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

Shimiao Liao,Ting Xu,Zongxiu Xu,Shouhong Guang,Xuezhu Feng,Demin Xu,Qile Jin,Xufei Zhou,Xiangyang Chen,Chengming Zhu

doi:10.1093/nar/gkab662

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.

Highlights

In eukaryotic cells, ribosomal RNAs are transcribed by RNA polymerase I into a single 47S polycistronic precursor in the nucleolus, which are processed and matured into 18S, 5.8S and 28S rRNAs; 5S rRNA is independently transcribed by RNA polymerase III in the nucleus
We previously described a forward genetic screening to search for suppressor of siRNA genes based on altered subcellular localization of NRDE-3 in C. elegans (Figure 1A) [5,14]
This screening identified a cytoplasmic localized exoribonuclease, SUSI-1(ceDis3L2), and a number of rRNA modifying and processing enzymes, including SUSI2(ceRRP8) (Figure 1B). susi-1 is the homologue of human DIS3-like exonuclease 2 (Dis3L2), the name disl2 is used hereafter. susi-2 is the homologue of yeast RRP8, the name rrp-8 is used hereafter

Summary

Introduction

Ribosomal RNAs (rRNAs) are transcribed by RNA polymerase I into a single 47S polycistronic precursor in the nucleolus, which are processed and matured into 18S, 5.8S and 28S rRNAs; 5S rRNA is independently transcribed by RNA polymerase III in the nucleus. The processing of ribosomal RNAs is extraordinarily complicated, in which defects of any steps could induce the accumulation of erroneous rRNAs [1,2]. Erroneous rRNAs are degraded from 3 to 5 by the RNA exosome complex. Erroneous rRNAs can be polyuridylated and degraded from 3 to 5 by the cytoplasmic exoribonuclease DISL-2 ( known as SUSI-1 in Caenorhabditis elegans) [5,6]

Methods

Results

Conclusion