P granules protect RNA interference genes from silencing by piRNAs

SUMMARYP granules are perinuclear condensates in C. elegans germ cells proposed to serve as hubs for self/non-self RNA discrimination by Argonautes. We report that a mutant (meg-3 meg-4) that does not assemble P granules in primordial germ cells loses competence for RNA-interference over several generations and accumulates silencing small RNAs against hundreds of endogenous genes, including the RNA-interference genes rde-11 and sid-1. In wild-type, rde-11 and sid-1 transcripts are heavily targeted by piRNAs, accumulate in P granules, but maintain expression. In the primordial germ cells of meg-3 meg-4 mutants, rde-11 and sid-1 transcripts disperse in the cytoplasm with the small RNA biogenesis machinery, become hyper-targeted by secondary sRNAs, and are eventually silenced. Silencing requires the PIWI-class Argonaute PRG-1 and the nuclear Argonaute HRDE-1 that maintains trans-generational silencing of piRNA targets. These observations support a “safe harbor” model for P granules in protecting germline transcripts from piRNA-initiated silencing.

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Germ Granules Functions are Memorized by Transgenerationally Inherited Small RNAs

10.1101/576934 ◽

2019 ◽

Cited By ~ 2

Author(s):

Itamar Lev ◽

Itai Antoine Toker ◽

Yael Mor ◽

Anat Nitzan ◽

Guy Weintraub ◽

...

Keyword(s):

Small Rna ◽

Small Rnas ◽

Early Embryo ◽

Small Rna Sequencing ◽

Wild Type ◽

P Granules ◽

Multiple Generations ◽

C Elegans ◽

The Core ◽

Germ Granules

AbstractInC. elegansnematodes, components of liquid-like germ granules were shown to be required for transgenerational small RNA inheritance. Surprisingly, we show here that mutants with defective germ granules (pptr-1,meg-3/4,pgl-1) can nevertheless inherit potent small RNA-based silencing responses, but some of the mutants lose this ability after many generations of homozygosity. Animals mutated inpptr-1, which is required for stabilization of P granules in the early embryo, display extremely strong heritable RNAi responses, which last for tens of generations, long after the responses in wild type animals peter out. The phenotype of mutants defective in the core germ granules proteins MEG-3 and MEG-4, depends on the genotype of the ancestors: Mutants that derive from maternal lineages that had functional MEG-3 and MEG-4 proteins exhibit enhanced RNAi inheritance for multiple generations. While functional ancestralmeg-3/4alleles correct, and even potentiates the ability of mutant descendants to inherit RNAi, defects in germ granules functions can be memorized as well; Wild type descendants that derive from lineages of mutants show impaired RNAi inheritance for many (>16) generations, although their germ granules are intact. Importantly, while P granules are maternally deposited, wild type progeny derived frommeg-3/4male mutants also show reduced RNAi inheritance. Unlike germ granules, small RNAs are inherited also from the sperm. Moreover, we find that the transgenerational effects that depend on the ancestral germ granules require the argonaute protein HRDE-1, which carries heritable small RNAs in the germline. Indeed, small RNA sequencing reveals imbalanced levels of many endogenous small RNAs in germ granules mutants. Strikingly, we find thathrde-1;meg-3/4triple mutants inherit RNAi, althoughhrde-1was previously thought to be essential for heritable silencing. We propose that germ granules sort and shape the RNA pool, and that small RNA inheritance memorizes this activity for multiple generations.

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Transformation of the germ line into muscle in mes-1 mutant embryos of C. elegans

Development ◽

10.1242/dev.121.9.2961 ◽

1995 ◽

Vol 121 (9) ◽

pp. 2961-2972 ◽

Cited By ~ 8

Author(s):

S. Strome ◽

P. Martin ◽

E. Schierenberg ◽

J. Paulsen

Keyword(s):

Cell Fate ◽

Germ Cells ◽

Germ Line ◽

Primordial Germ Cells ◽

Primordial Germ Cell ◽

Wild Type ◽

C Elegans ◽

Stem Cell Divisions ◽

Asymmetric Partitioning

Mutations in the maternal-effect sterile gene mes-1 cause the offspring of homozygous mutant mothers to develop into sterile adults. Lineage analysis revealed that mutant offspring are sterile because they fail to form primordial germ cells during embryogenesis. In wild-type embryos, the primordial germ cell P4 is generated via a series of four unequal stem-cell divisions of the zygote. mes-1 embryos display a premature and progressive loss of polarity in these divisions: P0 and P1 undergo apparently normal unequal divisions and cytoplasmic partitioning, but P2 (in some embryos) and P3 (in most embryos) display defects in cleavage asymmetry and fail to partition lineage-specific components to only one daughter cell. As an apparent consequence of these defects, P4 is transformed into a muscle precursor, like its somatic sister cell D, and generates up to 20 body muscle cells instead of germ cells. Our results show that the wild-type mes-1 gene participates in promoting unequal germ-line divisions and asymmetric partitioning events and thus the determination of cell fate in early C. elegans embryos.

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The Influence of Competition Among C. elegans Small RNA Pathways on Development

Genes ◽

10.3390/genes3040671 ◽

2012 ◽

Vol 3 (4) ◽

pp. 671-685 ◽

Cited By ~ 15

Author(s):

Jimmy J. Zhuang ◽

Craig P. Hunter

Keyword(s):

Rna Interference ◽

Caenorhabditis Elegans ◽

Small Rna ◽

Small Rnas ◽

Wild Type ◽

C Elegans ◽

Exogenous Rna ◽

Rna Pathways ◽

Sirna Pathway ◽

The Relationship

Small RNAs play a variety of regulatory roles, including highly conserved developmental functions. Caenorhabditis elegans not only possesses most known small RNA pathways, it is also an easy system to study their roles and interactions during development. It has been proposed that in C. elegans, some small RNA pathways compete for access to common limiting resources. The strongest evidence supporting this model is that disrupting the production or stability of endogenous short interfering RNAs (endo-siRNAs) enhances sensitivity to experimentally induced exogenous RNA interference (exo-RNAi). Here, we examine the relationship between the endo-siRNA and microRNA (miRNA) pathways, and find that, consistent with competition among these endogenous small RNA pathways, endo-siRNA pathway mutants may enhance miRNA efficacy. Furthermore, we show that exo-RNAi may also compete with both endo-siRNAs and miRNAs. Our data thus provide support that all known Dicer-dependent small RNA pathways may compete for limiting common resources. Finally, we observed that both endo-siRNA mutants and animals experiencing exo-RNAi have increased expression of miRNA-regulated stage-specific developmental genes. These observations suggest that perturbing the small RNA flux and/or the induction of exo-RNAi, even in wild-type animals, may impact development via effects on the endo-RNAi and microRNA pathways.

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Perinuclear P granules are the principal sites of mRNA export in adult C. elegans germ cells

Development ◽

10.1242/dev.044255 ◽

2010 ◽

Vol 137 (8) ◽

pp. 1305-1314 ◽

Cited By ~ 102

Author(s):

U. Sheth ◽

J. Pitt ◽

S. Dennis ◽

J. R. Priess

Keyword(s):

Germ Cells ◽

Mrna Export ◽

P Granules ◽

C Elegans

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

Development ◽

10.1242/dev.015552 ◽

2008 ◽

Vol 135 (5) ◽

pp. 983-993 ◽

Cited By ~ 49

Author(s):

C. A. Spike ◽

J. Bader ◽

V. Reinke ◽

S. Strome

Keyword(s):

Rna Interference ◽

Germ Cells ◽

C Elegans

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RNA Interference Mechanisms and Applications in Plant Pathology

Annual Review of Phytopathology ◽

10.1146/annurev-phyto-080417-050044 ◽

2018 ◽

Vol 56 (1) ◽

pp. 581-610 ◽

Cited By ~ 42

Author(s):

Cristina Rosa ◽

Yen-Wen Kuo ◽

Hada Wuriyanghan ◽

Bryce W. Falk

Keyword(s):

Plant Pathology ◽

Rna Interference ◽

Gene Function ◽

Small Rnas ◽

Plant Pathogens ◽

Common Ancestor ◽

Plant Protection ◽

Plant Diseases ◽

Antiviral Defense ◽

Endogenous Genes

The origin of RNA interference (RNAi), the cell sentinel system widely shared among eukaryotes that recognizes RNAs and specifically degrades or prevents their translation in cells, is suggested to predate the last eukaryote common ancestor ( 138 ). Of particular relevance to plant pathology is that in plants, but also in some fungi, insects, and lower eukaryotes, RNAi is a primary and effective antiviral defense, and recent studies have revealed that small RNAs (sRNAs) involved in RNAi play important roles in other plant diseases, including those caused by cellular plant pathogens. Because of this, and because RNAi can be manipulated to interfere with the expression of endogenous genes in an intra- or interspecific manner, RNAi has been used as a tool in studies of gene function but also for plant protection. Here, we review the discovery of RNAi, canonical mechanisms, experimental and translational applications, and new RNA-based technologies of importance to plant pathology.

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Identification of tissues and patterning events required for distinct steps in early migration of zebrafish primordial germ cells

Development ◽

10.1242/dev.126.23.5295 ◽

1999 ◽

Vol 126 (23) ◽

pp. 5295-5307 ◽

Cited By ~ 5

Author(s):

G. Weidinger ◽

U. Wolke ◽

M. Koprunner ◽

M. Klinger ◽

E. Raz

Keyword(s):

Germ Cells ◽

Primordial Germ Cells ◽

Cell Lineage ◽

Paraxial Mesoderm ◽

Molecular Characteristics ◽

Dorsoventral Patterning ◽

Wild Type ◽

Migration Process ◽

Early Migration ◽

Somatic Tissues

In many organisms, the primordial germ cells have to migrate from the position where they are specified towards the developing gonad where they generate gametes. Extensive studies of the migration of primordial germ cells in Drosophila, mouse, chick and Xenopus have identified somatic tissues important for this process and demonstrated a role for specific molecules in directing the cells towards their target. In zebrafish, a unique situation is found in that the primordial germ cells, as marked by expression of vasa mRNA, are specified in random positions relative to the future embryonic axis. Hence, the migrating cells have to navigate towards their destination from various starting positions that differ among individual embryos. Here, we present a detailed description of the migration of the primordial germ cells during the first 24 hours of wild-type zebrafish embryonic development. We define six distinct steps of migration bringing the primordial germ cells from their random positions before gastrulation to form two cell clusters on either side of the midline by the end of the first day of development. To obtain information on the origin of the positional cues provided to the germ cells by somatic tissues during their migration, we analyzed the migration pattern in mutants, including spadetail, swirl, chordino, floating head, cloche, knypek and no isthmus. In mutants with defects in axial structures, paraxial mesoderm or dorsoventral patterning, we find that certain steps of the migration process are specifically affected. We show that the paraxial mesoderm is important for providing proper anteroposterior information to the migrating primordial germ cells and that these cells can respond to changes in the global dorsoventral coordinates. In certain mutants, we observe accumulation of ectopic cells in different regions of the embryo. These ectopic cells can retain both morphological and molecular characteristics of primordial germ cells, suggesting that, in zebrafish at the early stages tested, the vasa-expressing cells are committed to the germ cell lineage.

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Analysis of RNA associated with P granules in germ cells of C. elegans adults

Development ◽

10.1242/dev.128.8.1287 ◽

2001 ◽

Vol 128 (8) ◽

pp. 1287-1298 ◽

Cited By ~ 4

Author(s):

J.A. Schisa ◽

J.N. Pitt ◽

J.R. Priess

Keyword(s):

Germ Cells ◽

Rna Binding ◽

Nuclear Surface ◽

Binding Motifs ◽

P Granules ◽

C Elegans ◽

Specific Mrnas ◽

Tubulin Mrna ◽

Protein Components ◽

Cytoplasmic Structures

P granules are cytoplasmic structures of unknown function that are associated with germ nuclei in the C. elegans gonad, and are localized exclusively to germ cells, or germ cell precursors, throughout the life cycle. All the known protein components of P granules contain putative RNA-binding motifs, suggesting that RNA is involved in either the structure or function of the granules. However, no specific mRNAs have been identified within P granules in the gonad. We show here that P granules normally contain a low level of RNA, and describe conditions that increase this level. We present evidence that several, diverse mRNAs, including pos-1, mex-1, par-3, skn-1, nos-2 and gld-1 mRNA, are present at least transiently within P granules. In contrast, actin and tubulin mRNA and rRNA are either not present in P granules, or are present at relatively low levels. We show that pgl-1 and the glh (Vasa-related) gene family, which encode protein components of P granules, do not appear essential for RNA to concentrate in P granules; these proteins may instead function in events that are a prerequisite for RNAs to be transported efficiently from the nuclear surface.

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Trans-generational epigenetic regulation in C. elegans primordial germ cells

Developmental Biology ◽

10.1016/j.ydbio.2010.05.363 ◽

2010 ◽

Vol 344 (1) ◽

pp. 513

Author(s):

Hirofumi Furuhashi ◽

Teruaki Takasaki ◽

Andreas Rechtsteiner ◽

Tengguo Li ◽

Hiroshi Kimura ◽

...

Keyword(s):

Germ Cells ◽

Epigenetic Regulation ◽

Primordial Germ Cells ◽

C Elegans

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Nuage condensates: accelerators or circuit breakers for sRNA silencing pathways?

RNA ◽

10.1261/rna.079003.121 ◽

2021 ◽

pp. rna.079003.121

Author(s):

John Paul Tsu Ouyang ◽

Geraldine Seydoux

Keyword(s):

Germ Cells ◽

Small Rnas ◽

Circuit Breakers ◽

C Elegans ◽

Silencing Pathways ◽

And Function

Nuage are RNA-rich condensates that assemble around the nuclei of developing germ cells. Many proteins required for the biogenesis and function of silencing small RNAs (sRNAs) enrich in nuage, and it is often assumed that nuage is the cellular site where sRNAs are synthesized and encounter target transcripts for silencing. Using C. elegans as a model, we examine the complex multi-condensate architecture of nuage and review evidence for compartmentalization of silencing pathways. We consider the possibility that nuage condensates balance the activity of competing sRNA pathways and serve to limit, rather than enhance, sRNA amplification to protect transcripts from dangerous run-away silencing.

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