CDE-1 suppresses the production of risiRNA by coupling polyuridylation and degradation of 26S rRNA

AbstractAntisense ribosomal siRNAs (risiRNAs) downregulate pre-rRNAs through the nuclear RNAi pathway in Caenorhabditis elegans. However, the biogenesis and regulation of risiRNAs remain obscure. Previously, we showed that 26S rRNAs are uridylated at the 3’-ends by an unknown terminal polyuridylation polymerase before the rRNAs are degraded by a 3’ to 5’ exoribonuclease SUSI-1(ceDIS3L2). There are three polyuridylation polymerases, CDE-1, PUP-2, and PUP-3, in C. elegans. Here, we found that CDE-1 is specifically involved in suppressing risiRNA production. CDE-1 localizes to perinuclear granules in the germline and uridylates both Argonaute-associated 22G-RNAs and 26S rRNAs at the 3’-ends. Immunoprecipitation followed by mass spectrometry (IP-MS) revealed that CDE-1 interacts with SUSI-1(ceDIS3L2). Consistent with those results, both CDE-1 and SUSI-1(ceDIS3L2) are required for the inheritance of RNAi. Therefore, this work identified a rRNA surveillance machinery of rRNAs that couples terminal polyuridylation and degradation.

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Genetic or Toxicant-Induced Disruption of Vesicular Monoamine Storage and Global Metabolic Profiling in Caenorhabditis elegans

Toxicological Sciences ◽

10.1093/toxsci/kfab011 ◽

2021 ◽

Author(s):

Joshua M Bradner ◽

Vrinda Kalia ◽

Fion K Lau ◽

Monica Sharma ◽

Meghan L Bucher ◽

...

Keyword(s):

Mass Spectrometry ◽

Caenorhabditis Elegans ◽

High Resolution ◽

High Resolution Mass Spectrometry ◽

Dopamine Neurons ◽

Vesicular Monoamine Transporter ◽

Monoamine Transporter ◽

C Elegans ◽

High Resolution Mass ◽

Resolution Mass

Abstract The proper storage and release of monoamines contributes to a wide range of neuronal activity. Here, we examine the effects of altered vesicular monoamine transport in the nematode Caenorhabditis elegans. The gene cat-1 is responsible for the encoding of the vesicular monoamine transporter (VMAT) in C. elegans and is analogous to the mammalian vesicular monoamine transporter 2 (VMAT2). Our laboratory has previously shown that reduced VMAT2 activity confers vulnerability on catecholamine neurons in mice. The purpose of this article was to determine whether this function is conserved and to determine the impact of reduced VMAT activity in C. elegans. Here we show that deletion of cat-1/VMAT increases sensitivity to the neurotoxicant 1-methyl-4-phenylpyridinium (MPP+) as measured by enhanced degeneration of dopamine neurons. Reduced cat-1/VMAT also induces changes in dopamine-mediated behaviors. High-resolution mass spectrometry-based metabolomics in the whole organism reveals changes in amino acid metabolism, including tyrosine metabolism in the cat-1/VMAT mutants. Treatment with MPP+ disrupted tryptophan metabolism. Both conditions altered glycerophospholipid metabolism, suggesting a convergent pathway of neuronal dysfunction. Our results demonstrate the evolutionarily conserved nature of monoamine function in C. elegans and further suggest that high-resolution mass spectrometry-based metabolomics can be used in this model to study environmental and genetic contributors to complex human disease.

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L-Ascorbate Biosynthesis Involves Carbon Skeleton Rearrangement in the Nematode Caenorhabditis elegans

Metabolites ◽

10.3390/metabo10080334 ◽

2020 ◽

Vol 10 (8) ◽

pp. 334

Author(s):

Yukinori Yabuta ◽

Ryuta Nagata ◽

Yuka Aoki ◽

Ayumi Kariya ◽

Kousuke Wada ◽

...

Keyword(s):

Mass Spectrometry ◽

Caenorhabditis Elegans ◽

Biosynthetic Pathway ◽

Carbon Skeleton ◽

Gas Chromatography Mass Spectrometry ◽

Knockout Mutant ◽

Nematode Caenorhabditis Elegans ◽

C Elegans ◽

Collagen Protein ◽

Ascorbate Biosynthesis

Ascorbate (AsA) is required as a cofactor and is widely distributed in plants and animals. Recently, it has been suggested that the nematode Caenorhabditis elegans also synthesizes AsA. However, its biosynthetic pathway is still unknown. To further understand AsA biosynthesis in C. elegans, we analyzed the incorporation of the 13C atom into AsA using gas chromatography-mass spectrometry (GC-MS) in worms fed with D-Glc (1-13C)-labeled Escherichia coli. GC-MS analysis revealed that AsA biosynthesis in C. elegans, similarly to that in mammalian systems, involves carbon skeleton rearrangement. The addition of L-gulono-1,4-lactone, an AsA precursor in the mammalian pathway, significantly increased AsA level in C. elegans, whereas the addition of L-galactono-1,4-lactone, an AsA precursor in the plant and Euglena pathway, did not affect AsA level. The suppression of E03H4.3 (an ortholog of gluconolactonase) or the deficiency of F54D5.12 (an ortholog of L-gulono-1,4-lactone oxidase) significantly decreased AsA level in C. elegans. Although N2- and AsA-deficient F54D5.12 knockout mutant worm (tm6671) morphologies and the ratio of collagen to non-collagen protein did not show any significant differences, the mutant worms exhibited increased malondialdehyde levels and reduced lifespan compared with the N2 worms. In conclusion, our findings indicate that the AsA biosynthetic pathway is similar in C. elegans and mammals.

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A deletion polymorphism in the Caenorhabditis elegans RIG-I homolog disables viral RNA dicing and antiviral immunity

eLife ◽

10.7554/elife.00994 ◽

2013 ◽

Vol 2 ◽

Cited By ~ 91

Author(s):

Alyson Ashe ◽

Tony Bélicard ◽

Jérémie Le Pen ◽

Peter Sarkies ◽

Lise Frézal ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Genome Wide Association Study ◽

Rna Virus ◽

Rnai Pathway ◽

Animal Cells ◽

C Elegans ◽

Base Pair Deletion ◽

A Genome ◽

Positive Strand Rna ◽

Viral Recognition

RNA interference defends against viral infection in plant and animal cells. The nematode Caenorhabditis elegans and its natural pathogen, the positive-strand RNA virus Orsay, have recently emerged as a new animal model of host-virus interaction. Using a genome-wide association study in C. elegans wild populations and quantitative trait locus mapping, we identify a 159 base-pair deletion in the conserved drh-1 gene (encoding a RIG-I-like helicase) as a major determinant of viral sensitivity. We show that DRH-1 is required for the initiation of an antiviral RNAi pathway and the generation of virus-derived siRNAs (viRNAs). In mammals, RIG-I-domain containing proteins trigger an interferon-based innate immunity pathway in response to RNA virus infection. Our work in C. elegans demonstrates that the RIG-I domain has an ancient role in viral recognition. We propose that RIG-I acts as modular viral recognition factor that couples viral recognition to different effector pathways including RNAi and interferon responses.

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Caenorhabditis elegans nuclear RNAi factor SET-32 deposits the transgenerational histone modification, H3K23me3

eLife ◽

10.7554/elife.54309 ◽

2020 ◽

Vol 9 ◽

Author(s):

Lianna Schwartz-Orbach ◽

Chenzhen Zhang ◽

Simone Sidoli ◽

Richa Amin ◽

Diljeet Kaur ◽

...

Keyword(s):

Chromatin Modification ◽

Epigenetic Inheritance ◽

Rnai Pathway ◽

C Elegans ◽

And Function ◽

Nuclear Rnai ◽

Factor Set

Nuclear RNAi provides a highly tractable system to study RNA-mediated chromatin changes and epigenetic inheritance. Recent studies have indicated that the regulation and function of nuclear RNAi-mediated heterochromatin are highly complex. Our knowledge of histone modifications and the corresponding histonemodifying enzymes involved in the system remains limited. In this study, we show that the heterochromatin mark, H3K23me3, is induced by nuclear RNAi at both exogenous and endogenous targets in C. elegans. In addition, dsRNA-induced H3K23me3 can persist for multiple generations after the dsRNA exposure has stopped. We demonstrate that the histone methyltransferase SET-32, methylates H3K23 in vitro. Both set-32 and the germline nuclear RNAi Argonaute, hrde-1, are required for nuclear RNAi-induced H3K23me3 in vivo. Our data poise H3K23me3 as an additional chromatin modification in the nuclear RNAi pathway and provides the field with a new target for uncovering the role of heterochromatin in transgenerational epigenetic silencing.

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

eLife ◽

10.7554/elife.35478 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 1

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.

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Erroneous ribosomal RNAs promote the generation of antisense ribosomal siRNA

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1800974115 ◽

2018 ◽

Vol 115 (40) ◽

pp. 10082-10087 ◽

Cited By ~ 13

Author(s):

Chengming Zhu ◽

Qi Yan ◽

Chenchun Weng ◽

Xinhao Hou ◽

Hui Mao ◽

...

Keyword(s):

Ribosome Biogenesis ◽

18S Rrna ◽

Rnai Pathway ◽

Rrna Processing ◽

Reverse Genetic ◽

Genetic Screens ◽

Multistep Process ◽

Quality Control Mechanism ◽

26S Rrna ◽

Nuclear Rnai

Ribosome biogenesis is a multistep process, during which mistakes can occur at any step of pre-rRNA processing, modification, and ribosome assembly. Misprocessed rRNAs are usually detected and degraded by surveillance machineries. Recently, we identified a class of antisense ribosomal siRNAs (risiRNAs) that down-regulate pre-rRNAs through the nuclear RNAi pathway. To further understand the biological roles of risiRNAs, we conducted both forward and reverse genetic screens to search for more suppressor of siRNA (susi) mutants. We isolated a number of genes that are broadly conserved from yeast to humans and are involved in pre-rRNA modification and processing. Among them, SUSI-2(ceRRP8) is homologous to human RRP8 and engages in m1A methylation of the 26S rRNA. C27F2.4(ceBUD23) is an m7G-methyltransferase of the 18S rRNA. E02H1.1(ceDIMT1L) is a predicted m6(2)Am6(2)A-methyltransferase of the 18S rRNA. Mutation of these genes led to a deficiency in modification of rRNAs and elicited accumulation of risiRNAs, which further triggered the cytoplasmic-to-nuclear and cytoplasmic-to-nucleolar translocations of the Argonaute protein NRDE-3. The rRNA processing deficiency also resulted in accumulation of risiRNAs. We also isolated SUSI-3(RIOK-1), which is similar to human RIOK1, that cleaves the 20S rRNA to 18S. We further utilized RNAi and CRISPR-Cas9 technologies to perform candidate-based reverse genetic screens and identified additional pre-rRNA processing factors that suppressed risiRNA production. Therefore, we concluded that erroneous rRNAs can trigger risiRNA generation and subsequently, turn on the nuclear RNAi-mediated gene silencing pathway to inhibit pre-rRNA expression, which may provide a quality control mechanism to maintain homeostasis of rRNAs.

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HERI-1 is a Chromodomain Protein that Negatively Regulates Transgenerational Epigenetic Inheritance

10.1101/384792 ◽

2018 ◽

Author(s):

Roberto Perales ◽

Daniel Pagano ◽

Gang Wan ◽

Brandon Fields ◽

Arneet L. Saltzman ◽

...

Keyword(s):

Gene Silencing ◽

Epigenetic Inheritance ◽

Normal Function ◽

Negative Regulator ◽

Rnai Pathway ◽

Transcriptional Gene Silencing ◽

Direct Role ◽

Transgenerational Epigenetic Inheritance ◽

C Elegans ◽

Nuclear Rnai

AbstractTransgenerational epigenetic inheritance (TEI) is the inheritance of epigenetic information for two or more generations. In most cases, TEI is limited to 2-3 generations. This short-term nature of TEI could be set by innate biochemical limitations to TEI or by genetically encoded systems that actively limit TEI. dsRNA-mediated gene silencing (RNAi) can be inherited in C. elegans (termed RNAi inheritance or RNA-directed TEI). To identify systems that might actively limit RNA-directed TEI, we conducted a forward genetic screen for factors whose mutation enhanced RNAi inheritance. This screen identified the gene heritable enhancer of RNAi (heri-1), whose mutation causes RNAi inheritance to last longer (>20 generations) than normal. heri-1 encodes a protein with a chromodomain and a kinase-homology domain that is expressed in germ cells and localizes to nuclei. In C. elegans, a nuclear branch of the RNAi pathway (nuclear RNAi or NRDE pathway) is required for RNAi inheritance. We find that this NRDE pathway is hyper-responsive to RNAi in heri-1 mutant animals, suggesting that a normal function of HERI-1 is to limit nuclear RNAi and that limiting nuclear RNAi may be the mechanism by which HERI-1 limits RNAi inheritance. Interestingly, we find that HERI-1 binds to genes targeted by RNAi, suggesting that HERI-1 may have a direct role in limiting nuclear RNAi and, therefore, RNAi inheritance. Surprisingly, recruitment of the negative regulator HERI-1 to genes depends upon that same NRDE factors that drive co-transcriptional gene silencing during RNAi inheritance. We therefore speculate that the generational perdurance of RNAi inheritance is set by competing pro- and anti-silencing outputs of the NRDE nuclear RNAi machinery.

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A transgenerational role of the germline nuclear RNAi pathway in repressing heat stress-induced transcriptional activation in C. elegans

Epigenetics & Chromatin ◽

10.1186/s13072-016-0052-x ◽

2016 ◽

Vol 9 (1) ◽

Cited By ~ 62

Author(s):

Julie Zhouli Ni ◽

Natallia Kalinava ◽

Esteban Chen ◽

Alex Huang ◽

Thi Trinh ◽

...

Keyword(s):

Heat Stress ◽

Transcriptional Activation ◽

Rnai Pathway ◽

C Elegans ◽

Nuclear Rnai

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

10.1101/260240 ◽

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.

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Quo Vadis Caenorhabditis elegans Metabolomics—A Review of Current Methods and Applications to Explore Metabolism in the Nematode

Metabolites ◽

10.3390/metabo11050284 ◽

2021 ◽

Vol 11 (5) ◽

pp. 284

Author(s):

Liesa Salzer ◽

Michael Witting

Keyword(s):

Mass Spectrometry ◽

Caenorhabditis Elegans ◽

Model Organism ◽

General Information ◽

Gas Chromatography Mass Spectrometry ◽

C Elegans ◽

Advantages And Disadvantages ◽

Chromatography Mass Spectrometry ◽

Quo Vadis ◽

Analytical Approaches

Metabolomics and lipidomics recently gained interest in the model organism Caenorhabditis elegans (C. elegans). The fast development, easy cultivation and existing forward and reverse genetic tools make the small nematode an ideal organism for metabolic investigations in development, aging, different disease models, infection, or toxicology research. The conducted type of analysis is strongly depending on the biological question and requires different analytical approaches. Metabolomic analyses in C. elegans have been performed using nuclear magnetic resonance (NMR) spectroscopy, direct infusion mass spectrometry (DI-MS), gas-chromatography mass spectrometry (GC-MS) and liquid chromatography mass spectrometry (LC-MS) or combinations of them. In this review we provide general information on the employed techniques and their advantages and disadvantages in regard to C. elegans metabolomics. Additionally, we reviewed different fields of application, e.g., longevity, starvation, aging, development or metabolism of secondary metabolites such as ascarosides or maradolipids. We also summarised applied bioinformatic tools that recently have been used for the evaluation of metabolomics or lipidomics data from C. elegans. Lastly, we curated metabolites and lipids from the reviewed literature, enabling a prototypic collection which serves as basis for a future C. elegans specific metabolome database.

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