Cavβ-subunit displacement is a key step to induce the reluctant state of P/Q calcium channels by direct G protein regulation

• Published in: Proceedings of the National Academy of Sciences - PNAS 2004-04, Vol.101 (16), p.6267-6272
• Main Authors: Sandoz, Guillaume, Lopez-Gonzalez, Ignacio, Grunwald, Didier, Bichet, Delphine, Altafaj, Xavier, Weiss, Norbert, Ronjat, Michel, Dupuis, Alain, De Waard, Michel
• Format: Article
• Language: English
• Subjects: Biological Sciences
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0306804101

P/Q Ca 2+ channel activity is inhibited by G protein-coupled receptor activation. Channel inhibition requires a direct Gβγ binding onto the pore-forming subunit, Ca v 2.1. It is characterized by biophysical changes, including current amplitude reduction, activation kinetic slowing, and an I-V curve shift, which leads to a reluctant mode. Here, we have characterized the contribution of the auxiliary β 3 -subunit to channel regulation by G proteins. The shift in I-V to a P/Q reluctant mode is exclusively observed in the presence of β 3 . Along with the observation that Gβγ has no effect on the I-V curve of Ca v 2.1 alone, we propose that the reluctant mode promoted by Gβγ corresponds to a state in which the β 3 -subunit has been displaced from its channel-binding site. We validate this hypothesis with a β 3 -I-II 2.1 loop chimera construct. Gβγ binding onto the I-II 2.1 loop portion of the chimera releases the β 3 -binding domain and makes it available for binding onto the I-II loop of Ca v 1.2, a G protein-insensitive channel. This finding is extended to the full-length Ca v 2.1 channel by using fluorescence resonance energy transfer. Gβγ injection into Xenopus oocytes displaces a Cy3-labeled β 3 -subunit from a GFP-tagged Ca v 2.1 channel. We conclude that β-subunit dissociation from the channel complex constitutes a key step in P/Q calcium channel regulation by G proteins that underlies the reluctant state and is an important process for modulating neurotransmission through G protein-coupled receptors.