High-resolution structures of the actomyosin-V complex in three nucleotide states provide insights into the force generation mechanism

  1. Sabrina Pospich
  2. H Lee Sweeney
  3. Anne Houdusse
  4. Stefan Raunser  Is a corresponding author
  1. Max Planck Institute of Molecular Physiology, Germany
  2. University of Florida, United States
  3. Institut Curie, Centre National de la Recherche Scientifique, France

Abstract

The molecular motor myosin undergoes a series of major structural transitions during its force-producing motor cycle. The underlying mechanism and its coupling to ATP hydrolysis and actin binding is only partially understood, mostly due to sparse structural data on actin-bound states of myosin. Here, we report 26 high-resolution cryo-EM structures of the actomyosin-V complex in the strong-ADP, rigor, and a previously unseen post-rigor transition state that binds the ATP analog AppNHp. The structures reveal a high flexibility of myosin in each state and provide valuable insights into the structural transitions of myosin-V upon ADP release and binding of AppNHp, as well as the actomyosin interface. In addition, they show how myosin is able to specifically alter the structure of F-actin.

Data availability

The atomic models and cryo-EM maps are available in the PDB (Burley et al., 2018) and EMDB databases (Lawson et al., 2011), under following accession numbers: aged PHD-stabilized actomyosin-V in the strong-ADP state: 7PM5, EMD-13521 (central 1er), 7PM6, EMD-13522 (central 3er/2er), 7PM7, EMD-13523 (class 2), 7PM8, EMD-13524 (class 3), 7PM9, EMD-13525 (class 4), 7PMA, EMD-13526 (class 5), 7PMB, EMD-13527 (class 6), 7PMC, EMD-13528 (class 7) ; aged PHD-stabilized actomyosin-V in the rigor state: 7PLT, EMD-13501 (central 1er), 7PLU, EMD-13502 (central 3er/2er), 7PLV, EMD-13503 (class 1), 7PLW, EMD-13504 (class 3) and 7PLX, EMD-13505 (class 4); aged PHD-stabilized actomyosin-V in the PRT state: 7PMD, EMD-13529 (central 1er), 7PME, EMD-13530 (central 3er/2er), 7PMF, EMD-13531 (class 1), 7PMG, EMD-13532 (class 3), 7PMH, EMD-13533 (class 4), 7PMI, EMD-13535 (class 5), 7PMJ, EMD-13536 (class 6), 7PML, EMD-13538 (class 8); young JASP-stabilized actomyosin-V in the rigor state: 7PLY, EMD-13506 (central 1er), 7PLZ, EMD-13507 (central 3er/2er), 7PM0, EMD-13508 (class 1), 7PM1, EMD-13509 (class 2), 7PM2, EMD-13510 (class 4); and young JASP-stabilized F-actin: 7PM3, EMD-13511. The datasets generated during the current study are available from the corresponding author upon reasonable request.

The following data sets were generated

Article and author information

Author details

  1. Sabrina Pospich

    Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5119-3039
  2. H Lee Sweeney

    Department of Pharmacology and Therapeutics and the Myology Institute, University of Florida, Gainesville, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6290-8853
  3. Anne Houdusse

    Structural Motility, Institut Curie, Centre National de la Recherche Scientifique, Paris, France
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8566-0336
  4. Stefan Raunser

    Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    For correspondence
    stefan.raunser@mpi-dortmund.mpg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9373-3016

Funding

Max-Planck-Gesellschaft

  • Stefan Raunser

European Commission (ERC-2019-SyG,856118)

  • Stefan Raunser

Agence Nationale de la Recherche (ANR-17-CE11-0029-01)

  • Anne Houdusse

National Institutes of Health (R01-DC009100)

  • H Lee Sweeney

Centre National de la Recherche Scientifique

  • Anne Houdusse

Studienstiftung des Deutschen Volkes

  • Sabrina Pospich

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

Reviewing Editor

  1. Samara L Reck-Peterson, University of California, San Diego, United States

Publication history

  1. Preprint posted: September 7, 2021 (view preprint)
  2. Received: September 8, 2021
  3. Accepted: November 22, 2021
  4. Accepted Manuscript published: November 23, 2021 (version 1)
  5. Version of Record published: January 5, 2022 (version 2)

Copyright

© 2021, Pospich et al.

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

  • 1,271
    Page views
  • 247
    Downloads
  • 4
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

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. Sabrina Pospich
  2. H Lee Sweeney
  3. Anne Houdusse
  4. Stefan Raunser
(2021)
High-resolution structures of the actomyosin-V complex in three nucleotide states provide insights into the force generation mechanism
eLife 10:e73724.
https://doi.org/10.7554/eLife.73724

Further reading

    1. Structural Biology and Molecular Biophysics
    Xavier Leray et al.
    Research Article Updated

    The acidic luminal pH of lysosomes, maintained within a narrow range, is essential for proper degrative function of the organelle and is generated by the action of a V-type H+ ATPase, but other pathways for ion movement are required to dissipate the voltage generated by this process. ClC-7, a Cl-/H+ antiporter responsible for lysosomal Cl- permeability, is a candidate to contribute to the acidification process as part of this ‘counterion pathway’ The signaling lipid PI(3,5)P2 modulates lysosomal dynamics, including by regulating lysosomal ion channels, raising the possibility that it could contribute to lysosomal pH regulation. Here, we demonstrate that depleting PI(3,5)P2 by inhibiting the kinase PIKfyve causes lysosomal hyperacidification, primarily via an effect on ClC-7. We further show that PI(3,5)P2 directly inhibits ClC-7 transport and that this inhibition is eliminated in a disease-causing gain-of-function ClC-7 mutation. Together, these observations suggest an intimate role for ClC-7 in lysosomal pH regulation.

    1. Structural Biology and Molecular Biophysics
    Josep Rizo et al.
    Research Article Updated

    Synaptic vesicles are primed into a state that is ready for fast neurotransmitter release upon Ca2+-binding to Synaptotagmin-1. This state likely includes trans-SNARE complexes between the vesicle and plasma membranes that are bound to Synaptotagmin-1 and complexins. However, the nature of this state and the steps leading to membrane fusion are unclear, in part because of the difficulty of studying this dynamic process experimentally. To shed light into these questions, we performed all-atom molecular dynamics simulations of systems containing trans-SNARE complexes between two flat bilayers or a vesicle and a flat bilayer with or without fragments of Synaptotagmin-1 and/or complexin-1. Our results need to be interpreted with caution because of the limited simulation times and the absence of key components, but suggest mechanistic features that may control release and help visualize potential states of the primed Synaptotagmin-1-SNARE-complexin-1 complex. The simulations suggest that SNAREs alone induce formation of extended membrane-membrane contact interfaces that may fuse slowly, and that the primed state contains macromolecular assemblies of trans-SNARE complexes bound to the Synaptotagmin-1 C2B domain and complexin-1 in a spring-loaded configuration that prevents premature membrane merger and formation of extended interfaces, but keeps the system ready for fast fusion upon Ca2+ influx.