1. Structural Biology and Molecular Biophysics

Ciliate mitoribosome illuminates evolutionary steps of mitochondrial translation

  1. Victor Tobiasson
  2. Alexey Amunts  Is a corresponding author
  1. Stockholm University, Sweden
https://doi.org/10.7554/eLife.59264
Share this article
Cite this article
  1. Victor Tobiasson
  2. Alexey Amunts
(2020)
Ciliate mitoribosome illuminates evolutionary steps of mitochondrial translation
eLife 9:e59264.
https://doi.org/10.7554/eLife.59264
  2. Download BibTeX
  3. Download .RIS

Abstract

To reveal steps in the evolution of translation, we identified ciliates as a model with high coding capacity of the mitochondrial genome and characterized its mitoribosomes by cryo-EM. It revealed a 94-protein and 4-rRNA assembly with an additional protein mass of ~700 kDa on the small subunit, while the large subunit lacks 5S rRNA. The structure shows that the small subunit head is constrained, tRNA binding sites are formed by mitochondria-specific protein elements, conserved protein bS1 is excluded, and bacterial RNA polymerase binding site is blocked. We provide evidence for intrinsic protein targeting system through visualization of mitochondria-specific mL105 by the exit tunnel that would facilitate recruitment of a nascent polypeptide. Functional protein uS3m is encoded by three complementary genes from the nucleus and mitochondrion, establishing a link between genetic drift and mitochondrial translation. Finally, we reannotated nine open reading frames in the mitochondrial genome that code for mitoribosomal proteins.

Data availability

The electron density maps have been deposited into EMDB, with accession codes EMD-11032 (monosome), EMD-11033 (LSU), EMD-11034 (SSU), EMD-11035 (CP), EMD-11036 (L7/L12 stalk), EMD-11037 (head), EMD-11038 (back protuberance). The model has been deposited in the PDB, with accession code 6Z1P.

The following data sets were generated

Article and author information

Author details

  1. Victor Tobiasson

    Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University, Solna, Sweden
    Competing interests
    The authors declare that no competing interests exist.

  2. Alexey Amunts

    Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University, Solna, Sweden
    For correspondence
    amunts@scilifelab.se
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5302-1740

Funding

Ragnar Söderbergs stiftelse (M44/16)

  • Alexey Amunts

Cancerfonden (2017/1041)

  • Alexey Amunts

H2020 European Research Council (ERC-2018-StG- 805230)

  • Alexey Amunts

Knut och Alice Wallenbergs Stiftelse (2018.0080)

  • Alexey Amunts

European Molecular Biology Organization (EMBO Young Investigator Program)

  • Alexey Amunts

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Cynthia Wolberger, Johns Hopkins University School of Medicine, United States

Version history

  1. Received: May 25, 2020
  2. Accepted: June 8, 2020
  3. Accepted Manuscript published: June 18, 2020 (version 1)
  4. Version of Record published: June 30, 2020 (version 2)

Copyright

© 2020, Tobiasson & Amunts

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 329
    downloads
  • 38
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Victor Tobiasson
  2. Alexey Amunts
(2020)
Ciliate mitoribosome illuminates evolutionary steps of mitochondrial translation
eLife 9:e59264.
https://doi.org/10.7554/eLife.59264

Share this article

https://doi.org/10.7554/eLife.59264

Categories and tags

Further reading

    1. Structural Biology and Molecular Biophysics

    The mechanism of mammalian proton-coupled peptide transporters

    Simon M Lichtinger, Joanne L Parker ... Philip C Biggin
    Research Article

    Proton-coupled oligopeptide transporters (POTs) are of great pharmaceutical interest owing to their promiscuous substrate binding site that has been linked to improved oral bioavailability of several classes of drugs. Members of the POT family are conserved across all phylogenetic kingdoms and function by coupling peptide uptake to the proton electrochemical gradient. Cryo-EM structures and alphafold models have recently provided new insights into different conformational states of two mammalian POTs, SLC15A1, and SLC15A2. Nevertheless, these studies leave open important questions regarding the mechanism of proton and substrate coupling, while simultaneously providing a unique opportunity to investigate these processes using molecular dynamics (MD) simulations. Here, we employ extensive unbiased and enhanced-sampling MD to map out the full SLC15A2 conformational cycle and its thermodynamic driving forces. By computing conformational free energy landscapes in different protonation states and in the absence or presence of peptide substrate, we identify a likely sequence of intermediate protonation steps that drive inward-directed alternating access. These simulations identify key differences in the extracellular gate between mammalian and bacterial POTs, which we validate experimentally in cell-based transport assays. Our results from constant-PH MD and absolute binding free energy (ABFE) calculations also establish a mechanistic link between proton binding and peptide recognition, revealing key details underpining secondary active transport in POTs. This study provides a vital step forward in understanding proton-coupled peptide and drug transport in mammals and pave the way to integrate knowledge of solute carrier structural biology with enhanced drug design to target tissue and organ bioavailability.

    1. Structural Biology and Molecular Biophysics

    Plasticity of the proteasome-targeting signal Fat10 enhances substrate degradation

    Hitendra Negi, Aravind Ravichandran ... Ranabir Das
    Research Article

    The proteasome controls levels of most cellular proteins, and its activity is regulated under stress, quiescence, and inflammation. However, factors determining the proteasomal degradation rate remain poorly understood. Proteasome substrates are conjugated with small proteins (tags) like ubiquitin and Fat10 to target them to the proteasome. It is unclear if the structural plasticity of proteasome-targeting tags can influence substrate degradation. Fat10 is upregulated during inflammation, and its substrates undergo rapid proteasomal degradation. We report that the degradation rate of Fat10 substrates critically depends on the structural plasticity of Fat10. While the ubiquitin tag is recycled at the proteasome, Fat10 is degraded with the substrate. Our results suggest significantly lower thermodynamic stability and faster mechanical unfolding in Fat10 compared to ubiquitin. Long-range salt bridges are absent in the Fat10 structure, creating a plastic protein with partially unstructured regions suitable for proteasome engagement. Fat10 plasticity destabilizes substrates significantly and creates partially unstructured regions in the substrate to enhance degradation. NMR-relaxation-derived order parameters and temperature dependence of chemical shifts identify the Fat10-induced partially unstructured regions in the substrate, which correlated excellently to Fat10-substrate contacts, suggesting that the tag-substrate collision destabilizes the substrate. These results highlight a strong dependence of proteasomal degradation on the structural plasticity and thermodynamic properties of the proteasome-targeting tags.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics

    Special Issue: Allosteric regulation of kinase activity

    Amy H Andreotti, Volker Dötsch
    Editorial

    The articles in this special issue highlight how modern cellular, biochemical, biophysical and computational techniques are allowing deeper and more detailed studies of allosteric kinase regulation.