Journal Article

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

Muñoz-Reyes, Daniel, Arroyo-Urea, Sandra, Mansilla, Alicia, Sprang, Stephen, McClelland, Levi J, Menendez, Margarita, Sánchez-Yepes, Sonia, García-Nafría, Javier, Sabín, Juan, Sanchez-Barrena, Maria Jose, Pérez-Suárez, Sara

2023

Overview

The neuronal calcium sensor 1 (NCS-1), an EF-hand Ca 2+ binding protein, and Ric-8A coregulate synapse number and probability of neurotransmitter release. Recently, the structures of Ric-8A bound to Gα have revealed how Ric-8A phosphorylation promotes Gα 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 Gα 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 Ca 2+ signals and phosphorylation events that promote the interaction of Ric-8A with Gα. Our data show that the binding of NCS-1 and Gα 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 guanine nucleotide exchange factor (GEF) activity toward Gα when complexed with NCS-1, and restoration of nucleotide exchange activity upon increasing Ca 2+ 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.

• Publisher: eLife Sciences Publications Ltd,eLife Sciences Publications, Ltd
• Subject: Binding sites / Biochemistry and Chemical Biology / Calcium - metabolism / calcium signaling / Calcium signalling / Calcium-binding protein / Casein / Casein kinase II / Cell division / EF-hand / Freq protein / guanine nucleotide exchange / Guanine nucleotide exchange factor / Guanine Nucleotide Exchange Factors - metabolism / Insects / Kinases / molecular chaperone / Molecular Chaperones - metabolism / Molecular modelling / Nervous system / Neurotransmitter release / Phosphorylation / protein-protein interaction / Proteins / Sensors / Signal Transduction / Structural Biology and Molecular Biophysics / synapse function regulation / Synapses / Therapeutic targets