Maternal Piwi regulates primordial germ cells to ensure the fertility of female progeny in Drosophila

ABSTRACT: PIWI proteins and their associated small noncoding piRNAs, which guide PIWI to target RNAs by base-pairing, are among the maternal components deposited into the germline of the early embryo in Drosophila. Piwi has been extensively studied in the adult ovary and testis, where it is required for transposon suppression, germline stem cell self-renewal, and fertility. Consequently, loss of Piwi in the adult ovary using piwi-null alleles or knockdown from early oogenesis results in complete sterility, limiting investigation into possible embryonic functions of maternal Piwi. In this study, we show that the maternal Piwi protein persists in the embryonic germline through gonad coalescence, suggesting that maternal Piwi can regulate germline development beyond early embryogenesis. Using a maternal knockdown strategy, we find that maternal Piwi is required for the fertility and normal gonad morphology of female, but not male, progeny. Following maternal Piwi knockdown, transposons were mildly derepressed in the early embryo but were fully repressed in the adult ovaries of progeny. Furthermore, the maternal piRNA pool is diminished, reducing the capacity of the PIWI/piRNA complex to target some zygotic genes during embryogenesis. Examination of embryonic germ cell proliferation and gene expression in the adult ovary showed that the germline of female progeny is partially masculinized upon maternal Piwi knockdown . This reveals a novel role for maternal Piwi in the germline development of female progeny and suggests that the PIWI/piRNA pathway is involved in germline sex determination in Drosophila.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE171951 | GEO | 2021/06/28

REPOSITORIES: GEO

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Project description:PIWI-clade Argonaute proteins silence transposon expression in animal gonads. Their target specificity is defined by bound ~23-30nt piRNAs that are processed from single-stranded precursor transcripts via two distinct pathways. Primary piRNAs are defined by the endo-nuclease Zucchini, while biogenesis of secondary piRNAs depends on piRNA-guided transcript cleavage and results in piRNA amplification. Here, we analyze the inter-dependencies between these piRNA biogenesis pathways in the developing Drosophila ovary. We show that secondary piRNA-guided target slicing is the predominant mechanism that specifies transcripts—including those from piRNA clusters—as primary piRNA precursors and that defines the spectrum of Piwi-bound piRNAs in germline cells. Post-transcriptional silencing in the cytoplasm therefore enforces nuclear, transcriptional target silencing, which ensures the tight suppression of transposons during oogenesis. As target slicing also defines the nuclear piRNA pool during mouse spermatogenesis, our findings uncover an unexpected conceptual similarity between the mouse and fly piRNA pathways. To understand the hierarchical order of primary versus secondary piRNA biogenesis in Drosophila ovaries, we sequenced piRNAs bound to total-Piwi, germline-Piwi, Aubergine and Argonaute3 from ovaries of germline specific knockdowns of control, piwi, aub, ago3 single knockdowns and aub/ago3 double knockdowns. To determine changes in Transposable Element (TE) transcription or TE RNA steady state in perturbed piRNA pathway conditions, we performed Pol2-ChIP-sequencing and polyA bound RNA-sequencing from ovaries of multiple germline knockdown genotypes. We also sequenced genomic DNA from ovaries of control knockdowns to experimentally estimate the TE copy number in our genetic background. Finally, we used CAP-seq from germline specific Piwi depletions to identify the Transcriptional Start Sites (TSS) in TEs in a deregulated background. Replicates are labeled with R1, R2, R3, R4 where indicated.

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Maternal Piwi regulates primordial germ cells to ensure the fertility of female progeny in Drosophila

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