PIWI-interacting RNAs (piRNAs) and their associated PIWI clade Argonaute proteins constitute the core of the piRNA pathway. In gonadal cells, this conserved pathway is crucial for genome defense, and its main function is to silence transposable elements. This is achieved through posttranscriptional and transcriptional gene silencing. Precursors that give rise to piRNAs require specialized transcription and transport machineries because piRNA biogenesis is a cytoplasmic process. The ping-pong cycle, a posttranscriptional silencing mechanism, combines the cleavage-dependent silencing of transposon RNAs with piRNA production. PIWI proteins also function in the nucleus, where they scan for nascent target transcripts with sequence complementarity, instructing transcriptional silencing and deposition of repressive chromatin marks at transposon loci. Although studies have revealed numerous factors that participate in each branch of the piRNA pathway, the precise molecular roles of these factors often remain unclear. In this review, we summarize our current understanding of the mechanisms involved in piRNA biogenesis and function.

The PIWI-interacting RNA (piRNA) pathway is a small RNA-based immune system that controls the expression of transposons and maintains genome integrity in animal gonads. In Drosophila, piRNA-guided silencing is achieved, in part, via co-transcriptional repression of transposons by Piwi. This depends on Panoramix (Panx); however, precisely how an RNA binding event silences transcription remains to be determined. Here we show that Nuclear Export Factor 2 (Nxf2) and its co-factor, Nxt1, form a complex with Panx and are required for co-transcriptional silencing of transposons in somatic and germline cells of the ovary. Tethering of Nxf2 or Nxt1 to RNA results in silencing of target loci and the concomitant accumulation of repressive chromatin marks. Nxf2 and Panx proteins are mutually required for proper localization and stability. We mapped the protein domains crucial for the Nxf2/Panx complex formation and show that the amino-terminal portion of Panx is sufficient to induce transcriptional silencing.

Transgenerational Epigenetic Inheritance (TEI) studies the transmission of alternative functional states through multiple generations in the presence of the same genomic DNA sequence. Very little is known on the principles and the molecular mechanisms governing this type of inheritance. Here, by transiently enhancing 3D chromatin interactions, we established stable and isogenic Drosophila epilines that carry alternative epialleles, defined by differential levels of the Polycomb-dependent H3K27me3 mark. Once established, epialleles can be dominantly transmitted to naïve flies and induce paramutation. Importantly, epilines can be reset to a naïve state by disrupting chromatin interactions. Finally, we show that environmental changes can modulate the expressivity of the epialleles and we extend our paradigm to naturally occurring phenotypes. Our work sheds light on how nuclear organization and Polycomb group proteins contribute to epigenetically inheritable phenotypic variability.

Metazoan genomes are highly organized inside the cell nucleus. Topologically associating domains (TADs) represent the building blocks of genome organization, but their linear modularity does not explain alone their spatial organization. Indeed, the chromatin type adorning a TAD can shape its structure and drives its nuclear positioning and its function. Genome-wide association studies revealed mainly four chromatin types: active chromatin, Polycomb-repressed chromatin, null chromatin and constitutive heterochromatin. In this review, we will describe the main three-dimensional features of each chromatin type and finally their relationships with TAD organization and epigenetic memory.

The oocyte is a unique cell type that undergoes extensive chromosome changes on its way to fertilization, but the chromatin determinants of its fate are unknown. Here, we show that Polycomb group (PcG) proteins of the Polycomb repressive complex 2 (PRC2) determine the fate of the oocyte in Drosophila. Mutation of the enzymatic PRC2 subunit Enhancer of zeste (E(z)) in the germline abolishes spatial and temporal control of the cell cycle and induces sterility via transdetermination of the oocyte into a nurse-like cell. This fate switch depends on loss of silencing of two PRC2 target genes, Cyclin E and the cyclin-dependent kinase inhibitor dacapo. By contrast, the PRC1 component Polycomb (Pc) plays no role in this process. Our results demonstrate that PRC2 plays an exquisite role in the determination of the oocyte fate by preventing its switching into an endoreplicative program.

The PIWI-interacting RNA (piRNA) pathway is a small RNA-based immune system that controls the expression of transposons and maintains genome integrity in animal gonads. In Drosophila, piRNA-guided silencing is achieved, in part, via co-transcriptional repression of transposons by Piwi. This depends on Panoramix (Panx); however, precisely how an RNA binding event silences transcription remains to be determined. Here we show that Nuclear Export Factor 2 (Nxf2) and its cofactor, Nxt1, form a complex with Panx and are required for cotranscriptional silencing of transposons in somatic and germline cells of the ovary. Tethering of Nxf2 or Nxt1 to RNA results in silencing of target loci and the concomitant accumulation of repressive chromatin marks. Nxf2 and Panx proteins are mutually required for proper localization and stability. We mapped the protein domains crucial for the Nxf2/Panx complex formation and show that the amino-terminal portion of Panx is sufficient to induce transcriptional silencing.

Zygotic genome activation (ZGA) is a crucial step of embryonic development. So far, little is known about the role of chromatin factors during this process. Here, we used an in vivo RNA interference reverse genetic screen to identify chromatin factors necessary for embryonic development in Drosophila melanogaster . Our screen reveals that histone acetyltransferases (HATs) and histone deacetylases are crucial ZGA regulators. We demonstrate that Nejire (CBP/EP300 ortholog) is essential for the acetylation of histone H3 lysine-18 and lysine-27, whereas Gcn5 (GCN5/PCAF ortholog) for lysine-9 of H3 at ZGA, with these marks being enriched at all actively transcribed genes. Nonetheless, these HATs activate distinct sets of genes. Unexpectedly, individual catalytic dead mutants of either Nejire or Gcn5 can activate zygotic transcription (ZGA) and transactivate a reporter gene in vitro. Together, our data identify Nejire and Gcn5 as key regulators of ZGA.

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