High-throughput sequencing reveals extraordinary fluidity of miRNA, piRNA, and siRNA pathways in nematodes

Abstract

The nematode Caenorhabditis elegans contains each of the broad classes of eukaryotic small RNAs, including microRNAs (miRNAs), endogenous small-interfering RNAs (endo-siRNAs), and piwi-interacting RNAs (piRNAs). To better understand the evolution of these regulatory RNAs, we deep-sequenced small RNAs from C. elegans and three closely related nematodes: C. briggsae, C. remanei, and C. brenneri. The results reveal a fluid landscape of small RNA pathways with essentially no conservation of individual sequences aside from a subset of miRNAs. We identified 54 miRNA families that are conserved in each of the four species, as well as numerous miRNAs that are species-specific or shared between only two or three species. Despite a lack of conservation of individual piRNAs and siRNAs, many of the features of each pathway are conserved between the different species. We show that the genomic distribution of 26G siRNAs and the tendency for piRNAs to cluster is conserved between C. briggsae and C. elegans. We also show that, in each species, 26G siRNAs trigger stage-specific secondary siRNA formation. piRNAs in each species also trigger secondary siRNA formation from targets containing up to three mismatches. Finally, we show that the production of male-and female-specific piRNAs is conserved in all four species, suggesting distinct roles for piRNAs in male and female germlines.

Specific down-regulation of spermatogenesis genes targeted by 22G RNAs in hybrid sterile males associated with an X-Chromosome introgression

“…Given the highly similar phenotypes in sperm as well as in the mRNA and TE expression patterns between the two sterile lines, we focused on small RNAs in one of the two sterile lines, ZZY10330. In C. briggsae, as in C. elegans, piRNAs are produced from individual loci located in clusters, with each individual piRNA associated with a specific upstream promoter identified by a highly conserved motif (the Ruby motif) (Ruby et al 2006;de Wit et al 2009;Shi et al 2013). The C. briggsae piRNA clusters have previously been shown to reside on Chromosomes I and IV (de Wit et al 2009;Shi et al 2013).…”

“…In C. briggsae, as in C. elegans, piRNAs are produced from individual loci located in clusters, with each individual piRNA associated with a specific upstream promoter identified by a highly conserved motif (the Ruby motif) (Ruby et al 2006;de Wit et al 2009;Shi et al 2013). The C. briggsae piRNA clusters have previously been shown to reside on Chromosomes I and IV (de Wit et al 2009;Shi et al 2013). We therefore aligned small RNAs from both C. nigoni parent males (hitherto wild type) and sterile hybrid males to the C. nigoni scaffolds "cn1" as described above.…”

Multiple small RNA pathways regulate the silencing of repeated and foreign genes in C. elegans

“…Data analysis was done as described (Shi et al 2013). Briefly, small RNA sequences were parsed and mapped to the C. elegans genome (WormBase release WS230) and transgene sequences.…”

A framework for understanding the roles of miRNAs in animal development

“…Sex determination pathways have evolved independently in different animals, suggesting that miRNAs could have become essential also at later points in evolution as they were co-opted for species-specific essential processes. Interestingly, other small RNAs have been implicated in sex determination, such as a Piwi-interacting RNA ( piRNA) in the silkworm (Kiuchi et al, 2014) and the miRNA let-7 in Drosophila (Fagegaltier et al, 2014), pointing to another potential link between the rapid evolution of certain biological processes and the cooption of evolutionarily fluid mechanisms such as small RNAmediated repression (Shi et al, 2013).…”