Early co-transcriptional events during eukaryotic ribosome assembly result in the formation of precursors of the small (40S) and large (60S) ribosomal subunits1. A multitude of transient assembly factors regulate and chaperone the systematic folding of pre-ribosomal RNA subdomains. However, owing to a lack of structural information, the role of these factors during early nucleolar 60S assembly is not fully understood. Here we report cryo-electron microscopy (cryo-EM) reconstructions of the nucleolar pre-60S ribosomal subunit in different conformational states at resolutions of up to 3.4 Å. These reconstructions reveal how steric hindrance and molecular mimicry are used to prevent both premature folding states and binding of later factors. This is accomplished by the concerted activity of 21 ribosome assembly factors that stabilize and remodel pre-ribosomal RNA and ribosomal proteins. Among these factors, three Brix-domain proteins and their binding partners form a ring-like structure at ribosomal RNA (rRNA) domain boundaries to support the architecture of the maturing particle. The existence of mutually exclusive conformations of these pre-60S particles suggests that the formation of the polypeptide exit tunnel is achieved through different folding pathways during subsequent stages of ribosome assembly. These structures rationalize previous genetic and biochemical data and highlight the mechanisms that drive eukaryotic ribosome assembly in a unidirectional manner.
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We thank M. Ebrahim and J. Sotiris for their support with data collection at the Evelyn Gruss Lipper Cryo-EM resource center, M. Tesic-Mark for analysis of mass spectrometry data, C. Cheng for help with the initial manual curation and analysis of the nucleolar pre-60S particles and members of the Walz laboratory for helpful discussions. L.M. is supported in part by NIH T32 GM115327-Tan. J.B. is supported by an EMBO long-term fellowship (ALTF 51-2014) and a Swiss National Science Foundation fellowship (155515). M.C.-M. is supported by a postgraduate scholarship from NSERC. S.K. is supported by the Robertson Foundation, the Irma T. Hirschl Trust, the Alexandrine and Alexander L. Sinsheimer Fund, the Rita Allen Foundation and an NIH New Innovator Award (1DP2GM123459). B.T.C. is supported by National Institute of Health Grant Nos. P41GM103314 and P41GM109824.
Extended data figures
Extended data tables
A 360° rotation of a composite cryo-EM map consisting of the 3.4 Å state 1 and the 3.7 Å state 2 maps. Densities for nucleolar pre-60S assembly factors and rRNA domains are colour-coded as in Figure 1 and ribosomal proteins are colored in grey. The rotation is paused at front and back view as shown in Figure 1.
About this article
Nature Reviews Drug Discovery (2018)