Biology subjects

Iwasaki, Y. W.

Publications and source records attributed to Iwasaki, Y. W..

7 recordsLinked to original sources

bioRxiv2026

SPIN1 selectively silences evolutionarily young transposable elements through chromatin regulation

Mammalian genomes encode multiple transposable element (TE) silencing pathways that distinguish their targets through different molecular features, with KRAB zinc finger proteins recognizing DNA sequence, the HUSH complex sensing intronless transcripts, and the PIWI-piRNA pathway using small RNA guides. How chromatin state itself contributes to TE recognition remains less defined. Here we identify Spindlin1 (SPIN1), a three-Tudor-domain histone reader implicated in germline piRNA-directed DNA methylation, as a transcriptional repressor of evolutionarily young TEs in mouse embryonic stem cells. SPIN1 selectively binds LINE and ERV loci carrying H3K4me3 and H3K9me3, a chromatin signature enriched at young, transcription-permissive elements, and recognition of these marks by Tudor domains 1 and 2 is required for TE targeting. SPIN1 engages SPINDOC as a Tudor 1 and 3-dependent cofactor whose loss phenocopies SPIN1 depletion, and associates with the H3K9 methyltransferases SETDB1 and G9a. SPIN1 loss reduces H3K9me3 and is accompanied by increased chromatin accessibility, without altering DNA methylation. Thus, SPIN1 uses a histone-state-based mechanism to identify and repress young TEs in pluripotent cells, mechanistically distinct from its germline mode in which SPIN1 cooperates with the PIWI-piRNA pathway to promote DNA methylation, illustrating how a single histone reader engages distinct silencing machineries across cellular contexts.

molecular biology↗

bioRxiv2025

EC-isHCR: a rapid method for in situ hybridization chain reaction in diverse animal samples

The in situ hybridization chain reaction (isHCR) visualizes RNA across multiple spatial scales, from organs to subcellular structures, in diverse samples. We previously proposed a rapid protocol, EC-isHCR, for Drosophila embryos and ovaries. Whether EC-isHCR retains the features of conventional isHCR, including wide-spatial-scale analyses in various samples, however, has remained unclear. Here, we show that EC-isHCR enables robust RNA detection in a broad range of samples, such as whole-mount fruit fly, parasitoid wasp, and aphid preparations; paraffin sections of trout; frozen mouse sections; and human cultured cells. Moreover, EC-isHCR enabled detection of subcellular RNA localization. EC-isHCR also visualized co-localization of RNA with phase-separated condensates in fruit fly embryos and detected the protrusion-enriched mRNA in HeLa cells. To broaden the applicability of EC-isHCR, we developed an automated probe design tool (https://github.com/ShuntaYorimoto/hcrkit). By combining this tool with EC-isHCR, we provide a fast and versatile framework to visualize mRNAs. This framework will help reduce the barrier to using fast isHCR and thereby facilitate research across diverse areas of the life sciences. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/696653v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1852584org.highwire.dtl.DTLVardef@621682org.highwire.dtl.DTLVardef@1b3bf5corg.highwire.dtl.DTLVardef@1d86841_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- EC-isHCR enables rapid acquisition of high-contrast images. - EC-isHCR preserves features of conventional isHCR, including versatile sample compatibility and high-resolution imaging. - An automated probe design tool was developed for EC-isHCR. - EC-isHCR/probe tool framework will help reduce the barrier to using fast isHCR.

bioengineering↗

bioRxiv2025

Flamenco plasticity tunes somatic piRNAs, rewiring host isoforms and opening a route to heritable transposon spread

Transposons drive genome innovation, yet how they evade somatic piRNA defenses, reach the germline, and rewire host genes with limited cost remains unclear. Using comparative long-read sequencing in Drosophila ovarian somatic cells (OSCs), we show that the LTR transposon Springer rewires gene expression by inserting into promoter-proximal introns at an AT-rich motif. Its 5' LTR initiates transcripts that splice into host exons, expanding isoform diversity without adding coding sequence. We catalog 72 Springer-driven isoforms, indicating broad mutagenic potential. In parallel, the Flamenco (Flam) uni-strand piRNA cluster undergoes structural remodeling that replaces antisense transposon fragments with forward-oriented copies, reshaping piRNA populations and eroding silencing of specific elements, including Springer. This relaxation of somatic repression creates conditions permissive for germ-cell entry, providing a plausible route to heritable genome change. Thus, Flam plasticity, contrasting with the relative stability of dual-strand clusters, mechanistically links transposon activity to genome rewiring and evolutionary innovation.

genetics↗

bioRxiv2022

Mod(mdg4) variants repress telomeric retrotransposonHeT-A by blocking subtelomeric enhancers

Telomeres in Drosophila are composed of sequential non-LTR retrotransposons: HeT-A, TART, and TAHRE. Although they are repressed by the piRNA pathway in the germline, how these retrotransposons are regulated in somatic cells is poorly understood. Here, we show that specific splice variants of Mod(mdg4) repress HeT-A by blocking subtelomeric enhancers in ovarian somatic cells. We found that the Mod(mdg4)-N variant represses HeT-A expression most efficiently among the variants. Mod(mdg4)-N mutant flies show elevated HeT-A expression and female sterility. Mod(mdg4)-N-binding subtelomeric sequences exhibit enhancer-blocking activity, and recruitment of RNA polymerase II (Pol II) on subtelomeres by Mod(mdg4)-N is essential for this enhancer-blocking. Moreover, Mod(mdg4)-N functions to form chromatin boundaries of higher-order chromatin conformation but this mechanism is independent of its Pol II recruitment activity at telomeres/subtelomeres. This study provides a link between enhancer-blocking and telomere regulation, and two different molecular mechanisms exhibited by an insulator protein to orchestrate precise gene expression.

molecular biology↗

bioRxiv2021

Production of functional oocytes requires maternally expressed PIWI genes and piRNAs in golden hamsters

Many animals have a conserved adaptive genome defense system known as the Piwi-interacting RNA (piRNA) pathway which is essential for germ cell development and function. Disruption of individual mouse Piwi genes results in male but not female sterility, leading to the assumption that PIWI genes play little or no role in mammalian oocytes. Here, we report generation of PIWI-defective golden hamsters, which reveals defects in the production of functional oocytes. The mechanisms involved vary among the hamster PIWI genes; lack of PIWIL1 has a major impact on gene expression, including hamster-specific young transposon de-silencing, whereas PIWIL3 deficiency has little impact on gene expression in oocytes, although DNA methylation was found to be reduced to some extent in PIWIL3-defecient oocytes. Our findings serve as the foundation for developing useful models to study the piRNA pathway in mammalian oocytes, including humans, which is not possible with mice.

developmental biology↗

bioRxiv2020

Hamster PIWI proteins bind to piRNAs with stage-specific size variations during oocyte maturation

In animal gonads, transposable elements (TEs) are actively repressed to preserve genome integrity through the Piwi-interacting RNA (piRNA) pathway. In mice, piRNAs are most abundantly expressed in male germ cells, and form effector complexes with three distinct PIWI proteins. The depletion of individual Piwi genes causes male-specific sterility owing to severe defects in spermatogenesis with no discernible phenotype in female mice. Unlike mice, most other mammals have four PIWI genes, some of which are expressed in the ovary. Here, purification of PIWI complexes from oocytes of the golden hamster revealed that the size of the piRNAs loaded onto PIWIL1 changed during oocyte maturation. In contrast, PIWIL3, an ovary-specific PIWI in most mammals, associates with short piRNAs only in metaphase II oocytes, which coincides with intense phosphorylation of the protein. An improved high-quality genome assembly and annotation revealed that PIWIL1- and PIWIL3-associated piRNAs appear to share the 5'- ends of common piRNA precursors and are mostly derived from unannotated sequences with a diminished contribution from TE-derived sequences, most of which correspond to endogenous retroviruses (ERVs). Although binding sites for the transcription factor A-Myb are identified in the transcription start site regions of the testis piRNA clusters, the piRNA clusters in the ovary show no well-defined binding motifs in their upstream regions. These results show that hamster piRNA clusters are transcribed by different transcriptional factors in the ovary and testis, resulting in the generation of sex-specific piRNAs. Our findings show the complex and dynamic nature of biogenesis of piRNAs in hamster oocytes, and together with the new genome sequence generated, serve as the foundation for developing useful models to study the piRNA pathway in mammalian oocytes. Highlights- The size of PIWIL1-associated piRNAs changes during oocyte maturation - Phosphorylation of PIWIL3 in MII oocytes coincides with its association with small 19-nt piRNAs - Improved high-quality genome assembly and annotation identifies young endogenous retroviruses as major targets of piRNAs in hamster oocytes - PIWIL1- and PIWIL3-associated piRNAs share the 5'-ends of the common piRNA precursors in oocytes

molecular biology↗

bioRxiv2019

Nuclear RNA export factor variant initiates piRNA-guided co-transcriptional silencing

The PIWI-interacting RNA (piRNA) pathway preserves genomic integrity by repressing transposable elements (TEs) in animal germ cells. Among PIWI-clade proteins in Drosophila, Piwi transcriptionally silences its targets through interactions with cofactors, including Panoramix (Panx) and forms heterochromatin characterized by H3K9me3 and H1. Here, we identified Nxf2, a nuclear RNA export factor (NXF) variant, as a protein that forms complexes with Piwi, Panx, and p15. Panx-Nxf2-p15 complex formation is necessary in the silencing by stabilizing protein levels of Nxf2 and Panx. Notably, ectopic targeting of Nxf2 initiates co-transcriptional repression of the target reporter in a manner independent of H3K9me3 marks or H1. However, continuous silencing requires HP1a and H1. In addition, Nxf2 directly interacts with target TE transcripts in a Piwi-dependent manner. These findings suggest a model in which the Nxf2-Panx-p15 complex enforces the association of Piwi with target transcripts to trigger co-transcriptional repression, prior to heterochromatin formation in the nuclear piRNA pathway.\n\nHighlightsO_LINxf2 plays an essential role in the Piwi-piRNA pathway\nC_LIO_LIFormation of Piwi-Panx-Nxf2-p15 (PPNP) complexes stabilizes both Panx and Nxf2\nC_LIO_LIThe PPNP complex triggers transcriptional silencing before heterochromatin formation\nC_LIO_LINxf2 directly binds to target transcripts in a Piwi-dependent manner\nC_LI

molecular biology↗