Grishok A (2012) Worm "Endogenous RNAi and adaptation to environment in C. elegans."

Grishok A

[Worm, 2012]

The contributions of short RNAs to the control of repetitive elements are well documented in animals and plants. Here, the role of endogenous RNAi and AF10 homolog ZFP-1 in the adaptation of C. elegans to the environment is discussed. First, modulation of insulin signaling through regulation of transcription of the PDK-1 kinase (Mansisidor et al., PLoS Genetics, 2011) is reviewed. Second, an siRNA-based natural selection model is proposed in which variation in endogenous siRNA pools between individuals is subject to natural selection similarly to DNA-based genetic variation. The value of C. elegans for the research of siRNA-based epigenetic variation and adaptation is highlighted.

Grishok A (2021) DNA (Basel) "Small RNAs Worm Up Transgenerational Epigenetics Research."

Grishok A

[DNA (Basel), 2021]

DNA is central to the propagation and evolution of most living organisms due to the essential process of its self-replication. Yet it also encodes factors that permit epigenetic (not included in DNA sequence) flow of information from parents to their offspring and beyond. The known mechanisms of epigenetic inheritance include chemical modifications of DNA and chromatin, as well as regulatory RNAs. All these factors can modulate gene expression programs in the ensuing generations. The nematode <i>Caenorhabditis elegans</i> is recognized as a pioneer organism in transgenerational epigenetic inheritance research. Recent advances in <i>C. elegans</i> epigenetics include the discoveries of control mechanisms that limit the duration of RNA-based epigenetic inheritance, periodic DNA motifs that counteract epigenetic silencing establishment, new mechanistic insights into epigenetic inheritance carried by sperm, and the tantalizing examples of inheritance of sensory experiences. This review aims to highlight new findings in epigenetics research in <i>C. elegans</i> with the main focus on transgenerational epigenetic phenomena dependent on small RNAs.

Grishok A et al. (2005) Genes Dev "Transcriptional silencing of a transgene by RNAi in the soma of C. elegans."

Sinskey JL, Grishok A, Sharp PA

[Genes Dev, 2005]

The silencing of transgene expression at the level of transcription in the soma of Caenorhabditis elegans through an RNAi-dependent pathway has not been previously characterized. Most gene silencing due to RNAi in C. elegans occurs at the post-transcriptional level. We observed transcriptional silencing when worms containing the elt-2::gfp/LacZ transgene were fed RNA produced from the commonly used L4440 vector. The transgene and the vector share plasmid backbone sequences. This transgene silencing depends on multiple RNAi pathway genes, including dcr-1, rde-1, rde-4, and rrf-1. Unlike post-transcriptional gene silencing in worms, elt-2::gfp/LacZ silencing is dependent on the PAZ-PIWI protein Alg-1 and on the HP1 homolog Hpl-2. The latter is a chromatin silencing factor, and expression of the transgene is inhibited at the level of intron-containing precursor mRNA. This inhibition is accompanied by a decrease in the acetylation of histones associated with the transgene. This transcriptional silencing in the soma can be distinguished from transgene silencing in the germline by its inability to be transmitted across generations and its dependence on the rde-1 gene. We therefore define this type of silencing as RNAi-induced Transcriptional Gene Silencing (RNAi-TGS). Additional chromatin-modifying components affecting RNAi-TGS were identified in a candidate RNAi screen.

Grishok A et al. (2005) Proc Natl Acad Sci U S A "Negative regulation of nuclear divisions in Caenorhabditis elegans by retinoblastoma and RNA interference-related genes."

Sharp PA, Grishok A

[Proc Natl Acad Sci U S A, 2005]

Short RNA regulatory molecules, microRNAs, and short interfering RNAs participate in a range of developmental gene networks by base-pairing with their target sequences. Consistent with these findings, genes required for the biogenesis and function of short interfering RNAs and microRNAs, dicer (dcr-1 in Caenorhabditis elegans) and argonaute homologs, are essential for development in diverse organisms, including C. elegans. We demonstrate that genes required for the function of short RNAs synergize with the retinoblastoma tumor suppressor homolog lin-35 in negative regulation of the nuclear divisions in the intestine of C. elegans. The level of cyclin E (cye-1) expression is critical for nuclear divisions in the intestine and is elevated in double mutants in lin-35 and RNA interference pathway genes. We propose that RNA interference-related pathways cooperate with retinoblastoma in transcriptional repression of endogenous genes, an example being cyclin E.

Grishok A et al. (2008) Proc Natl Acad Sci U S A "RNA interference and retinoblastoma-related genes are required for repression of endogenous siRNA targets in Caenorhabditis elegans."

Sharp PA, Hoersch S, Grishok A

[Proc Natl Acad Sci U S A, 2008]

In Caenorhabditis elegans, a vast number of endogenous short RNAs corresponding to thousands of genes have been discovered recently. This finding suggests that these short interfering RNAs (siRNAs) may contribute to regulation of many developmental and other signaling pathways in addition to silencing viruses and transposons. Here, we present a microarray analysis of gene expression in RNA interference (RNAi)-related mutants rde-4, zfp-1, and alg-1 and the retinoblastoma (Rb) mutant lin-35. We found that a component of Dicer complex RDE-4 and a chromatin-related zinc finger protein ZFP-1, not implicated in endogenous RNAi, regulate overlapping sets of genes. Notably, genes a) up-regulated in the rde-4 and zfp-1 mutants and b) up-regulated in the lin-35(Rb) mutant, but not the down-regulated genes are highly represented in the set of genes with corresponding endogenous siRNAs (endo-siRNAs). Our study suggests that endogenous siRNAs cooperate with chromatin factors, either C. elegans ortholog of acute lymphoblastic leukemia-1 (ALL-1)-fused gene from chromosome 10 (AF10), ZFP-1, or tumor suppressor Rb, to regulate overlapping sets of genes and predicts a large role for RNAi-based chromatin silencing in control of gene expression in C. elegans.

Grishok A et al. (2001) Cell "Genes and mechanisms related to RNA interference regulate expression of the small temporal RNAs that control C. elegans developmental timing."

Pasquinelli AE, Grishok A, Ha I, Li N, Parrish S, Baillie DL, Fire A, Ruvkun G, Mello CC, Conte D

[Cell, 2001]

RNAi is a gene-silencing phenomenon triggered by double-stranded (ds) RNA and involves the generation of 21 to 26 nt RNA segments that guide mRNA destruction. In Caenorhabditis elegans, lin-4 and let-7 encode small temporal RNAs (stRNAs) of 22 nt that regulate stage-specific development. Here we show that inactivation of genes related to RNAi pathway genes, a homolog of Drosophila Dicer (dcr-1), and two homologs of rde-1 (alg-1 and alg-2), cause heterochronic phenotypes similar to lin-4 and let-7 mutations. Further we show that dcr-1, alg-1, and alg-2 are necessary for the maturation and activity of the lin-4 and let-7 stRNAs. Our findings suggest that a common processing machinery generates guide RNAs that mediate both RNAi and endogenous gene regulation.

Sharp PA et al. (2000) Science "Molecular biology. RNA interference."

Zamore PD, Sharp PA

[Science, 2000]

The mechanical sounds of robots and the electronic hum of computers are features of 21st-century life. Despite the joys of increasing automation, simple experiments still produce wonderful new discoveries in molecular biology. The realization several years ago that double-stranded RNA (dsRNA) is a key player in one form of posttranscriptional gene silencing indicated that a new field of great importance was about to be created (1). The study of how RNA regulates gene expression called RNA interference (RNAi)- has an exciting future, and the report by Grishok et al. on page 2494 of this issue (2) provides further support for such optimism.

Cecere G et al. (2014) Bio Protoc "RNA Chromatin Immunoprecipitation (RNA-ChIP) in Caenorhabditis elegans."

Grishok A, Cecere G

[Bio Protoc, 2014]

The RNA chromatin immunoprecipitation assay (RNA-ChIP) allows detection and quantification of RNA-protein interactions using in vivo cross-linking with formaldehyde followed by immunoprecipitation of the RNA-protein complexes. Here we describe the RNA-ChIP protocol that we have adapted for Caenorhabditis elegans (C. elegans) to detect interaction between the nuclear Argonaute CSR-1 (chromosome segregation and RNAi deficient) protein and its target nascent RNAs. We have used a transgenic strain expressing a recombinant long isoform of CSR-1 protein fused with N-terminal 3x FLAG epitope.

Esse R et al. (2020) Cells "Caenorhabditis elegans Deficient in DOT-1.1 Exhibit Increases in H3K9me2 at Enhancer and Certain RNAi-Regulated Regions."

Grishok A, Esse R

[Cells, 2020]

The methylation of histone H3 at lysine 79 is a feature of open chromatin. It is deposited by the conserved histone methyltransferase DOT1. Recently, DOT1 localization and H3K79 methylation (H3K79me) have been correlated with enhancers in <i>C. elegans</i> and mammalian cells. Since earlier research implicated H3K79me in preventing heterochromatin formation both in yeast and leukemic cells, we sought to inquire whether a H3K79me deficiency would lead to higher levels of heterochromatic histone modifications, specifically H3K9me2, at developmental enhancers in <i>C. elegans.</i> Therefore, we used H3K9me2 ChIP-seq to compare its abundance in control and <i>dot-1.1</i> loss-of-function mutant worms, as well as in <i>rde-4; dot-1.1</i> and <i>rde-1; dot-1.1</i> double mutants. The <i>rde-1</i> and <i>rde-4</i> genes are components of the RNAi pathway in <i>C. elegans</i>, and RNAi is known to initiate H3K9 methylation in many organisms, including <i>C. elegans</i>. We have previously shown that <i>dot-1.1(-)</i> lethality is rescued by <i>rde-1</i> and <i>rde-4</i> loss-of-function. Here we found that H3K9me2 was elevated in enhancer, but not promoter, regions bound by the DOT-1.1/ZFP-1 complex in <i>dot-1.1(-)</i> worms. We also found increased H3K9me2 at genes targeted by the ALG-3/4-dependent small RNAs and repeat regions. Our results suggest that ectopic H3K9me2 in <i>dot-1.1(-)</i> could, in some cases, be induced by small RNAs.

Tabara H et al. (1998) Science "RNAi in C. elegans: soaking in the genome sequence."

Grishok A, Tabara H, Mello CC

[Science, 1998]

The completion of the Caenorhabditis elegans genome sequence represents a major milestone in a journey initiated by Sydney Brenner some 30 years ago. The goal then as now was to discover how genetic information specifies the development, anatomy, and behavior of a simple animal. Bringing the full potential of the genome sequence to bear on this goal will require facile new reverse genetic tools for converting sequence information into functional information. Here, we briefly describe progress toward understanding and using one such tool termed RNA interference" or "RNAi".