Antisense ribosomal siRNAs inhibit RNA polymerase I-directed transcription in C. elegans

Abstract

Eukaryotic cells express a wide variety of endogenous small regulatory RNAs that function in the nucleus. We previously found that erroneous rRNAs induce the generation of antisense ribosomal siRNAs (risiRNAs) which silence the expression of rRNAs via the nuclear RNAi defective (Nrde) pathway. To further understand the biological roles and mechanisms of this class of small regulatory RNAs, we conducted forward genetic screening to identify factors involved in risiRNA generation in Caenorhabditis elegans. We found that risiRNAs accumulated in the RNA exosome mutants. risiRNAs directed the association of NRDE proteins with pre-rRNAs and the silencing of pre-rRNAs. In the presence of risiRNAs, NRDE-2 accumulated in the nucleolus and colocalized with RNA polymerase I. risiRNAs inhibited the transcription elongation of RNA polymerase I by decreasing RNAP I occupancy downstream of the RNAi-targeted site. Meanwhile, exosomes mislocalized from the nucleolus to nucleoplasm in suppressor of siRNA (susi) mutants, in which erroneous rRNAs accumulated. These results established a novel model of rRNA surveillance by combining ribonuclease-mediated RNA degradation with small RNA-directed nucleolar RNAi system.

The conserved helicase ZNFX-1 memorializes silenced RNAs in perinuclear condensates

“…This response requires the nuclear Argonaute HRDE-1 and may reflect stalling of RNA polymerase II and/or pre-mRNA processing, causing nascent transcripts to accumulate at the locus. Stalling of RNA polymerase has previously been implicated in nuclear RNAi 36,37 and several lines of evidence have linked RNAi and splicing, including apparent co-evolution of the RNAi and splicing machineries 38 , splicing factors identified as HRDE-1 interactors 39,40 , sRNA defects associated with mutations or knock down of spliceosome components 41,42 and insensitivity to nuclear RNAi of an endogenous transcript whose introns were removed by genome editing 43 .…”

Hypodermal ribosome synthesis inhibition induces a nutrition-uncoupled organism-wide growth quiescence in C. elegans

“…Strains expressing degron-GFP-integrated RPOA-2, RRB-1 or TSR-2 are homozygous viable and phenotypically wild-type with nucleolar RPOA-2 [37], nucle(ol)ar RRB-1 and nuclear TSR-2 localization patterns ( Figure-1B, Figure-S2A ), demonstrating that the degron-GFP tags are compatible with organism growth. To test the AID system, we then crossed strains expressing degron-GFP-integrated ribosome biogenesis factors (RPOA-2, RRB-1, TSR-2) with strains ubiquitously expressing TIR1 from the eft-3 promoter.…”

A ZTF-7/RPS-2 complex mediates the cold-warm response inC. elegans

“…Meanwhile, after knocking down these genes by RNAi, a class of antisense ribosomal siRNAs (risiRNAs) were enriched, suggesting that the proper nucleolar localization of exosomes may be important for the suppression of risiRNA production. In addition, we also found that environmental stimuli, such as low temperature, induced the accumulation of risiRNAs that subsequently guide the nuclear Argonaute proteins NRDE-3 and NRDE-2 to the nucleoli, which are associated with pre-rRNAs and silence RNA polymerase I transcription (ZHOU et al 2017;LIAO et al 2021). Whether and how environmental stimuli could induce the translocation of exosome proteins in distinct subcellular compartments is intriguing.…”

“…In C. elegans, EXOS-10 and DIS-3 are ubiquitously expressed in all of the cells and enriched in the nucleus. Interestingly, EXOS-10 is enriched in nucleoli, while DIS-3 is enriched in the nucleoplasm but depleted in nucleoli (LIAO et al 2021). The RNA exosome functions both in the nucleus and cytoplasm to process noncoding RNAs (ncRNAs) and degrade improperly processed (faulty) RNAs in RNA surveillance pathways (HOUSELEY et al 2006;BUTLER AND MITCHELL 2010;SCHAEFFER et al 2010;SCHNEIDER AND TOLLERVEY 2013;KILCHERT et al 2016).…”