Piwi Nuclear Localization and Its Regulatory Mechanism in Drosophila Ovarian Somatic Cells.

In Drosophila ovarian somatic cells (OSCs), Piwi represses transposons transcriptionally to maintain genome integrity. Piwi nuclear localization requires the N terminus and PIWI-interacting RNA (piRNA) loading of Piwi. However, the underlying mechanism remains unknown.

Hierarchical roles of mitochondrial Papi and Zucchini in Bombyx germline piRNA biogenesis.

PIWI-interacting RNAs (piRNAs) are small regulatory RNAs that bind to PIWI proteins to control transposons and maintain genome integrity in animal germ lines. piRNA 3' end formation in the silkworm Bombyx mori has been shown to be mediated by the 3'-to-5' exonuclease Trimmer (Trim; known as PNLDC1 in mammals), and piRNA intermediates are bound with PIWI anchored onto mitochondrial Tudor domain protein Papi. However, it remains unclear whether the Zucchini (Zuc) endonuclease and Nibbler (Nbr) 3'-to-5' exonuclease, both of which have pivotal roles in piRNA biogenesis in Drosophila, are required for piRNA processing in other species.

Immuno-Electron Microscopy and Electron Microscopic In Situ Hybridization for Visualizing piRNA Biogenesis Bodies in Drosophila Ovaries.

Immuno-electron microscopy and electron microscopic in situ hybridization are powerful tools to identify the precise subcellular localization of specific proteins and RNAs at the ultramicroscopic level. Here we describe detailed procedures for how to detect the precise location of a specific target labeled with both fluorescence and gold particles. Although they have been developed for the analysis of Drosophila ovarian somatic cells, these techniques are suitable for a wide range of biological applications including human, primate, and rodent analysis.

TAF11 Assembles the RISC Loading Complex to Enhance RNAi Efficiency.

Assembly of the RNA-induced silencing complex (RISC) requires formation of the RISC loading complex (RLC), which contains the Dicer-2 (Dcr-2)-R2D2 complex and recruits duplex siRNA to Ago2 in Drosophila melanogaster. However, the precise composition and action mechanism of Drosophila RLC remain unclear. Here we identified the missing factor of RLC as TATA-binding protein-associated factor 11 (TAF11) by genetic screen.

Respective functions of two distinct Siwi complexes assembled during PIWI-interacting RNA biogenesis in Bombyx germ cells.

PIWI-interacting RNA (piRNA) biogenesis consists of two sequential steps: primary piRNA processing and the ping-pong cycle that depends on reciprocal Slicer-mediated RNA cleavage by PIWI proteins. However, the molecular functions of the factors involved remain elusive. Here, we show that RNAs cleaved by a Bombyx mori PIWI, Siwi, remain bound to the protein upon cleavage but are released by a DEAD box protein BmVasa.

Yb integrates piRNA intermediates and processing factors into perinuclear bodies to enhance piRISC assembly.

PIWI-interacting RNAs (piRNAs) direct Piwi to repress transposons and maintain genome integrity in Drosophila ovarian somatic cells. piRNA maturation and association with Piwi occur at perinuclear Yb bodies, the centers of piRNA biogenesis. Here, we show that piRNA intermediates arising from the piRNA cluster flamenco (flam) localize to perinuclear foci adjacent to Yb bodies, termed Flam bodies.