Structural Heterogeneity in Pre-40S Ribosomes

Matthew C Johnson; Homa Ghalei; Katelyn A Doxtader; Katrin Karbstein; M Elizabeth Stroupe

doi:10.1016/j.str.2016.12.011

Structural Heterogeneity in Pre-40S Ribosomes

Structure. 2017 Feb 7;25(2):329-340. doi: 10.1016/j.str.2016.12.011. Epub 2017 Jan 19.

Authors

Matthew C Johnson¹, Homa Ghalei², Katelyn A Doxtader², Katrin Karbstein², M Elizabeth Stroupe³

Affiliations

¹ Department of Biological Science, Institute of Molecular Biophysics, Florida State University, 91 Chieftain Way, Tallahassee, FL 32306, USA.
² Department of Cancer Biology, The Scripps Research Institute, Jupiter, FL 33458, USA.
³ Department of Biological Science, Institute of Molecular Biophysics, Florida State University, 91 Chieftain Way, Tallahassee, FL 32306, USA. Electronic address: mestroupe@bio.fsu.edu.

Abstract

Late-stage 40S ribosome assembly is a highly regulated dynamic process that occurs in the cytoplasm, alongside the full translation machinery. Seven assembly factors (AFs) regulate and facilitate maturation, but the mechanisms through which they work remain undetermined. Here, we present a series of structures of the immature small subunit (pre-40S) determined by three-dimensional (3D) cryoelectron microscopy with 3D sorting to assess the molecule's heterogeneity. These structures demonstrate an extensive structural heterogeneity of interface AFs that likely regulates subunit joining during 40S maturation. We also present structural models for the beak and the platform, two regions where the low resolution of previous studies did not allow for localization of AFs and the rRNA, respectively. These models are supported by biochemical analyses using point variants and suggest that maturation of the 18S 3' end is regulated by dissociation of the AF Dim1 from the subunit interface, consistent with previous biochemical analyses.

Keywords: cryo-EM; pre-40S ribosome; ribosome assembly; ribosome biogenesis; small subunit assembly.

MeSH terms

Amino Acid Motifs
Binding Sites
Cloning, Molecular
Cryoelectron Microscopy
Crystallography, X-Ray
Escherichia coli / genetics
Escherichia coli / metabolism
Gene Expression
Genetic Heterogeneity*
Humans
Models, Molecular
Nuclear Proteins / chemistry
Nuclear Proteins / genetics
Nuclear Proteins / metabolism
Protein Binding
Protein Interaction Domains and Motifs
Protein Structure, Secondary
RNA, Ribosomal / chemistry
RNA, Ribosomal / genetics
RNA, Ribosomal / metabolism
Recombinant Proteins / chemistry
Recombinant Proteins / genetics
Recombinant Proteins / metabolism
Ribonucleoprotein, U5 Small Nuclear / chemistry
Ribonucleoprotein, U5 Small Nuclear / genetics
Ribonucleoprotein, U5 Small Nuclear / metabolism
Ribosomal Proteins / chemistry*
Ribosomal Proteins / genetics
Ribosomal Proteins / metabolism
Ribosome Subunits, Small, Eukaryotic / metabolism
Ribosome Subunits, Small, Eukaryotic / ultrastructure*
Saccharomyces cerevisiae / metabolism
Saccharomyces cerevisiae / ultrastructure*
Saccharomyces cerevisiae Proteins / chemistry
Saccharomyces cerevisiae Proteins / genetics
Saccharomyces cerevisiae Proteins / metabolism
Thermodynamics

Substances

NOB1 protein, S cerevisiae
Nuclear Proteins
PNO1 protein, S cerevisiae
RNA, Ribosomal
RPS3 protein, human
Recombinant Proteins
Ribonucleoprotein, U5 Small Nuclear
Ribosomal Proteins
Saccharomyces cerevisiae Proteins
TXNL4A protein, human
Tsr1 protein, human

Abstract

MeSH terms

Substances

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