Chromatin Compaction by Small RNAs and the Nuclear RNAi Machinery in C. elegans

Chromatin Compaction ◽

C Elegans ◽

Regulation Mechanisms ◽

AbstractDNA is organized and compacted into higher-order structures in order to fit within nuclei and to facilitate proper gene regulation. Mechanisms by which higher order chromatin structures are established and maintained are poorly understood. In C. elegans, nuclear-localized small RNAs engage the nuclear RNAi machinery to regulate gene expression and direct the post-translational modification of histone proteins. Here we confirm a recent report suggesting that nuclear small RNAs are required to initiate or maintain chromatin compaction states in C. elegans germ cells. Additionally, we show that experimentally provided small RNAs are sufficient to direct chromatin compaction and that this compaction requires the small RNA-binding Argonaute NRDE-3, the pre-mRNA associated factor NRDE-2, and the HP1-like protein HPL-2. Our results show that small RNAs, acting via the nuclear RNAi machinery and an HP1-like protein, are capable of driving chromatin compaction in C. elegans.

Chromatin Compaction by Small RNAs and the Nuclear RNAi Machinery in C. elegans

Scientific Reports ◽

10.1038/s41598-019-45052-y ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 7

Author(s):

Brandon D. Fields ◽

Scott Kennedy

Keyword(s):

Small Rnas ◽

Rnai Machinery ◽

Chromatin Compaction ◽

C Elegans ◽

E. coli OxyS non-coding RNA does not trigger RNAi in C. elegans

10.1101/013029 ◽

2014 ◽

Author(s):

Alper Akay ◽

Peter Sarkies ◽

Eric Alexander Miska

Keyword(s):

Small Rnas ◽

Biological Function ◽

Rapid Development ◽

Rnai Pathway ◽

Double Stranded Rna ◽

E Coli ◽

C Elegans ◽

Non Coding Rna ◽

Rnai Response

The discovery of RNA interference (RNAi) in C. elegans has had a major impact on scientific research, led to the rapid development of RNAi tools and has inspired RNA-based therapeutics. Astonishingly, nematodes, planaria and many insects take up double-stranded RNA (dsRNA) from their environment to elicit RNAi; the biological function of this mechanism is unclear. Recently, the E. coli OxyS non-coding RNA was shown to regulate gene expression in C. elegans when E. coli is offered as food. This was surprising given that C. elegans is unlikely to encounter E. coli in nature. To directly test the hypothesis that the E. coli OxyS non-coding RNA triggers the C. elegans RNAi pathway, we sequenced small RNAs from C. elegans after feeding with bacteria. We clearly demonstrate that the OxyS non-coding RNA does not trigger an RNAi response in C. elegans. We conclude that the biology of environmental RNAi remains to be discovered.

KH domain containing RNA-binding proteins coordinate with microRNAs to regulate Caenorhabditis elegans development

G3 Genes|Genome|Genetics ◽

10.1093/g3journal/jkab013 ◽

2021 ◽

Vol 11 (2) ◽

Author(s):

Dustin Haskell ◽

Anna Zinovyeva

Keyword(s):

Gene Expression ◽

Caenorhabditis Elegans ◽

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

C Elegans ◽

Binding Domains ◽

Kh Domain ◽

Regulate Gene

Abstract MicroRNAs (miRNAs) and RNA-binding proteins (RBPs) regulate gene expression at the post-transcriptional level, but the extent to which these key regulators of gene expression coordinate their activities and the precise mechanisms of this coordination are not well understood. RBPs often have recognizable RNA binding domains that correlate with specific protein function. Recently, several RBPs containing K homology (KH) RNA binding domains were shown to work with miRNAs to regulate gene expression, raising the possibility that KH domains may be important for coordinating with miRNA pathways in gene expression regulation. To ascertain whether additional KH domain proteins functionally interact with miRNAs during Caenorhabditis elegans development, we knocked down twenty-four genes encoding KH-domain proteins in several miRNA sensitized genetic backgrounds. Here, we report that a majority of the KH domain-containing genes genetically interact with multiple miRNAs and Argonaute alg-1. Interestingly, two KH domain genes, predicted splicing factors sfa-1 and asd-2, genetically interacted with all of the miRNA mutants tested, whereas other KH domain genes showed genetic interactions only with specific miRNAs. Our domain architecture and phylogenetic relationship analyses of the C. elegans KH domain-containing proteins revealed potential groups that may share both structure and function. Collectively, we show that many C. elegans KH domain RBPs functionally interact with miRNAs, suggesting direct or indirect coordination between these two classes of post-transcriptional gene expression regulators.

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 ◽

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.

Arginine methylation promotes siRNA-binding specificity for a spermatogenesis-specific isoform of the Argonaute protein CSR-1

Nature Communications ◽

10.1038/s41467-021-24526-6 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Dieu An H. Nguyen ◽

Carolyn M. Phillips

Keyword(s):

Small Rnas ◽

Rna Binding ◽

Regulatory Mechanism ◽

Expression Patterns ◽

Arginine Methylation ◽

Argonaute Protein ◽

C Elegans ◽

Argonaute Proteins ◽

Preferential Binding ◽

Temporal And Spatial

AbstractCSR-1 is an essential Argonaute protein that binds to a subclass of 22G-RNAs targeting most germline-expressed genes. Here we show that the two isoforms of CSR-1 have distinct expression patterns; CSR-1B is ubiquitously expressed throughout the germline and during all stages of development while CSR-1A expression is restricted to germ cells undergoing spermatogenesis. Furthermore, CSR-1A associates preferentially with 22G-RNAs mapping to spermatogenesis-specific genes whereas CSR-1B-bound small RNAs map predominantly to oogenesis-specific genes. Interestingly, the exon unique to CSR-1A contains multiple dimethylarginine modifications, which are necessary for the preferential binding of CSR-1A to spermatogenesis-specific 22G-RNAs. Thus, we have discovered a regulatory mechanism for C. elegans Argonaute proteins that allows for specificity of small RNA binding between similar Argonaute proteins with overlapping temporal and spatial localization.

Arginine methylation promotes siRNA-binding specificity for a spermatogenesis-specific isoform of the Argonaute protein CSR-1

10.1101/2020.06.08.134908 ◽

2020 ◽

Cited By ~ 1

Author(s):

Dieu An H. Nguyen ◽

Carolyn M. Phillips

Keyword(s):

Small Rnas ◽

Rna Binding ◽

Regulatory Mechanism ◽

Expression Patterns ◽

Arginine Methylation ◽

Argonaute Protein ◽

C Elegans ◽

Argonaute Proteins ◽

Preferential Binding ◽

Temporal And Spatial

SummaryCSR-1 is an essential Argonaute protein that binds to a subclass of 22G-RNAs targeting most germline-expressed genes. Here we show that the two isoforms of CSR-1 have distinct expression patterns; CSR-1B is ubiquitously expressed throughout the germline and during all stages of development while CSR-1A expression is restricted to germ cells undergoing spermatogenesis. Furthermore, CSR-1A associates preferentially with 22G-RNAs mapping to spermatogenesis-specific genes whereas CSR-1B-bound small RNAs map predominantly to oogenesis-specific genes. Interestingly, the exon unique to CSR-1A contains multiple dimethylarginine modifications, which are necessary for the preferential binding of CSR-1A to spermatogenesis-specific 22G-RNAs. Thus, we have discovered a regulatory mechanism for C. elegans Argonaute proteins that allows for specificity of small RNA binding between similar Argonaute proteins with overlapping temporal and spatial localization.

KH domain containing RNA-binding proteins coordinate with microRNAs to regulate Caenorhabditis elegans development

10.1101/2020.08.03.235127 ◽

2020 ◽

Author(s):

D Haskell ◽

A Zinovyeva

Keyword(s):

Gene Expression ◽

Caenorhabditis Elegans ◽

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

C Elegans ◽

Binding Domains ◽

Kh Domain ◽

Regulate Gene

ABSTRACTmicroRNAs (miRNAs) and RNA binding proteins (RBPs) regulate gene expression at the post-transcriptional level, but the extent to which these key regulators of gene expression coordinate and the precise mechanisms of their coordination are not well understood. RNA binding proteins often have recognizable RNA binding domains that correlate with specific protein function. Recently, several RBPs containing K Homology (KH) RNA binding domains were shown to work with miRNAs to regulate gene expression, raising the possibility that KH domains may be important for coordinating with miRNA pathways in gene expression regulation. To ascertain whether additional KH domain proteins functionally interact with miRNAs during Caenorhabditis elegans development, we knocked down twenty-four genes encoding KH-domain proteins in several miRNA sensitized genetic backgrounds. Here, we report that a majority of the KH domain-containing genes genetically interact with multiple miRNAs and Argonaute alg-1. Interestingly, two KH domain genes, predicted splicing factors sfa-1 and asd-2, genetically interacted with all of the miRNA mutants tested, while other KH domain genes exhibited functional interactions only with specific miRNAs. Our domain architecture and phylogenetic relationship analyses of the C. elegans KH domain-containing proteins revealed potential groups that may share both structure and function. Collectively, we show that many C. elegans KH domain RBPs functionally interact with miRNAs, suggesting direct or indirect coordination between these two classes of post-transcriptional gene expression regulators.

Cooperation between Caenorhabditis elegans COMPASS and condensin in germline chromatin organization

10.21203/rs.3.rs-17459/v1 ◽

2020 ◽

Author(s):

Marion Herbette ◽

Valérie Robert ◽

Aymeric Bailly ◽

Loïc Gely ◽

Robert Feil ◽

...

Keyword(s):

Gene Expression ◽

Topoisomerase Ii ◽

Resonance Energy Transfer ◽

Resonance Energy ◽

Higher Order ◽

Chromosome Organization ◽

Fluorescence Lifetime Imaging ◽

Chromatin Compaction ◽

C Elegans ◽

Meiotic Chromosomes

Abstract Background Histone-modifying activities play important roles in gene expression and influence higher-order genome organization. SET1/COMPASS (Complex Proteins Associated with Set1) deposits h istone H3 lysine 4 (H3K4) methylation at promoter regions and is associated with context-dependent effects on gene expression. Whether it also contributes to higher-order chromosome organization has not been explored. Results Using a quantitative FRET (Förster resonance energy transfer)-based fluorescence lifetime imaging microscopy (FLIM) approach to assay nanometer scale chromatin compaction in live animals, we reveal a novel role for SET1/COMPASS in structuring meiotic chromosomes in the C. elegans germline . Inactivation of SET-2, the C. elegans homologue of SET1, strongly enhanced chromosome organization defects and loss of fertility resulting from depletion of condensin-II, and aggravated defects in chromosome morphology resulting from inactivation of topoisomerase II, another major structural component of chromosomes. Loss of CFP-1, the chromatin targeting subunit of COMPASS, similarly affected germline chromatin compaction measured by FLIM-FRET and enhanced condensin-II knock-down phenotypes. Conclusions The data presented here are consistent with a role of SET1/ COMPASS in shaping meiotic chromosomes in the C. elegans germline. This new insight has important implications for how c hromatin-modifying complexes and histone modifications may cooperate with non histone-proteins to achieve proper chromosome organization, not only in meiosis, but also in mitosis.

HRPK-1, a conserved KH-domain protein, modulates microRNA activity during Caenorhabditis elegans development

10.1101/569905 ◽

2019 ◽

Author(s):

Li Li ◽

Isana Veksler-Lublinsky ◽

Anna Y. Zinovyeva

Keyword(s):

Gene Expression ◽

Messenger Rna ◽

Binding Proteins ◽

Rna Binding ◽

Rna Binding Proteins ◽

Genetic Interaction ◽

Transcriptional Level ◽

C Elegans ◽

Multiple Levels

AbstractmicroRNAs (miRNAs) are potent regulators of gene expression that function in diverse developmental and physiological processes. Argonaute proteins loaded with miRNAs form the miRNA Induced Silencing Complexes (miRISCs) that repress gene expression at the post-transcriptional level. miRISCs target genes through partial sequence complementarity between the miRNA and the target mRNA’s 3’ UTR. In addition to being targeted by miRNAs, these mRNAs are also extensively regulated by RNA-binding proteins (RBPs) through RNA processing, transport, stability, and translation regulation. While the degree to which RBPs and miRISCs functionally interact to regulate gene expression is likely extensive, we have only begun to unravel these functional interactions. An RNAi-based screen of putative ALG-1 Argonaute interactors has identified a role for a conserved RNA binding protein, HRPK-1, in modulating miRNA activity during C. elegans development. Here, we report the physical and genetic interaction between HRPK-1 and ALG-1/miRNAs. Specifically, we report the genetic and molecular characterizations of hrpk-1 and its role in C. elegans development and miRNA-mediated target repression. We show that loss of hrpk-1 causes numerous developmental defects and enhances the mutant phenotypes associated with reduction of miRNA activity, including those of lsy-6, mir-35-family, and let-7-family miRNAs. In addition to hrpk-1 genetic interaction with these miRNA families, hrpk-1 is required for efficient regulation of lsy-6 target cog-1. We report that hrpk-1 may play a role in miRNA processing but is not globally required for mature miRNA biogenesis or ALG-1/AIN-1 miRISC assembly and confirm HRPK-1 ability to co-precipitate with ALG-1. We suggest that HRPK-1 may functionally interact with miRNAs on multiple levels to enhance miRNA/miRISC gene regulatory activity and present several models for its activity.Author summarymicroRNAs are small non-coding RNAs that regulate gene expression at the post-transcriptional level. The core microRNA Induced Silencing Complex (miRISC), composed of Argonaute, mature microRNA, and GW182 protein effector, assembles on the target messenger RNA and inhibits translation or leads to messenger RNA degradation. RNA binding proteins interface with miRNA pathways on multiple levels to coordinate gene expression regulation. Here, we report identification and characterization of HRPK-1, a conserved RNA binding protein, as a physical and functional interactor of miRNAs. We confirm the physical interaction between HRPK-1, an hnRNPK homolog, and Argonaute ALG-1. We report characterizations of hrpk-1 role in development and its functional interactions with multiple miRNA families. We suggest that HRPK-1 promotes miRNA activity on multiple levels in part by contributing to miRNA processing and by coordinating with miRISC at the level of target RNAs. This work contributes to our understanding of how RNA binding proteins and auxiliary miRNA cofactors may interface with miRNA pathways to modulate miRNA gene regulatory activity.

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 ◽

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.