The neuronal calcium sensor NCS-1 regulates the phosphorylation state and activity of the Ga chaperone and GEF Ric-8A
- Institute of Physical-Chemistry Blas Cabrera, Spain
- University of Montana, United States
- University of Zaragoza, Spain
- Hospital Universitario Ramón y Cajal, Spain
- Software 4 Science Developments, Spain
- Institute of Physical-Chemistry, Spain
- Institute of Physical Chemistry Blas Cabrera, Spain
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
Maria Jose Sanchez-Barrena
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.
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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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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