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Licensing RNA-Guided Genome Defense TIBS-1023; No. of Pages 10 Review Recognizing the enemy within: licensing RNA-guided genome defense Phillip A. Dumesic and Hiten D. Madhani Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158, USA How do cells distinguish normal genes from transpo- RNAi-related RNA silencing pathways are deeply con- sons? Although much has been learned about RNAi- served genome defense mechanisms that act in organisms related RNA silencing pathways responsible for genome from protist to human to recognize and suppress transpo- defense, this fundamental question remains. The litera- sons [8]. In all of these pathways, 20–30 nucleotide small ture points to several classes of mechanisms. In some RNAs act in complex with Argonaute family proteins to cases, double-stranded RNA (dsRNA) structures pro- silence complementary transcripts. Depending on the duced by transposon inverted repeats or antisense inte- pathway and context, silencing proceeds by a variety of gration trigger endogenous small interfering RNA mechanisms, which include RNA degradation, translation- (siRNA) biogenesis. In other instances, DNA features al repression, and the establishment of repressive histone associated with transposons – such as their unusual modifications [9,10]. The pathways also differ in the means copy number, chromosomal arrangement, and/or chro- by which they generate small RNAs. In the canonical RNAi matin environment – license RNA silencing. Finally, re- pathway, RNase-III-type Dicer enzymes convert long cent studies have identified improper transcript dsRNA into small interfering RNA (siRNA). In contrast, processing events, such as stalled pre-mRNA splicing, PIWI-interacting RNA (piRNA) pathways do not require as signals for siRNA production. Thus, the suboptimal Dicer, and instead generate small RNA from single-strand- gene expression properties of selfish elements can en- ed precursors. Still other pathways, such as the endoge- able their identification by RNA silencing pathways. nous siRNA pathways of worm and plant, require RNA- dependent RNA polymerases (RdRPs) for small RNA bio- RNA silencing pathways suppress transposons to genesis; these enzymes are thought to produce small RNAs protect genome integrity directly or to generate long dsRNA substrates for Dicer. Transposons have parasitized nearly all eukaryotic gen- The diversity of small RNA biogenesis mechanisms per- omes, including the human genome, over half of which is mits a wide variety of RNA sequences to template the derived from transposon sequences. In so doing, transpo- production of small RNAs for genome defense. sons have shaped eukaryotic evolution, in part by contrib- The adaptability of RNA silencing pathways, whose uting regulatory and coding information to nearby host targeting depends primarily on the sequences of their genes [1]. In fact, 4% of human protein-coding open read- small RNA guides, makes them well suited to defend ing frames contain transposon-derived sequences [2], as do against transposons, which differ extensively in sequence 25% of promoter regions [3]. Remarkably, host organisms and distribution among eukaryotic species. Indeed, ortho- have even co-opted the protein activities encoded by trans- logous Argonaute proteins silence different transposon poson genes, as in the case of Recombination activating families in different host organisms, suggesting that gene 1 (RAG1), a transposon-derived endonuclease that RNA silencing pathways can adapt to recognize novel now serves to catalyze V(D)J recombination in humans transposons that a host has not previously encountered [4]. Despite these positive contributions to host biology, [11,12]. This adaptability raises a central question in the transposons are primarily selfish elements that threaten study of these pathways: how does RNA silencing specifi- the integrity of host genomes. Transposon mobilization can cally distinguish transposons from host genes? Here, we disrupt host genes and promote deleterious chromosomal review our current understanding of principles by which rearrangements [5,6], and these perturbations contribute to transposons can be recognized as non-self by RNA silenc- Mendelian disease and cancer in humans [5,7]. To combat ing pathways of fungi, plants, and metazoans. this threat, host organisms have evolved multiple genome defense mechanisms to suppress transposon mobility. The challenges of transposon recognition RNA silencing pathways must overcome several properties Corresponding author: Madhani, H.D. ([email protected]). of transposons in order to target them specifically. First, like Keywords: transposon; genome defense; small RNA; RNAi; small interfering RNA; host genes, transposons reside in the nuclear genome and PIWI-interacting RNA. utilize host gene expression machinery. Thus, transposons 0968-0004/$ – see front matter are not broadly distinguished by the use of alternative gene ß 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.tibs.2013.10.003 expression mechanisms. Second, transposons themselves differ widely in sequence, with many families exhibiting no homology to each other, which hinders sequence-specific transposon recognition mechanisms (Box 1). Finally, even Trends in Biochemical Sciences xx (2013) 1–10 1 TIBS-1023; No. of Pages 10 Review Trends in Biochemical Sciences xxx xxxx, Vol. xxx, No. x Box 1. Common types of eukaryotic transposable elements Studies of RNA silencing pathways have begun to reveal strategies by which these pathways recognize transposons Transposons are nucleic acid elements that can mobilize to new while avoiding inappropriate silencing of host sequences. chromosomal locations. They co-opt the host gene expression machinery to produce their own protein products, which facilitate These strategies are not mutually exclusive, and an indi- mobilization. Although common in these respects, transposons are vidual RNA silencing pathway sometimes utilizes multiple remarkably diverse in sequence and transposition mechanism, which strategies. One general strategy recognizes transposons by hinders their identification by cellular genome defense pathways. virtue of their tendency to generate dsRNA. A second Transposons can be broadly divided into retrotransposons (Class strategy distinguishes transposons by their unique ability I) and DNA transposons (Class II) Figure I [85,86]. Retrotransposons mobilize through an RNA intermediate, which is reverse transcribed to mobilize, which makes them more likely to exist in to allow its integration into the genome. This ‘copy and paste’ unusual chromosomal arrangements or in high copy num- mechanism does not alter the original transposon locus and ber. A third class of mechanism exploits the suboptimal therefore acts to increase transposon copy number. Some retro- gene expression properties of transposons, which might transposons encode long terminal repeats (LTRs), which act as promoters and polyadenylation signals. Like their retrovirus rela- arise due to their distinct evolutionary histories, to distin- tives, these transposons undergo reverse transcription in virus-like guish them from host genes. A fourth class of mechanism particles in the cytoplasm. Other retrotransposons, such as LINEs, licenses small RNA production against transposons based 0 also encode their own promoters and 3 end formation signals, but on the prior capture of transposon sequences by specialized lack LTRs. These elements undergo reverse transcription in the chromatin niches. These four transposon recognition strat- nucleus using nicked genomic DNA as a primer. By contrast, short interspersed nuclear elements (SINEs) mobilize in a manner similar egies are discussed in turn in this review. to that of LINEs, but do not themselves encode the proteins required for mobilization. They are therefore nonautonomous, and depend Transposons are identified by their production of on LINE-encoded factors. dsRNA DNA transposons do not utilize an RNA intermediate, but instead Early studies of Caenorhabditis elegans RNAi have indi- generally mobilize through a ‘cut and paste’ mechanism in which the original transposon locus is excised and reinserted in a new cated that some mutants defective in gene silencing location. These transposons generally encode terminal inverted triggered by exogenous dsRNA are also defective in sup- repeat (TIR) sequences, which recruit transposase to the transposon pressing transposons, raising the possibility that endoge- DNA locus in order to initiate its excision. nous dsRNA initiates transposon silencing [14,15]. In such a model, transposon-derived dsRNA is processed by Dicer Class I: retrotransposons enzymes to yield siRNA, which then acts to repress homol- LTRGAG PRO RT RH INT LTR LTR ogous transposon sequences throughout the genome. An (e.g., Gypsy) attractive feature of this model is that transposons exhibit several properties that might increase their likelihood of ORF1 ORF2 p(A) LINE generating dsRNA, thereby enabling them to be distin- (e.g., L1) guished from host genes [16]. For instance, some transpo- son families encode repeats and antisense promoters that p(A) SINE (non-autonomous) can produce dsRNA. Furthermore, the mobilization of transposons into existing host transcriptional units may Class II: DNA transposons lead to the production of antisense transposon transcripts. Finally, the repetitiveness of transposon sequences in the TIR Transposase TIR TIR genome, together with their tendency to undergo rearran- (e.g., Mutator) gements, may promote the formation of
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