PRDE-1 is a nuclear factor essential for the biogenesis of Ruby motif-dependent piRNAs in C. elegans

Abstract

Piwi-interacting RNAs (piRNA) are small regulatory RNAs with essential roles in maintaining genome integrity in animals and protists. Most Caenorhabditis elegans piRNAs are transcribed from two genomic clusters that likely contain thousands of individual transcription units; however, their biogenesis is not understood. Here we identify and characterize prde-1 (piRNA silencing-defective) as the first essential C. elegans piRNA biogenesis gene. Analysis of prde-1 provides the first direct evidence that piRNA precursors are 28- to 29-nucleotide (nt) RNAs initiating 2 nt upstream of mature piRNAs. PRDE-1 is a nuclear germline-expressed protein that localizes to chromosome IV. PRDE-1 is required specifically for the production of piRNA precursors from genomic loci containing an 8-nt upstream motif, the Ruby motif. The expression of a second class of motif-independent piRNAs is unaffected in prde-1 mutants. We exploited this finding to determine the targets of the motif-independent class of piRNAs. Together, our data provide new insights into both the biogenesis and function of piRNAs in gene regulation.

piRNAs: from biogenesis to function

“…There has been some debate regarding the overall length of the precursors: Cecere et al have detected a >60 nt capped transcript using 5′ RACE, whereas Gu et al detected shorter 26 nt putative precursor species by small RNA sequencing (Cecere et al, 2012;Gu et al, 2012b). We and others have recently refined this finding showing that it is indeed short species of 28 or 29 nt that are made from most piRNA loci (Goh et al, 2014;Weick et al, 2014).…”

“…Our laboratory has identified piRNA defective 1 (PRDE-1) as a novel factor that is essential for the generation of Ruby-motif dependent piRNA precursors and accumulates in pachytene germ cell nuclei on chromosome IV. PRDE-1 is either involved in generating these precursors by direct or indirect interaction with the motif or may stabilise the precursors once they have been transcribed from the motif (Weick et al, 2014). Furthermore, using an RNAi based genome-wide screen, the Hannon lab has identified several other factors required for C. elegans piRNA biogenesis, which they collectively refer to as TOFUs for 'twenty-one U fouled up' (Goh et al, 2014).…”

“…1C) (Billi et al, 2013;Cecere et al, 2012). Of note, a subset of small RNAs bound to PRG-1 are generated from sites outside the canonical clusters and do not depend on the Ruby motif (Gu et al, 2012b;Weick et al, 2014). The 21U-RNA precursor transcripts carry a 5′ 7-methylguanylate cap and are likely generated by POL II, with transcription starting exactly 2 nt upstream of the 5′ U of the mature piRNA.…”

Diversity of the piRNA pathway for nonself silencing: worm-specific piRNA biogenesis factors

“…In fact, most of piRNA biogenesis factors identified in other animals, including the endonuclease Zucchini/MitoPLD, are not conserved in worms. In this issue of Genes & Development, three groups (de Albuquerue et al 2014;Goh et al 2014;Weick et al 2014) report on their discoveries of novel factors that are involved in the production of 21U-RNAs in C. elegans. Goh et al (2014) performed an RNAi screen covering >80% of annotated C. elegans protein-coding genes and identified 22 genes that affect mature 21U-RNA levels (named TOFUs).…”

“…piRNA silencing defective 1 (prde-1) Weick et al (2014) also conducted EMS mutagenesis screening using a 21U-RNA sensor and identified PRDE-1. PRDE-1 has a protein kinase-like domain, but amino acid residues critical for ATP binding in the kinase domain are not conserved.…”