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

Gerald Ryan R Aquino,Nicolai Krogh,Henning Urlaub,Kuan-Ting Pan,Henrik Nielsen,Anthony K Henras,Philipp Hackert,Katherine E Bohnsack,Mariam Jaafar,Markus T Bohnsack

doi:10.1038/s41467-021-26208-9

Abstract

Early pre-60S ribosomal particles are poorly characterized, highly dynamic complexes that undergo extensive rRNA folding and compaction concomitant with assembly of ribosomal proteins and exchange of assembly factors. Pre-60S particles contain numerous RNA helicases, which are likely regulators of accurate and efficient formation of appropriate rRNA structures. Here we reveal binding of the RNA helicase Dbp7 to domain V/VI of early pre-60S particles in yeast and show that in the absence of this protein, dissociation of the Npa1 scaffolding complex, release of the snR190 folding chaperone, recruitment of the A3 cluster factors and binding of the ribosomal protein uL3 are impaired. uL3 is critical for formation of the peptidyltransferase center (PTC) and is responsible for stabilizing interactions between the 5′ and 3′ ends of the 25S, an essential pre-requisite for subsequent pre-60S maturation events. Highlighting the importance of pre-ribosome remodeling by Dbp7, our data suggest that in the absence of Dbp7 or its catalytic activity, early pre-ribosomal particles are targeted for degradation.

Highlights

Pre-60S ribosomal particles are poorly characterized, highly dynamic complexes that undergo extensive ribosomal RNAs (rRNAs) folding and compaction concomitant with assembly of ribosomal proteins and exchange of assembly factors
Recent evidence from bacteria highlighted a role of transient interactions of r-proteins in regulating rRNA folding and in eukaryotes a subset of small nucleolar ribonucleoprotein complexes are proposed to act as rRNA chaperones[9,10,11,12,13,14,15,16,17,18]
Dbp[7] is a putative RNA helicase localized in the nucleolus and implicated in LSU biogenesis (Supplementary Fig. 1a)[52], but its catalytic activity and function(s) during subunit assembly have remained poorly characterized

Summary

Introduction

Pre-60S ribosomal particles are poorly characterized, highly dynamic complexes that undergo extensive rRNA folding and compaction concomitant with assembly of ribosomal proteins and exchange of assembly factors. It is believed that the earliest assembly steps involve the association of early r-proteins and AFs to the solvent-exposed structural domains I and II, leading to the folding and stabilization of these domains followed by those of domain VI3,35,36 This results in the formation and clustering of the respective root helices, which is an essential pre-requisite for subsequent maturation steps including downstream processing of the LSU pre-rRNAs and recruitment of later r-proteins. Our data support pre-rRNA remodeling by Dbp[7] to release snR190, a subset of other snoRNAs and the Npa[1] complex from early pre-60S particles, thereby allowing incorporation of uL3 to stabilize domain I and VI interactions and license downstream 60S maturation events. Our findings provide important insights into how early remodeling events are coordinated by AFs to initiate compaction of the nascent 60S

Methods

Results

Conclusion