scholarly journals A team of heterochromatin factors collaborates with small RNA pathways to combat repetitive elements and germline stress

2017 ◽  
Author(s):  
Alicia N. McMurchy ◽  
Przemyslaw Stempor ◽  
Tessa Gaarenstroom ◽  
Brian Wysolmerski ◽  
Yan Dong ◽  
...  
Keyword(s):  
Small Rna ◽  
Germ Line ◽  
Large Fraction ◽  
Repetitive Sequences ◽  
Genotoxic Stress ◽  
Repetitive Elements ◽  
C Elegans ◽  
Genes Encoding ◽  
Rna Pathways ◽  
Nuclear Rnai

AbstractRepetitive sequences derived from transposons make up a large fraction of eukaryotic genomes and must be silenced to protect genome integrity. Repetitive elements are often found in heterochromatin; however, the roles and interactions of heterochromatin proteins in repeat regulation are poorly understood. Here we show that a diverse set of C. elegans heterochromatin proteins act together with the piRNA and nuclear RNAi pathways to silence repetitive elements and prevent genotoxic stress in the germ line. Mutants in genes encoding HPL-2/HP1, LIN-13, LIN-61, LET-418/Mi-2, and H3K9me2 histone methyltransferase MET-2/SETDB1 also show functionally redundant sterility, increased germline apoptosis, DNA repair defects, and interactions with small RNA pathways. Remarkably, fertility of heterochromatin mutants could be partially restored by inhibiting cep-1/p53, endogenous meiotic double strand breaks, or the expression of MIRAGE1 DNA transposons. Functional redundancy among these factors and pathways underlies the importance of safeguarding the genome through multiple means.

scholarly journals A team of heterochromatin factors collaborates with small RNA pathways to combat repetitive elements and germline stress

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Alicia N McMurchy ◽  
Przemyslaw Stempor ◽  
Tessa Gaarenstroom ◽  
Brian Wysolmerski ◽  
Yan Dong ◽  
...  
Keyword(s):  
Small Rna ◽  
Germ Line ◽  
Large Fraction ◽  
Repetitive Sequences ◽  
Genotoxic Stress ◽  
Repetitive Elements ◽  
C Elegans ◽  
Genes Encoding ◽  
Rna Pathways ◽  
Nuclear Rnai

Repetitive sequences derived from transposons make up a large fraction of eukaryotic genomes and must be silenced to protect genome integrity. Repetitive elements are often found in heterochromatin; however, the roles and interactions of heterochromatin proteins in repeat regulation are poorly understood. Here we show that a diverse set of C. elegans heterochromatin proteins act together with the piRNA and nuclear RNAi pathways to silence repetitive elements and prevent genotoxic stress in the germ line. Mutants in genes encoding HPL-2/HP1, LIN-13, LIN-61, LET-418/Mi-2, and H3K9me2 histone methyltransferase MET-2/SETDB1 also show functionally redundant sterility, increased germline apoptosis, DNA repair defects, and interactions with small RNA pathways. Remarkably, fertility of heterochromatin mutants could be partially restored by inhibiting cep-1/p53, endogenous meiotic double strand breaks, or the expression of MIRAGE1 DNA transposons. Functional redundancy among factors and pathways underlies the importance of safeguarding the genome through multiple means.

scholarly journals The TRIM-NHL protein NHL-2 is a co-factor in the nuclear and somatic RNAi pathways in C. elegans

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Gregory M Davis ◽  
Shikui Tu ◽  
Joshua WT Anderson ◽  
Rhys N Colson ◽  
Menachem J Gunzburg ◽  
...  
Keyword(s):  
Small Rna ◽  
Rna Binding ◽  
Meiotic Chromosome ◽  
Rna Stability ◽  
Rnai Pathway ◽  
Regulatory Pathways ◽  
C Elegans ◽  
Bona Fide ◽  
Rna Pathways ◽  
Nuclear Rnai

Proper regulation of germline gene expression is essential for fertility and maintaining species integrity. In the C. elegans germline, a diverse repertoire of regulatory pathways promote the expression of endogenous germline genes and limit the expression of deleterious transcripts to maintain genome homeostasis. Here we show that the conserved TRIM-NHL protein, NHL-2, plays an essential role in the C. elegans germline, modulating germline chromatin and meiotic chromosome organization. We uncover a role for NHL-2 as a co-factor in both positively (CSR-1) and negatively (HRDE-1) acting germline 22G-small RNA pathways and the somatic nuclear RNAi pathway. Furthermore, we demonstrate that NHL-2 is a bona fide RNA binding protein and, along with RNA-seq data point to a small RNA independent role for NHL-2 in regulating transcripts at the level of RNA stability. Collectively, our data implicate NHL-2 as an essential hub of gene regulatory activity in both the germline and soma.

scholarly journals The TRIM-NHL protein NHL-2 is a Novel Co-Factor of the CSR-1 and HRDE-1 22G-RNA Pathways

2018 ◽  
Author(s):  
Gregory M. Davis ◽  
Shikui Tu ◽  
Rhys N. Colson ◽  
Joshua W. T. Anderson ◽  
Menachem J. Gunzburg ◽  
...  
Keyword(s):  
Small Rna ◽  
Rna Binding ◽  
Meiotic Chromosome ◽  
Rna Stability ◽  
Rnai Pathway ◽  
Regulatory Pathways ◽  
C Elegans ◽  
Bona Fide ◽  
Rna Pathways ◽  
Nuclear Rnai

ABSTRACTProper regulation of germline gene expression is essential for fertility and maintaining species integrity. In the C. elegans germline, a diverse repertoire of regulatory pathways promote the expression of endogenous germline genes and limit the expression of deleterious transcripts to maintain genome homeostasis. Here we show that the conserved TRIM-NHL protein, NHL-2, plays an essential role in the C. elegans germline, modulating germline chromatin and meiotic chromosome organization. We uncover a role for NHL-2 as a co-factor in both positively (CSR-1) and negatively (HRDE-1) acting germline 22G-small RNA pathways and the somatic nuclear RNAi pathway. Furthermore, we demonstrate that NHL-2 is a bona fide RNA binding protein and, along with RNA-seq data point to a small RNA independent role for NHL-2 in regulating transcripts at the level of RNA stability. Collectively, our data implicate NHL-2 as an essential hub of gene regulatory activity in both the germline and soma.

scholarly journals Nematode Small RNA Pathways in the Absence of piRNAs

2021 ◽  
Author(s):  
Maxim Zagoskin ◽  
Jianbin Wang ◽  
Ashley T. Neff ◽  
Giovana M.B. Veronezi ◽  
Richard E. Davis
Keyword(s):  
Small Rna ◽  
Small Rnas ◽  
Parasitic Nematode ◽  
Repetitive Sequences ◽  
C Elegans ◽  
Early Embryos ◽  
Specific Mrnas ◽  
Rna Pathways ◽  
Pirna Pathway ◽  
Mature Mrnas

Small RNA pathways play diverse regulatory roles in the nematode C. elegans. However, our understanding of small RNA pathways, their conservation, and their roles in other nematodes is limited. Here, we analyzed small RNA pathways in the parasitic nematode Ascaris. Ascaris has ten Argonautes with five worm-specific Argonautes (WAGOs) that are associated with secondary 5'-triphosphate small RNAs (22-24G-RNAs). These Ascaris WAGOs and their small RNAs target repetitive sequences (WAGO-1, WAGO-2, WAGO-3, and NRDE-3) or mature mRNAs (CSR-1, NRDE-3, and WAGO-3) and are similar to the C. elegans mutator, nuclear, and CSR-1 small RNA pathways. Ascaris CSR-1 likely functions to "license" gene expression in the absence of an Ascaris piRNA pathway. Ascaris ALG-4 and its associated 26G-RNAs target and appear to repress specific mRNAs during meiosis in the testes. Notably, Ascaris WAGOs (WAGO-3 and NRDE-3) small RNAs change their targets between repetitive sequences and mRNAs during spermatogenesis or in early embryos illustrating target plasticity of these WAGOs. We provide a unique and comprehensive view of mRNA and small RNA expression throughout nematode spermatogenesis that illustrates the dynamics and flexibility of small RNA pathways. Overall, our study provides key insights into the conservation and divergence of nematode small RNA pathways.

scholarly journals The RNA phosphatase PIR-1 regulates endogenous small RNA pathways in C. elegans

2020 ◽  
Author(s):  
Daniel A. Chaves ◽  
Hui Dai ◽  
Lichao Li ◽  
James J. Moresco ◽  
Myung Eun Oh ◽  
...  
Keyword(s):  
Small Rna ◽  
C Elegans ◽  
Rna Pathways

scholarly journals The Conserved Intron Binding Protein EMB-4 Plays Differential Roles in Germline Small RNA Pathways of C. elegans

2017 ◽  
Vol 42 (3) ◽  
pp. 256-270.e6 ◽  
Author(s):  
Katarzyna M. Tyc ◽  
Amena Nabih ◽  
Monica Z. Wu ◽  
Christopher J. Wedeles ◽  
Julia A. Sobotka ◽  
...  
Keyword(s):  
Small Rna ◽  
Binding Protein ◽  
C Elegans ◽  
Rna Pathways

scholarly journals Author response: A team of heterochromatin factors collaborates with small RNA pathways to combat repetitive elements and germline stress

2017 ◽  
Author(s):  
Alicia N McMurchy ◽  
Przemyslaw Stempor ◽  
Tessa Gaarenstroom ◽  
Brian Wysolmerski ◽  
Yan Dong ◽  
...  
Keyword(s):  
Small Rna ◽  
Repetitive Elements ◽  
Author Response ◽  
Rna Pathways

Concepts and functions of small RNA pathways in C. elegans

2020 ◽  
Author(s):  
F. René Ketting ◽  
Luisa Cochella
Keyword(s):  
Small Rna ◽  
C Elegans ◽  
Rna Pathways

scholarly journals GTSF-1 is required for the formation of a functional RNA-dependent RNA Polymerase complex in C. elegans

2018 ◽  
Author(s):  
Miguel Vasconcelos Almeida ◽  
Sabrina Dietz ◽  
Stefan Redl ◽  
Emil Karaulanov ◽  
Andrea Hildebrandt ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Gene Silencing ◽  
Small Rna ◽  
Small Rnas ◽  
Zinc Fingers ◽  
Specific Factor ◽  
Rna Dependent Rna Polymerase ◽  
C Elegans ◽  
Argonaute Proteins ◽  
Rna Pathways

AbstractIn every domain of life, Argonaute proteins and their associated small RNAs regulate gene expression. Despite great conservation of Argonaute proteins throughout evolution, many proteins acting in small RNA pathways are not widely conserved. Gametocyte-specific factor 1 (Gtsf1) proteins, characterized by two tandem CHHC zinc fingers and an unstructured, acidic C-terminal tail, are conserved in animals and act in small RNA pathways. In fly and mouse, they are required for fertility and have been shown to interact with Piwi clade Argonautes. We identified T06A10.3 as the Caenorhabditis elegans Gtsf1 homolog and named it gtsf-1. Given its conserved nature and roles in Piwi-mediated gene silencing, we sought out to characterize GTSF-1 in the context of the small RNA pathways of C. elegans. Like its homologs, GTSF-1 is required for normal fertility. Surprisingly, we report that GTSF-1 is not required for Piwi-mediated gene silencing. Instead, gtsf-1 mutants show strong depletion of a class of endogenous small RNAs, known as 26G-RNAs, and fully phenocopy mutants lacking RRF-3, the RNA-dependent RNA Polymerase that synthesizes 26G-RNAs. We show, both in vivo and in vitro, that GTSF-1 specifically and robustly interacts with RRF-3 via its tandem CHHC zinc fingers. Furthermore, we demonstrate that GTSF-1 is required for the assembly of a larger RRF-3 and DCR-1-containing complex, also known as ERIC, thereby allowing for 26G-RNA generation. We propose that GTSF-1 homologs may similarly act to drive the assembly of larger complexes that subsequently act in small RNA production and/or in imposing small RNA-mediated silencing activities.

scholarly journals RppH can faithfully replace TAP to allow cloning of 5’-triphosphate carrying small RNAs

2018 ◽  
Author(s):  
Miguel Vasconcelos Almeida ◽  
António Miguel de Jesus Domingues ◽  
Hanna Lukas ◽  
Maria Mendez-Lago ◽  
René F. Ketting
Keyword(s):  
Small Rna ◽  
Small Rnas ◽  
Small Rna Sequencing ◽  
Cloning Efficiency ◽  
Library Preparation ◽  
Small Interfering Rnas ◽  
Rdrp Activity ◽  
C Elegans ◽  
Rna Pathways ◽  
Small Rna Species

AbstractRNA interference was first described in the nematode Caenorhabditis elegans. Ever since, several new endogenous small RNA pathways have been described and characterized to different degrees. Much like plants, but unlike Drosophila and mammals, worms have RNA-dependent RNA Polymerases (RdRPs) that directly synthesize small RNAs using other transcripts as a template. The very prominent secondary small interfering RNAs, also called 22G-RNAs, produced by the RdRPs RRF-1 and EGO-1 in C. elegans, maintain the 5’ triphosphate group, stemming from RdRP activity, also after loading into an Argonaute protein. This creates a technical issue, since 5’PPP groups decrease cloning efficiency for small RNA sequencing. To increase cloning efficiency of these small RNA species, a common practice in the field is the treatment of RNA samples, prior to library preparation, with Tobacco Acid pyrophosphatase (TAP). Recently, TAP production and supply was discontinued, so an alternative must be devised. We turned to RNA 5’ pyrophosphohydrolase (RppH), a commercially available pyrophosphatase isolated from E. coli. Here we directly compare TAP and RppH in their use for small RNA library preparation. We show that RppH-treated samples faithfully recapitulate TAP-treated samples. Specifically, there is enrichment for 22G-RNAs and mapped small RNA reads show no small RNA transcriptome-wide differences between RppH and TAP treatment. We propose that RppH can be used as a small RNA pyrophosphatase to enrich for triphosphorylated small RNA species and show that RppH- and TAP-derived datasets can be used in direct comparison.

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