Elimination of 01/A′–A0 pre-rRNA processing by-product in human cells involves cooperative action of two nuclear exosome-associated nucleases: RRP6 and DIS3

Abstract

Pre-rRNA processing generates mature 18S, 5.8S, and 28S/25S rRNAs through multistage removal of surrounding 5 ′ ′ ′ ′ ′ -ETS/ 3 ′ ′ ′ ′ ′ -ETS and intervening ITS1/ITS2 segments. Endonucleolytic activities release by-products, which need to be eliminated. Here, we investigated the interplay of exosome-associated 3 ′ ′ ′ ′ ′ -5 ′ ′ ′ ′ ′ exonucleases DIS3 and RRP6 in rRNA processing and by-product elimination in human cells. In agreement with previous reports, we observed accumulation of 5.8S and 18S precursors upon dysfunction of these enzymes. However, none of these phenotypes was so pronounced as previously overlooked accumulation of short RNA species derived from 5 ′ ′ ′ ′ ′ -ETS (01/A ′ ′ ′ ′ ′ -A0), in cells with nonfunctional DIS3. We demonstrate that removal of 01/A ′ ′ ′ ′ ′ -A0 is independent of the XRN2 5 ′ ′ ′ ′ ′ -3 ′ ′ ′ ′ ′ exonucleolytic activity. Instead, it proceeds rapidly after A0 cleavage and occurs exclusively in the 3 ′ ′ ′ ′ ′ -5 ′ ′ ′ ′ ′ direction in several phases-following initiation by an unknown nuclease, the decay is executed by RRP6 with some contribution of DIS3, whereas the ultimate phase involves predominantly DIS3. Our data shed new light onto the role of human exosome in 5 ′ ′ ′ ′ ′ -ETS removal. Furthermore, although 01/A ′ ′ ′ ′ ′ -A0 degradation involves the action of two nucleases associated with the exosome ring, similarly to 5.8S 3 ′ ′ ′ ′ ′ -end maturation, it is likely that contrary to the latter process, RRP6 acts prior to or redundantly with DIS3.

RNA exosome mutations in pontocerebellar hypoplasia alter ribosome biogenesis and p53 levels

“…Abbreviations: CH, ceratohyal; M, Meckel's cartilage. EXOSC3, EXOSC8, EXOSC9, EXOSC10, or DIS3 depletion causes the appearance of mis-processed 5.8S pre-rRNA species and accumulation of larger rRNA precursors (Tomecki et al, 2010;Lubas et al, 2011;Tafforeau et al, 2013;Sloan et al, 2013bSloan et al, , 2014Kobylecki et al, 2018;Davidson et al, 2019). Moreover, a recent study showed that neurological disease-associated mutations of EXOSC3 indeed caused pre-rRNA processing defects when introduced into Rrp40, the EXOSC3 homologue in budding yeast (Gillespie et al, 2017).…”

Loss of Tsc1 in cerebellar Purkinje cells induces transcriptional and translation changes in FMRP target transcripts

“…DIS3 was also found to be upregulated due to ER stress in Caenorhabditis elegans ( Sakaki et al, 2012 ), and we have found that TSC2-deficient neurons show evidence of ER stress ( Di Nardo et al, 2009 ). In addition, the RNA exosome complex is involved in ribosomal RNA maturation ( Kobyłecki et al, 2018 ; Pirouz et al, 2019 ), and mTORC1 is known to increase ribosomal biogenesis ( Gentilella et al, 2015 ). Therefore, it is possible that mTORC1 may induce the RNA exosome by stimulating the production of ribosomes, and the RNA decay phenomenon that we observe could be a side effect of this process.…”

Loss ofTsc1in cerebellar Purkinje cells induces transcriptional and translation changes in FMRP target transcripts

“…DIS3 was also found to be upregulated due to ER stress in C. elegans (Sakaki et al, 2012), and we have found that TSC2-deficient neurons show evidence of ER stress (Di Nardo et al, 2009). In addition, the RNA exosome complex is involved in ribosomal RNA maturation (Kobylecki et al, 2018; Pirouz et al, 2019), and mTORC1 is known to increase ribosomal biogenesis (Gentilella et al, 2015). Therefore, it is possible that mTORC1 may induce the RNA exosome by stimulating the production of ribosomes, and the RNA decay phenomenon that we observe could be a side effect of this process.…”