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

The neuronal calcium sensor NCS-1 regulates the phosphorylation state and activity of the Ga chaperone and GEF Ric-8A

  1. Daniel Muñoz-Reyes
  2. Levi J McClelland
  3. Sandra Arroyo-Urea
  4. Sonia Sánchez-Yepes
  5. Juan Sabín
  6. Sara Pérez-Suárez
  7. Margarita Menendez
  8. Alicia Mansilla
  9. Javier García-Nafría
  10. Stephen Sprang
  11. Maria Jose Sanchez-Barrena  Is a corresponding author
  1. Institute of Physical-Chemistry Blas Cabrera, Spain
  2. University of Montana, United States
  3. University of Zaragoza, Spain
  4. Hospital Universitario Ramón y Cajal, Spain
  5. Software 4 Science Developments, Spain
  6. Institute of Physical-Chemistry, Spain
  7. Institute of Physical Chemistry Blas Cabrera, Spain
https://doi.org/10.7554/eLife.86151
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Cite this article
  1. Daniel Muñoz-Reyes
  2. Levi J McClelland
  3. Sandra Arroyo-Urea
  4. Sonia Sánchez-Yepes
  5. Juan Sabín
  6. Sara Pérez-Suárez
  7. Margarita Menendez
  8. Alicia Mansilla
  9. Javier García-Nafría
  10. Stephen Sprang
  11. Maria Jose Sanchez-Barrena
(2023)
The neuronal calcium sensor NCS-1 regulates the phosphorylation state and activity of the Ga chaperone and GEF Ric-8A
eLife 12:e86151.
https://doi.org/10.7554/eLife.86151
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Abstract

The Neuronal Calcium Sensor 1, an EF-hand Ca2+ binding protein, and Ric-8A coregulate synapse number and probability of neurotransmitter release. Recently, the structures of Ric-8A bound to Ga have revealed how Ric-8A phosphorylation promotes Ga recognition and activity as a chaperone and guanine nucleotide exchange factor. However, the molecular mechanism by which NCS-1 regulates Ric-8A activity and its interaction with Ga subunits is not well understood. Given the interest in the NCS-1/Ric-8A complex as a therapeutic target in nervous system disorders, it is necessary to shed light on this molecular mechanism of action at atomic level. We have reconstituted NCS-1/Ric-8A complexes to conduct a multimodal approach and determine the sequence of Ca2+ signals and phosphorylation events that promote the interaction of Ric-8A with Ga. Our data show that the binding of NCS-1 and Ga to Ric-8A are mutually exclusive. Importantly, NCS-1 induces a structural rearrangement in Ric-8A that traps the protein in a conformational state that is inaccessible to Casein Kinase II-mediated phosphorylation, demonstrating one aspect of its negative regulation of Ric-8A-mediated G-protein signaling. Functional experiments indicate a loss of Ric-8A GEF activity towards Ga when complexed with NCS-1, and restoration of nucleotide exchange activity upon increasing Ca2+ concentration. Finally, the high-resolution crystallographic data reported here define the NCS-1/Ric-8A interface and will allow the development of therapeutic synapse function regulators with improved activity and selectivity.

Data availability

The atomic coordinates and structure factors have been deposited in the Protein Data Bank, https://www.pdb.org/. PDB with codes: Structure 1 (8ALH), Structure 2 (8AHY), Structure 3 (8ALM).All data generated or analysed during this study are included in the manuscript and supporting file; Source Data files have been provided for Figure 2, Figure 3, Figure 4, Figure 5

Article and author information

Author details

  1. Daniel Muñoz-Reyes

    Department of Crystallography and Structural Biology, Institute of Physical-Chemistry Blas Cabrera, Madrid, Spain
    Competing interests
    The authors declare that no competing interests exist.

  2. Levi J McClelland

    Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Sandra Arroyo-Urea

    Institute for Biocomputation and Physics of Complex Systems, University of Zaragoza, Zaragoza, Spain
    Competing interests
    The authors declare that no competing interests exist.

  4. Sonia Sánchez-Yepes

    Department of Neurobiology, Hospital Universitario Ramón y Cajal, Madrid, Spain
    Competing interests
    The authors declare that no competing interests exist.

  5. Juan Sabín

    AFFINImeter Scientific & Development, Software 4 Science Developments, Santiago de Compostela, Spain
    Competing interests
    The authors declare that no competing interests exist.

  6. Sara Pérez-Suárez

    Department of Crystallography and Structural Biology, Institute of Physical-Chemistry Blas Cabrera, Madrid, Spain
    Competing interests
    The authors declare that no competing interests exist.

  7. Margarita Menendez

    Department of Biological Physical-Chemisty, Institute of Physical-Chemistry, Madrid, Spain
    Competing interests
    The authors declare that no competing interests exist.

  8. Alicia Mansilla

    Department of Neurobiology, Hospital Universitario Ramón y Cajal, Madrid, Spain
    Competing interests
    The authors declare that no competing interests exist.

  9. Javier García-Nafría

    Institute for Biocomputation and Physics of Complex Systems, University of Zaragoza, Zaragoza, Spain
    Competing interests
    The authors declare that no competing interests exist.

  10. Stephen Sprang

    Center for Biomolecular Structure and Dynamics, University of Montana, Missoula, United States
    Competing interests
    The authors declare that no competing interests exist.

  11. Maria Jose Sanchez-Barrena

    Department of Crystallography and Structural Biology, Institute of Physical Chemistry Blas Cabrera, Madrid, Spain
    For correspondence
    xmjose@iqfr.csic.es
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5986-1804

Funding

Spanish National Plan for Scientific and Technical Research and Innovation (PID2019-111737RB-I00)

  • Maria Jose Sanchez-Barrena

Spanish National Plan for Scientific and Technical Research and Innovation (PID2019-106608RB-I00)

  • Alicia Mansilla

Spanish National Plan for Scientific and Technical Research and Innovation (PDC2022-133775-I00)

  • Alicia Mansilla

Spanish National Plan for Scientific and Technical Research and Innovation (RTI2018-099985-B-I00)

  • Margarita Menendez

Spanish National Plan for Scientific and Technical Research and Innovation (PID2020-113359GA-I00)

  • Javier García-Nafría

Spanish National Plan for Scientific and Technical Research and Innovation (RYC-2017-22392)

  • Alicia Mansilla

Spanish National Plan for Scientific and Technical Research and Innovation (RYC2018-025731-I)

  • Javier García-Nafría

National Institutes of Health (P30GM140963)

  • Stephen Sprang

National Institutes of Health (R01GM105993)

  • Stephen Sprang

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

Reviewing Editor

  1. Randy B Stockbridge, University of Michigan, United States

Version history

  1. Received: January 12, 2023
  2. Accepted: November 24, 2023
  3. Accepted Manuscript published: November 29, 2023 (version 1)

Copyright

© 2023, Muñoz-Reyes 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.

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  1. Daniel Muñoz-Reyes
  2. Levi J McClelland
  3. Sandra Arroyo-Urea
  4. Sonia Sánchez-Yepes
  5. Juan Sabín
  6. Sara Pérez-Suárez
  7. Margarita Menendez
  8. Alicia Mansilla
  9. Javier García-Nafría
  10. Stephen Sprang
  11. Maria Jose Sanchez-Barrena
(2023)
The neuronal calcium sensor NCS-1 regulates the phosphorylation state and activity of the Ga chaperone and GEF Ric-8A
eLife 12:e86151.
https://doi.org/10.7554/eLife.86151

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