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DEPS-1 promotes P granule assembly and RNA interference in C. elegans germ cells

ABSTRACT: P granules are germ-cell-specific cytoplasmic structures containing RNA and protein and required for proper germ cell development in C. elegans. A new P-granule-associated protein, DEPS-1, whose loss disrupts P granule structure and function was analysed in this study. Genome wide analysis of gene expression in deps-1 mutant germ lines identified additional targets of DEPS-1 regulation, many of which are also regulated by the RNAi factor RDE-3. Our studies suggest that DEPS-1 is a key component of the P granule assembly pathway and that its roles include promoting accumulation of some mRNAs, such as glh-1 and rde-4, and reducing accumulation of other mRNAs, perhaps by collaborating with RDE-3 to generate endogenous short interfering RNAs (endo-siRNAs). Keywords: Mutant analysis of deps-1 dissected C. elegans gonads

ORGANISM(S): Caenorhabditis elegans

PROVIDER: GSE9993 | GEO | 2008/02/01

SECONDARY ACCESSION(S): PRJNA104013

REPOSITORIES: GEO

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DEPS-1 promotes P granule assembly and RNA interference in C. elegans germ cells

Project description:P granules are germ-cell-specific cytoplasmic structures containing RNA and protein and required for proper germ cell development in C. elegans. A new P-granule-associated protein, DEPS-1, whose loss disrupts P granule structure and function was analysed in this study. Genome wide analysis of gene expression in deps-1 mutant germ lines identified additional targets of DEPS-1 regulation, many of which are also regulated by the RNAi factor RDE-3. Our studies suggest that DEPS-1 is a key component of the P granule assembly pathway and that its roles include promoting accumulation of some mRNAs, such as glh-1 and rde-4, and reducing accumulation of other mRNAs, perhaps by collaborating with RDE-3 to generate endogenous short interfering RNAs (endo-siRNAs). We isolated dissected gonads from deps-1 mutant adults and compared each to wild type dissected gonads. RNA was linearly amplified prior to labeling for all genotypes. The comparisons were done in quadruplicate on independently grown and isolated animals.

2010-05-26 | E-GEOD-9993 | biostudies-arrayexpress

Transcriptome profiling of P-granule-depleted or P-granule mutant gonads versus control gonads over time

Project description:The germ cells of multicellular organisms protect their developmental potential through specialized mechanisms. A shared feature of germ cells from worms to humans is the presence of non-membrane-bound ribonucleoprotein organelles called germ granules. Depletion of germ granules in Caenorhabditis elegans (i.e., P granules) leads to sterility and in some germlines expression of the neuronal transgene unc-119::gfp and the muscle myosin MYO-3. Thus, P granules are hypothesized to maintain germ cell totipotency by preventing somatic development, although the mechanism by which P granules carry out this function is unknown. In this study, we performed transcriptome and single molecule RNA-FISH analyses of dissected P-granule-depleted gonads at different developmental stages. Our results demonstrate that P granules are necessary for adult germ cells to down-regulate spermatogenesis RNAs and to prevent the accumulation of numerous soma-specific RNAs. P-granule-depleted gonads that express the unc-119::gfp transgene also express many other genes involved in neuronal development and concomitantly lose expression of germ cell fate markers. Finally, we show that removal of either of two critical P-granule components, PGL-1 or GLH-1, is sufficient to cause germ cells to express UNC-119::GFP and MYO-3 and to display RNA accumulation defects similar to those observed after depletion of P granules. Our data identify P granules as critical modulators of the germline transcriptome and guardians of germ cell fate.

2016-12-22 | GSE92690 | GEO

Germ granule compartments and small RNA production

Project description:In this project, we identified an uncharted subcompartment of the C. elegans germ granule, which we termed the E granule. We show that the E granule is required for the synthesis of a specialized class of 22G-RNAs.

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The TDRD5/7-like protein, LOTR-1, functions within Z granules to balance epigenetic signals in the Caenorhabditis elegans germline

Project description:LOTUS and Tudor domain containing proteins have critical roles in the germline. Proteins that contain these domains, such as Tejas/Tapas in Drosophila, help localize Vasa to the germ granules and facilitate piRNA transposon mediated silencing. The homologous proteins in mammals, TDRD5 and TDRD7, are required during spermiogenesis. Until now, LOTUS + Tudor domain proteins in C. elegans have remained elusive. Here we describe LOTR-1 (D1081.7), which derives its name from its LOTUS and Tudor domains. Interestingly, LOTR-1 docks next to P granules to colocalize with the Z-granule component ZNFX-1. LOTR-1’s Z-granule association requires its Tudor domain, but both LOTUS and Tudor deletions affect brood size when coupled with a knockdown of the Vasa homolog glh-1. LOTR-1 IP-mass spectrometry confirmed a Tudor-dependent association with Z-granule proteins ZNFX-1 and WAGO-1, but also with germ-granule proteins DEPS-1, the piRNA Argonaute PRG-1, and other WAGO-class Argonautes. Like znfx-1, lotr-1 mutants redistribute the coverage of 22G-RNAs toward the 5’ end of mutator targets and impact transgenerational epigenetic inheritance. Unlike znfx-1, the 5’ shift in 22G-RNA coverage does not extend to CSR-1 targets. Combined, these results suggest that LOTR-1 facilitates interactions between PRG-1/WAGO-class Argonautes, ZNFX-1 and target 3’UTRs to balance 22G-RNA distribution across mutator targets.

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GLH/VASA helicases promote germ granule formation to ensure the fidelity of piRNA-mediated transcriptome surveillance

Project description:The ability to distinguish non-self from self is the key characteristic for any defense system. piRNAs function as guardians of the genome by silencing non-self nucleic acids and transposable elements in animals. Many piRNA factors are enriched in perinuclear germ granules, but whether their localization is required for piRNA biogenesis or function is not known. Here we show that GLH/VASA helicase mutants exhibit defects in forming perinuclear condensates containing PIWI and other small RNA cofactors. These mutant animals produce largely normal levels of piRNA but are defective in triggering piRNA silencing. Strikingly, while many piRNA targets are activated in GLH mutants, we observed that hundreds of endogenous genes are aberrantly silenced by piRNAs. This defect in self versus non-self recognition was also observed in other mutants where perinuclear P granules are disrupted. Together, our results argue that perinuclear germ granules function critically to promote the fidelity of piRNA-based transcriptome surveillance in C. elegans and preserve self versus non-self distinction.

2022-08-05 | PXD033506 | Pride

A family of C. elegans VASA homologs control Argonaute pathway specificity and promote transgenerational silencing

Project description:Germline Argonautes direct transcriptome surveillance within peri-nuclear membraneless organelles called nuage. In C. elegans, a family of Vasa-related Germ Line Helicase (GLH) proteins localize in, and promote the formation of nuage called P granules. Previous studies have implicated GLH proteins in inherited silencing but direct roles in amplification of small RNAs, or in target mRNA or Argonatue binding have not been identified. Here we show that GLH proteins compete with each other to control Argonaute pathway specificity, bind directly to Argonaute-target mRNAs and act to promote the amplification of small RNAs required for transgenerational inheritance. We show that the ATPase cycle of GLH-1 regulates its direct binding to the Argonaute WAGO-1 which engages amplified small RNAs. Our findings support a dynamic and direct role for GLH proteins in inherited silencing beyond their role as structural components of nuage.

2022-01-30 | GSE195536 | GEO

A family of C. elegans VASA homologs control Argonaute pathway specificity and promote transgenerational silencing [IP small-RNA seq]

2022-03-10 | GSE198101 | GEO

Caenorhabditis elegans

Project description:DEPS-1 promotes P granule assembly and RNA interference in C. elegans germ cells

| PRJNA104013 | ENA

RNA self-assembly contributes to stress granule formation and defining the stress granule transcriptome

Project description:Ribonucleoprotein (RNP) granules are non-membrane bound organelles thought to assemble by protein-protein interactions of RNA-binding proteins. We present evidence that a wide variety of RNAs self-assemble in vitro into either liquid-liquid phase separations or more stable assembles, referred to as RNA tangles. Self-assembly in vitro is affected by RNA length, ionic conditions, and sequence. Remarkably, self-assembly of yeast RNA in vitro under physiological salt mimics the composition of mRNAs within yeast stress granules. This suggests that the biophysical principles that drive RNA self-assembly in vitro contribute to determining the stress granule transcriptome. Consistent with RNA self-assembly contributing to RNP granule formation, an excess of purified RNA injected into C. elegans syncytium coalesces into droplet-like assemblies. We suggest that diverse RNA-RNA interactions in trans contribute to RNP granule formation and may explain the prevalence of large RNA-protein assemblies in eukaryotic cells.

2017-05-23 | GSE99170 | GEO

A Ribonuclease Coordinates siRNA Amplification and mRNA Cleavage during RNAi

Project description:Effective silencing by RNA-interference (RNAi) depends on mechanisms that amplify and propagate the silencing signal. In some organisms, small-interfering (si) RNAs are amplified from target mRNAs by RNA-dependent RNA polymerase (RdRP). Both RdRP recruitment and mRNA silencing require Argonaute proteins, which are generally thought to degrade RNAi targets by directly cleaving them. However in C. elegans, the enzymatic activity of the primary Argonaute, RDE-1, is not required for silencing activity. We show that RDE-1 can instead recruit an endoribonuclease, RDE-8, to target RNA. RDE-8 can cleave RNA in vitro and is needed for the production of 3′ uridylated fragments of target mRNA in vivo. We also find that RDE-8 promotes RdRP activity, thereby ensuring amplification of siRNAs. Together, our findings suggest a model in which RDE-8 cleaves target mRNAs to mediate silencing, while generating 3’ uridylated mRNA fragments to serve as templates for the RdRP-directed amplification of the silencing signal. We examined the role of rde-8 in C. elegans small RNA biogenesis pathways, including endogenous and exogenous RNAi pathways. We performed 3' RACE seq from the sel-1 target mRNA and correlate with small RNAs from wild type, rde-8 and rde-8 transgenic strains after sel-1(RNAi) for different lengths of time.

2015-01-29 | E-GEOD-59300 | biostudies-arrayexpress

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