Ancient and Novel Small RNA Pathways Compensate for the Loss of piRNAs in Multiple Independent Nematode Lineages

Abstract

Small RNA pathways act at the front line of defence against transposable elements across the Eukaryota. In animals, Piwi interacting small RNAs (piRNAs) are a crucial arm of this defence. However, the evolutionary relationships among piRNAs and other small RNA pathways targeting transposable elements are poorly resolved. To address this question we sequenced small RNAs from multiple, diverse nematode species, producing the first phylum-wide analysis of how small RNA pathways evolve. Surprisingly, despite their prominence in Caenorhabditis elegans and closely related nematodes, piRNAs are absent in all other nematode lineages. We found that there are at least two evolutionarily distinct mechanisms that compensate for the absence of piRNAs, both involving RNA-dependent RNA polymerases (RdRPs). Whilst one pathway is unique to nematodes, the second involves Dicer-dependent RNA-directed DNA methylation, hitherto unknown in animals, and bears striking similarity to transposon-control mechanisms in fungi and plants. Our results highlight the rapid, context-dependent evolution of small RNA pathways and suggest piRNAs in animals may have replaced an ancient eukaryotic RNA-dependent RNA polymerase pathway to control transposable elements.

Pan-arthropod analysis reveals somatic piRNAs as an ancestral defence against transposable elements

“…The RdRPs expressed in the chelicerates and S. maritima may generate dsRNA precursors which can then be processed by Dicer to generate siRNAs, similar to RdRPs in basal nematodes 22 , while species lacking an RdRP would require bidirectional transcription by RNA polymerase II to generate dsRNA. To test this idea, we sequenced long RNA (RNA-Seq) and small RNA from all species (Supplementary Table 1).…”

“…The rapid evolution of small RNA pathways makes inferences drawn from detailed studies of individual model organisms misleading 22 . Our results suggest that the best studied arthropods, concentrated in a small region of the phylogenetic tree, are not representative of the entire phylum (Fig.…”

“…There is also considerable divergence in downstream pathways linked to piRNA silencing—for example, in C. elegans where piRNAs act upstream of an RNA-dependent RNA polymerase (RdRP) pathway that generates secondary siRNAs antisense to piRNA targets. Moreover, many nematode species have lost the piRNA pathway altogether, with RNAi-related mechanisms assuming the role of TE suppression 22 . These examples highlight the need for further characterisation across animals to better understand the diversity of the piRNA pathway.…”

DNA Methylation Influences the Expression of DICER-LIKE4 Isoforms, Which Encode Proteins of Alternative Localization and Function

“…Dicer of Schizosaccharomyces pombe, which directs heterochromatin formation, was shown to shuttle between the nucleus and cytoplasm through motifs that mediate nuclear export and retention (Emmerth et al, 2010). Caenorhabditis elegans Dicer can relocalize to the nucleus following ERK-mediated phosphorylation (Drake et al, 2014), while the nematode Trichinella spiralis evolved a Dicer with an N-terminal NLS, which can likely mediate RdDM to repress transposons (Sarkies et al, 2015). Finally, metazoan Dicer is cytoplasmic, but human Dicer encodes a noncanonical nuclear localization signal, normally masked in the folded protein, and was shown to shuttle into the nucleus where it represses spurious interferon response activation (White et al, 2014;Doyle et al, 2013;Burger and Gullerova, 2015).…”

No evidence for extensive horizontal gene transfer in the genome of the tardigrade Hypsibius dujardini

“…H. dujardini is a parthenogen, with first division restitution of ploidy (22) and therefore is intractable for traditional genetic analysis, although reverse genetic approaches are being developed (17). H. dujardini has become a genomic model system, revealing the pattern of ecdysozoan phylogeny (3,4) and the evolution of small RNA pathways (23). H. dujardini is poorly cryptobiotic (24), but serves as a useful comparator for good cryptobiotic species (9).…”