Comparative analysis of piRNA sequences, targets and functions in nematodes

Abstract

Piwi proteins and Piwi-interacting RNAs (piRNAs) are best known for their roles in suppressing transposons and promoting fertility. Yet piRNA biogenesis and its mechanisms of action differ widely between distantly related species. To better understand the evolution of piRNAs, we characterized the piRNA pathway in C. briggsae , a sibling species of the model organism C. elegans . Our analyses define 25,883 piRNA producing-loci in C. briggsae . piRNA sequences in C. briggsae are extremely divergent from their counterparts in C. elegans , yet both species adopt similar genomic organization that drive piRNA expression. By examining production of Piwi-mediated secondary small RNAs, we identified a set of protein-coding genes that are evolutionarily conserved piRNA targets. In contrast to C. elegans , small RNAs targeting ribosomal RNAs or histone transcripts are not hyper-accumulated in C. briggsae Piwi mutants. Instead, we found that transcripts with few introns are prone to small RNA overamplification. Together our work highlights evolutionary conservation and divergence of the nematode piRNA pathway and provides insights into its role in endogenous gene regulation.

C. elegans germ granules sculpt both germline and somatic RNAome

“…β–elimination was performed as described 54 . Small RNA-sequencing libraries were constructed following an previously established protocol 106 . Briefly, Small RNA samples were treated with the polyphosphatase PIR-1 to remove γ and β phosphates from 5′-triphosphorylated RNAs 107 .…”

Spatially clustered piRNA genes promote the transcription of piRNAs via condensate formation of the H3K27me3 reader UAD-2

“…1c). The published C. briggsae piRNA high-throughput sequencing data 54 showed that, similar to that of C. elegans , piRNA genes located within clusters exhibited explicitly higher expression compared to those of out-cluster regions. The average expression level of clustered piRNAs bound to the PIWI protein was ∼50 RPM, while out-cluster piRNAs showed an average of ∼3.1 RPM (Extended Data Fig.…”

“…To further test whether the clustering of piRNA genes is important for piRNA expression, we analyzed the distribution and expression of piRNAs in another nematode, C. briggsae . The C. briggsae genome contains over 25,000 piRNAs across 6 chromosomes 54 , clustering into one locus on LG Ⅰ and two loci on LG Ⅳ, which are termed Clusters 1, 2, and 3, respectively (Extended Data Fig. 1c).…”

Pre-piRNA trimming safeguards piRNAs against erroneous targeting by RNA-dependent RNA Polymerase

“…Mature piRNAs are commonly referred to as 21U-RNAs due to their strong propensity for a 5' uridine (U) and a primary length of 21-nucleotides (nts) (15)(16)(17)(18)(19). Type I and II piRNAs are found in C. elegans and its sibling species C. briggsae, with Type I piRNAs being notably much more abundant (16,20). Type I 21U-RNAs in C. elegans are encoded by over 15,000 individual mini-genes organized within two large clusters at Chromosome IV (15)(16)(17)(18).…”