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  • Review Article
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RNA interference in the nucleus: roles for small RNAs in transcription, epigenetics and beyond

Key Points

  • Small RNAs with roles in the nucleus include those that are generated mainly by Dicer proteins and loaded into Argonaute proteins (small interfering RNAs (siRNAs)) and also those that are Dicer-independent (largely generated by the ping-pong cycle) and are loaded into PIWI proteins (PIWI-interacting RNAs (piRNAs)).

  • The subcellular localization where siRNA biogenesis occurs is variable among organisms. However, cytoplasmic Argonaute loading might be conserved.

  • Nuclear RNA interference (RNAi) directs heterochromatic modifications at target loci including methylation of histone H3 at lysine 9 (H3K9; in Schizosaccharomyces pombe) and DNA methylation (in Arabidopsis thaliana). These reduce transcription, facilitating transcriptional gene silencing (TGS).

  • Examples in A. thaliana and S. pombe show that transcription by RNA polymerase is required to produce nascent RNA that is targeted by nuclear RNAi; this process is termed co-transcriptional gene silencing (CTGS).

  • There is evidence in the somatic cells of metazoans that endogenous siRNA pathways are involved in co-transcriptional regulation and heterochromatin formation, suggesting a conserved nuclear role for RNAi.

  • RNAi has a crucial germline role in silencing transposons both post-transcriptionally and transcriptionally.

  • In both metazoans and plants, transposons are revealed for transcription and produce small RNAs that target transposons in the germ cells to maintain silencing through nuclear RNAi.

  • In mammals, piRNAs can direct de novo cytosine methylation in the germ line and have been shown to do so at an imprinted locus. This suggests that nuclear RNAi might have another conserved role in parent-of-origin imprinting.

  • The piRNAs of Caenorhabditis elegans (21U small RNAs) can direct transcriptional silencing by H3K9 methylation in the germ line that is heritable and dependent on nuclear RNAi (NRDE) pathway members.

  • Roles are emerging for small RNAs in DNA repair and genome maintenance, through both the maintenance and regulation of heterochromatic domains (such as centromeres and telomeres) and through direct involvement in DNA repair: for example, at double-strand breaks.

Abstract

A growing number of functions are emerging for RNA interference (RNAi) in the nucleus, in addition to well-characterized roles in post-transcriptional gene silencing in the cytoplasm. Epigenetic modifications directed by small RNAs have been shown to cause transcriptional repression in plants, fungi and animals. Additionally, increasing evidence indicates that RNAi regulates transcription through interaction with transcriptional machinery. Nuclear small RNAs include small interfering RNAs (siRNAs) and PIWI-interacting RNAs (piRNAs) and are implicated in nuclear processes such as transposon regulation, heterochromatin formation, developmental gene regulation and genome stability.

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Figure 1: Generalized pathways for the biogenesis of nuclear small RNAs.
Figure 2: Co-transcriptional gene silencing in Schizosaccharomyces pombe.
Figure 3: The RNA-dependent DNA methylation pathway in Arabidopsis thaliana.
Figure 4: RNA-interference-mediated transposon silencing in the germ line.
Figure 5: The 21U small RNA pathway in the Caenorhabditis elegans germ line.

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Acknowledgements

We thank the members of the Martienssen laboratory for discussion. S.E.C. is a Cashin Scholar of the Watson School of Biological Sciences and is supported by a Natural Sciences and Engineering Research Council of Canada Post Graduate Scholarship.

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Correspondence to Robert A. Martienssen.

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Glossary

RNA interference

(RNAi). Silencing at both the post-transcriptional and transcriptional levels that is directed by small RNA molecules.

Post-transcriptional gene silencing

(PTGS). Silencing achieved by the degradation and/or prevention of translation of a transcript targeted by small RNAs.

Transcriptional gene silencing

(TGS). Silencing achieved by the formation of a repressive chromatin environment at a locus targeted by small RNA, making it inaccessible to transcriptional machinery.

Argonaute

The effector proteins of RNA inteference that are composed of three characteristic domains, a PAZ domain and a MID domain, which bind the 3′ and 5′ ends of small interfering RNA respectively, and a PIWI domain, which may possess RNase-H-like slicer activity if the protein is catalytically active.

Co-transcriptional gene silencing

(CTGS). The coupling of repressive epigenetic modification with transcription by an RNA polymerase that produces a nascent RNA molecule targeted by small RNAs.

Pericentromeric regions

Sites of constitutive heterochromatin that flank the central kinetochore-binding region of the centromere and are necessary for proper centromere function.

Histone H3 methylated at lysine 9

(H3K9me). H3K9 can be mono-, di- or tri-methylated. Methylation is catalysed by a histone methyltransferase and is highly enriched in repressive heterochromatin. This mark acts as a binding site for heterochromatin protein 1 (HP1; known as Swi6 in Schizosaccharomyces pombe), the presence of which is the defining feature of heterochromatic loci.

RNA-induced transcriptional silencing complex

(RITSC). The effector of nuclear RNA interference in Schizosaccharomyces pombe. It is composed of an Argonaute protein and other cofactors that may aid in localization to chromatin.

Passenger strand

The antisense small RNA strand in the double-stranded RNA molecule initially loaded by an Argonaute. The passenger strand is released by the catalytic 'slicing' activity of the Argonaute protein (like homologous RNA targets), whereas the guide strand is retained and acts to determine the specificity of the silencing complex.

Position effect variegation

Refers to the variegated expression pattern of a gene that is stochastically inactivated by the spreading of a nearby heterochromatic domain. For example, a pericentromere and an inserted nearby reporter gene.

Cytosine methylation

Covalent modification of a cytosine base catalysed by a DNA methyltransferase that often associates with heterochromatic loci. It can occur in various sequence contexts, including CG, CHG and CHH, which influence establishment and inheritance.

NRDE pathway

In Caenorhabditis elegans, components of the nuclear RNA interference pathway are termed NRDE for 'nuclear RNAi defective' owing to the phenotype of mutants (nrde).

Transposable element

Genetic elements that can move their positions within the genome. The mechanism of transposition varies and defines transposon families.

Companion cells

Cells in the germ line of plants that will not contribute genetically to progeny but are produced by meiosis. These are the vegetative nucleus in the male germ line and the central cell in the female germ line. The central cell is fertilized to produce the endosperm that acts as a supportive tissue to the developing embryo.

Cohesin

Large protein rings that predominantly localize to heterochromatic regions of the genome. They function to keep sister chromatids connected during mitosis, facilitate spindle attachment to chromosomes and are involved in DNA repair through recombination.

Recombination

The joining of similar or identical DNA sequences to produce a novel molecule. Homologous recombination is used as a mechanism to repair damaged DNA in cells; however, at repetitive regions, it can be detrimental by leading to copy number changes of repetitive elements.

Double-strand breaks

(DSBs). A deleterious form of DNA damage that occurs when the covalent bonds of both strands of a double helix are broken at a locus. It can be repaired by homologous recombination or by error-prone non-homologous end joining.

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Castel, S., Martienssen, R. RNA interference in the nucleus: roles for small RNAs in transcription, epigenetics and beyond. Nat Rev Genet 14, 100–112 (2013). https://doi.org/10.1038/nrg3355

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