Research Article

Architecture of the yeast small subunit processome

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Science  13 Jan 2017:
Vol. 355, Issue 6321, eaal1880
DOI: 10.1126/science.aal1880

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A machine for building ribosomes

The ribosome is a very large protein and RNA complex responsible for the difficult process of synthesizing proteins. Construction of the ribosome itself involves several molecular machines and an army of helper proteins and RNAs. Chaker-Margot et al. determined the structure of one of those machines, the yeast small subunit processome. The structure reveals how the processome helps in the maturation of individual domains of the ribosome and suggests that the mechanism involves a molecular motor to drive conformational changes.

Science, this issue p. 10.1126/science.aal1880

Structured Abstract

INTRODUCTION

The eukaryotic ribosome is assembled through an intricate process that involves in excess of 250 nonribosomal proteins and small nucleolar RNAs (snoRNAs). Ribosomal RNA (rRNA) that later gives rise to the small ribosomal subunit is initially transcribed as a long precursor, the 35S pre-rRNA, which also contains a 5′ external transcribed spacer (5′ ETS). Ribosome biogenesis factors assemble cotranscriptionally on nascent pre-rRNA and coordinate the folding and cleavage of the 35S pre-rRNA, forming large terminal knobs, which can be observed in Miller spreads. The identity of these large terminal structures, referred to as small and large subunit processomes, remained elusive for decades. Recent studies revealed that the small subunit (SSU) processome is a large ribonucleoprotein particle composed of approximately 70 nonribosomal proteins, pre-rRNA, and the U3 snoRNA. It organizes the assembly of the eukaryotic small ribosomal subunit at the early stages by coordinating the folding and modification of nascent pre-rRNA. In addition, the SSU processome facilitates the cleavage of the precursor RNA at distinct sites in the 5′ ETS (A0 and A1) and the internal transcribed spacer 1 (ITS1; A2) to give rise to the mature SSU.

RATIONALE

For decades, a lack of structural information has precluded a mechanistic understanding of early ribosome biogenesis. Mean- while, important genetic and biochemical insights into eukaryotic ribosome assembly have been obtained by using the model organism Saccharomyces cerevisiae. To bridge this gap of knowledge, we sought to determine the structure of the SSU processome from S. cerevisiae in order to elucidate the architecture of the functional core of this particle.

RESULTS

Here, we present the cryo–electron microscopy (cryo-EM) structure of the yeast SSU processome at 5.1-Å resolution. We describe the organization of the 5′ ETS and its role, together with the U3 snoRNA, in providing a structural blueprint for the entire particle. This very 5′ end of the pre-rRNA folds into several helices that coordinate the recruitment of large ribosome biogenesis complexes, such as UtpA and UtpB, to the SSU processome. UtpA, UtpB, and the U3 snoRNA, which pairs at two sites with the base 5′ ETS, bridge the A0-cleaved 5′ ETS with the 18S precursor RNA. In conjunction with many other essential ribosome biogenesis factors, this structure forms an intertwined RNA-protein assembly platform for the 18S rRNA. This platform guarantees the spatial segregation of 18S rRNA domains, facilitating the recruitment of enzymes and other assembly factors required for their maturation. Several large helical repeat proteins mediate long-range interactions between distant domains of the SSU processome.

We discovered structural similarities between the subcomplexes UtpA and UtpB, such as a conserved helical tetramerization domain, that suggest that these complexes share a common evolutionary origin. The strategic placement of the 135-kDa guanosine triphosphatase Bms1 at the center of the particle near long architectural helices suggests a mechanism for mediating conformational changes within this giant particle. These motions could facilitate subsequent cleavage reactions and rearrangements required in the maturation of the small ribosomal subunit.

CONCLUSION

The architecture of the yeast SSU processome allows us to rationalize a wealth of biochemical and genetic data available for the model organism S. cerevisiae. Most notably, it sheds light on the central role of the 5′ ETS and the U3 snoRNA in initiating and organizing SSU processome assembly. In addition, the structural information allows us to contextualize biochemical data on SSU processome assembly as a function of transcription. Proteins that bind within the 5′ ETS are located at the base of the SSU processome structure, whereas factors associated with 18S rRNA domains form the core of the particle. Last, proteins recruited only after 18S rRNA completion form the outer tier of the structure.

This study provides an improved structural framework for a mechanistic understanding of eukaryotic ribosome assembly in S. cerevisiae. Targeted functional studies will now be possible to further elucidate the individual roles of many ribosome assembly factors in the model organism for ribosome biogenesis.

Architecture of the S. cerevisiae SSU processome.

Front view of the segmented cryo-EM density map of the complete particle, with color-coded protein and RNA elements (left), and an outline of a 10-Å low-pass filtered map, with only RNA elements shown in full color (right).

Abstract

The small subunit (SSU) processome, a large ribonucleoprotein particle, organizes the assembly of the eukaryotic small ribosomal subunit by coordinating the folding, cleavage, and modification of nascent pre–ribosomal RNA (rRNA). Here, we present the cryo–electron microscopy structure of the yeast SSU processome at 5.1-angstrom resolution. The structure reveals how large ribosome biogenesis complexes assist the 5′ external transcribed spacer and U3 small nucleolar RNA in providing an intertwined RNA-protein assembly platform for the separate maturation of 18S rRNA domains. The strategic placement of a molecular motor at the center of the particle further suggests a mechanism for mediating conformational changes within this giant particle. This study provides a structural framework for a mechanistic understanding of eukaryotic ribosome assembly in the model organism Saccharomyces cerevisiae.

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