Deep sequencing reveals unique small RNA repertoire that is regulated during head regeneration in Hydra magnipapillata

Abstract

Small non-coding RNAs such as miRNAs, piRNAs and endo-siRNAs fine-tune gene expression through post-transcriptional regulation, modulating important processes in development, differentiation, homeostasis and regeneration. Using deep sequencing, we have profiled small non-coding RNAs in Hydra magnipapillata and investigated changes in small RNA expression pattern during head regeneration. Our results reveal a unique repertoire of small RNAs in hydra. We have identified 126 miRNA loci; 123 of these miRNAs are unique to hydra. Less than 50% are conserved across two different strains of Hydra vulgaris tested in this study, indicating a highly diverse nature of hydra miRNAs in contrast to bilaterian miRNAs. We also identified siRNAs derived from precursors with perfect stem–loop structure and that arise from inverted repeats. piRNAs were the most abundant small RNAs in hydra, mapping to transposable elements, the annotated transcriptome and unique non-coding regions on the genome. piRNAs that map to transposable elements and the annotated transcriptome display a ping–pong signature. Further, we have identified several miRNAs and piRNAs whose expression is regulated during hydra head regeneration. Our study defines different classes of small RNAs in this cnidarian model system, which may play a role in orchestrating gene expression essential for hydra regeneration.

Cnidarian microRNAs frequently regulate targets by cleavage

“…The high abundance of piRNAs in our libraries is in agreement with previous studies of Cnidaria (Grimson et al 2008;Krishna et al 2013). In addition to piRNAs, we found two types of small RNAs, which originate from predicted genomic inverted repeats.…”

“…The unusual features of pre-miRs 2024b, c, and d and their low abundance relative to other miR-2024 variants suggest that they may be degenerating, providing unprecedented examples of pre-miRNAs transitioning into sources of siRNAs. We found the conserved and experimentally confirmed miR-2022 (Chapman et al 2010;Krishna et al 2013; this study) to be a false negative in our initial annotation, because it did not pass one of the many stringent criteria (it folded less stably than another RNA hairpin located in close proximity).…”

“…Thus, the previous observation indicating that the vast majority of miRNAs evolved independently in Cnidaria and Bilateria was not biased by sequencing depth or the samples used (Grimson et al 2008). Comparing our annotated Nematostella miRNAs to the genome of the reef-building coral Acropora digitifera and to sequenced small RNAs from Hydra magnipapillata (Shinzato et al 2011;Krishna et al 2013) enabled us to reconstruct the gain and loss history of miRNAs in Cnidaria (Fig. 1D).…”

PIWI proteins and PIWI-interacting RNAs function in Hydra somatic stem cells

“…These features are identical to the ping-pong signature of biogenesis that was first described in Drosophila (14,26) and also observed in mice and zebrafish (16,23,27). Previous sequencing of total Nematostella vectensis and Hydra RNAs identified putative piRNAs (28,29). Here we have identified bona fide cnidarian piRNAs bound to specific PIWI proteins, thus allowing for comparisons between piRNAs bound to different PIWI proteins.…”

Characterization of the piRNA pathway during development of the sea anemone Nematostella vectensis

“…Recently, piRNAs and PIWI proteins were identified in the cnidarian Hydra. [14][15][16] Hywi and Hyli (the two paralogous Hydra PIWI proteins) are predominantly expressed in the germline as well as in the interstitial cell lineage (the so-called i-cells) including multipotent progenitor stem cells and germline stem cells, but also in epithelial stem cells. Hydra piRNAs comprise a large fraction among their entire repertoire of small RNAs and map to transposons.…”