Abstract
Eukaryotic ribosome biogenesis is initiated with the transcription of pre-ribosomal RNA at the 5' external transcribed spacer, which directs the early association of assembly factors but is absent from the mature ribosome. The subsequent co-transcriptional association of ribosome assembly factors with pre-ribosomal RNA results in the formation of the small subunit processome. Here we show that stable rRNA domains of the small ribosomal subunit can independently recruit their own biogenesis factors in vivo. The final assembly and compaction of the small subunit processome requires the presence of the 5' external transcribed spacer RNA and all ribosomal RNA domains. Additionally, our cryo-electron microscopy structure of the earliest nucleolar pre-ribosomal assembly - the 5' external transcribed spacer ribonucleoprotein - provides a mechanism for how conformational changes in multi-protein complexes can be employed to regulate the accessibility of binding sites and therefore define the chronology of maturation events during early stages of ribosome assembly.
Highlights
In yeast, the mature ribosome is a 3.2 MDa ribonucleoprotein complex composed of 79 proteins and four ribosomal RNAs assembled into the small and the large subunit
We show that the 5’ ETS and stable ribosomal RNAs (rRNA) domains are functionally independent in their ability to recruit their own assembly factors, and the strict order of events is enforced through molecular switches within the 5’ ETS ribonucleoprotein (RNP) that require the presence of all rRNA domains for the final SSU processome to be formed
Each MS2 aptamer-tagged pre-rRNA domain was expressed in a strain containing a tagged ribosome assembly factor that was known to associate with it (Utp10, Esf1, Kri1 and Mrd1 for 5’ ETS, 5’ domain, central domain and 3’ major domain, respectively)
Summary
The mature ribosome is a 3.2 MDa ribonucleoprotein complex composed of 79 proteins and four ribosomal RNAs (rRNA) assembled into the small and the large subunit. Ribosome assembly starts with the RNA polymerase-I driven transcription of the 35S precursor ribosomal RNA (pre-rRNA) in the nucleolus. This primary transcript contains rRNAs of the small ribosomal subunit (SSU, 18S) and the large ribosomal subunit (LSU, 5.8S and 25S) flanked by external transcribed spacers (5’ ETS, 3’ ETS) and separated by internal transcribed spacers (ITS1, ITS2). Factors are co-transcriptionally recruited and associate with the nascent pre-rRNA, resulting in a chronology of biogenesis that starts at the 5’ end, where the 5’ ETS and rRNA for the small subunit reside. The first structurally defined particle that assembles co-transcriptionally is the small subunit processome, a giant precursor of the small subunit (Dragon et al, 2002)
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