Association of snR190 snoRNA chaperone with early pre-60S particles is regulated by the RNA helicase Dbp7 in yeast

Abstract

Synthesis of eukaryotic ribosomes involves the assembly and maturation of precursor particles (pre-ribosomal particles) containing ribosomal RNA (rRNA) precursors, ribosomal proteins (RPs) and a plethora of assembly factors (AFs). Formation of the earliest precursors of the 60S ribosomal subunit (pre-60S r-particle) is among the least understood stages of ribosome biogenesis. It involves the Npa1 complex, a protein module suggested to play a key role in the early structuring of the pre-rRNA. Npa1 displays genetic interactions with the DExD-box protein Dbp7 and interacts physically with the snR190 box C/D snoRNA. We show here that snR190 functions as a snoRNA chaperone, which likely cooperates with the Npa1 complex to initiate compaction of the pre-rRNA in early pre-60S r-particles. We further show that Dbp7 regulates the dynamic base-pairing between snR190 and the pre-rRNA within the earliest pre-60S r-particles, thereby participating in structuring the peptidyl transferase center (PTC) of the large ribosomal subunit.

The RNA helicase Dbp7 promotes domain V/VI compaction and stabilization of inter-domain interactions during early 60S assembly

“…5a). Strikingly, snR190, which is 12fold enriched in the Dbp7-HTP PAR-CRAC data, is suggested to chaperone rRNA folding rather than guiding rRNA 2′-Omethylation 61 and has a pre-rRNA basepairing site in close proximity to the identified Dbp7-HTP pre-rRNA cross-linking sites. Notably, several, but not all of the other enriched snoRNAs, also have basepairing sites in the vicinity of the Dbp7-HTP pre-rRNA cross-linking sites.…”

“…4a,b). snR190 was initially predicted to 2′-O-methylate 25S-G2395 close to the Dbp7 pre-rRNA cross-linking sites, but no methylation of this site has been detected 55,61,62 . Instead, predicted snR190 basepairing sites in domains I and V of the 25S rRNA have led to the suggestion that the snoRNA may function as a pre-rRNA folding chaperone (Fig.…”

“…Instead, predicted snR190 basepairing sites in domains I and V of the 25S rRNA have led to the suggestion that the snoRNA may function as a pre-rRNA folding chaperone (Fig. 4c, d) 38,61 . The distribution of sequencing reads mapping to snR190 (and snR34 and snR42) were examined to determine whether Dbp7 physically associates with the regions of snR190 involved in basepairing with the 25S rRNA.…”

“…Yeast strains lacking snR34 or snR42, or carrying a mutation in box C of snR190 that destabilizes the snoRNA, were therefore generated in the wild-type or Δdbp7 background (Supplementary Fig. 6a-d) 61 . Growth analysis of these strains demonstrated that lack of snR34, snR42, or snR190 did not strongly affect growth (Fig.…”

“…Alongside protein scaffolds, several non-methylating snoRNPs, such as U3, U14, snR10, and snR190 tether linearly distant pre-rRNA sequences in favorable folding conformations and/or prevent formation of aberrant basepairing interactions 13,19,61 . Lack of Dbp7 or its catalytic activity lead to accumulation of snR190 in pre-60S particles, suggesting that pre-ribosome remodeling by Dbp7 contributes to the release of snR190.…”