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Small RNA sorting: matchmaking for Argonautes

Key Points

  • Small RNAs are defined by their size (~20–30 nucleotides in length) and their association with members of the Argonaute family. They impact nearly every biological process in eukaryotic cells, directly or indirectly.

  • MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are born from dsRNA precursors, whereas Piwi-interacting RNAs (piRNAs) originate from single-stranded transcripts.

  • To perform their myriad roles, different classes of small RNAs must not only be generated in a precise manner, but must also be sorted into specific Argonaute complexes. An Argonaute protein primed with a single-stranded small RNA is called an RNA-induced silencing complex (RISC).

  • Eukaryotic organisms often encode several Argonaute proteins that function in distinct pathways. They typically show various preferences for the small RNAs they accept, comprising loading determinants that include the identity of terminal nucleotides, small RNA duplex structure and thermodynamic properties.

  • Small RNA duplexes are usually not incorporated into Argonaute proteins without assistance from additional protein factors, known as the RISC-loading machinery.

  • miRNAs and siRNAs arise from small RNA duplexes and are loaded into Argonaute as dsRNA molecules. Thus, during RISC maturation, one strand must be selected specifically, whereas the other strand must be lost or degraded.

  • Mature RISC regulates targets through sequence complementarity. The ultimate impact of accurate strand selection and sorting is that an active RISC is formed, imbued with the ability to regulate target transcripts.

  • Biogenesis and sorting of small RNAs in animals and plants share some key mechanistic features, but have also evolved myriad variations and adaptations.

Abstract

Small RNAs directly or indirectly impact nearly every biological process in eukaryotic cells. To perform their myriad roles, not only must precise small RNA species be generated, but they must also be loaded into specific effector complexes called RNA-induced silencing complexes (RISCs). Argonaute proteins form the core of RISCs and different members of this large family have specific expression patterns, protein binding partners and biochemical capabilities. In this Review, we explore the mechanisms that pair specific small RNA strands with their partner proteins, with an eye towards the substantial progress that has been recently made in understanding the sorting of the major small RNA classes — microRNAs (miRNAs) and small interfering RNAs (siRNAs) — in plants and animals.

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Figure 1: MicroRNA biogenesis in Drosophila melanogaster.
Figure 2: Production of small interfering RNAs.
Figure 3: Small RNA sorting and RNA-induced silencing complex assembly in flies.

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Acknowledgements

The authors thank O. Tam, F. Muerdter, J.-W. Wang and R. Zhou for comments on the manuscript. The authors are greatly indebted to J. Duffy for assistance with figures. B.C. is supported by a Ph.D fellowship from the Boehringer Ingelheim Fonds. This work was supported by grants from the National Institutes of Health and a kind gift from K. W. Davis. G.J.H. is an investigator of the Howard Hughes Medical Institute.

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Glossary

RNase III protein

A member of a family of ribonucleases that process dsRNA, leaving 5′ monophosphates and 2-nt 3′ overhangs with hydroxyl ends. Drosha and Dicer are examples of such ribonucleases.

RNA-induced silencing complex

A regulatory multi-protein complex containing an Argonaute protein bound to a single-stranded small RNA that regulates gene expression through sequence complementarity between the guide RNA and the target transcript.

Guide strand

During RISC loading, one strand of an siRNA is selected and stabilized. This is termed the guide strand, and it confers target specificity. miRNA guide strands are termed miR strands.

Passenger strand

The non-incorporated strand of the siRNA duplex that is degraded during the assembly of RISC. Non-incorporated strands of miRNAs are called miR* strands.

Stem–loop structure

A region of dsRNA (stem) connected by an unpaired region (loop) in a single RNA molecule. This is a structure typical for miRNA precursors.

Mirtron

A miRNA that originates from a very short intron and is excised to form a pre-miRNA by the splicing machinery (and occasionally subsequent trimming), therefore bypassing the Drosha processing step.

Dicing

Refers to the cleavage events carried out by the RNase III family nuclease Dicer.

RNA-dependent RNA polymerase

An RNA polymerase that uses ssRNA as a template to synthesize dsRNA.

Trans-acting siRNA

A plant small RNA that primarily associates with AGO2. ta-siRNA biogenesis depends on miRNA-mediated cleavage of precursors that are further processed by DCL4 and other siRNA machinery factors.

Natural antisense transcript-derived siRNA

A stress-induced small RNA produced by DCL1 and DCL2 that originates from dsRNA formed by convergent transcription.

Heterochromatic siRNA

A highly abundant plant small RNA that arises from transposons and repeats. hc-siRNAs depend on DCL3 and mainly load into AGO4.

RNase H

A conserved family of endonucleases that cleave the RNA strand of RNA:DNA hybrid duplexes. AGO proteins contain RNase H-like domains.

Seed region

A region consisting of nucleotides 2–8 counted from the 5′ end of miRNAs that participates in the interaction between a small RNA and target transcript.

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Czech, B., Hannon, G. Small RNA sorting: matchmaking for Argonautes. Nat Rev Genet 12, 19–31 (2011). https://doi.org/10.1038/nrg2916

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